Ion Chromatography (Journal of Chromatography Library)

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JOURNAL OF CHROMATOGRAPHY LIBRARY - volume 46

ion chromatography principles and applications

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JOURNAL OF CHROMATOGRAPHY LIBRARY- volume 46

ion chromatography principles and applications

Paul R. Haddad and Peter E. Jackson Department of Analytical Chemistry, University of New South Wales, P.O. Box 7, Kensington, N.S. W. 2033, Australia

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ELSEVIER SCIENCE B.V. Sara Burgcrliartstraat25 P.O.Box 2 I I, loOn AE Arnstcrdam, Thc Ncthcrlands

C 1990 Elscvicr Scicncc B.V. All rights racwcd. This work is protcctcd undcr copyright by Elscvicr Scicncc. and thc following t c m and conditions apply to its USC: Photocopying Single photocopies of single chapters may be made for personal use ns allowed by national copyright laws. Pcrmission of thc Publishcr and paynmt of a fcc is quircd for all othcr photocopying. including multiplc or systcmaticcopying, copying for advertising or promotional pwposcs. rcsalc. and all f i s ofdocumcnt clclivcry. Special tatcs arc available for educational institutions that wish to make pliutocopiw for lion-profit cducational classroom use. Pcrmissions may bc sought dircctly fiorn Elscvicr's Scicncc & Technology RighLs Dcparhncnt in Oxford, UK: phone:

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Clt8loqing-in-Publication Data

Hadda.3, Paul P. I m chroma:,yraphy : p r i n c i p l c o and applications I Paul R. Haddad and Filter E. J a c k s o n . p. crn. -- IJc-urnal o f chromarography l i b r a r y ; v . 4 6 ) 111 .lucfes t.ibliuqx+~.hic.dl r e t e r e n c e r . I r i c l u 3 c u iridex. Tfl?}! 0-444-8873?-6 1 . Ion nxchangc chr.mnatcqraphy. I. J a c k s o n , P c t c r E. 11. Title. 111. S e r i e s . 91873. :4531133 133'2 90- 3 60 0 1

543' .OEg3--,jc:o

CIP

First cditioti I990 Second imprcssion 1994

Third imprcssion 2003 ISBN:

0444-8x2324

@. The p a p - uscd in this publication mccts thc rcquircmcnts of ANSlMlSO 239.48-1992 (Pcrmancncc of P a p ) . Printed in TIic Netherlands.

To Simone, Lianne, Christopher and, most of all, to Kerry


vii

Preface Ion chromatography (IC) was first introduced in 1975 and since that time, the technique has grown in usage at a phenomenal rate. The reasons for this are clear; IC offers the only simple, reliable and inexpensive means for the simultaneous separation and determination of inorganic (and organic) ions in complex mixtures. The growth of IC has been accompanied by a blumng of the original definition of the technique, so that it now embraces a very wide range of separation and detection methods, many of which bear little resemblance to the initial concept of ion-exchange separation coupled with conductivity detection. We have chosen to define IC to encompass those modem column liquid chromatographic methods which can be used for the determination of inorganic anions and cations, water-soluble organic acids and bases, and ionic organometallic compounds. Using this definition, the separation methods which are applicable to IC include ion-exchange chromatography, ion-interaction chromatography, ion-exclusion chromatography, and some further, miscellaneous techniques. Appropriate detection methods include conductivity, amperometry, coulometry, polarography, potentiometry, spectrophotometry, atomic spectroscopy, refractive index measurements, luminescence techniques and post-column reactions. This text is structured into five logical parts, each of which groups chapters on related topics. Accordingly, Part I (Chapters 2-5) discusses ion-exchange separation methods, whilst Part I1 (Chapters 6-8) treats the remainder of the separation methods listed above. Part 111 (Chapters 9-13) is devoted solely to detection methods, and Part IV (Chapters 14,15) concerns some practical aspects of IC. Part V (Chapters 16-22) comprises a detailed, structured listing of experimental conditions for the numerous applications of IC. The applications section is a unique feature of this book and we hope it will become a valuable reference source for practitioners of the technique. Throughout the book, we have attempted to integrate suppressed and non-suppressed IC methods in the discussion of each aspect: that is, we have emphasized the similarities between these two approaches, rather than their differences. In writing this book, we have endeavoured to provide a m l y comprehensive text which treats the principles and applications of IC and is suitable as a reference work for both researchers and those involved with the use of IC in practical situations. Every effort has been made to ensure that the literature coverage is as complete as possible for the period 1975 to the early part of 1989, but we have also included some references which are more recent than this. Throughout the text, each topic is discussed both from a descriptive standpoint, in order to illustrate the underlying principles, and also from a theoretical standpoint, in which a full mathematical treatment is provided. Theoretical aspects, such as retention models and detector response equations, are presented either as separate chapters or as clearly identified Sections. In this way, the theory can be examined in depth or bypassed, according to the interests of the reader. The text has been amplified by liberal usage of chromatograms and Tables showing practical examples of each of the principles of IC, as they are discussed. We hope that this

viii

approach assists in the understanding of each point. Schematic overviews are also used extensively so that the organization of each topic is clear. Abbreviations and acronyms have been kept to a minimum and those that have been used are collected for ease of reference in Appendix R, together with all of the symbols employed in the book. Special attention has been given to the compilation of a comprehensive, detailed and crossreferenced Index. Writing a book of this size is an arduous task which cannot be accomplished by the authors alone. We are greatly indebted to our many friends and colleagues who provided assistance along the way and we would like to thank them sincerely for their time and effort. Martin Dudman used his impeccable drafting skills to produce all of the chromatograms and Helen Shumsky performed all of the photographic work required for preparation of the Figurcs. The staff of the Physical Sciences Library of the University of New South Wales, and of the Technical Library at Millipore Waters, cheerfully procured hundreds of journal articles. Only through their capable efforts have we been able to achieve a comprehensive coverage of the literature. Numerous colleagues, both within Australia and overseas, provided valuable and enlightened comments on the manuscript, or helped with proof reading. In this regard, thanks are due especially to Jim Fritz and Petr Jandik, and to the following students in the IC research group at the University of New South Wales; Roy Foley, Chris Cowie, John Brayan, Andrew Sosirnenko and Peter Fagan. We gratefully acknowledge those publishers and authors who have granted permission for us to reproduce Figures from their publications. Finally, this book would never have been completed without the support, encouragement and love of the members of the fladdad family; Simone made the coffee, Lianne filed the reprints, Christopher shared his bedroom with a computer and Kerry's patience and acceptance kept everything and everyone together over two years with a part-time husband and father. Paul R. Haddnd, Peter E. Jackson Kensington, I990

Technical nofe: Text and artwork (with the exception of chromatograms) for this book were produced on an Apple Macintosh@ IIcx microcomputer, using Microsoft@ Word 4.0 (Microsoft Corporation), McDraw (Apple@ Computer Inc.), Cricket Graph (Cricket Software) and MathWriteP (Cooke Publications) software packages, and was printed in camera-ready form on an Apple Laserwrite@.

ix

Short Contents Chapter I PART I

Introduction

1

ION-EXCHANGE SEPARATION METHODS

An Introduction to Ion-Exchange Methods Ion-Exchange Stationary Phases for Ion Chromatography Eluents for Ion-Exchange Separations Retention Models for Ion-Exchange

Chapter 2 Chapter 3 Chapter 4 Chapter 5 PART 11

15 29 79 133

ION-INTERACTION, ION-EXCLUSION AND MISCELLANEOUS SEPARATION METHODS

Chapter 6 Chapter 7 Chapter 8

Ion-Interaction Chromatography Ion-Exclusion Chromatography Miscellaneous Separation Methods

165 195 223

PART III DETECTION METHODS

Chapter 9 Chapter 10 Chapter I1 Chapter 12 Chapter 13 PART IV

Chapter 14 Chapter 15

Conductivity Detection Electrochemical Detection (Amperometry, Voltammetry and Coulometry) Potentiometric Detection Spectroscopic Detection Methods Detection by Post-Column Reaction

245 29 1 323 343 387

PRACTICAL ASPECTS

Sample Handling in Ion Chromatography Methods Development

409 463

PART V APPLICATIONS OF ION CHROMATOGRAPHY

Overview of the Applications Section Chapter 16- Environmental Applications Chapter I7 Industrial Applications , Chapter 18 Analysis of Foods and Plants Chapter 19 Clinical and Pharmaceutical Applications Chapter 20 Analysis of Metals and Metallurgical Solutions Chapter 21 Analysis of Tkated Waters Chapter 22 Miscellaneous Applications

487 489 543 593 633 667 695 717

Appendix A Statistical Information on Ion Chromatography Publications Appendix B Abbreviations and Symbols

735 745

Index

751


xi

Full Contents Chapter 1

Introduction 1.1 BACKGROUND TO ION CHROMATOGRAPHY

Hardware and software components The chromatographic separation process Thechromatogram Mechanisms of interaction 1.2 WHAT IS ION CHROMATOGRAPHY? 1.2.1 Historical aspects 1.1.1 1.1.2 1.1.3 1.1.4

1.2.2 Definition 1.3 ORGANIZATION OF THIS BOOK 1.4 SOME FUNDAMENTAL CHROMATOGRAPHIC CONCEPTS 1.4.1 Chromatographic efficiency 1.4.2 Factors affecting band broadening Eddy diffusion, 10. Longitudinal diffusion, 10. Resistance to mass transfer, 10. Extra-column band broadening, 11. 1.5 REFERENCES

PART I

1 1 2 2 4 5 6 6 6 7 8 8 10

12

ION-EXCHANGE SEPARATION METHODS

Chapter 2 An Introduction to Ion-Exchange Methods 2.1 INTRODUCTION TO ION-EXCHANGE

15 15 15 17

h c i p l e s of ion-exchange Configuration of an ion-exchange chromatographic system Open-column(chsical)ion-exchange,17. Modern ionexchange chromatography,18. Eluent characterirtics, 19. 2.1.3 Types of ion-exchange materials 2.1.4 Characteristics of ion-exchangers Ion-exchange capacity, 21. Swelling characterirtics,22. Ion-exchange selectivity,22. 2.2 CLASSIFICATION OF ION CHROMATOGRAPHIC METHODS EMPLOYING ION-EXCHANGE SEPARATION 2.2.1 Non-suppressed ion chromatography 2.2.2 Suppressed ion chromatography 2.2.3 Similarities between non-suppressed and suppressed methods 2.3 REFERENCES

24 24 25 26 27

Chapter 3 Zon-Exchange Stationary Phases for Zon Chromatography

29

3.1 INTRODUCTION 3.2 SILICA-BASED ION-EXCHANGE MATERIALS 3.2.1 Types of silica-based ion-exchangers

29 29 29

2.1.1 2.1.2

20 21

'

Contents

xii

3.3

3.4

3.5

3.6

3.7

3.2.2 Functionalized silica ionexchangers 3.2.3 Polymer-coated silica ionexchangers 3.2.4 Advantages and limitationsof silica-basedTC packings Chromatographic mciency, 33. pH limitariom, 35. Retention of metal ions on silica anion-exchangers,36. Sample size, 37. RESIN-BASED ION-EXCHANGERS 3.3.1 Polymerization reactions 3.3.2 Surface-functi0naliz.dcation-exchange resins Synthesis, 40. Characteristics,42. 3.3.3 Surface-funcaonalizedanion-exchangeresins Synthesis, 45. Characteristics,47. Effect of functional group, 48. Nature of the polymeric substrate,52. 3.3.4 Advantages and disadvantagesof resin-based ion-exchangers pH tolerance,53. Chromatographic efficiency,53. Pressure limitations,55. Operating conditions,55. AGGLOMERATED ION-EXCHANGE RESINS 3.4.1 Description 3.4.2 Synthesis Electrostatic binding, 57. Hydrophobic binding, 59. Mechanical binding, 60. 3.4.3 Characteristics of agglomerated ion-cxchangers Chromatographicefficiency, 60. Stability, 63. Selectivity, 63. HYDROUS OXIDE ION-EXCHANGERS 3.5.1 Introduction 3.5.2 Silica 3.5.3 Alumina Ion-exchange characteristics,67. Effect of pH, 69. Selectivity,69. SIMULTANEOUS SEPARATION OF ANIONS AND CATIONS 3.6.1 Tandem anion- and cation- exchange columns 3.6.2 Mixed-bed ion-exchangers 3.6.3 Bi-functional ionexchange materials REFERENCES

Chapfer 4

Eluents for Ion-Exchange Separations

4.1 INTRODUCTJON 4.2 ELUENT CHARACI'ERISTICS 4.2.1 Compatibility with the detection mode 4.2.2 Nature and concentration of the competing ion 4.2.3 Effect of eluent pH 4.2.4 Complexation characteristics 4.2.5 Use of organic solvents in ion-exchange separdtions 4.3 ELUENTS FOR NON-SUPPRESSED ION CHROMATOGRAPHY 4.3.1 Eluents for anion separation in non-suppressed IC Aromatic carhxylic acids and their salts, 84. Aliphatic carboxylic acids, 88. Aromatic and aliphatic sulfonic acids, 88. Potarsiwn hydroxide, 90. Polyol-borate complexes, 91. ErhyIenediaminetetraacetic acid, 94. Inorganic eluents, 95. 4.3.2 Eluents for cation separation in non-suppressed 1C

30 31 33

37 38

40 45

53

56 56 57

60 66 66 67 67 70 71 72 73 74 79

19 79 79 80 81 83 83 84 84

98

Contents

4.4

4.5

4.6

4.7

xiii

Inorganic acidr, 98. Organic bases, 98. lnorganic eluents, 99. Complexing eluents, 101. ELUENTS FOR SUPPRESSED ION CHROMATOGRAPHY 4.4.1 Eluent requirements for use with suppressors 4.4.2 Eluents for anion separations in suppressed IC 4.4.3 Eluents for cation separations in suppressed IC GRADIENT ELUTION IN ION-EXCHANGE SEPARATIONS 4.5.1 Principles of gradient elution 4.5.2 Gradient elution using detection methods other than conductivity 4.5.3 Gradient elution with conductivity detection High capacity suppressors, 115. Isoconductivegradients, 119. Baseline balancing methodr, 121. EXTRANEOUS (SYSTEM)PEAKS IN ION-EXCHANGE IC 4.6.1 Introduction 4.6.2 Extraneous peaks in non-suppressed IC The injection peak, 124. The system peak, 124. 4.6.3 Extraneous peaks in suppressed IC REFERENCES

106 106 107 109 114 114 115 115

123 123 123 126 128

Chapter 5 Retention Models f o r Ion-Exchange

133

5.1 INTRODUCTlON 5.2 RETENTION MODELS FOR ANION-EXCHANGE 5.2.1 Single eluent competing anion Effects of KAE, Q and w/Vm,137. Effect of charges on solute and competing anions, 138. Effect of [EL], 139. 5.2.2 Multiple eluent competing anions Dominant equilibrium approach, 140. Competing anion "effective charge" approach, 140. Dual eluent species approach, 142. 5.2.3 Gradient elution in anion-exchange IC 5.3 RETENTION MODELS FOR CATION-EXCHANGE 5.3.1 Single eluent competing cation 5.3.2 Complexing eluents Separation by cation-exchange, 156. Separation by anionexchange, 160. 5.4 REFERENCES

133 133 133

PART II

Chapter 6

140

149 152 152 156

161

ION-INTERACTION, ION-EXCLUSION AND MISCELLANEOUS SEPARATION METHODS Ion-Interaction Chromatography

6.1 INTRODUCIlON 6.2 MECHANISM 6.2.1 Trends in solute retention in ion-interaction chromatography 6.2.2 The ion-pair model 6.2.3 The dynamic ion-exchangemodel 6.2.4 The ion-interaction model

165 165 166 166 166 168 168

Contents

xiv

6.2.5 Evaluation of mechanistic models in retention of inorganic ions 6.3 STATIONARY PHASES AND ELUENTS 6.3.1 Stationary phases 6.3.2 Type of ion-interaction reagent Rquirements of the IIR, 173. " D y m ' ccoating" ion-interaclhn chromatography, I73. "Permanentcwn'ng" ion-interaction chromorography,I75. 6.3.3 Role of the countdon of the IIR 6.3.4 S u m of eluent and stationary phasc effects 6.3.5 Guidelines for eluent selection in ion-inmactiOn chromatography Anion separations, 181. Catwn separations, 181. 6.4 RETENTION MODELS FOR DYNAMIC COATING ION-INTERACTION

CHROMATOGRAPHY 6.4.1 Model for anion retention Effect of [A-j m and p-j,,,,183. Effect of the concentration of IIR in the eluent, 184. 6.4.2 Model for cation retention

169 171 171 173

177 179 181 182 182

6.5 APPLICATIONS 6.6 REFERENCES

186 188 190

Chapter 7

195

Zon-Exclusion Chromatography

7.1 INTRODUCllON 7.1.I Basic principles 7.2 STATIONARY PHASES 7.3 ELUENTS 7.3.1 Waterelwnts 7.3.2 Acideluents 7.3.3 Complexing eluents 7.4 FACTORS INFLUENCING RETENTION IN ION-EXCLUSION CHROMA TOG^ 7.4.1 Degree of ionization of the solute Dependence of solute retention on pK,, 203. Dependence of solute retention on eluent pH, 205. 7.4.2 Molecular size of the solute 7.4.3 Hydrophobic interactions between the solute and stationary phase Use of organic modfiers in the eluent, 207. 7.4.4 Ion-exchange capacity of the stationary phase 7.4.5 Ionic form of the ion-exchange resin 7.4.6 Temperature

7.4.7 summary 7.5 RETENTION MODEL FOR ION-EXCLUSION CHROhlATOGRAPHY 7.6 APPLICATIONS OF ION-EXCLUSION CHROMATo<jRAPHY 7.6.1 Carboxylic acids 7.6.2 Weak inorganic acids and bases 7.6.3 Water 7.7 REFERENCES

195 195 197 199 199 199 202 202 202

205 206

209 210 211 21 1 212 215 215 216 218 220

Contents

xv

Chapter 8 Miscellaneous Separation Methods

223

8.1 INTRODUCI'ION 8.2 REVERSED-PHASE LIQUID CHROMATOGRAPHY 8.2.1 Coordination cornpounds Dithiocarbmate complexes, 225. Formation of the chelate,225. 8.2.2 Organometallicmpounds 8.2.3 Carboxylic acids (Ion-suppression) 8.3 CHELATING STATIONARY PHASES 8.3.1 Chemically-boundligands 8.3.2 Crown ether stationary phases Synrhesis of stationaryphases. 233. Chromatographicproperties, 235. Applications,236. Gradient elruion on macrocyclic stationary phares, 237. 8.4 MICELLE EXCLUSION CHROMATOGRAPHY 8.4.1 Introduction 8.4.2 Micelle exclusion chmatography of anions 8.4.3 Micelle exclusion chromatographyof cations 8.5 REFERENCES

223 224 224

PART III

DETECTION METHODS

Chapter 9

Conductivity Detection

9.1 INTRODUCTION 9.2 PRINCIPLES OF CONDUCl'IVITY DETJZCI'ION 9.2.1 Nature of electrical conductivity of electrolyte solutions 9.2.2 Factors influencing limitingequivalent ionic conductance 9.2.3 Theory of conductivity response 9.3 MODES OF CONDUCI'MTYDETECTION 9.3.1 Direct and indirectconductivity detection Simultaneour direct and indirect conductiviry detection, 254. 9.3.2 Magnitude of conductance change on sample elution 9.4 ELECTRONIC CIRCUITRY AND CELL DESIGN FOR CONDUCTIVITY DETECTION 9.4.1 Introduction 9.4.2 AC conductance bridge 9.4.3 Bipolar-pulse circuitry 9.4.4 Four-electrode conductance measurement 9.4.5 Differentialconductivity detection 9.5 SUPPRESSORS IN IC 9.5.1 Function of the suppressor 9.5.2 Packedcolurhn suppressors 9.5.3 Hollow-fibre membrane suppressors Operating principle,264. Regeneranu,266. Packed-fibre suppressors, 266. 9.5.4 Micromembrane suppressor Design of the micromembrane suppressor,268. aperating principles, 269. Advantagesof the micromembrane suppressor,269.

228 230 232 232 233

237 237 238 240 240

245 245 245 245 249 249 252 252

254 255 255 256 258 258 260 261 261 262 264

267

Contents

xvi 9.5.5 Post-suppressors Design of post-suppressors,270. Advantages of post-suppression,271. 9.5.6 Other post-column signal enhancing devices Electrochemical suppressor, 271. Signal enhancement devices for ion-exclusionchromatogrqhy, 273. 9.5.7 Response equation for suppressed conductivity detection 9.5.8 Suppression based on precipitation or chelation reactions 9.6 PERFORMANCE CHARACTERISTICS OF CONDUCI'MTY DETECTORS 9.6.1 Non-linearity of calibration plots in suppressed IC Effect of hydrogen ions in the sample band, 277. Practical considerationof the hydrogen ion effect, 279. Effect of suppression mciency on linearity of calibration,280. 9.6.2 Temperature effects in conductivity detection 9.6.3 Conductivity detection without the use of standards 9.6.4 Utilization of the injection peak in conductivity detection 9.7 APPLICATIONS OF CONDUCTIVITY DETECTION IN IC 9.7.1 Anions 9.7.2 Cations 9.7.3 Weakly ionized species 9.8 REFERENCES

Chapter 10

Electrochemical Detection (Amperometry, Voltammetry and Coulometry)

10.1 INTRODUCIlON 10.1.1 Definitions Voltammetry andpolarography,291. Amperometry and coulometry, 291. 10.1.2 Interrelationships between voltammetry, amperometry, and coulometry Potential window, 292. Addition of an electroactivesolute, 293. Hydrodynamic voltammograms,294. Analysis of mixtures of electroactive species, 294. 10.I .3 Basic instrumentation for electrochemical detection in IC 10.1.4 Usage patterns for electrochemical detection in IC 10.2 MODES OF OPERATION OF ELECIROCHEMJCAL DJTECTORS 10.2.1 Direct and indirect detection Direcr elec~mhemicaidetection, 297. lndirect electrochemical detection,298. 10.2.2 Amperomemc detection with pulsed potential Problems with conventional DC amperometry,301. Pulsed amperomerric detection (PAD),302. Reverse-pulsemethods, 303. 10.3 ELECTRODES FOR AMPEROMETRIC DETECTION 10.3.1 Reference and auxiliary electrodes 10.3.2 Working electrode materials Mercury, 306. Carbon,306. Silver, platinum and gold, 307. Criteriafor selection of the working electrode material,308. 10.4 FLOW-CELL DESIGN AND RESPONSE EQUATIONS 10.4.1 Thin-layer cells 10.4.2 Row-through, highefficiency cells

270 27 1

275 276 277 277

28 1 282 283 285 285 285 285 288

29 1 29 1 29 1

292

294 295 297 297

30 1

304 304 305

309 310 311

Contents

xvii

10.4.3 Wall-jet cells 10.4.4 Polarographic cells 10.4.5 Multi-functional cells 10.4.6 Dual-electrode ampemmeuic cells 10.5 APPLICATIONS OF ELECTROCHEMICALDETECTION 10.6 REFERENCES

312 313 313 314 319 319

Chapter 11

Polentiornetrie Detection

11.1 INTRODUCTION 11.2 PRINCIPLES OF POTENTIOMETRIC DETECTION 11.2.1 General response equation 11.2.2 Instrumental considerations 11.3 INDICATOR ELECTRODES AND RESPONSE PROFILES 11.3.1 Ion-selective indicator electrodes pH electrodes, 326. Solid-state ion-selectiveelectrodes, 327. Liquid membrane electrodes, 327. 1 1.3.2 Coated-wire indicator electrodes Coating materials, 328. Calibration and response characteristics,.329. 1 1.3.3 Metallic copper indicator electrode Modes of operation of a metallic copper indicator electrode,331. 1 1.4 FLOW-CELLS FOR POTENTIOMETRIC DE"ION 1 1.4.1 Flow-cellsfor cylindrical ISEs 11.4.2 Other flow-cells 1 1.5 APPLICATIONS OF POTENTIOMETRIC DETECTION IN IC 11.5.1 Halides and pseudohalides 1 1.5.2 Weak acid anions 11.5.3 Cations 11S . 4 Applications of the metallic copper indicator electrode Direct detection,337. Indirect detection,338. 11.6 REFERENCES

323 323 324 324 324 325 325

328 329 333 333 334 336 336 337 337 337 34 1

Spectroscopic Detection Methods

343

12.1 INTRODUCTION 12.1.1 Types of spectroscopic detection in IC 12.1.2 Direct and indirect spectroscopic detection 12.2 UV-VISIBLESPECTROPHOTOMETRICDETECTION IN IC METHODS USING ION-EXCHANGE SEPARATIONS 12.2.1 Detection response equations for ion-exchange 12.2.2 Direct spectrophotomeaic detection in ion-exchange IC Direct detection of anions, 347. Direct detection of cations, 348. Replacement IC with spectrophotometric detection,350. 12.2.3 Indirect spectrophotometric detection in ionexchange IC Sensitivity of indirect spectrophotometric detection, 351. Indirect spectrophotometric detection of anions, 354. Indirect spectrophotometric detection of cations, 356. Simultaneour indirect detection of anions and cations,358. Indirect spectrophotometric detection without standards,360.

343 343 343

Chapter 12

344 344 347

35 1

xviii

Contents

12.3 SPECIROPHOTOMETRICDETECTION IN IC METHODS USING ION-INTERACTIONSEPARATIONS 12.3.1 Direct and indirect detection in ion-interactionchron%ttography Spectrophoromenic detection in permmently coated ioninteraction chromntogrophy, 361. Specwophoromenic detection in dynamically coated ion-interactionchromatography, 361. 12.3.2 W visualization detection in ion-interaction chromatography Mechanism of W visualization,363, Applications of W visualizanon in IC, 366. 12.4 SPEClXOPHOTOMETRICDETECTION IN IC METHODS USING ION-EXCLUSION SEPARATIONS 12.5 REFRACTIVEINDEX DEECTION 12.5.1 Direct refractive index detection 12.5.2 indirect refractive index detection 12.6 PHOTOLUMINESCENCE DETECTION IN IC 12.6.1 Direct fluorescenccdetection 12.6.2 Indirect fluorescence detection 12.6.3 Indirect phosphorescence detection 12.7 ATOMIC SPECTROSCOPICDETECTION IN 1C 12.7.1 Atomic absorption specaoscopy Discrete sampling, 376. Flow-rate matching, 376. Indirect detection using AAS, 377. 12.7.2 Atomic emission spectroscopy Plarma excitation,379. Replacement IC with spectroscopicdetection 379. 12.8 REFERENCES

360 360

363

369 369 369 370 37 1 37 1 372 374 376 376

378 380

Detection by Post-Column Reaction

387

13.1 INTRODUCTION 13.1.1 T p s of PCR systems Solution PCR. 387. Packed-bed PCR, 388. 13.2 HARDWARE FOR POST-COLUMNREACTTON 13.2.1 Pumps for PCR Effect ofpump pulsations, 389. Materialsfor pump construction, 3 PO. 13.2.2 Mixing chambers Tee-piece mixers, 391. Membrane reactor, 393. Comparison of dizerent mixing chambers,394. 13.2.3 Reactors 13.3 PCR DETECTION OF INORGANIC ANIONS 13.3.1 Phosphorus oxo-anions 13.3.2 Fe(CiO4)j as a postcolumn reagent 13.3.3 Other PCR methods for anions 13.4 PCR DETECTION OF INORGANIC CATIONS 13.4.1 Choice of the PCR reagent 13.4.2 PCR detection of transition mtals using PAR 13.4.3 PCR detection of lanthanides using Arsenazo dyes 13.4.4 PCR detection of other metal ions 13.5 PCR DETECTION OF ORGANIC SPECIES 13.6 REFERENCES

387 387

Chapter I 3

389 389 391

394 394 394 395 398 398 398 400 402 403 403 404

Contents

PART IV

xix

PRACTICAL ASPECTS

Chapter 14 Sample Handling in Ion Chromatography

409

14.1 INTRODUCTION 14.2 SAMPLE COLLECTION PROCEDURES 14.2.1 General 14.2.2 Sampling of gases, aerosols and particulates for IC analysis Filter media, 410. Impingers, 411. Solid aakorbents, 413. Diffusion denuders, 415. 14.3 EXTRACTION OF IONIC SPECIES FROM SAMPLES 14.3.1 Introduction 14.3.2 Simple extraction methods 14.3.3 Aciddigestion 14.3.4Alkali fusion 14.3.5 Combustion methods Ashing, 419. Schoenigerjlask combustion,419. Parr oxygen bomb combustion,420. Furnace combustion,421. Composition of absorbing solution, 421. 14.4 SAMPLE CLEANUP METHODS 14.4.1 Introduction 14.4.2 Sample filtration 14.4.3 Chemical modification of the sample using ioncxchange resins 14.4.4 Chemical modification of the sample using membranes Passive dialysis,425, Donnan dialysis,426. Electrochemical dialysis, 431. 14.4.5 Chemical modification of the sample with disposable cartridge columns Modes of operation, 432. Practical aspects,433. 14.4.6 Chemical modification of the sample by pre-columnreaction 14.5 CONTAMINATION EFFECI'S 14.5.1 Introduction 14.5.2 Contamination from physical handling of the sample 14.5.3 Contamination from filtration devices and cartridge columns 14.5.4 Contamination fmm chromatographic hardware components 14.5.5 Contamination of the column Contamination by organic species,440. Contamination by metal ions, 440. 14.6 SAMPLE HANTILING FOR ULTRA-TRACE ANALYSIS 14.6.1 Introduction 14.6.2 Large injection volumes 14.6.3 Trace enrichment with preconcenmtion columns Hardware considerations, 444. Choice of eluent, 446. Sample loading &em, 449. Concentrator column characteristics, 449. Preconcenrration of samples of low ionic strength, 451. Preconcentration of samples of high ionic strength, 452. Conclusions,453. 14.6.4 Dialytic trace enrichment methods 14.7 MATRIX ELIMINATION METHODS 14.7.1 Introduction 14.7.2 Oncolumn maaix elimination Matrix elimination with concentrator columns, 454.

409 409 409

410

417 417 417 418 418 419

423 423 423 424 425 432 435 435 435 435 436 437 440 441 441 442 443

453 454 454 454

Contents

xx

14.7.3 Post-column matrix elimination 14.8 REFERENCES

456 457

Chapter 15

463

Methods Development

15.1 INTRODUCTION 15.2 SELECTION OF APPROPRIAE CHROMATOGRAPHIC PARAMETERS 15.2.1 The separation method Anion separations, 465. Cation separations, 465. 15.2.2 Thedetectionmode 15.2.3 The nature of the eluent 15.3 OPTlMlZATION OF THE ELUENT COMPOSITION 15.3.1 Empirical selection of eluent composition 15.3.2 Principlesof computer optimization procedures Defining the optimizationproblem, 469. The search area, 469. Oprimization strategies. 469. Optimizationcriterion, 471. Response sutface, 471. 1 5.3.3 Application of computer optimization to IC Factorial design and Simplex methods, 472. Interpretive met&, 474. Summary, 475. 15.4 MULTI-DIMENSIONAL(COUPLED) IC METHODS 15.4.1 Introduction 15.4.2 Column switching methods 15.4.3 Coupled ion-exclusion / ion-exchange IC 15.5 AUTOMATION IN IC 1 5.6 REFERENCES

PART V

463 465 465 467 467 468 468 468

47 1

476 476 476 477 48 1 482

APPLICATIONS OF ION CHROMATOGRAPHY

Overview of the Applications Section

487

En vironm e n ta 1 Applications

489

16.1 OVERVLEW Table 16.1 Acid rain and rainwaters Table 16.2 Seawater and brines Table 16.3 River, stream, pond and lakewaters Table 16.4 Other natural waters Table 16.5 Air, aerosols and airborne particulates Table 16.6 Soils and soil extracts Table 16.7 Geological materials 16.2 REFERENCES

489 490 495 500 506 513 523 528 534

Chapter I 7

543

Chapter I 6

Industrial Applications

17.1 OVERVIEW Table 17.1 industrial wastewaters and effluents Table 17.2 Other industrial waters Table 17.3 Organic compounds Table 17.4 Pulp and paper liquors

543 544 550 559

566

xxi

Contents

Table 17.5 Acids and bases Table 17.6 Detergents and polymers Table 17.7 Fuels, oils and engine products 17.2 REFERENCES

570 576 582 585

Chapter 18 Analysis of Foods and Plants

593

18.1 OVERVIEW Table 18.1 Foods Table 18.2 Beverages Table 18.3 Plants and plant products 18.2 REFERENCES

593 594 605 617 627

Chapter 19

Clinical and Pharmaceutical Applications

633

19.1 OVERVIEW Table 19.1 Blood, serum and plasma Table 19.2 Urine Table 19.3 Other clinical and biological samples Table 19.4 Pharmaceuticals 19.2 REFERENCES

633 634 641 646 651 662

Chapter 20 Analysis of Metals and Metallurgical Solutions

667

20.1 OVERVIEW Table 20.1 Metal plating solutions Table 20.2 Metallurgical processing solutions 20.2 REFERENCES

667 668 683 69 1

Chapter 21

Analysis of Treated Waters

695

21.1 OVERVIEW Table 21.1 Drinking water Table 21.2 High purity waters 2 1.2 REFERENCES

695 696 706 713

Chapter 22

717

Miscellaneous Applications

22.1 OVERVIEW Table 22.1 Chemicals, chemical products and reaction mixtures Table 22.2 Photographic solutions and explosives Table 22.3 Miscellaneous applications 22.2 REFERENCES

Appendix A Statistical Information on Ion Chromatography Public ation s Appendix B Abbreviations and Symbols Index Journal of Chromatography Library (other volumes in the series)

717 718 725 728 732

735 745 75 1 773


1

Chapter 1 Introduction 1.1

BACKGROUND TO ION CHROMATOGRAPHY

Ion chromatography (abbreviated throughout this text as IC) is an analytical technique for the separation and determination of ionic solutes. A more detailed definition is presented later in Section 1.3. IC falls into the general classification of liquid-solid chromatographic methods, in which a liquid (called the eluent) is passed through a solid stationary phase and then to a flow-through detector. The stationary phase is usually in the form of small-diameter (e.g. 5 pm), uniform particles which are packed into a cylindrical column. This column is constructed from a rigid material (such as stainless steel or plastic) and is generally 5-30 cm long, with an internal diameter in the range 4-9 mm. The stationary phase particles are packed uniformly into the column and are retained by means of porous frits located at each end of the column. A high-pressure pump is required to force the eluent through the column at typical flowrates of 1-2 ml/min. The sample to be separated is introduced into the flowing eluent stream by means of an injection device inserted into the flow-path prior to the column. The detector usually contains a low volume (e.g. 4 pl) cell through which the eluent flows. The components of an ion chromatograph are shown schematically in Fig. 1.1. The ion chromatograph is operated in the elution mode. That is, a discrete amount of the sample is applied to the top of the column (via the injector), whilst the eluent is passed continuously through the column. The sample components progress through the column at different rates and therefore enter the detector at different times. The detector senses the sample components and produces the chromatogram. This process is illustrated schematically in Fig. 1.2.

Eluent delivery

Sample injection

Fig. 1.1 Components of an ion chromatograph.

Separation

Detection

2

Chapter 1

Eluent

Sample A+B

A

B

Time Fig. 1.2 Schematic illustration of the separation of two components (A and B) by IC.

1.1.1 Hardware and software components

Any chromatographic system of the type shown in Fig. 1.1 can be divided into hardware and software components. The hardware comprises the instrumentation (pump, injector, detector and data station), whereas the software consists of the variable components of the system (principally the nature of the stationary phase, the composition of the eluent and the flow-rate). This book concentrates on the software of IC. Hardware components will not be discussed, since these components are derived from (and in many cases are identical to) instrumentation developed for High Performance Liquid Chromatography (HPLC). The construction and operation of HPLC equipment has been treated thoroughly elsewhere 1e.g. 11. We need only note that a typical IC instrument uses a loop injector, a dual reciprocating piston pump (often with pulse dampeners to reduce pressure pulsations) and a chart recorder or computing integrator. The nature of the detector used is variable, but conductivity detectors are employed most frequently. We can further note that the hardware components in contact with the eluent can be constructed from either a polymeric material (e.g. PTFE) or stainless steel. The former material may be preferable when trace metal determinations are to be performed, since there is some possibility of contamination of the sample and eluent by metal ions produced from corrosion of stainless steel. 1.1.2

The chromatographic separation process

When a sample is introduced into an IC system, an equilibrium is established for each sample component between the eluent and stationary phases. Thus, for a component A we can write:

3

Introduction

Fig. 1.3 Schematic illustration of the distribution of solute molecules (represented by the black

dots) between the stationary and mobile phases when the distribution coefficient DA is (a) low and (b) high.

Am % A, where the subscript m refers to the mobile (eluent) phase and r refers to the stationary phase. The distribution of component A between the two phases is given by the distribution coefficient, DA,where:

The value of DA is dependent on the size of the population of molecules of component A in the stationary and eluent phases. This is represented schematically in Fig. 1.3, which shows the distribution of component A between the stationary and eluent phases for a small (Fig. 1.3(a)) and a large (Fig. 1.3(b)) value of DA. Since the equilibrium shown in eqn. (1.1) is dynamic, there is a continual, rapid interchange of molecules of component A between the two phases. The fraction of time, fm, that an average molecule of A spends in the mobile phase is given by:

fm =

amount of A in mobile phase total amount of A

(1.3)

That is, (1.4)

(1.5)

where Vm is the volume of the mobile phase and w is the weight of stationary phase (this

Chapter I

4

latter parameter is more convenient, but perhaps less logical, to use than the volume of the stationary phase). If we now define a new variable called the cupucityfuctor, k', as:

k' = DA($)

(1.6)

then eqn. (1.5) can be rewritten as:

f, =

1 1 +k'

(1.7)

Now the rate of travel of component A through the chromatographic system can be obtained by multiplying fm by the average linear velocity of the mobile phase, u, to give:

Rate of travel of A = u

(1L)

(1.8)

The rate of travel will therefore be determined by the average velocity of the mobile phase, the volume of mobile phase, the weight of stationary phase and the distribution coefficient for the particular component. If the chromatographic parameters remain constant, the only factor which changes from one sample component to the next is the distribution ratio. Provided that each component has a unique value of the distribution ratio, then each component will have a different (and characteristic) rate of travel through the stationary phase. That is, each component will emerge from the column at a characteristic retention time.

1.1.3 The chromatogram A representative chromatogram for two sample components is shown in Fig. 1.4. This chromatogram shows three peaks; one for each sample component (A and B) and a so-called "solvent" or "injection" peak. The first of these peaks contains the solvent in which the sample is dissolved, together with those components of the sample which show no interaction with the stationary phase. The sample solvent and unretained components take some time to pass through the column and interconnecting tubing, even though they do not interact with the stationary phase. This time is designated as the void time, to. Similarly, the time at which each retained solute appears is known as the retention time, tR, for that component. The values of to and tR will depend on the flow-rate of the mobile phase and the physical dimensions of the column and connecting tubing. It is therefore often convenient to express the retention of a component in terms of its capacity factor, which is expressed as: (1.9)

5

Introduction

Solvent

lime

-

Fig 1.4 A typical ion chromatogram resulting when a mixture of two sample components (1 and 2) is injected. 1.1.4

Mechanisms of interaction

Up to this stage, we have not specified the manner in which the sample components interact with the stationary and mobile phases. The actual mechanism by which this interaction occurs is one simple means for classifying chromatographic methods into broad groups. For example, the sample components may be retained on the stationary phase through the interaction of permanent dipoles on the component with permanent dipoles on the stationary phase. This results in an adsorption mechanism and gives rise to the general class of adsorption chromatographic methods in which polar stationary phases are employed. Similarly, partition chromatographic methods may be classified as those which involve the distribution of sample components between two immiscible fluid phases, one of which is immobilized to form the stationary phase. Subdivision of these general classes can then be made.according to the physical states of the stationary and mobile phases. For example, gas-liquid chromatography uses a gaseous mobile phase and a liquid stationary phase; liquid-solid chromatography (which embraces IC) uses a liquid mobile phase and a,solid stationary phase. It is unfortunate that IC does not lend itself readily to such a simple definition, because a wide range of different interaction mechanisms is employed. In the following Section, we will provide a working definition of IC which will be used to define the scope of this book.

6

1.2

Chapter 1

WHAT IS ION CHROMATOGRAPHY?

1.2.1 Historical aspects The foundations for modem IC were laid down by Small, Stevens and Bauman in 1975, when they described a novel ion-exchange chromatographic method for the separation and conductimetric detection of anionic and cationic species [2]. Their method employed a low-capacity ion-exchange stationary phase for the separation step, together with a second column and conductivity detector which comprised the detection system for continuous monitoring of the eluted ions. The second column was called a "stripper" column (which was later called a "suppressor") and this column served to reduce the background conductance of the eluent in order to enhance the detectability of the eluted ions. The term "Ion Chromatography" was introduced only when this technology was licensed to the Dionex Corporation for commercial development. For this reason. IC was originally interpreted to mean the specific use of the same combination of separation and detection modes as reported in the original publication. The growth of IC was very rapid because it provided, for the first time, a reliable and accurate method for the simultaneous determination of many simple inorganic anions and cations. In the early stages of its development, IC was seen essentially as a tool for inorganic analysis, but this perception changed with the realization that many organic solutes could also be analyzed using the same approach. This, in turn, led to the use of different stationary phases and detectors, so the original definition of IC became somewhat inaccurate. Later developments in 1979 by Gjerde, Fritz and Schmuckler [3-51 showed that the stripper column was not essential to successful conductivity detection, provided the correct types of stationary phase and eluent were used. At about the same time, the separation of inorganic ions on traditional c18 reversed-phase columns was demonstrated [e.g. 61. Although not specifically suggested by the authors of these papers, the new methods became commonly referred to as IC. 1.2.2 Definition

We are therefore left with the situation where the technique of 1C is very loosely defined, both in terms of the manner in which the solutes are separated and the types of solutes applicable to the technique. For this reason, it is pertinent to begin this book by delineating the range of techniques and applications which are to be included in our working definition of IC. We will consider IC to encompass those column liquid chromatographic techniques which can be used for the determination of Inorganic anions, such as Cl-, B r , Sod2-,efc. Inorganic cations, including alkali metal, alkaline earth, transition metal and rare earth ions, but not including neutral metal chelates. (iii) Low molecular weight (water soluble) carboxylic acids, such as formic acid, acetic acid, etc., and organic sulfonic acids, including detergents. (iv) Low molecular weight (water soluble) organic bases. (v) Ionic organometallic compounds, such as tributyltin, etc-.

(i) (ii)

Intrdction

7

It can be noted that amino acids and carbohydrates are not included, despite the fact that these species are sometimes considered to be part of IC, especially when ionexchange separations are used [7]. The justification for excluding these compounds is that, at best, they fall at the periphery of IC and as the reader is now aware, this text has already reached a considerable size. The column liquid chromatographic techniques applicable to the separation of the species listed in (i)-(v) above will be used to define the IC separation modes. These are: (i) Ion-exchange chromatography. (ii) Ion-interaction (or "ion-pair") chromatography using reversed-phase columns. (iii) Ion-exclusion chromatography. (iv) Miscellaneous separation methods, such as reversed-phase liquid chromatography, the use of chelating stationary phases, etc. Each of these techniques can be coupled with one or more of the following detection methods: (i) (ii) (iii) (iv) (v)

Conductivity detection. Electrochemical (amperometric or coulometric) detection. Potentiometric detection. Spectroscopicdetection. Post-column reaction detection.

Naturally, many of these separation and detection procedures are used routinely for the determination of species other than those listed earlier. Thus, the core of our definition of IC is the types of solutes to be considered, rather than the more conventional chromatographic approach of classification according to the interaction mechanism or the physical states of the stationary and mobile phases.

1.3

ORGANIZATION OF THIS BOOK

The separation and detection methods used in IC are manifold and in most cases, each separation method can be used with a number of different detection methods. We have elected to begin by discussing each of the separation methods used in IC, and to follow this with discussion of the detection methods. Accordingly, Part I (Chapters 2-5) of this book is devoted solely to ion-exchange separations, with separate chapters discussing stationary phases, eluents and retention models. Other separation methods, such as ion-interaction chromatography, ion-exclusion chromatography and reversedphase chromatography are covered in the chapters comprising Part I1 (Chapters 6-8). All of the detection methods listed above are then treated sequentially in Part 111 (Chapters 9-13). Part IV (Chapters 14 and 15) is concerned with some practical considerations in IC, including sample handling and methods development procedures. Finally, an extensive listing of applications is presented in Part V (Chapters 16-22).

Chapter 1

8

1.4

SOME FUNDAMENTAL CHROMATOGRAPHIC CONCEPTS

In this section we will provide a brief overview of some of the more important definitions applicable to modem liquid chromatography. The interested reader seeking further background information on chromatographic concepts is referred to the many excellent texts available on this topic (e.g. [8]). 1.4.1

Chromatographic efficiency

As sample components traverse a chromatographic column, the width of the band travelling through the stationary phase increases. This process is known as band broadening and is evident in Fig. 1.2, where the band due to each component in the mixture is more compact near the head of the column than it is when it reaches the detector. The degree to which two components are separated on a chromatographic system is governed, amongst other factors, by the amount of band broadening which has occurred. The best chromatographic systems are those in which band broadening is kept to a minimum. The "Plate Theory" of chromatography relates band broadening to solute migration [9, 101. This theory uses the concept of theoretical plates, in which the column is considered to consist of a series of thin sections, or "plates", each of which permits a solute to equilibrate between the mobile and stationary phases. The movement of a solute along the column is viewed as a stepwise transfer from one plate to the next. An efficient chromatographic system is one in which there is a large number of theoretical plates, or looked at another way, one in which each theoretical plate occupies only a very short portion of the overall column length. We therefore introduce the term N for the number of theoretical plates and HEW for the height equivalent to a theoretical plate (usually expressed in mm). These terms are related as follows:

HETP =

L N

(1.10)

where L is the length of the column (in mm). The value of N can be calculated from a chromatographic peak by considering the peak to be a population of solute molecules, each with a discrete retention time. This population can be characterized by the mean retention time of the molecules comprising the population (which is the retention time at the peak maximum) and the standard deviation, 0 , of the population. The latter parameter will define the width of the peak. N is given by:

[z) 2

N =

(1.11)

From eqn. ( l . l l ) , it can be seen that a high value of N (and hence an efficient chromatographic system) occurs when the standard deviation of the peak remains small while the retention time is large. The value of 0 can be estimated in a number of ways if

Introducrion

9

Fig. 1.5 Methods for estimating the standard deviation, (T. of a peak in which the retention times of the component molecules follow a Normal distribution.

the population of solute molecules is considered to be a Normal (Gaussian) distribution. Fig. 1.5 shows that the width of the peak at half height (wso) is equal to 2.3450. the width of the peak between tangents drawn to each side of the peak (WT)is equal to 40, and the width of the peak at 4.4% of peak height (w4.4) is equal to 50. Thus, 0 for a Gaussian peak is given by: (1.12)

So that N can be calculated by the following relationships:

(1.13)

For a number of reasons, values of N are meaningful only if they are calculated for peaks with moderately large values of capacity factor (e.g. k' > 5). We can express the separation of two peaks in terms of the resolution, Rs,which can be calculated from the retention time and standard deviation of each peak: (1.14)

10

Chapter 1

Using eqn. (1.12) we can rewrite eqn. (1.14) in terms of the width of each peak between tangents: (1.15)

We should note that all of the above relationships are valid only for Gaussian peaks, but for convenience, they are usually applied also to non-symmetrical peaks. 1.4.2 Factors affecting band broadening

We have already seen that the standard deviation, 0, of a chromatographic peak can be used to estimate the degree of band broadening which has occurred for that particular solute. It is more common to express the width of a Gaussian distribution in terns of the variance, 02. In this Section, wc will examine briefly the physical processes which contribute to the obscrvcd variance. These processes must be considered in the correct design of any chromatographic system so that variance (and hence peak width) can be minimized and efficiency therefore maximized. Eddy diffusion The solute molecules traversing a packed chromatographic column can follow a multitude of paths through the column. Each of these paths is of a different length, so that different molecules of the same component will take slightly varying times to travel through the column. This band broadening process is independent of the flow-rate (average linear velocity) of the mobile phase. The eddy diffusion effect can be minimized if the column is packed uniformly with particles of constant size. Longitudinal diffusion Molecules constituting a band of sample component in a chromatographic system will tend to diffuse out of the sample band during passage through the column. This diffusion occurs both in the direction of flow of the mobile phase, and in the opposite direction. Since diffusion is a time-dependent process, the longitudinal diffusion effect increases at low mobile phase flow-rates. Resistance to mass transfer In an ideal chromatographic system, the interchange of solute molecules between the mobile and stationary phases (i.e. the muss transfer process) would be instantaneous. This does not occur in practice. Additionally, different molecules of the same sample component may spend different amounts of time in the stationary and mobile phases. This leads to a band broadening effect which increases as the mobile phase flow-rate increases. Band broadening due to mass transfer can be. minimized through the use of packing materials which are either of small diameter (and therefore have short diffusion paths) or which have an active layer of stationary phase confined to the outer surface of the particle.

11

introduction

-E0.2 -

k E

W

I

0.1-

I

I

I

0.5 1.a Eluent flow-rate (mllmin)

1

1.5

Fig. 2.6 Plot of HETP versus mobile phase flow-rate for typical modern IC columns. The particle size for the packing material in A is 7 pm and in B is 10 pm.

Extra-column band broadening The sample band will undergo further broadening as a result of diffusion and mixing processes occurring outside the chromatographic column. These effects will be present in the injector, the detector flow-cell, the tubing used to interconnect the chromatographic components, etc. These areas are known as "dead volume" since they do not contribute to the separation process. In all types of chromatographic systems, dead volume should be minimized. The total measured variance for a solute peak can be considered to be the sum of the variances arising from each of the above sources. That is, (1.16)

where the subscripts ed, Id, mt and ec indicate eddy diffusion, longitudinal diffusion, mass transfer and extra-column variances, respectively. When a well-constructed ion chromatograph is used with a column containing uniformly packed, small diameter particles, the most significant of the terms in eqn. (1.16) is the variance arising from mass transfer effects. It is informative to assess the performance of a column in terms of a plot of HETP (which is directly proportional to the measured variance) versus the mobile phase flow-rate. Examples of such plots (often called van Deemter plots) for typical modem IC columns are given in Fig. 1.6, for two different particle sizes of packing material. There is an increase in HETP as the flow-

Chapter I

12

rate increases because of mass transfer effects, but it can be seen that the rate of increase of HETP with flow-rate is lower for the particles of smaller diameter. This permits the use of relatively high flow-rates without undue loss of chromatographic efficiency and shows why there is a trend towards smaller diameter packing materials.

1.5

9 10

REFERENCES Bristow P.A., LC in Practice, HETP,1976. Small H.. Stevens T.S. and Bauman W.C., Anal. Chem., 47 (1975) 1801. Gjerde D.T., Fritz J.S. and Schmuckler G., J. Chromarogr., 186 (1979) 509. Gjerde D.T., Schmuckler G. and Fritz J.S., J. Chromarogr., 187 (1980) 35. Fritz J.S., Gjerde D.T. and Schmuckler G., US Purenf,4,272,246 (1981). Molnar I., Knauer H. and Wilk D., J . Chromafogr.,201 (1980) 225. Weiss J., Handbook of Ion Chromatography,Dionex Corporation, Sunnyvale, CA, 1986. Snyder L.R. and Kirkland J.J., Introduction to Modern Liquid Chromatography, 2nd Ed., Wiley, New York, 1979. Martin A.J.P. and Synge R.L.M., Biochem:J., 35 (1941) 1358. van Deemter J.J., Zuideweg F.J. and Kiinkenberg A., Cbem. Eng. Sci., 5 (1956) 271.

Ion-Exchange Separation Methods

14

Principles Ion-exchangers

Classical 5 Modern Capacity Characteristics f Swelling Selectivity

INTRODUCTION (Chap 2)

Ion chromatSuppressed _I Non-suppressed ography

II

- Surfacefunctionalized

Resin-based ion-exchangers Agglomerated ion-exchangers

STATIONARY PHASES (Chap 3)

r Electrostatically bound

t

Hydrophobically bound Mechanically bound

Hydrous oxide ion-exchangers Carboxylic acids Sulfonic acids

KOH

Anions IONEXCHANGE

-

Polyol-borate EDTA I- Inorganic eluents Non-suppressed I r Nitric acid Organic bases Inorganic elueirts Complexing eluents I

, .

ELUENTS (Chap 4)

--

Anions

I

Suppressed

Gradient elution

F t

G & n a t e buffers Borate Phenate Amino acids Nitric acid High capacity suppressors lsoconductive gradients Baseline balancing

-E

Single eluent competing anion

-

RETENTION MODELS (Chap 5 )

Schematic overview of Part 1.

Cation-exchange

I

competing anions Single eluent competing cation Complexing eluenls

15

Chapter 2 An Introduction to Ion-Exchange Methods 2.1 INTRODUCTION TO ION-EXCHANGE This section introduces the fundamental terminology and principles of ion-exchange chromatography and is intended to provide an overview of the technique. The structures, classifications and important characteristics of ion-exchangers are described. From the basis given in this introduction, detailed treatment of specific ion-exchangers and eluent types used in IC will be provided in subsequent chapters of Part I.

2.1.1

Principles of ion-exchange

An ion-exchanger in aqueous solution consists of anions, cations and water, where either the cations or the anions are chemically bound to an insoluble matrix. The chemically bound ions are referred to as the fixed ions and the ions of opposite charge are referred to as the counter-ions. The insoluble matrix may be inorganic or may be a polymeric organic resin, and especially in older types of ion-exchangers, is generally porous in nature. These pores contain water from the aqueous solution, together with a sufficient concentration of counter-ions to render the whole exchanger electrically neutral. Counter-ions may move through the matrix either by diffusion or under the influence of an electric field and in the ion-exchange process itself, are replaced by ions of the same charge from the external solution. The ion-exchanger is classified as a cation-exchange material when the fixed ion carries a negative charge, and as an anionexchange material when the fixed ion carries a positive charge. The ion-exchange process can be illustrated by considering an anion-exchange material, for which the counter-ion is E-. The exchanger can therefore be represented as M+E-, where M+ denotes the insoluble matrix material containing the fixed (positive) ion. When a solution containing a different anion, A-, is brought into contact with the ion-exchanger, an equilibrium is established between the two mobile ions E- and A- as follows:

M+E- + A-

+ M+A- + E-

(2.1)

Since the electroneutrality of the solution must be maintained during the ionexchange process, the exchange is stoichidmetric, such that a single monovalent anion Adisplaces a single monovalent counter-anion E-.Eqn. (1) can be generalized for y moles of AX- exchanging with x moles (i.e. the stoichiometric amount) of EY- to give:

Chapter2

16

where the subscript m denotes the mobile (i.e. solution) phase and r denotes the stationary (or resin) phase. It should be noted that eqn. (2.2) is an equilibrium and under conditions where the exchanged ions remain in contact with the ion-exchange matrix, complete exchange will not be attained. Furthermore, the solution phase contains a co-ion of the same charge as the fixed ion, but this co-ion plays no part in the ion-exchange process and is therefore not shown. The equilibrium constant for the reaction shown in eqn. (2.2) is called the selectivity coefficient, and is given by:

where the parentheses indicate the activity of the designated species. Since the ion activity in the resin phase cannot be determined, KA,E is not a thermodynamically defined equilibrium constant but a coefficient which is defined according to practical requirements. Under conditions where the activity coefficients approximate unity, eqn. (2.3) can be simplified to:

where the brackets indicate molar or molal concentrations, or equivalent fraction units. For convenience, concentrations are often expressed in the units molesA for the solution phase, and millimoles/g for the matrix phase. The selectivity coefficient derives its name from the information it provides on the likelihood of exchange between two particular ions. In the above example, if K=l, then the ion-exchange matrix shows no selectivity for anion AX-over Ev-;that is, the ratios of the concentrations of these ions in the matrix and solution phases are equal. If K is greater than unity, the matrix (or resin) phase will contain a higher concentration of ion AX- than the solution phase, and will select AX- preferentially over Ey-. The reverse situation applies for values of K less than one. Clearly, a competition for the ionexchange sites exists between the two ions AX-and Ey-.It is convenient to designate EYas the competing anion and to describe AX-as the solute anion. For cation-exchange equilibria, a similar series of equations can be derived. Thus, for an ion-exchange reaction between the ion Ax+ and ion EY+, the equilibrium can be written:

YA,

X+

+ XE, Y+

+ YA:++

Y+ XE,

(2.5)

Introduction to Ion-Exchange

17

and after assuming that the activity coefficients are close to unity, the selectivity coefficient is given by: (2.6)

A more detailed discussion of retention in ion-exchange chromatography, as applied to modern ion chromatographic ion-exchangers, is presented in Chapter 5. Another parameter commonly used to express the position of an ion-exchange equilibrium is the distribution coefficient, DA, which is defined as the ratio of the concentrations of the ionic species in the exchanger phase and in the solution phase. Using the equilibrium shown in eqn. (2.2) as an example, the weight distribution coefficient for the solute ion AX- is given by:

Note that different symbols (e.g. KD)may be used for the distribution coefficient. The concentration units used in eqn. (2.7) are the same as those for eqn. (2.4). Distribution coefficients are concentration dependent, but under conditions where the exchanged ion exists in trace amounts in the presence of a higher concentration of competing ion, the distribution coefficient becomes constant. 2.1.2 Configuration of an ion-exchange chromatographic system Open-column (classical) ion-exchange chromatography Historically, ion-exchange separations have been performed in the open-column mode, wherein the ion-exchange matrix is loosely packed as small particles into a glass column of 1-2 cm diameter. The mobile phase, or eluent as it is usually called in ionexchange chromatography, contains the competing ion and is passed continuously into the column and percolates through it under gravity. When a separation is to be performed, the flow of eluent is stopped and a small amount of the sample mixture is applied to the top of the column and allowed to pass into the bed of ion-exchange material. Eluent flow is then resumed and fractions of eluent are collected at regular intervals from the column outlet, to be later analyzed for the solute components. The results of these analyses are then combined to provide an elution profile showing the volume of eluent required,to elute each sample component from the column. Fig. 2.1 shows a schematic representation of the apparatus used and a typical set of results obtained. This analysis procedure is described as the elution mode of chromatography, and whilst different alternatives are possible, this mode is the most commonly used in open column ion-exchange chromatography. Open column ion-exchange is not ideal for trace analysis for a number of reasons. First, the process is very slow because of the low eluent flow-rates employed, and attempts to increase the flow-rate (e.g. by raising the height of the eluent container

18

n

Chapter2

B

Fraction Number

Fig. 2.1 Apparatus for classical open column ion-exchange chromatographyand the type of result typically obtained. A and B are two components eluted from the column.

above the head of the column) invariably result in poorer separation efficiency. This effect is primarily a result of the poor mass-transfer characteristics of the relatively large particles used for the column packing. Second, the chromatographic efficiency attained is rather poor and some important separations cannot be achieved using this method. Third, it is inconvenient to collect and separately analyze fractions of eluent to determine the elution volumes of the sample components.

Modern ion-exchange chromatography Modem IC using ion-exchange separations circumvents these difficulties through the use of high efficiency ion-exchange materials combined with continuous flowthrough detection. Separations are performed in the column mode, where the ionexchanger exists as particles of uniform size packed into a column housing constructed of rigid material. The size of this column is dependent on a number of factors, including the chromatographic efficiency of the ion-exchange material and its mechanical stability. Typical columns are 15 cm in length, with an internal diameter of 3-5 mm. The particles of ion-exchange material are generally very much smaller than those used for classical open column ion-exchange chromatography. The further instrumental components of a chromatographic system used for ion-exchange follow the general outline presented in Fig. 1.1. The eluent must be pumped through the column since the small particle size of the stationary phase precludes appreciable flow under gravity. The sample mixture is introduced into the eluent via the injection port and is


19

separation takes place. Finally, the separated ions are detected with a flow-through detection device.

Eluent characteristics The eluent used in ion-exchange chromatography generally consists of an aqueous solution of a suitable salt or mixture of salts, with a small percentage of an organic solvent being sometimes added. The salt mixture may itself be a buffer, or a separate buffer can be added to the eluent if required. The prime component of the eluent is the Competing ion, which has the function of eluting sample components through the column within a reasonable time. The three foremost properties of the eluent affecting the elution characteristics of solute ions are: (i) The eluent pH. (ii) The nature of the competing ion. (iii) The concentration of the competing ion. The eluent pH can have profound effects on the form in which the functional group on the ion-exchange matrix exists, and also on the forms of both the eluent and solute ions. The selectivity coefficient existing between the competing ion and a particular solute ion will determine the degree to which that competing ion can displace the solute ion from the stationary phase. Since different competing ions will have different selectivity coefficients, it follows that the nature of the competing ion will be a prime factor in determining whether solute ions will be eluted readily. Finally, the concentration of the Competing ion can be seen to exert a major effect by influencing the position of the equilibrium point for ion-exchange equilibria, such as those depicted in eqns. (2.2) and (2.5). The higher the concentration of competing ion in the eluent, the more effectively the eluent displaces solute ions from the stationary phase and thus the more rapidly is the solute eluted from the column. In addition to the above three factors, elution of the solute is influenced by the eluent flow-rate and the temperature. Faster flow-rates lead to lower elution volumes because the solute ions have less opportunity to interact with the fixed ions. Temperature has a less predictable effect, which is somewhat dependent on the type of ion-exchange material used. An elevated temperature increases the rate of diffusion within the ion-exchange matrix, generally leading to increased interaction with the fixed ions and therefore larger elution volumes. Chromatographic efficiency is usually improved at higher temperatures. Prior to sample injection, the column must be equilibrated with eluent so that all the exchange sites on the stationary phase contain the same counter-ion. A point of terminology arises here, when one considers that equilibration of a column with a solution of competing ion (i.e. the eluent) results in the counter-ions associated with the fixed ions being completely replaced with competing ions. When the column is in this condition, the competing ions become the new counter-ions at the ion-exchange sites and the column is said to be in the forin of that particular ion. For example, an anionexchange column which is fully equilibrated with il NaOH eluent is in the hydroxide form. Reproducible elution volumes are obtained only when the column is converted to the same form prior to the injection of each sample and this becomes especially

Chapter2

20

ION-EXCHANGERS

I SILICA-BASEDMATERIALS polymer-coated silica+ functionalised silica*

I

I 1 ORGANIC MATERIALS INORGANICMATERIALS synthetic polymeric resins* aluminosilicates celluloses insoluble salts

dextrans

heteropolyacids clays hydrous oxides'

Fig. 2.2 Classification of ion-exchangers.

important when the eluent is changed. The time required for a column to equilibrate to a new eluent depends on the selectivity coefficient for the competing ion in that eluent over the previous competing ion, and also on the concentration of the competing ion in the new eluent. 2.1.3 Types of ion-exchange materials Ion-exchangers are characterized both by the nature of the ionic species comprising the fixed ion and by the nature of the insoluble ion-exchange matrix itself. The matrix types used for ion-exchange chromatography can be subdivided broadly into inorganic materials and organic (polymeric) materials, as illustrated in Fig. 2.2. The four materials shown in bold face type. and marked with an asterisk are those that are used for IC, and these will be discussed in detail in Chapter 3. However, it is pertinent to include here a brief general description of some of the more important properties common to all ionexchange materials, so that the special characteristicsof the four materials used for IC can be better appreciated when these materials are discussed more fully. With the exception of the hydrous oxide materials, the fixed ion in the above ionexchangers forms part of afuncrional group which is bound covalently to the surface of the material. The fixed ion in hydrous oxide exchangers is introduced by protonation or deprotonation reactions. Table 2.1 shows the types of functional groups commonly encountered in synthetic ion-exchangers. The chemistry involved in the introduction of these functional groups onto silica and synthetic polymers is discussed in Chapter 3. Cation-exchange resins are classified into srrong acid and weak acid types. The former retain the negative charge on the fixed ion over a wide pH range, whereas the latter type are ionized (and hence act as cation-exchangers) only over a much narrower pH range. Sulfonic acid exchangers are strong acid types, whilst the remaining cationexchange functional groups in Table 2.1 are weak. The weak acid types require a sufficiently high pH for use, and this is exemplified by the use of a NaOH eluent with a carboxylic acid cation-exchanger.

Resin-COOH + NaOH % Resin-COO-Na'

+ 30

(2.8)


21

TABLE 2.1 FUNCTIONALGROUPS FOUND ON SOME TYPICAL SYNTHETICION-EXCHANGE MATERIALS

Cation-exchangers Type

Functional group

Sulfonic acid carboxylic acid

Phosphonic acid Phosphinic acid Phenolic Arsonic acid Selenonic acid

-SO3- H+ -COO-H+ -PO3H- H+ -P02H- H+ -0H+ -As03R H+ -SeO3- H+

Anionexchangers

Type Quaternaryamine Quaternary amine Tertiary amine Secondary amine Primaryamine

Functional group -N(CH3)3+ OH-N(CH3)2(EtOH)+ OH-NH(CH3)2+ OH-NH2(CH3)+ OH-NH3+ OH-

Similarly, the anion-exchangers are classified as strong base and weak base exchangers. Quaternary amine functional groups form strong base exchangers, whilst less substituted amines form weak base exchangers. A weak base material will function only when the pH is sufficiently low to protonate the nitrogen atom in the functional group. This is illustrated below for a primary amine functionality in the presence of HCl eluent.

Resin-NH2 + HC1 % Resin-NHiCl-

(2.9)

Whilst a diverse range of ion-exchange functionalities exists, most IC separations with silica and organic ion-exchangers are performed on strong acid cation-exchangers of the sulfonic acid type, and on strong base anion-exchangers of the quaternary ammonium type. These strong cation-exchangers and strong anion-exchangersare often labelled SCX and SAX,respectively. 2.1.4

Characteristics of ion-exchangers

Ion-exchange capacity

The ion-exchange capacity of an ion-exchanger is determined by the number of functional groups per unit weight of the resin. It may be measured in a variety of units, the most common of which are milliequivalents (of charge) per gram of dry resin, or milliequivalents per millilitre of wet resin. In the latter case, it is usual to state the type of counter-ion present on the resin since this affects the degree of swelling of the resin and hence its volume. The ion-exchange capacity is often measured by saturating a known weight of resin with a particular ion, followed by washing the resin and then quantitative displacement of this ion. The number of moles of the displaced ion can then be determined. It should be noted that the capacity measured in this way is often somewhat higher than that applicable when the resin is packed as the stationary phase in

22

Chapter 2

a chromatographic column. The ion-exchange capacity of a resin plays a large role in determining the concentration of competing ion used in an eluent to be employed with that resin. Higher capacity resins generally require the use of more concentrated eluents, and as will become evident in Chapter 4, the eluent concentration is of paramount importance in IC. Classical ion-exchangeresins of the types described above have capacities which typically fall into the range 3-5mequivlg.

Swelling characteristics Organic resin exchangers consist of cross-linked polymer chains containing ionic functionalities. When such a material comes into contact with water, the outermost functional groups are solvated and the randomly arranged polymer chains unfold to accomodate the larger solvated ions. A very concentrated internal solution of fixed ions and counter-ions therefore exists and the mobile counter-ions tend to diffuse out of the exchanger into the external aqueous solution. The fixed ions cannot diffuse, and as a result, external water molecules are forced into the resin in an attempt to reduce the internal ionic concentration in the resin. The cross-linking of the resin provides mechanical stability which prevents dissolution of the resin, but swelling persists as a result of the equilibrium pressure due to the differences in concentration between the external and internal ionic solutions. The swelling pressure may be as high as 300 atmospheres for a polymeric resin of high ion-exchange capacity [l]. It is clear from the above description that the degree of swelling of the resin is dependent on the composition of the solution with which it is equilibrated. Thus, changes of eluent are accompanied by changes in the level of swelling and this effect has important ramifications on the types of resins suitable for use as stationary phases in chromatographic columns of fixed volume. Resins of low cross-linking (c 2%) exist as soft gels in aqueous solution and exhibit large volume changes when the eluent is altered; for this reason, they are unsuitable as stationary phases for high performance applications where the eluent is delivered under pressure. Macroporous resins (which are discussed in detail in Section 3.3) are more rigid due to their high cross-linking and their resistance to swelling effects renders them more suitable as chromatographic stationary phases for column packing purposes. Ion-exchange selectivity Selectivity coefficients (eqn. (2.3)) provide a means for determining the relative affinities of an ion-exchanger for different ions. It might be considered that a welldefined affinity series for anions and cations could be obtained by simple experiment, but in reality, the relative affinities show considerable variation with the type of ionexchanger and the conditions under which it is used. In some cases, simple ion-exchange may not be the sole retention mechanism operating; for example, partitioning of solute ions between the eluent and the pores of the stationary phase may occur, or the solute ion could be adsorbed onto the surface of the ion-exchange matrix. In view of these factors, it is possible to provide only approximate guidelines for the relative affinities of ionexchangers for different ions.


23

Selectivity coefficients for the uptake of cations by a strong acid cation-exchange resin are generally in the following order [2]:

Pu4+ >> La3+> Ce3+> Pr3+> Eu3+ > Y3+> Sc3+> A13+ >> Ba2+> Pb2+> Sr2+> Caz+> NiZ+> Cd2+> Cu2+ > Co2+> Zn2+> Mg2+> U022+ >> Tl+ > Ag+ > Cs+ > Rb+ > K+> N&+ > Na+ > H+ > Li+ It follows from this series that a cation-exchange eluent of 0.1 M KC1 will be stronger than that containing 0.1 M NaCl, provided other factors are equal. Selectivity coefficients for unions on strong base anion-exchangersfollow the general order:

> W0d2- > M 0 0 4 ~> - C104~- > C ~ 0 4>~ citrate > salicylate > C104- > SCN- > I- > S~o3~S042- > SO-j2- > HP042- > NO3- > Br- > NOz- > CN- > C1- > HCO3- > H2P04- > CH3COO- > 103- > HCOO- > BrO3- > ClO3- > F > OHSome general rules can be offered to assist in the prediction of the affinity order. These are based on a number of properties of the solute and the ion-exchanger and include: (i) (ii) (iii) (iv) (v) (vi) (vii)

The charge on the solute ion. The solvated size of the solute ion. The degree of cross-linking of the ion-exchange resin. The polarizability of the solute ion. The ion-exchange capacity of the ion-exchanger. The functional group on the ion-exchanger. The degree to which the solute ion interacts with the ion-exchange matrix.

An increase in the charge on the solute ion increases its affinity for an ionexchanger through increased coulombic interactions. This trend is known as electroselectivity and becomes more pronounced as the external solution in contact with the ion-exchanger becomes more dilute. Electroselectivity may be explained in terms of the Donnan potential, which is the potential difference arising because of the imbalance in the ionic concentra;ions in the resin bead and in the external solution [3]. An exchange involving the replacement of two bound monovalent ions with a single divalent ion causes this imbalance to be diminished, and is thus a favourable process. Electroselectivity is reflected in the following series of selectivity coefficients: Pu4+ >> La3+ >> Ba2+ >> T1+. For cations, the trend is very strong, such that the selectivity differences between these four ions are large. The size of the solvated solute ion also exerts a significant effect, with ions of smaller solvated size showing greater binding affinity than larger ions. Thus the selectivity sequence Cs+ > Rb+ > K+ > Na+ > H+ > Li+ is exactly the reverse of the sequence of ionic radii for the hydrated' ions, and follows the well-known lyotropic series with the most strongly hydrated ion, Li+, being held most weakly. This behaviour is related directly to swelling of the resin, since a smaller ion is more easily

Chapter2

24

accommodated in the resin pores. Thus, the higher the degree of cross-linking, the greater the preference of the resin for smaller solute ions. The combination of factors (i) and (ii) above suggests that binding affinity should increase with increasing polarizing power; that is, for ions with a high charge and small hydrated radius. Ion-exchange selectivity coefficients increase with the degree of polarizability of the solute ion. Thus, sulfonic acid fixed ions show greater affinity for the more polarizable Ag+ and Tl+ than for the alkali metal ions. Similarly, I- is more strongly retained on an anion-exchanger than Br- or Cl-.However, polarization does not explain why C l o g has a higher anion-exchange affinity than I-. The strong retention of anions such as Clod-, which are large, have low charge and a~ weak bases, can be attributed to the interaction of these ions with the water structure at the resin surface. Large, polarizable ions with a diffuse charge do not easily form a well-orientated layer of water molecules at their surface, and so tend to disrupt the surrounding water structure. This leads to an increase in free energy, which is the driving force for these ions to bind (that is, to form an ion-pair) with the fixed ion of an ion-exchanger, thereby diminishing both the disruption to the water structure and the free energy. This binding process is called water-structure induced ion-pairing 14). The trends evident from the remaining factors ((v) - vii)) listed above are not as clear-cut. The ion-exchange capacity of the ion-exchanger can affect the selectivity coefficients for some anions and cations, but for most ions, the selectivity coefficient remains essentailly constant as the ionexchange capacity is decreased. Similarly, the nature of the functional group exerts little effect for most ions, but significantly affects the selectivity coefficients for other ions. Large, polarizable anions such as BF4', I-, ClO4- and C l o g show changes in selectivity as the alkyl- substituents in trialkylammonium strong-base anion-exchangers are varied. This may be related to the waterstructure induced ion-pairing mechanism discussed above, with the larger functional groups (i.e. those with the largest alkyl- substituents) causing greatest disruption to the water structure, causing them to bind large, polarizable ions more strongly than smaller functional groups (5.61. Interactions between the solute ion and the ion-exchange matrix are difficult to predict, and are of course specific to individual ions. Factors (v)-(vii) above will be discussed later in Part 1 as a component of the detailed treatment of those ion-exchangers developed specifically for IC. 2.2

CLASSIFICATION OF ION CHROMATOGRAPHIC METHODS EMPLOYING ION-EXCHANGE SEPARATION

Ion-exchange methods have always formed the basis of IC, and due to a combination of historical development and commercial marketing influences, these methods have been divided somewhat arbitrarily into two main groups. 2.2.1 Non-suppressed ion chromatography

The first of these groups comprises all of the methods in which an ion-exchange column is used to separate a mixture of ions, with the separated solutes being passed directly to the detector. The hardware configuration employed is shown schematically


25

I

I Recorder or integrator Fig. 2.3 Block diagram showing the instrumental components used in non-suppressed IC.

in Fig. 2.3, from which it can be seen that this configuration parallels the traditional HPLC approach in which the chromatographic column is coupled directly to the detector. Some of the names proposed for this technique are [7-111: (i) "non-suppressedion chromatography", (ii) "single-columnion chromatography". (iii) "electronically-suppressedion chromatography". The first two names indicate that only a single chromatographic column is employed and that the eluent is not chemically modified prior to entering the detector, whereas the last name pertains to the fact that the background conductance of the eluent can be nulled electronically by certain types of conductivity detectors, "Non-suppressed IC" is the most frequently used term and will be employed throughout this text.

2.2.2

Suppressed ion chromatography

The second group of ion-exchange methods consists of those in which an additional device, called the suppressor, is inserted between the ion-exchange separator column and the detector, as shown in Fig. 2.4. The function of the suppressor is to modify both the eluent and the solute in order to improve the detectability of the solutes with a

7 Regenerant

I

Recorder integrator OrI

Fig. 2.4 Block diagram showing the instrumental components used for suppressed IC.

Chapter2

26

conductivity detector. The suppressor requires a regenerant (or scavenger) solution to enable it to operate for extended periods. Methods using the configuration shown in Fig. 2.4 are referred to as [7-131: (i) (ii) (iii) (iv)

"suppressed ion chromatography", "chemically-suppressedion chromatography", "eluent-suppressed ion chromatography", "dual-column ion chromatography".

The last of these names is misleading because modem suppressors are not columns, but rather flow-through membrane devices. The term "suppressed IC" is preferable and will be used in this text.

2.2.3

Similarities between non-suppressed and suppressed methods

The contention that the division of ion-exchange procedures into non-suppressed and suppressed methods is arbitrary merits further discussion. The first point to be made is that the detection mode employed in IC is the chief factor which determines the type of eluent and column used. Conductivity detection gives excellent sensitivity when the conductance of the eluted solute ion is measured in an eluent of low background conductance. This suggests that dilute eluents should be preferred, and in order for such eluents to act as effective competing ions (and so elute solute ions within a reasonable time), the ion-exchange capacity of the column should be low when conductivity detection is used. This principle applies equally well to both the non-suppressed and suppressed IC methods, although in the latter case, somewhat higher column capacities (and hence less dilute eluents) can be used because the suppressor serves to decrease the eluent conductance. With the exception of the suppressor itself, the only real distinctions which can be made between suppressed and non-suppressed methods are the small difference in column capacities and the use of a specialised group of eluents. These distinctions become pertinent only when conductivity detection is employed. This situation is quite common, but as Part I11 of this book will show, there are many alternative detection methods applicable to IC which do not require the use of a suppressor. The second point is that the suppressor is a device designed to improve detection (and will hence be discussed in detail in Part 111) and therefore exerts no influence on the ion-exchange separation, except for the fact that the eluent components must be compatible with the suppressor. The ion-exchange columns used with the nonsuppressed and suppressed approaches have very similar separation characteristics and this permits them to be used satisfactorily with either technique, provided that attention is paid to the ion-exchange capacities and pH limitations of each column. Indeed, the literature of IC abounds with examples of this practice. The above discussion suggests that little difference exists between non-suppressed and suppressed IC from a separation standpoint, and the stationary phases used for both approaches will be therefore be discussed together in Chapter 3. However, some aspects of suppressed and non-suppressed IC differ when eluents are considered, and the distinction between these approaches will be made in the discussion of eluents in Chapter


27

4. It is one of the prime aims of this book that non-suppressed and suppressed IC will be treated together wherever possible, so that the reader is directed towards the common elements of these approaches, rather than towards their differences.

2.3 1 2 3 4 5 6 7 8 9 10 12 13

REFERENCES Samuelson 0.. Ion Exchangers in Analytical Chemistry, Wiley, New York, 1953. Peters D.G.,Hayes J.M. and Heiftje G.M., Chemical Separations and Measurements, Saunders, Philadelphia, 1974, p. 583. Paterson R., An Introduction to Ion Exchange, Heyden, London, 1970, p. 30. Diamond R.M., J. Phys. Chem., 67 (1963) 25 13. Barron R.E. and Fritz J.S., J . Chromatogr., 316 (1984) 201. Barron R.E. and Fritz J.S.,J . Chromatogr., 284 (1984) 13. Fritz J.S., Gjerde D.T. and Pohlandt C., Ion Chromatography, Huthig, Heidelberg, 1982. Fritz J.S. and Gjerde D.T.,Ion Chromatography, 2nd edn., Huthig, Heidelberg, 1987. Smith F.C. and Chang R.C., The Practice of Ion Chromatography, Wiley, New York, 1983. Tarter J.G. (Ed.), Ion Chromatography, Marcel Dekker, New York. 1987. Weiss J.. Handbook of Ion Chromatography, Dionex Corporation, Sunnyvale, CA, 1986. Smith R.E., Ion ChromarographyApplications, CRC Press. Boca Raton, FI, 1988.


29

Chapter 3 Ion-Exchange Stationary Phases for Ion Chromatography 3.1

INTRODUCTION

The main factor which differentiates the ion-exchange materials used in IC from the conventional ion-exchangers discussed in Section 2.1.3 is their ion-exchange capacity. Ion-exchange separations in IC are generally performed on ion-exchangers with low ionexchange capacity, typically in the range 10-100 pequiv/g. As discussed earlier, this characteristic can be attributed chiefly to the fact that IC was developed originally for use with conductivity detection, which introduces a preference for eluents of low background conductance. The diversity of detection methods currently available (see Part 111) now makes it possible to use columns of much higher ion-exchange capacity, but because conductivity detection is still the most commonly employed detection mode, the majority of separations continue to be performed on low capacity materials. It is convenient to discuss the different types of ion-exchangers used in IC according to the groupings which were shown in Fig. 2.2.

3.2

SILICA-BASED ION-EXCHANGE MATERIALS

3.2.1 Types of silica-based ion-exchangers Silica-based materials constitute one of the most important classes of ionexchangers used in chromatography. Two distinct groups of materials can be recognized. The first comprises polymer-coated materials, in which a silica particle is first coated with a layer of polymer, such as polystyrene, silicone or fluorocarbon, and this layer is then derivatized to introduce functional groups of the types listed in Table 2.1. The main advantage of such a particle is that diffusion within the thin layer of polymer is very much faster than that occurring with totally polymeric particles. This leads to favourable mass-transfer characteristics and hence improved chromatographic efficiency for polymer-coated particles. The second group of silica-based ionexchangers comprisesfunctionalized silica materials, where a functional group (which acts as the fixed ion) is chemically bonded directly to a silica particle. The silica particles used for both polymer-coated and functionalized silica ionexchangers can be either pellicular or microparticulate. Pellicular materials have a solid, inner core and ion-exchangers formed from these particles have the functional

Chapter3

30

groups confined to the outer surface of the particle, or to a thin outer surface layer. Superficially-porous particles are one example of pellicular materials and these consist of solid, spherical glass beads of relatively large diameter (e.g. 30 pm), with a thin (approximately 1 pm) layer of porous silica on the surface. Zipax (Du Pont) is the chief example of such a pellicular, superficially porous material. Microparticulate materials are small diameter (e.g. 5 pm), fully porous particles, which can be irregular or spherical in shape. Ion-exchangers formed from microparticulate particles have functional groups distributed throughout the internal pore structure. Pellicular and microparticulate materials provide similar chromatographic efficiencies, but the pellicular materials are restricted to the separation of small amounts of sample because of their low active surface area. On the other hand, pellicular materials are more easily packed into columns than are microparticulate materials. The chromatographic properties of spherical and irregular microparticulate particles are similar.

3.2.2 Functionalized silica ion-exchangers ion-exchangers produced by chemically bonding the ion-exchange functional groups to a silica backbone were initially the main type of column packings commercially available for non-suppressed IC. Typically, quaternary ammonium functionalities or sulfonic acid groups are bound to microparticulate silica to produce strong base anion-exchangers or strong acid cation-exchangers, respectively. Functionalized silica IC packings have found most application in anion separations. Weak-base exchangers with a primary amine functional group can also be employed provided the eluent is sufficiently acidic to generate adequate anion-exchange capacity in order to give suitable retention of sample anions. Functionalized silica ion-exchangers can be produced by a variety of synthetic methods. Well-established and reliable procedures exist for the chemical bonding process and these reactions form the basis for the preparation of stationary phases for reversed-phase HPLC. The most commonly used method involves reaction of the silanol groups on silica with organochlorosilanes or organoalkoxysilanes. as shown in Fig. 3.1. This produces a siloxane-type (Si-0-Si-C) bonded phase, where the bonded layer can be monomolecular or polymeric in nature. The field of HPLC has provided a vast amount of experience in the preparation and handling of silica-based stationary phases. Many different types of functionalized silica ion-exchangers were in use in 1979, when non-suppressed IC was first introduced [l].

I -SOH

I I

-%OH

I

I + SOCI, - b - - S i - C I

H2NC%CH2NH2

I

+ CISiR3 or ROSiR3

b

I

-Si-NH-CH2CH2NH2 I I

-Si-O-SiR3 I

Fig. 3.1 Reaction schemes for the preparation of functionalized silica ion-exchangers.

Ion-ExchangeStationary Phases

31

These materials were of the moderate to high capacity type and, as such, were unsuitable for use with conductivity detection because of the high eluent concentrations required. For this reason, specially designed low-capacity exchangers were produced, with ionexchange capacities of about 100 pequiv/g. Table 3.1 lists the characteristics of some functionalized silica IC packing materials, together with those of the polymer-coated silica IC materials discussed in the next Section. When employed with dilute eluents, these columns give excellent separations of common anions, together with adequate sensitivity using conductivity detection. Fig. 3.2(a) shows a typical separation obtained with a low-capacity silica-based anion exchanger (TSKgel IC-Anion-SW), whilst a separation of inorganic cations on a lowcapacity functionalized silica cation-exchanger (Vydac 400 IC) is illustrated in Fig. 3.2(b). The traditional types of strong or weak silica-based ion-exchangers can also be employed for IC, especially when a detection method other than conductivity is used. Chromatograms obtained on high capacity functionalized silica SAX and SCX exchangers are shown in Fig. 3.3, whilst Fig. 3.4 illustrates the use of a weak-base amino column.

3.2.3

Polymer-coated silica ion-exchangers

Pellicular ion-exchangers formed by coating an impervious silica core with a polymeric ion-exchange material have found widespread application in non-suppressed IC. These packing, marketed under the name of Zipax, are of low ion-exchange capacity, have rapid radial mass transfer characteristics [26] and their relatively large particle size (ca. 35 pm) provides for ease of packing. Fig. 3.5 shows representative separations of anions and cations on polymer-coated silica ion-exchangers. The characteristics of some polymer-coated IC materials are included in Table 3.1. Polymer-coated silica cation-exchangers for IC can be synthesized by depositing varying film thicknesses of a pre-polymer onto 5 pm porous silica, with subsequent immobilization of the polymer by in-situ cross-linking reactions using radical starters or y-radiation [46, 471. Poly(butadiene-maleic acid) (PBDMA, see below) is the preferred polymer and the film thickness can be varied to regulate the ion-exchange capacity of the final product. Ionization of the carboxylate functionalities in the polymer is significant above pH 3, so weak acid eluents can be used to separate inorganic cations. Fig. 3.6 illustrates the simultaneous separation of monovalent and divalent cations on PBDMAcoated silica.

COOH . .

COOH Poly(butadiene-maleicacid) (PBDMA).

.E

-

+

32

El

0

mi=

;-

Q-n

In-

- E,

i -

-!e -$!

;=

.-C

- E .-

-a--

-* -0

I,

I D

.-c E

-

-2 -L"

-0'

c

.-E

-a'

-m

-0

Chapter 3

33


TABLE 3.1 SOME COMMERCIALLY AVAILABLE SILICA-BASEDIC COLUMN PACKINGS Column

Silica type

Vydac 302.IC Vydac 300.IC Wescan ANon/S TSKgel IC Anion SW Nucleosil SB PartisillO SAX Zorbax NH2 zipax SAX Vydac 400.IC vydac sc Wescan CatiodS TSK gel IC Cation SW Nucleosil 10 SA PartisillO SCX Zipax SCX Nucleosil-5-100-PBDMAa

Functionalized Functionalized Functionalised Functionalized Functionalized Functionalized Functionalized Polymer-coated Functionalized Functionalized Functionalised Functionalized Functionalized Functionalized Polymer-coated Polymer-coated

Class

Particle size (PI

Anion Anion

8 5.5 10

Anion

Anion Anion Anion Anion Anion Cation Cation Cation Cation Cation Cation Cation Cation

5-8

5

Capacity (Wg) 100 250 400 lo00

10 10 25-37 4.5 30-44 10

500

5

450 lo00

10 10 15-37

12

Refsb

2-5

6-8 9-12 13-16 17-20

21-23 24 25,26 6

100

500

5

5

27-29 30-33 34-36 37 38-41 42-45 46,47

a PBDMA = polyfiutadiene-maleic acid). b This listing is not comprehensive, but provides some examples of the use of each column.

Nafion, a cation-exchange material manufactured by Du Pont, may be coated onto an octadecylsilyl reversed-phase column by simple hydrophobic interaction [49]. The resultant material can then be used to pack ion-exchange columns of unusual selectivity. This approach has been demonstrated for the separation of aromatic base cations and may prove to be applicable also to the separation of inorganic cations. 3.2.4

Advantages and limitations of silica-based IC packings

Chromatographic efficiency The prime advantage of 1C columns packed with silica-based materials is the favourable chromatographic efficiency they produce when correctly packed into chromatographic columns. As an illustration of this efficiency, the peak for nitrate ion in a typical separation gave 16,652 theoretical plates per metre using the 5 0 method of calculation [50]. Silica can be obtained as small diameter particles with a very narrow size distribution, and being a non-swelling, rigid material, can be packed at high pressure to produce a uniform and stable ,chromatographic bed which is not subject to stringent pressure or flow-rate limitations during usage. For this reason, columns packed with silica-based ion-exchangers may be relatively long (e.g. 30 cm) and tolerate backpressures as high as 4,000 psi. Moreover, organic modifiers can be used freely with

34

Chapter3

I

0

I

1

1

I

I

2

3

6

I

1

5 6 7 Time ( m i n )

I

I

8

I

9

I

1

0

Fig. 3.4 IC on an amino column. Zorbax NH2 (250 x 4.6 m m ID)column using 0.03 M H3P04 at pH 3.2 as eluent. Solute concentrations 25-100 ppm. Spectrophotometricdetection at 205 nm was used. Reprinted from [24] with permission. Time (min) 0

1

I

I

I

1

0

2

4

6

0

Time Imin) (01

I

10

I

20

1

lb)

Fig. 3.5 IC on polymer-coated silica ion-exchangers. (a) Zipax-SAX (200 x 4.5 m m ID)

column, using 2 mM sodium adipate as eluent. Solute concentrations: 2-20 ppm. Conductivity detection was used. Reprinted from [26] with permission. (b) Zipax-SCX (250 x 4.6 mm ID) column using 2.5 mM copper sulfate as eluent. The peaks are. in the negative direction because indirect spectrophotometric detection was employed (see Ch. 12). Reprinted from [45]with permission.


35

Na*

I

0

I

4

I

0

I

12

I

16

I

20

I

21

1

27

Time (rnin]

Fig 3.6 Separation of monovalent and divalent cations on a poly(butadiene-maleic acid)-coated silica cation-exchange (125 x 4.5 mm ID) column. The eluent was 10 mM tartaric acid, and conductivity detection was employed. Reprinted from [46] with permission.

functionalized silica materials (but not with polymer-coated silica exchangers) to manipulate ion-exchange selectivities or to reduce column fouling by organic sample components. The cost of silica-based IC columns is generally significantly less than for alternative packings. A further advantage of silica-based materials is that the retention mechanism operating is frequently more simple than with other materials because of the low probability of secondary interactions between solute ions and the silica substrate [45]. This means that rctention times are often shorter on silica columns than on other packings having similar ion-exchange capacities. p H limitations A number of serious drawbacks exists with the use of silica-based IC materials. The first of these is the restricted pH range over which the columns can be operated. At pH values below 2.0, the covalent bond linking the ion-exchange functionality to the silica substrate may become unstable and prolonged usage at low pH can result in a progressive loss of ion-exchange capacity as the functional groups are cleaved. Eluents or samples of alkaline pH must also be avoided because of dissolution of the silica matrix itself. The upper limit of pH which can be tolerated by silica-based IC columns varies with the type of column, however severely curtailed column lifetime is often observed when 'eluents of pH greater than 7 are used. This pH limitation is more severe than

36

Chapter3

Zn

0

5

Time (min)

10

Fig. 3.7 Elution of cations from an anion-exchange column. A Vydac 302.IC (250 x 4.1 m m ID) column was used with 2mM potassium phthalate at pH 5.0 as eluent. Detection: conductivity. Solute concentrations: 4 mM for the metal ions. Reprinted fnnn [54]with permission.

that commonly applied to reversed-phase HPLC packing materials, but this can be rationalized by the more exposed nature of the silica surface in the very lightly functionalized IC packings. When this form of column degradation has occurred, column regeneration is not possible [51]. Wide variation in ion-exchange capacities of new silica-based IC columns (from 0.19 to 0.66 mequiv/column for Vydac 302.IC columns) has been reported [52], leading to variation in retention times between columns used under the same eluent conditions. The same authors have devised a simple procedure for compensating for these capacity variations.

Retention of metal ions on silica aninn-exchangers Metal ions are retained on silica-based anion-exchange IC columns and may cause interference with anions by eluting in the same time frame. This phenomenon has been observed for Cu2+,Pb2+and Zn2+ on a Vydac column operated with phthalate eluents in the pH range 4-5 [53]. Under the same conditions, Na+, K+,Ca2+, Mgz+, Ni2+, Mn2+ and Cd2+were unretained, whilst Fe3+,A13+ and Hg2+ were not eluted from the column. Silica itself can act as both an anion- or cation-exchanger, as discussed in Section 3.5. Although silica-based IC columns are often "end-capped" (i.e. reacted with trialkylsilane reagents) to reduce the number of silanol (SOH) sites present at the stationary phase surface, it is to be expected that some cation-exchange capacity would exist when neutral or alkaline eluents are employed. However, studies have shown [54] that metal ion


37

retention is not dependent on the concentration of free silanol groups at the silica surface and it has been proposed that the metal ions showing moderate retention on the silica column are retained by adsorption of their anionic complexes formed with eluent species. Fig. 3.7 shows the elution of metal ions and nitrate in the same chromatogram.

Sample size The final limitation of silica-based IC columns is their susceptibility to overloading. This results in severe distortion of the chromatographic peaks and is a direct result of the combination of low ion-exchange capacity and use of a dilute eluent. When the concentration of solute ions in the sample increases beyond a critical point, the solute ions are no longer confined to a discrete, well-shaped band on the Stationary phase. Some ions move along the column at a relatively faster rate than the main band of solute ions, producing a "fronted" peak at the detector. This limitation is common to any ionexchanger of low capacity and is therefore not specific to silica-based materials.

3.3 RESIN-BASED ION-EXCHANGERS Low-capacity ion-exchange resins were the stationary phases with which nonsuppressed IC was initially introduced to the scientific community by Gjerde, Fritz and Schmuckler in 1979 [l, 551. These materials are produced by chemical derivatization of synthetic organic polymers. Fig. 3.8 shows the manner in which ion-exchange resins may be classified.

I IC ION-EXCHANGE RESINS1

+

Macro porous

Microporous

SURFACE-FUNCTIONALIZED MATERIALS I

I

1

Surface sulfonated

Surface aminated

AGGLOMERATED MATERIALS

binding

binding

U Polystyrenedivinylbenzene

Polymethacryiate

Fig. 3.8 Classification of resin-based ion-exchangers for IC.

Mechanical binding

38

Chapter3

Fig. 3.9 Reaction of styrene (1) with divinylbenzene (2) to produce styrene-divinylbenzene copolymers (3).

Organic materials, in the form of synthetic polymeric resins, are the most widely used types of ion-exchangers. They are manufactured by first synthesizing a polymer with suitable physical and chemical properties, and this polymer is then further reacted to introduce the functional group which acts as the fixed ion in the ion-exchange process. The polymer used as the backbone of the ion-exchange resin must be mechanically stable and pxsess the required degree of insolubility for use in aqueous systems, even after the derivatization process which invariably incorporates polar functionalities onto the resin. Whilst a number of polymers satisfy these criteria, most ion-exchange resins are made from copolymers produced from styrene and divinylbenzene, with a small number consisting of copolymers of divinylbenzene and acrylic or methacrylic acid. 3.3.1

Polymerization reactions

In polystyrene-divinylbenzene(PS-DVB) copolymers, the styrene is cross-linked with itself and with divinylbenzene into a polymeric network, according to the reaction scheme shown in Fig. 3.9. The reaction of methacrylic acid with divinylbenzene to produce cross-linked polymethacrylate resin is shown in Fig. 3.10. Of the two types of copolymer depicted in Figs. 3.9 and 3.10, the PS-DVB is by far the most commonly used. It can be seen from Fig. 3.9 that reaction of styrene with itself produces linear chains, whilst the difunctional divinylbenzene causes these chains to become cross-linked. This cross-linking imparts mechanical stability to the polymer and increases the average molecular weight. The degree of cross-linking is determined by the percentage of divinylbenzene in the reaction mixture. The reaction is carried out by suspending the reactants as droplets in an aqueous phase. Ethylbenzene is usually present in the divinylbenzene and the reaction proceeds via a free-radical chain mechanism. Radical-producing initiators such as benzoyl peroxide are commonly used, together with a polymerization inhibitor (such as sodium dichromate) and a protective colloid (such as inorganic clays, alumina or carboxymethylcellulose) [56]. When the reaction mixture is stirred at an appropriate rate, oil droplets are formed, which on heating polymerize to


CH=CH,

39

-6-

CH,-

I

I

COOH

CH3 CH=CH,

-C-

I

COOH

CH,-CH-CH,-

I I COOH C-

Fig. 3.10 Reaction of rnethacrylic acid (1) with divinylbenzene (2) to produce methacrylatedivinylbenzene copolymer (3).

form the polymer beads. The characteristics of the polymeric resin bead are regulated by the reaction conditions. Particle size is determined to a large degree by the oil-water ratio in the reaction, the rate of agitation, the geometry of the reaction vessel, and the nature of the protective colloid. The resin beads are essentially solid. Resins prepared in this way are described as microporous to indicate that they have low porosity. Microporous resins are relatively rigid (depending on the degree to which they are cross-linked) and have low surface area due to the lack of a significant internal pore structure. The surface area of the resin can be increased enormously if internal pores are introduced into the resin, particularly if these pores are of sufficient size for penetration by solvents. This may bc achieved by conducting the polymerization reaction in a solvent system in which the solvent will dissolve the monomer but tends to precipitate the polymerized material. Alcohols and hydrocarbons are examples of solvents suitable for this purpose with PS-DVB polymerization. When this solvent is removed after polymerization, a proliferous material containing an open, porous structure is produced. Such polymers are called macroporous, or macroreticular (this latter name was originally applied to some of the resins produced by Rohm Rr Haas) to describe their open network structure. Porous resins can also be produced if the cross-links in the polymer are introduced after polymerization by a chemical reaction in the preformed resin bead. Performance of this reaction in the presence of a solvent which will dissolve the monomer and non-cross-linked polymer, leaves a solid resin which is permeated with channels of uniform size. Toluene and xylene are suitable solvents for PS-DVB copolymers. Resins manufactured in this way have been called isoporous. All types of porous resins require a sufficient degree of cross-linking to ensure that the structure does not collapse. Macroporous resins have well-regulated pore sizes and materials can be produced with pore diameters up to lWA. The surface areas of macroporous resins are often in excess of 500 m2/g. Fig. 3.11 shows a schematic representation of a microporous and a macroporous resin bead.

40

Chapter3

Schematic cross-sectionai structure of (a) microporous and (b) macroporous (macroreticular) resin beads.

Fig. 3.1 I

The procedures by which these polymers are converted into functional ionexchangers, and the chromatographic characteristics of the resultant materials, are discussed in the following Sections.

3.3.2

Surface-functionalized

cation-exchaneeresins

Synthesis Conversion of polymeric resins into cation-exchange materials can be achieved by chemically treating the surface of the resin to introduce the functional groups which form the fixed ions of the ion-exchanger. Some of the main classes of functional groups which can be introduced onto polymeric resins were listed in Table 2.1, with H+being shown as the counter-ion in each case. The sulfonic acid functional group produces a strong type of exchanger, whilst the rest are classified as weak. Sulfonic acid groups can be introduced onto PS-DVB copolymers by reaction with sulfuric acid, sulfur trioxide, fuming sulfuric acid or chlorosulfonic acid, as shown in Fig. 3.12. A potential problem with these reactions is to ensure intimate contact between the sulfonating agent and the hydrophobic polymer surface. However, this problem is generally not severe since as the surface of the beads become sulfonated, the bead swells in the aqueous reagent solution, thereby increasing contact with the sulfonating reagent. The end result of this process is that sulfonation occurs in a relatively uniform manner throughout the bead. Sulfuric acid is the most commonly employed sulfonation reagent and is used at elevated temperatures, sometimes with a catalyst. If the reaction is allowed to go to completion, monosulfonation of the aromatic rings occurs, with the functional groups being located in the orrho, mera or puru positions relative to the bond between the ring and the hydrocarbon chain. The benzene rings from both the styrene and the divinylbenzene become sulfonated. Resins with low cross-linking do not become fully functionalized, even under severe reaction conditions, because they do not swell sufficiently to ensure adequate contact with the sulfonating reagent. In these cases, the use of organic swelling agents, such as chlorinated hydrocarbons, results in improved physical properties of the finished product and increased reaction rates. The pre-formed


41

Fig. 3.12 Reaction schemes for the introduction of sulfonic acid functional groups onto styrenedivinylbenzenecopolymers.

polymer bead is swollen by direct contact with the solvent in aqueous suspension, after which the solvent-swollen bead is sulfonated. The solvent leaves the bead during or prior to sulfonation, or may react with the sulfonating agent and in so doing, become a better swelling agent for the sulfonated polymer. Sulfuric acid may itself serve as the slurtying agent during sulfonation, but must be present in excess concentration in order to remove the water produced in the reaction. Organic solvents are generally used as the reaction medium when chlorosulfonic acid or sulfur trioxide are used as the sulfonating agents. Sulfur trioxide may also be employed in the gaseous phase, but this process is only applicable to macroporous resins which do not require the use of a swelling agent. The ion-exchange capacity of sulfonated resins can be regulated by controlling the sulfuric acid concentration and also the reaction temperature and time. Correct design of the reaction apparatus permits the resin-sulfuric acid reaction mixture to be immediately separated on a vacuum-assisted filter system once the prescribed reaction time is completed 1571. The reaction can then be quenched by placing the resin in a large volume of cold water, or by washing the resin with water, ethanol and acetone [58]. Table 3.2 lists the ion-exchange capacities reported for various reaction conditions. Further details of the synthesis of cation-exchangers are available elsewhere [57,59-611. The utility of the above sulfonation procedures is not restricted to PS-DVBresins. For example, phenyl-modified Kel-F (polychlorotrifluoroethylenereacted with phenyllithium) has been sulfonated [62] to give a cation-exchange capacity of 20 pequiv/g.

Chapter3

42 TABLE 3.2

TYPICAL REACTION COhQITIONS FOR THE SYNTHESIS OF SLJLFONATED CATIONEXCHANGE RESINS

Ion-exchange capacity (pequiv/g)

Reaction time

Reaction temperature

(h)

(OC)

XAD-2

9.7

54

6.5% PS-DVB 8% PS-DVB 8% PS-DVB 8% PS-DVB 12%PS-DVB

47

10 1.5 90 90 35 103 103

90

XAD-2 4% PS-DVB

Base resin

88

70

70 25 25

90

-10

80 80 90

80 60 80

[H2S04] (%)

Ref

70 90 96 96

59 59 60

100 100 100

96

60 57 57 57

60

Characteristics The low-capacity cation-exchangers produced by partial sulfonation of PS-DVB have characteristics which differ markedly from classical ion-exchange resins, where the sulfonation reaction is permitted to go to completion. The classical materials attain capacities of about 5 mcquiv/g by the introduction of sulfonic acid groups throughout the macroporous structure of the resin. This results in poor chromatographic mass-transfer characteristics because of the requirement for solute ions to traverse long diffusion paths, thereby leading to the elution of relatively broad bands of solute. Mass-transfer is improved markedly if the sulfonic acid groups are confined to the outer surface of the resin bead, producing a material which is essentially pellicular in nature. This can be achieved by limiting the degree of sulfonation, which at the same time decreases the ionexchange capacity. Such materials are often described as surface-sulfonated cationexchange resins. Several studies have examined the manner in which the sulfonate functional groups are distributed on the resin bead [57, 60.631. Stevens and Small [63] performed an experimental optimization of the depth of the sulfonation layer. Working on the assumption that the bead is composed of a 100% sulfonated layer ( i . e . every benzene ring has one sulfonic acid group) residing on a non-sulfonated core, they found that the optimal thickness of the sulfonated layer was 200 A for a 50 prn bead. Hajos and Inczedy 1571 used secondary electron image scanning electron microscopy to demonstrate that sulfur (from sulfonic acid groups) was present only on the surface of the bead. In a similar experiment, Sevenich and Fritz [60] obtained transmission electron micrographs of thin sections of functionalized resin which had been saturated with uranyl ions in order to provide high electron capture at the functional group sites. This study showed that the sulfonatcd zone consisted of a band of complete sulfonation extending some 200 A into the bead. The density of sulfonate groups in this layer was similar to that existing throughout the entire bead of a typical high capacity, fully sulfonated cation-exchange

Ion-ExchangeStationary Phares

43

Fig. 3.Z3 Schematic representation of the cross-section of a surface-sulfonated cation-exchange resin. The negative charges represent sulfonic acid groups which are located on the surface of the resin bead. Note that the interior of the bead is not sulfonated as occurs in high capacity, fully functionalized materials. resin. These results confirm that the resins are indeed surface-sulfonated and are pellicular in nature. Fig. 3.13 gives a schematic representation of a surface-sulfonated cation-exchange resin. There is a significant side-effect resulting from this partial sulfonation of PS-DVB. The relatively sparse distribution of the functional groups on the resin surface means TABLE3.3 SOME COMMERCIALLY AVAILABLE SURFACE SULFONATED RESIN-BASED CATlONEXCHANGERS USED FOR IC

Polymer

Column

Particle size

Capacity (Pew

Refsa

5 10 12 35 100 560

66-68 69-7 1 72-75 76 77 78 79 80 81-85

(Pm)

Dionex HPlC CS- 1 Dionex HPIC CS-2 Waters 1C Pak C Hamilton PRP-X200 Interaction ION-200 Interaction ION-210 Wescan Cation/R Wescan SACb Aminex A5 Aminex A9 Mitsubishi MCI CPK-08 a This listing is not

PS-DVB PS-DVB PS-DVB PS-DV,B Polyvinyl aromatic PS-DVB PS-DVB PS-DVB PS-DVB PS-DVB PS-DVB

20 15 10 I0 10

5 10 10 13 11 20

40 5000 5000 1700

41, 85

96

comprehensive, but provides some examples of the use of each column. anion- and cation-exchange capabilities.

b This column has both

44

Chapter3

( Li-Cs

Li+

1

la+

l g 2' K'

1[

0.3pS

IHZ

Rb*

L-

0

L

8

12 16 20 Time (min)

L

b

I

I

0

L

I

I

8 12 l i m e lmin)

1

16

Ib)

Fig. 3.14 Typical chromatograms obtained with surface sulfonated resins. (a) Dionex HPICCSI column using 5 mM HCI as eluent and conductivity detection. Reprinted from [64]with permission. (b) Column packed with surface-sulfonated 35-55 pm resin, with 2.5 mM mphenylenediaminedihydrochloride and 2.5 mM HN@ as eluent. Conductivity detection was used. Reprinted from [65] with permission. that a significant portion of the surface remains as unfunctionalized resin, which can contribute to solute retention by simple adsorption effects. This behavior has been shown to be particularly important in the retention of organic cations [59], where solutes such as (m-ninobenzy1)trimethyl-ammoniumions are retained by a combination of ionexchange and surface adsorption processes. Under some circumstances, surface adsorption may also play an important role in the retention of inorganic cations, especially when they are chromatographed as complexes with an organic ligand. Low-capacity surface-sulfonated cation-exchange relrins have proved to be very successful in routine applications. Fig. 3.14 shows typical separations of monovalent or divalent cations obtained with these materials. Some commercially available IC resins of this type are listed in Table 3.3; all the materials shown exhibit similar chromatographic selectivities because the functional group is the same in each case.

Ion-ExchangeSmtionary Phases

45

Fig. 3.25 Two step reaction scheme for the production of anion-exchangers from styrenedivinylbenzenecopolymers.

3.3.3 Surface-functionalized anion-exchange resins Synthesis

Reaction of PS-DVB copolymer to produce a strong-base anion-exchange resin generally proceeds via chloromethylation, which can be accomplished readily using chloromethylmethylether in the presence of a suitable catalyst. This reagent also serves as a swelling agent which promotes intimate contact between the resin and reagents, leading to more uniform and rapid reaction. The reaction may be carried out at moderate temperature (58 OC) with a mild catalyst such as zinc chloride, or at lower temperatures with more active catalysts. This procedure results in high yields, but has the disadvantages of the extreme toxicity of chloromethylmethylether, and difficulty in controlling the degree of chloromethylation. After chloromethylation, a second reaction with an amine produces the required anion-exchange material. The complete reaction scheme is shown in Fig. 3.15. The ion-exchange capacity is regulated both by the time permitted for the chloromethylation reaction to continue and by the reaction temperature. Table 3.4 shows some capacities obtained by reacting 3 g portions of various resins with 10 ml of chloromethylmethylether, 10 ml of methylene chloride, 3 ml of nitromethane and 3 g of zinc chloride at room temperature for varying amounts of time. These results show clearly the dependence of the reaction on the type of base polymer employed. Whilst Fig. 3.15 shows the method for the introduction of quaternary ammonium functionalities onto a PS-DVB polymer, the same reactions are applicable to other types of polymer with an aromatic moiety. The type of amine used in the second step of the above synthesis determines the nature of the functional group formed on the resin. Some typically used amination reagents and the functional groups formed on the resin are listed in Table 3.5. Alternative chloromethylation procedures have been reported in which the use of highly carcinogenic chloromethylmethyletheris avoided. Barron and Fritz [88] used paraformaldehyde and aqueous hydrochloric acid in the absence of a catalyst. This approach simplifies control of the reaction and eliminates secondary cross-linking reactions found in the presence of Lewis acid catalysts. Furthermore, reaction in

46

Chapter 3

TABLE 3.4 ANION-EXCHANGE CAPACITIES OF POLYMERS CHLOROMETHYLATEDFOR VARYING AMOUNTS OF TIME. DATA TAKEN FROM REFS [ 11 AND [87] Resin

Surface area (m2/g)

XAD- 1

Pore diameter

(Angstroms)

Chloromethylation time (min)

205

100

XAD-2

300

90

XAD-4

784

50

Capacity (wquiv/g)

7 which will not undergo any significant hydrolysis and therefore will not lead to significant changes in bdseline conductance as their concentration in the

118

Chapter4

11

19

L

r

0

I

5

I

10

I

15

1

20

I

25

1

30

Time (min)

Fig. 4.23 Gradient elution using conductivity detection and a high capacity suppressor. A Dionex HPIC-ASSA column was used. The eluents used were eluent A, 0.75 mM NaOH and eluent B, 100 mM NaOH. The gradient program is shown below. Peak identities: 1= fluoride, 2 = ahydroxybutyrate, 3 = acetate, 4 = glycolate, 5 = butyrate, 6 = gluconate, 7 = a-hydroxyvalerate, 8 = formate, 9 = valerate, 10 = pyruvate, 1 1 = monochloroacetate, 12 = bromate, 13 = chloride, 14 = galacturonate, 15 = nimte, 16 = glucuronate, 17 = dichloroacetate, 18 = trifluoroacetate, 19 = phosphite, 20 = selenite, 21 = bromide, 22 = nitrate, 23 = sulfate, 24 = oxalate, 25 = selenate, 26 = a-ketoglutarate, 27 = fumarate, 28 = phthalate, 29 = oxaloacetate, 30 = phosphate, 31 = arsenate, 32 = chromate, 33 = citrate, 34 = isocitrate, 35 = cis-aconitate, 36 = trans-aconitate. Reprinted from [135] with permission. 0 5 15 30 Gradient program Time(min) %A 100 100 70 14 %B 0 0 30 86

suppressed eluent builds up. Using this criterion, suitable eluents for gradient elution include NaOH, sodium tetraborate, salts of phenates (such as p-cyanophenate) and anions forming zwitterions in the suppressor. It is noteworthy that the HC03-/C032- buffers, which have proved so versatile for isocratic elution in suppressed IC, have limited suitability for gradient elution because the suppressor product (H2CO3) has pK,1 = 6.35 and so is too strong an acid. However, it has been shown [I551 that a porous polypropylene tubular membrane (called a "post-suppressor"), incorporated between the suppressor and the detector, results in removal of dissolved C 0 2 and hence permits gradient elution with carbonate buffers. In addition, gradients which involve only small

Eluents for Ion-ExchangeSeparations

119

increases in the concentration of a carbonate buffer have been used [ 1581. Sodium hydroxide is a more useful eluent for gradient elution because the suppressor product is water, regardless of the original concentration of NaOH in the eluent. The anion micromembrane suppressor can fully suppress 100 mM NaOH at a flow-rate of 1.0 ml/min [129], so concentration gradients up to this level are possible. Fig 4.23 shows the separation of 36 anions using a 0.75-100 mM NaOH gradient. The major problem with NaOH as an eluent for gradient elution is contamination by C032produced from the absorption of carbon dioxide from the atmosphere. This can cause a drift in the conductance of the suppressed eluent. NaOH gradients are therefore successful only when suitable precautions are taken in eluent preparation and storage. Indeed, eluent purity is an inherent problem in gradient elution because even trace levels of impurities build up at the head of the column in the early portion of the gradient and are eluted only when the eluent concentration increases.

Isoconductive gradients An alternative approach to gradient elution with conductivity detection is the use of two eluents which have different elution strengths, but the same background conductance. Such eluents may be described as isoconductive an'd the use of these eluents has been reported for anion-exchange separations in non-suppressed IC [ 157, 1591. The background conductance of an anion-exchange eluent formed from a competing anion E- and a cation M+ is calculated by taking into account the limiting equivalent ionic conductances and concentrations of both E- and M+ (see Section 9.2). It follows that two eluents formed from the same concentration of E-, but with different cations, will have different conductances. Viewed another way, it should then be possible for two eluents comprising different concenlrations of E and different cations, to have the same conductance, i.e. to be isoconductive. The weaker eluent used at the start of the gradient has a low concentration of E- in the presence of a highly conducting cation, such as MI+. On the other hand, the stronger eluent used later in the gradient has a higher concentration of E- in the presence of a less-conducting cation such as M2+. Fig. 4.24 gives a schematic representation of an isoconductive gradient, and Fig. 4.25 shows a gradient separation of anions using this method. The limitation of isoconductive gradients is that only a relatively small range in the concentration of competing anion is accessible, In other words, the difference in eluting strengths between the starting and finishing eluents in the gradient is quite small. This can be illustrated by considering two isoconductive eluents, the first (weaker eluent) containing c1 mM of the salt MIE, and the second (stronger eluent) containing c2 mM of , as in the salt M2E. The attainable gradient strength is given by the ratio C ~ C I calculated eqn. (4.21) [ 1571. (4.21)

Chapter 4

Eluent A

Eluent B

MlE c1 mM

M2E c2mM

A

B

Eluent

.

Fig. 4.24 Schematic representation of the composition of an isoconductive gradient for anionexchange separations. The two isoconductive eluents are formed from MIE (cl mM) and M2E (c2 mM), where the eluent competing anion is E-, MI+is a strongly conducting cation, and M2+ is a weakly conducting cation. Modified from [ 1591.

1

0

~

~

~

'

5

"

"

'

Time ( m i d

l

10

'

'

'

'

1s

l

Fig. 4.25 Anion-exchange separation using an isoconductive gradient. A Waters IC Pak Anion HR column was used. The column was equilibrated with eluent A (8.25 mM boric acid, 1.11 mM gluconic acid, 3.08 mM cesium hydroxide, 0.48 mM glycerin and 12% acetonimlef and a step gradient to eluent B (12.65 mM boric acid, 1.70 mM gluconic acid, 4.72 mM lithium hydroxide, 0.75 mM glycerin and 12% acetonitrile) was initiated at the moment of injection. Note that with the pump used, there was a considerable lag time before the gradient reached the column. Reprinted from [ 1571 with permission.

Eluentsfor Ion-ExchangeSeparations

121

where X represents the limiting equivalent ionic conductance. The gradient strength is therefore dependent chiefly on the difference in X values between the cations MI+and M2+, and also on the X value for the eluent anion. Values of X for Rb+, K+,Li+ and tetrabutylammonium (TBA+) are 78, 73,37 and 20, respectively. Typical combinations of cations for isoconductive gradients would therefore be Rb+/TBA+ or K+/Li+. The gradient strengths attainable with these cation combinations are shown in Fig. 4.26 for a range of different eluent anions. It can be seen that isoconductive gradients formed from Rb+ and TBA+ salts of the gluconate-borate complex anion offer the highest gradient strength for any of the eluent anions shown.

Baseline balancing methods Gradients with conductivity detection are possible if careful steps are taken to balance the changing conductance of the eluent in some way. Two methods by which this may be achieved have been suggested. In the first method, a step gradient is used between two carbonate buffer eluents which give identical concentrations of H2CO3 in the suppressor, and hence identical background conductances [156]. The simplest case is to use equimolar concentrations of NaHC03 and Na2C03 as the two eluents and the gradient is achieved as a result of the greater eluting strength of the C032- eluent. This approach can be extended to the use of NaHC03/Na2C03 buffers as the two eluents, provided that the total molarities of the two eluents are the same. The second method of baseline balancing is to counteract the increasing conductance encountered during the gradient with another species added to the weaker

‘n 3

RblTBA

Limiting equivalent ionic conductance I X ) of eluent anion Fig. 4.26 Isoconductive gradient strengths (given by cz/cl - see text) attainable for Rb+/TBA+ and K+/Li+ cation combinations with different eluent anions. Reprinted from [159] with

permission.

122

r 0

Chapter 4

lil

17

I

5

I

10 Time (min)

I

I

15

20

Fig. 4.27 Gradient elution by suppressed 1C with baseline balancing using borate-mannitol. A Dionex HPIC-AS6 column was used with a gradient of ammoniump-cyanophenate. Mannitol is added to the eluent and borate forms pan of the regenerant in the suppressor. The following eluents were used. Eluent A, 35 mM pcyanophenol, 50 mM ammonium hydroxide, 2% acetonitrile; eluent B, 50 mM mannitol, 2% acetonitrile; eluent C, 2% acetonitrile. The gradient program is shown below. Peak identities: 1 = 2 = acetate, 3 = formate, 4 = pyruvate, 5 = monochloroacetate, 6 = BrOg-, 7 = Cl-, 8 = NOz', 9 = HP042-, 10 = HA SO^^-, 11 = glutarate, 12 = succinate, 13 = maleate, 14 = S042-, 15 = NOg-, 16 = oxalate, 17 = fumarate, 18 = trichloroacetate, 19 = oxaloacctate, 20 = pyrophosphate, 21 = citratc, 22 = iswitrate. Reprinted from [I351 with permission. Gradient program: Time(&) 0 3 3.1 7 13 15 53 100 %A 30 53 7 15 40 30 25 0 %B 45 40 17 22 0 30 %C 48 45 F-9

eluent. One successful approach to this in suppressed IC is to add mannitol to the weaker eluent, whilst at the same time adding boric acid to the regenerant [135]. Mannitol is a neutral polyalcohol and has little effect on either separation or detection, but can react with neutral boric acid permeating from the suppressor regenerant into the eluent to form a conductive anionic species. If the mannitol concentration entering the suppressor is decreased in inverse proportion to the increase in eluent concentration in the gradient, a steady baseline can be achieved. Fig. 4.27 shows a separation achieved with such a gradient. Baseline balancing can also be accomplished with a computer which automatically subtracts the baseline signal for a blank gradient from the conductance signal produced during the gradient separation of a sample.


123

INJECTION PEAK

I

0

I

I

G

I

I

8

I

Time Imin)

I

12

I

1

16

Fig. 4.28 Illustration of the injection and system peaks observed in non-suppressed IC.

4.6 EXTRANEOUS (SYSTEM) PEAKS IN ION-EXCHANGE IC 4.6.1

Introduction

When a solute is injected into an IC system, additional peaks sometimes appear in the chromatogram. These peaks, which are often detectable only with certain types of detectors, are referred to by names such as system peaks, pseudo peaks, ghost peaks, vacuncy peaks or inducedpeuks. Most analysts treat these peaks as a nuisance because of the possibility of interference with solute peaks.

4.6.2 Extraneous peaks in non-suppressed IC System peaks are most commonly observed in anion-exchange non-suppressed IC, where the phenomenon can be seen by looking at a chromatogram resulting from the injection of a solute Na+S- into an eluent formed from a weak acid, HE. Fig. 4.28 shows that a peak directly attributable to the solute (in this case, nitrite) is observed, together with two additional peaks. The first of these is eluted at the column void volume and will be referred to as the injection peak (sometimes also called the solvent peak). The second additional peak is eluted later in the chromatogram and will be referred to as the system peak. It should be emphasized that the injection peak occurs in almost every chromatogram, whilst the system peak is present only when the eluent and sample meet certain conditions, and only with some detection methods, Both the injection and system peaks can be positive or negative in direction. The origins of these peaks have been studied by numerous authors [31. 58, 1601671 and some of the conclusions reached are summarized below. For simplicity, we will consider only the situation where a conductivity detector is used with an anionexchange IC system.

chapter 4

124

The injection peak We assume that the sample consists of an aqueous solution of Na+S-and the column is equilibrated with an eluent HE, containing the competing anion, E-. When the sample reaches the head of the column, the solute anions become adsorbed onto the stationary phase by displacing E- ions. The sample band therefore contains Na+ ions and E-, together with some HE. This band moves through the column and since it has a different composition to that of the bulk eluent, gives rise to a detector signal. The resultant injection peak has the following characteristics [166,167]: The retention time is constant regardless of the eluent pH or the concentration of the injected solute. (ii) The conductance of the injection peak increases in proportion to the concentration of the injected solute. (iii) The conductance of the injection peak relative to that of the eluent shows little variation with changes in eluent composition. (iv) The conductance of the injection peak may be greater or less than that of the eluent, depending on the sample concentration and the eluent composition. The direction of the injection peak can therefore be positive or negative.

(i)

The injection peak can be attributed to the solute cations which are excluded from the anion-exchange resin, together with displaced eluent anions. Hershcovitz et al. 11671 have used characteristic (ii) above to show that when a mixture of solute anions is injected, the area of the injection peak is related stoichiometrically to the combined areas of the solute peaks. They have demonstrated that it is possible to predict accurately the area of a nominated solute peak if the areas of the injection peak and the other solute peaks are known. Further studies [ 1681 have indicated that the injection peak can be used for a range of analytical purposes, which are discussed in Section 9.6.4.

The system peak The following characteristics have been noted for the system peak in nonsuppressed IC [ 160, 1661. The retention time of the system peak is dependent on the eluent composition and on the type of stationary phase used in the column. (ii) The height of the system peak varies with the eluent pH and is also dependent on the difference in pH between the eluent and sample. (iii) The height of the system peak depends on both the volume and concentration of the injected sample.

(i)

These characteristics indicate clearly that the system peak is the result of some kind of disturbance of column equilibria caused by injection of the sample. The fact that the size of the system peak is greater for some solutes than others has led to the erroneous conclusion by some early workers that the system peak was due to the elution of a specific anion. This confusion is particularly evident when HCO3- is the solute and an eluent of pH=5 is used. Under these conditions, HCO3- forms H2CO3 and is therefore


Hydrophobic surface

125

Ionexchange site

Fig. 4.29 Schematic illustration of eluent equilibria at the surface of an anion-exchange stationary

phase. unretained, appearing in the chromatogram as part of the injection peak. The latereluted system peak may therefore be mistaken easily for the HCO3- peak. There are several schools of thought regarding the origin of the system peak, but the essential aspects of most of these can be incorporated into the following model. The sample injection causes a disruption of the equilibria existing between eluent species in the flowing solution and eluent species adsorbed onto the stationary phase. This is followed by a relaxation process which results in a decrease in the velocity of the solute (which initially enters the column at the velocity of the mobile phase) towards the equilibrium velocity dictated by the distribution coefficient for that particular solute. At the same time, the velocities are also decreased for mobile phase components whose concentrations are different to those present in the bulk eluent. We note that both an excess or deficiency of an eluent component (relative to the concentration of that component present in the bulk eluent) will move through the column at the same speed, which is determined by the distribution coefficient for that particular species. The final result is a chromatogram which contains a peak for the solute, as well as peaks for some or all of the eluent components. This effect will be observed for any chromatographic system in which the eluent contains more than one component [163, 1641 and we can expect peaks to appear at the characteristic retention times for each eluent component. The column equilibria are represented schematically in Fig. 4.29. All samples, including water, will cause some disturbance of these equilibria. For example, sample injection may cause a change in the concentration of an anionic competing ion in the eluent. This change may be an increase or a decrease in concentration of the competing anion, relative to that in the bulk eluent. However, a decrease is the more probable outcome. The changed concentration of competing ion will appear at the characteristic ion-exchange retention time for the competing anion [165]. This mechanism is readily illustrated using a phthalate eluent which also contains some C1-. Injection of any solute will produce a peak for that solute, as well as a peak at the retention time of C1- [165, 1691. The same model can apply if we consider an eluent which contains some of the neutral eluent acid (i.e. HE in the example under consideration). Adsorption of HE onto

126

Chapter4

the unfunctionalized parts of the ion-exchange material can be expected to occur (see Fig. 4.29). It should be remembered here that PS-DVB resins show strong reversedphase properties and there is ample evidence for adsorption of neutral or ionic eluent components onto the resin [170-1721. There is an equilibrium concentration of HE both on the resin and in the eluent. These concentrations are altered by sample injection and once again, there follows a relaxation process which produces a change in eluent composition at the detector. The retention time of this change (and hence the system peak) is determined by the retention time of the neutral eluent species, HE [26, 1601. Eluents formed from aromatic carboxylic acid salts are particularly prone to system peaks, but these are generally eluted late in the chromatogram where interference with solute peaks is minimal. System peaks can be eliminated with these eluents by adjusting the pH so that the eluent acid is completely ionized (e.g. phthalate eluents above pH 6.5). Under these conditions, the eluent therefore contains a single competing anion. Apart from the above general mechanism, some system peaks can be attributed to specific chemical interactions existing in certain eluents. For example, gluconate-borate eluents give a negative system peak when Ca2+ is present in the injected sample. This peak has been assigned to an anionic complex of calcium with a borate-gluconate diester present in the eluent [%I. The complex is retained by an anion-exchange mechanism and being less conducting than the eluent itself, gives a negative baseline change. Removal of Ca2+ from the sample with an ion-exchange pre-column was shown to eliminate this system peak. 4.6.3 Extraneous peaks in suppressed IC Extraneous peaks also appear in the chromatograms produced in suppressed IC. These occur despitc the fact that the eluent composition is normalized to some extent after passage through the suppressor. Fig. 4.30 shows typical peaks appearing in the early part of a chromatogram using suppressed IC with a carbonate-bicarbonate eluent [ 1731. It can be seen that injection of a HC03-/C032- sample of higher concentration than the eluent produces two positive peaks (Fig. 4.30(a)), whereas samples of lower concentration than the eluent produce two negative peaks (Fig. 4.30(c) and (d)). Samples with the same concentration as the eluent show no peaks (Fig. 4.30(b)). It is the second and larger of the two extraneous peaks which can cause problems in suppressed IC, chiefly by distorting the peaks due to early eluted solutes such as F- and C1-. This peak is most often negative in direction since most samples in IC contain relatively low levels of HCO3- and C032-. For this reason, it is often referred to as the water dip. The water dip is produced by the same mechanism as that encountered in non-suppressed IC for the formation of the injection peak. That is, injection of the sample onto the column results in adsorption of the solute anions of the sample and dcsorption of the competing anions, HCO3- and C032-, from the stationary phase. The injected volume therefore consists of HCO3- and C032-, along with the cations from the sample. This band passes through the separator column to the suppressor, where the usual suppressor reactions occur. l h a t is, the sample cations are exchanged for H+ and H2CO3 is formed. The concentration of H2CO3 produced in this manner is directly proportional to the concentration of adsorbable solute anions in the sample, and the water dip can therefore also be termed the ionic concentration peak [ 1731. The second,

Eluents for Ion-Exchange Separations

0-

127

Retention volume fmlJ

Fig. 4.30 Extraneous peaks produced from injection of Na~C03-NaHC03mixtures into a suppressed IC system with a packed-bed suppressor. The eluent is 3 mM NaHC03 + 2.4 mM Na2C03. The total injected concentrations were (a) 10.8 mM, (b) 5.4 mM, (c) 2.7 mM, (d) distilled water. Reprinted from [I731 with permission.

smaller peak in Fig. 4.30 has been attributed tentatively to the temperature change resulting from the heat of neutralization produced as the sample encounters the first part of the suppressor, which is rich in hydrogen ions [173]. A further complication arises when a packed-bed suppressor column is used. The retention time of the band comprising the sample solvent, the sample cations and the displaced HCO3- and C032- in the suppressor column will depend slightly on the degree of depletion of the suppressor. The reason for this is that the band will be unretained (because of Donnan exclusion) on the parts of the suppressor which have been depleted of H+ ions, but as soon as the active zone in the suppressor is reached and the suppression reaction occurs, the H2CO3 produced can be adsorbed by the resin. This occurs because H2CO3 is a neutral species and there is no Donnan exclusion effect to prevent its retardation. Keeping in mind that it is this H2CO3 which leads to the water dip, then it can be appreciated that the actual elution time of the water dip will change (and hence interference effects will vary) with the degree of suppressor depletion. This effect does not occur with fibre or micromembrane suppressors which are continuously regenerated, since their degree of depletion is constant for a specified eluent and flowrate.

Chapter4

128

4.7 1 2 3 4 5 6

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Eluents for Ion-ExchangeSeparations 122 123 124 125 126 127 128 129 130 131 131 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 151 152 153 154 155 156 157 158 159 160

161 162 163

131

Dionex Technical Note 24. Sevenich G.J. and Fritz J.S., Anal. Chem., 55 (1983) 12. Haddad P.R., Alexander P.W. and Trojanowicz M., J . Chromatogr., 294 (1984) 397. Haddad P.R., Alexander P.W. and Trojanowicz M.. J . Chromatogr.. 324 (1985) 319. Yan D. and Schwedt G.,Fres. Z . Anal. Chem., 320 (1985) 121. Sato H. and Miyanaga A., Anal. Chem., 61 (1989) 122. Weiss J., Handbook of ion Chromatography, Dionex Corporation, Sunnyvale, CA, 1986. Stillian J., LC, 3 (1985) 802. Bouyoucos S.A., J. Chromatogr., 242 (1982) 170. Nonomura M., Anal. Chem., 59 (1987) 2073. Watanabe T., Nagaoka M. and Yamazaki S., Ryusan to Kogyo, 38 (1985) 162. Stevens T.S., Davis J.C. and Small H., Anal. Chem., 53 (1981) 1488. Small H., Stevens T.S. and Bauman W.C., Anal. Chem., 47 (1975) 1801. Rocklin R.D., Pohl C.A. and Schibler J.A., J . Chromatogr., 411 (1987) 107. Zolotov Y.A., Shpigun O.A., Pazukhina Y.E. and Voloshik I.N.,Int. J. Environ. Anal. Chem., 31 (1987) 99. Ivey J.P., J. Chromatogr., 287 (1984) 128. Pohl C.A. and Johnson E.L., J. Chromatogr. Sci., 18 (1980) 442. Slingsby R.W. and Riviello J.M., LC, 1 (1983) 354. Mulik J.D., Estes E. and Sawicki E., in Sawicki E., Mulik J.D. and Wittgenstein E., (Eds.) Ion Chromatographic Analysis of Environmental Pollutants, Vol. I, Ann Arbor Sci. Publ., Ann Arbor, MI, 1978, p. 41. Wimberley J.W., Anal. Chem., 53 (1981) 2137. Dionex Application Note 17. Nordmeyer F.R., Hansen L.D., Eatough D.J., Rollins D.K. and Lamb J.D., Anal. Chem., 52 (1980) 852. Lamb J.D., Hansen L.D., Patch G.G.and Nordmeyer F.R., Anal. Chem., 53 (1981) 749. Franklin G.O., Am. Lab., 17 (1985) 65. Kifune I. and Oikawa K., Niigara Rihgaku, 5 (1979) 9. Jenke D.R., Mitchell P.K. and Pagenkopf G.K., Anal. Chim. Acta, 155 (1983) 279. Elchuk S. and Cassidy R.M., Anal. Chem., 51 (1979) 1434. Heberling S.S., Riviello J.M. Shifen M. and Ip A.W., Res. & Dev., September (1987) 74. Mazzucotelli M., Dadone A., Frache R. and Baffi F., Chromatographia, 15 (1982) 697. Wang W., Chen Y.and Wu M., Analyst (London), 109 (1984) 281. Dionex Technical Note 23. Ramstad T. and Weaver M.J., Anal. Chim. Acta, 204 (1988) 95. Yamaguchi H., Nakamura T., Hirai Y. and Ohashi S., J. Chromatogr., 172 (1979) 131. Shintani H. and Dasgupta P.K., Anal. Chem., 59 (1987) 802. Sunden T., Lindgren M., Cedergren A. and Siemer D.D., Anal. Chem., 55 (1983) 2. Jones W.R., Jandik P. and Heckenberg A.L., Anal. Chem., 60 (1988) 1977. Tarter J.G, Anal. Chem., 56. (1984) 1264. Jones W.R. and Jandik P., Res. & Dev., September (1988) 92. Jackson P.E. and Haddad P.R., J . Chromatogr., 346 (1985) 125. Brandt G., Vogler P. and Kettrup A. , ires. Z. Anal. Chem., 325(1986) 252. Brandt G., Matuschek G. and Kettrup A.. Fres. Z. Anal. Chem., 321 (1985) 653. Levin S. and Grushka E., Anal. Chem., 58 (1986) 1602.

132

Chapter4

164 Levin S. and Grushka E., And. Chem., 59 (1987) 1157. 165 Papp E., J . Chromutogr., 402 (1987) 211. 166 Okada T. and Kuwamoto T., Anal. Chem., 56 (1984) 2073. 167 Hershcovitz H., Yarnitzky C. and Schmuckler G., J . Chromurogr., 244 (1982) 217. 168 Strassburg R., Fritz J.S.,Berkowitz J. and Schmuckler G., J . Chrornatogr.. 482 (1989) 343. 169 Hertz J. and Baltensperger U.. LC,2 (1984) 600. 170 Afrashtehfar S. and Cantwell F.F.. Anal. Chem., 54 (1982) 2422. 171 Cantwell F.F. and Puon S., Anal. Chem., 51 (1979) 623. 172 Hux R.A. and Cantwell F.F., Anal. Chem., 56 (1984) 1258. 173 Doury-Berthod M.. Stammose D. and Poitrenaud C., Reucr. Polymers, 2 (1984) 37.

133

Chapter 5 Retention Models for Ion-Exchange 5.1

INTRODUCTION

The equilibria which govern ion-exchange separations are well understood, and detailed retention models providing a quantitative description of the factors which govern solute retention can therefore be developed for the various types of ion-exchange methods discussed in the previous two Chapters. For the purpose of clarity, these models are discussed together in this Chapter, along with experimental data used to validate each model.

5.2 RETENTION MODELS FOR ANION-EXCHANGE 5.2.1 Single eluent competing anion

The most straightforward situation in anion-exchange IC is where the eluent contains a single type of competing anion. Derivation of a retention model proceeds as follows [1-6]: The ion-exchange equilibrium for binding of a solute anion, AX-. to a stationary phase which has been conditioned with an eluent containing a competing anion, Ey-, is given by: (5.1)

where the subscripts m and r refer to the mobile and stationary phases, respectively. The selectivity coefficient for the system is given by:

where the parentheses represent the activity of the particular species. When activity coefficients are included, eqn. (5.2) becomes:

(5.3)

134

Chapter 5

As the ensuing discussion will show, i t is generally not necessary to consider activity effects in IC because of the very dilute eluents and low sample concentrations which are routinely used. Moreover, the inclusion of activity factors would require knowledge of the activities of species in the resin phase, and these cannot be determined. For these reasons, and also in the interests of simplicity of presentation, no further reference to activity coefficients will be made in the subsequent derivations. However, the reader should be aware of the implied assumption that activity coefficients are very close to unity in all the cases to be considered. Notwithstanding these comments, there will be a number of occasions on which it will be necessary to reconsider activity effects. The weight distribution coefficient for solute A is designated as DA and is given by:

In general, the weight distribution coefficient is related to the capacity factor (kA') for solute AX-by the expression:

where w is the weight of the stationary phase and V m is the volume of mobile phase. From eqns. (5.4) and (5.5) we obtain:

Substituting into eqn. (5.3),and neglecting the activity coefficients, gives:

If we assume that the eluent ion, Ey-, occupies y ion-exchange sites on the stationary phase (we will re-evaluate this assumption at a later stage), then the ion-exchange capacity of the column, Q, is given by:

Eqn. (5.7) now becomes:

Retention Models for Ion-Exchange

135

Rearranging gives:

(5.10) Taking logarithms provides the relationship: (5.1 1)

Eqn. (5.11) is of fundamental importance in IC, since it provides a quantitative relationship between capacity factor and some measurable column and eluent parameters. If an experiment is conducted in which the concentration of competing anion in the eluent is varied and the column and solute anion are the same, then KA,E, Q, w and Vm can be considered to be constant. Eqn. (5.1 1) therefore reduces to: (5.12)

where C1 is a constant, Eqn. (5.12) predicts that when the results of the above experiment are presented as a plot of log k' versus log [EmY-1, a straight line should result with a negative slope equal to x/y. We can now consider the ion-exchange equilibrium existing between a second solute ion, Bz-and the same eluent competing anion, Ey-. This equilibrium can be written: ZYzx B m + zEr % x B r

+ zE,Y-

(5.13)

where the subscripts r and m have the same meanings as previously used. The selectivity coefficient for solute BZ-is K B ~which , is given by:

We can also write the ion-exchange equilibrium existing between the two solutes AX-and B", as follows: (5.15)

and the selectivity coefficient between the two solute anions is given by:

Chapter5

136

(5.16)

Eqn. (5.6) can be rewritten for solute B2- and rearranged to give: (5.17)

The chromatographic separation factor for solutes AX- and B2- is given by which is calculated according to:

aA3.

(5.18)

Eqn. (5.18) can be manipulated and rewritten as:

(5.19)

Substitution of eqns. (5.16) and (5.17) into eqn. (5.19) gives:

(5.20)

In a similar manner we can obtain: x-2

(5.21)

Eqn. (5.20) can be rewritten in a logarithmic form to give: (5.22)

When the two solutes have the same charge, i.e. x = z, eqn. (5.22) reduces to:

Retention Models for ion-Exchange

137

Eqns. (5.11). (5.12). (5.22) and (5.23) are of great significance to IC and allow the following predictions to be made for the situation in which only a single type of competing anion is present in the eluent: When a solute anion, AX-. is eluted with a competing anion, Ey-, eqn. (5.11) shows that the capacity factor for AX- is determined by the selectivity coefficient, KA,E, the ion-exchange capacity of the column, Q, the ratio of stationary to mobile phases, WNm, and the concentration of competing anions in the eluent, [EYm]. Increases in KA,E,Q or wNm lead to increased capacity factors. whilst increasing [EY'm] leads to decreased capacity factors. Increased eluent charge leads to decreased capacity factors, whilst increased solute charge leads to increased capacity factors. Under conditions where the same solute anion, AX-.is chromatographed on the same column with varying cbncentrations of competing anion (Ey-) in the eluent, then eqn. (5.12) predicts that a plot of log kA' versus log [Epm] will be a straight line with a slope equal to -x/y. From eqn. (5.231, it can be concluded that the separation factor, (XA,B, for two anions of the same charge depends only on the selectivity coefficient KA,B and on the charge of the solute anions. Providing that the selectivity coefficient remains constant, the separation factor does not depend on the concentration, charge or type of competing anion in the eluent. When two solutes have different charges, the separation factor depends on the capacity factor of one of the solute anions (note that the capacity factors of the solute anions are not independent). The capacity factor can be altered by changing the parameters listed in (i) and (ii) above. We can now examine the validity of these predictions.

Effects of KA,E,Q and wN, By definition, an increase in KA,E indicates enhanced binding of the solute anion to the ion-exchange stationary phase, which must result in increased capacity factors. Similarly, the phase ratio, W N m , has an obvious effect on capacity factor, with higher phase ratios giving longer retention. This explains why pellicular or surfacefunctionalized anion-exchangers provide smaller values of k' than porous, fully functionalized materials. The effect of resin capacity, Q,is of particular importance to IC. If it is assumed that the selectivity coefficient does not change significantly with Q (and this has been shown to be a valid assumption [7]) and the phase ratio is assumed constant, then the following proportionality holds: (5.24)

138

Chapter 5

41

3

3

Y 2

4

Slope -2 Slope -1

1

Slope -1 Slope - 0 . 5

Fig. 5.1 Plots of log k’ versus log [EYarn] for different combinations of singly and doubly charged solute and competing anions.

That is, constant capacity factors can be obtained when Q is decreased only by proportionally decreasing the concentration of competing anions in the eluent. Experimental evidence supports this relationship [7]. It is for this reason that the dilute eluents required in 1C are applicable only to columns of low ion-exchange capacity.

Effect of charges on solute and competing anions As the charge on the solute anion increases, electroselectivity effects suggest that the solute will be more strongly held on an anion-exchanger (i.e. KA,E increases). This effect can be seen from Fig. 4.1, where sulfate had a longer retention time than the monovalent solute anions. By analogy, increasing the charge on the eluent competing anion will cause a reduction in Ir‘ because KA,Edecreases. These effects are evident from eqn. (5.10), which can be simplified by assuming that the column capacity and phase ratio are constant, to give the proportionality:

k,’ a

KA, -

(5.25)

lE~Jx’y

Isocapacitivc elucnts (i.e. those giving identical capacity factors) can be produced by simultaneously increasing the charge and decreasing the concentration of the eluent competing anion, or vice-versa.

Retention Moakls for ton-Exchange

139

log [OH-]

Fig. 5.2 Plots of log kA‘ versus log [EY-m] for NaOH eluents of varying concentration. A

Dionex HPIC-ASSA column was used. Reprinted from [S] with permission.

Effect of [EJ’-m] Eqn. (5.12) shows that the manner in which k‘ is affected by the concentration of competing anions in the eluent is dependent on the charges on both the solute and competing anions. Prediction (iii) above is that a plot of log k’ versus log [EY-m] should be a straight line with a slope of -x/y. The magnitude of the intercept of such a plot is determined by KA,E,Q and wN,. Fig. 5.1 shows schematic plots for singly and doubly charged solute anions, eluted with singly or doubly charged eluent anions. It can be seen that changing the concentration of a competing anion has a greater effect on the retention of a doubly charged solute in comparison to a singly charged solute, and that singly charged competing anions cause the largest changes in retention. The behaviour illustrated in Fig. 5.1 has been verified in numerous practical studies [e.g. 1, 2, 3, 5, 6 , 8, 91. A typical plot of log k’ versus log [Em] is shown in Fig. 5.2. The slopes of the plots are generally quite close to those predicted from eqn. (5.12), and the observed differences between theory and practice can be attributed to the assumptions made in the derivation. This is especially true of activity effects, which are most evident for the triply charged citrate ion. Under the conditions used in Fig. 5.2, activity coefficients vary between 0.755 - 0.900 for monovalent anions, between 0.355 - 0.660 for divalent anions, and between 0.1 15 0.405 for trivalent anions [5]. It is therefore not surprising that citrate shows a large deviation from the slope of -3 predicted by eqn. (5.12). 4

Chapter 5

140

5.2.2 Multiple eluent competing anions Many anion separations in IC use an eluent which contains more than one type of competing anion. Sometimes two quite separate components are added to the eluent, but it is more common that the multiple eluent species are different dissociated forms of the same weak acid. For example, carbonate buffers contain both HCO3-and C0s2- as competing anions, whilst phthalate eluents at suitable pH values can contain both the singly and doubly charged anions produced by dissociation of phthalic acid. For simplicity, we will consider initially only the case where the eluent contains two competing anions, which will be designated HP-and P2-. The solute will again be represented by AX-, so the relevant ion-exchange equilibria are:

A,

X-

+ XHP; %

2p;b'+ 2-

P,

xP:-

+ 2HP;

4-+ x H P i X-

% 2A, 2-

% P,

+ xP,2-

+ 2HPm

(5.26)

(5.27)

(5.28)

There are three approaches to developing a retention model for the above situation, namely the dominant equilibrium approach, the competing ion "effective charge" approach, and the dual eluent species approach. These are discussed separately below.

Dominant equilibrium approach This method assumes that the equilibrium shown in eqn. (5.28) lies well to the right because the P2- competing anion would be bound most strongly to the ion-exchange resin on the basis of electroselectivity The outcome of this assumption is that the doubly charged form of the competing anion is responsible solely for the elution of the solute, even when some H P is present. The charge on the competing anion is therefore -2. This approach has been found to yield satisfactory results with succinate eluents, where slopes of log k versus log [P2-] were -0.55 for monovalent anions and-1.15 for divalent anions using eluents which contained only 18% of the doubly charged succinate anion [31. Competing anion "effective charge '' approach When the two eluent competing anions are in rapid protolytic equilibrium (such as HP-and P2-), we can define the "effective charge" (-y) of a mixture of these species using the following relationship [2. 6, 81:

y = a1

+

2a2

(5.29)

where a1 and a2 are the fractions of total eluent species present as HP- and P2-, respectively. Eqn. (5.29) is a simplified version of eqn. (4.6).

Retention Modelsfor lon-Exchange

141

1.o

0.8

0.6

OA log k '

czop I-

0.2

5201-

0

5012HZPOi

Br-

-0.2

-0.L -27

a-26

-2.5

-2L -2.3 log [eluent]

-2.2

-2.1

-2.0

Fig. 5.3 Plots of log kA' versus log CEfor 2.0 - 10.0 mM phthalate eluents at pH 5.3. At this pH, the eluent contains 47.6% H P and 52.2% P2-. Reprinted from 121 with permission.

Eqn. (5.12) can be rewritten to include the effective charge on the competing anion, to give: (5.30)

where C2 is a constant and CEis the total eluent concentration, which is equal to:

CE = [HP-]

+ [P2-]

(5.31)

It is assumed in this approach that both competing anions have similar selectivities for the solute anion. Fig. 5.3 shows some experimental data [2] for plots of log k versus log CE for phthalate eluents containing both singly and doubly charged forms of phthalate, and Table 5.1 gives the slopes of these plots. Shown also are the theoretical slopes calculated using the dominant equilibrium approach (i.e. assuming that only P2- contributes to elution) and the competing anion "effective charge" approach. Using eqn. (5.29), we can calculate that the "effective charge" in this case is -1.52. It can be seen that linear plots were observed, as predicted from eqn. (5.30), but the observed slopes do not agree for all solutes with those predicted by either of the above approaches. The dominant

142

Chapter5

TABLE 5.1 OBSERVED AM) PREDICTED SLOPES OF LOG k VERSUS LOG CE PLOTS USING 2.010.0 mM PHTHALATE ELUENTS AT pH 5.3 Slope

Ion

Observed

-0.63 -0.63 -0.67 -0.60 -1.13 -1.10 -0.98

Dominant equilibrium

Effective charge

method y=2

method y = 1.52

-0.5 -0.5 -0.5 -0.5 -1.0 -1.0 -1.0

-0.66 -0.66 -0.66 -0.66 - 1.33 -1.33 -1.33

equilibrium approach gives best agreement for divalent solutes, whilst the effective charge approach gives best agreement for monovalent solute anions. It can be concluded that, for phthalate eluents at least, the IP-competing anion contributes to the elution of monovalent solutes on an approximately equal basis with P2-, whereas divalent solutes are eluted almost entirely by P2-.

Dual eluent species approach It is evident that neither of the above approaches gives reliable results for all solutes and this can be attributed to the fact that the different selectivity coefficients for the solute with both compcting anions are not considered. The dual eluent species approach, suggested by Hoover [ 101 2nd Jenke and Pagenkopf [ 11-13], is a more rigorous method which takes into consideration all of the relevant equilibria listed in eqns. (5.26) - (5.28). From eqn. (5.28) we can write: (5.32)

which can be rearranged to give: (5.33)

143

Retention Modelsfor Ion-Exchange

If we continue to assume that the eluent contains only two competing anions, HPand P2-, then the column capacity, Q, is given by:

Q = [HPJ

+ 2[P:-]

(5.34)

Combination of eqns. (5.33) and (5.34) yields:

(5.35)

which is a quadratic equation in [HP;]. Solution of this equation by the quadratic formula gives:

(5.36)

From eqn. (5.26), we can write:

(5.37)

which, from eqn. (5.4), can be rewritten:

(5.38)

Combination of eqns. (5.5), (5.38) and (5.36) gives: X

(5.39)

which can be simplified to give:

Chapter 5

144

Use of eqn. (5.40) requires knowledge of the concentrations of both Hp-and P2-. It is more common in IC to describe the eluent in terms of the total concentration of eluent acid (which we have designated above as CE)and the eluent pH. Eqn. (5.40) can be made more practically applicable if we consider the protolytic dissociation reaction of HP-,that is: 2-

+

HPi%PP,+H

(5.41)

From eqn. (5.41) we can write: (5.42)

which can be rearranged to give: (5.43)

If we assume that the eluent acid, H2P, is present only as HP- and P2-, then eqn. (5.31) holds, which combined with eqn. (5.43) gives: (5.44)

Substituting eqns. (5.43)and (5.44) into eqn. (5.40) gives:

(5.45)

Under circumstances where the eluent contains some undissociated H2P, eqn. (5.31) must be rewritten as:


[HPJ

+ IPkI = (1 - a

~ ~ CE p )

145

(5.46)

and eqn. (5.45)then becomes:

(5.47)

It can be noted in passing that eqn. (5.47)does not include the term K A , (i.e. ~ the selectivity coefficient for exchange between the solute AX-and the competing anion P2-). The reason for this is that K A , is ~ not independent of KA,HP and K p ~ pand so its inclusion is not necessary. The selectivity factor a A , B for the two solutes AX-and BZ-istherefore given by:

(5.48)

Jenke and Pagenkopf have successfully used eqn. (5.47)to predict the retention behaviour of a wide range of solutes using phthalate eluents with surface functionalized silica, surface functionalized resin or agglomerated ion-exchangers [l1, 12, 141. Retention data for each solute anion are obtained for a variety of different eluent concentrations (CE) and pH values, and these data are substituted into eqn. (5.47)to give a series of simultaneous equations with KA,HPand K p , ~ pas the only unknowns (correct values of Vm,w and Q are not necessary to the solution and arbitrary values are used). This process gives values of KA,HPand KP,HPfor each solute anion and Table 5.2 lists some typical results. Monovalent anions give very similar results for KPJP, and so do divalent solute anions, but the values determined for each group differ. This is a result of eqn. (5.47) taking a different form for divalent anions (i.e. when x = 2) to that used for monovalent anions (i.e. when x = I). The values of KA,HPfor monovalent anions follow the same sequence as the retention times for these solutes, with a larger value of KA,HP corresponding to longer retention. The same applies to the divalent solutes, but again it is not possible to directly compare selectivity coefficients for monovalent and divalent anions because of the different form of eqn. (5.47) used for their calculation. The minimized errors for the calculation of KA,HP values are shown in Table 5.2,from which it can be seen that this error varies from 2.57% for C1- to 6.13% for Sod2-.

3.0I I

= Br0 = CI-

"0 Calculated retention time iminl

2

4 6 8 10 12 1L 16 Calculated retention time (mint

18

20

Ibl

Fig. 5.4 Comparison of observed and predicted (eqn. 5.48) retention times for (a) monovalent and (b) divalent solute anions using 2 mM phthalate eluents with pH values in the range 4.0-5.5. Under these conditions, the eluent contains both singly and doubly charged phthalate ions. Reproduced from [13] with permission.

bl


147

TABLE 5.2 SELECTIVITY COEFFICIENTS AND MODEL FIT PARAMETERS FOR THE MULTIPLE ELUENT COMPETING ION MODEL, APPLIED TO PHTHALATE ELUENTS USED ON A HAMILTON PRP-X100 COLUMN. REPRINTED FROM [ 131 WITH PERMISSION

Solute

Selectivity coefficients KPJIP

so42~2032-

N0-jB r-

c1-

3.14 2.95 1.70 1.63 1.70

10-3 10-3 10-3 10-3

10-3

KA,IIP

3.59 5.89 4.61 3.90 2.65 x

10-5

10-5 10-3 103

lW3

%RSD in KA,HP

Retention time error (%RSD)

6.13 5.78 2.86 3.12 2.57

2.65 3.22 3.22 1S O 1.61

Model fit parameters: slope = 0.980, intercept = 0.07, correlation coefficient = 0.9973.

The values of KA,HPand K p , ~ pdetermined above can now be used to calculate predicted retention times for solute anions under a variety of eluent and column conditions. For this calculation, V m , w and Q must be known. Fig. 5.4 shows the agreement obtained between predicted and observed retention times and illustrates that excellent correlation is observed (see model fit parameters in Table 5.2). The average error in the determination of retention times for individual solutes is also listed in Table 5.2, which shows that a maximum error of 3.22% was observed. These results are not surprising since the predictions are made on the basis of selectivity coefficients determined using experimental conditions which are very similar to those used to test the model. Nevertheless, the multiple eluent competing ion model gives the most reliable description of retention behaviour when the eluent contains two competing anions. The multiple eluent competing ion model has also been used for eluents which contain three competing anions [lo, 13, 151. A typical example would be the use of an alkaline carbonate buffer in suppressed IC, where OH-, HCO3- and C032- all act as competing anions in the eluent. Under these circumstances, the derivation of a retention model proceeds along the same lines as for eluents containing only two competing anions. The capacity factors for monovalent and divalent solute anions, designated as kl' and k i , respectively, are given by the following equations:

(5.49)

148


149

(5.50)

where HP- and P2- represent HCO3- and C032-, respectively, whilst A- and A2- are monovalent and divalent solute anions. Again, the selectivity coefficients in eqns. (5.49) and (5.50) are determined by substituting experimental retention data for known eluent compositions and solving the resultant simultaneous equations. Table 5.3 lists values of the selectivity coefficients obtained for three solute anions on a Dionex HPIC-AS1 column. It is evident that the C032- component of the eluent dominates the elution of solutes (as indicated by the high selectivity coefficient for this species) and the selectivity coefficients for the solutes parallel their elution order. Fig. 5.5 shows the excellent agreement obtained between measured retention times and those predicted by eqns. (5.49) and (5.50) for a three component eluent system.

5.2.3 Gradient elution in anion-exchange IC The retention models developed above can be extended to gradient elution separations in which the concentration of the competing anion in the eluent is varied over the course of the separation. For simplicity, the discussion of gradient elution will be limited to the case where the eluent contains a single competing anion, Ey-. The following derivation is based on that reported by Rocklin et al. [5]. Assuming a linear gradient, beginning at zero concentration of the competing anion and increasing with time, the instantaneous concentration of the competing anion at the column inlet at any given time is:

TABLE5.3

SELECTIVITY COEFFICIENTS FOR SOLUTES INAN OH-~HCO~-/CO~~ELUENT SYSTEM, CALCULATEDFROMEQNS.(5.49)AND(5.50).DATAFROM[15] Selectivitycoefficients

Solute ion

c1NQso42a

KP,HP

KOH,HP

KA,HP

5.0 5.0 5.0

0.01 0.01

3.02 16.8

0.01

18.5

P = ~ 0 3 2 - ,HP = HCO~-.A = solute anion.

Chapter5

150

[EL-] = RV

(5.51)

where V is the volume of eluent pumped since the gradient was initiated and R is the slope of the gradient ramp (e.g. in mM/ml). If a solute, AX-, was injected onto the column and the composition of the eluent was kept constant until that solute eluted (i.e. the solute was eluted isocratically), the capacity factor, k A ' , would be given by a combination of eqns. (5.10) and (5.51):

Under gradient elution conditions, kA' must be integrated from injection to elution. This can be achieved by considering k A ' as dV/dx, where a fractional volume of eluent, dV, passes over the band maximum for the solute and moves the band a distance of dx down the column. It should be noted here that x is expressed as the column void volume, rather than as a length. This gives:

(5.53) If we consider w, Vm, KA,E and Q to remain constant during the separation, then eqn. (5.53) reduces to: (5.54)

where C3 is a constant. Eqn. (5.54) can be rearranged and integrated as follows:

It is convenient to describe retention in terms of (VR - V,)/Vm (where VR is the retention volume of the solute) instead of kA' (note that in isocratic elution, the two are equal). Solving eqn. (5.55) and rearranging gives:

(5.56)

We can collect the constant terms on the right hand side to give a single constant,

C,, which can be referred to as the gradient constant.


151

t

Citrate

-0.77

-

Fumarate 0.65

-0.47

- 0.67

\r Chloride 0.21

0.2

I

0.L

I

1

0.8

0.6

I

1.0

- 0.50

I 1.2

log R

Fig. 5.6 Plot of log (VR - Vm)/Vm) versus log R for gradient elution using NaOH eluents. A Dionex HPIC-ASSA column was used. Reprinted from [5] with permission.

(5.57)

Expressing eqn. (5.57) in logarithmic form gives:

Eqn. (5.58) is very similw in form to eqn. (5.12), which was developed for isocratic elution when a single competing anion was present in the eluent. From eqn. (5.58), we can predict that a plot of log ((VR - Vm)/V,) versus log R should be a straight line with a slope determined by the charges on the solute and competing anions. When the competing anion has a single charge, the slopes are predicted to be -0.5, -0.66 and -0.75 for mono-, di- and trivalent solute anions, respectively. Eqn. (5.58) has been evaluated experimentally using NaOH gradients with varying gradient ramps [5]. Fig. 5.6 shows the required plots and Table 5.4 presents pertinent data taken from these plots. Excellent linearity was obtained and the slopes showed good agreement with the predicted values. Moreover, it was even possible to predict the values of C, for most solutes on the basis of C3 values determined from isocratic elution of the solutes (see Fig. 5.2), together with values for the remaining constant terms in eqn. (5.56).

Chapter5

152

T A B E 5.4 PARAMETERS FROM THE GRADIENT PLOTS SHOWN IN FIG. 5.6.DATA FROM [5]

Solute anion

PallMEter ~

Measured slope Predicted slope (eqn. (5.58)) c, (measured) C, (calculated)

Chloride

Nitrate

Sulfate

Fumarate

Citrate

-0.50 -0.50 7.4 7.0

-0.47

-0.67 -0.67 20 19

-0.65 -0.67 26

-0.77 -0.75 42

24

65

-0.50 15 14

It can be concluded from the above results that the retention model in eqn. (5.58) gives a satisfactory description of the behaviour of solutes under gradient elution conditions in which the gradient is generated by linearly increasing the concentration of the competing anion in an eluent which contains no other competing anion.

5.3 5.3.1

RETENTION MODELS FOR CATION-EXCHANGE Single eluent competing cation

We begin by considering the simplest case in cation-exchange IC, in which the eluent contains a single type of competing cation. The ion-exchange equilibrium for to a stationary phase which has been conditioned with an binding of a solute cation, Mx+, eluent containing a competing cation, Ey+, is given by: (5.59)

where the subscripts m and r refer to the mobile and resin phases, respectively. If activity coefficients are assumed to be equal to unity, the selectivity coefficient, KMJ, can be written:

If we now follow the same derivation used in Section 5.2.1 for the development of retention equations for anions, we can produce the following important equations for the retention of cations with a single type of eluent competing cation:


153

Eqn. (5.10) gives us:

(5.61)

Eqn. (5.1 1) gives: (5.62)

Eqn. (5.12) gives: (5.63)

Eqns. (5.20) and (5.21) give:

for two solutes Mix+ and M2z+. And finally, from eqn. (5.22) we can write: (5.65)

Since eqns. (5.61) - (5.65) take the same form as the corresponding anion-exchange equations derived earlier, we can make the same generalizations regarding retention behaviour which were made for anions in Section 5.2.1. That is, solutes are eluted earlier with eluent competing cations of higher charge, whilst retention increases with increasing charge on the solute cation. These trends have been confirmed in practice [16]. The major test of the above derivation is the validity of eqn. (5.63), which predicts a linear relationship between log kM' and log [EY+,,,].We can begin by examining this relationship for monovalent cations eluted with a nitric acid eluent [17]. Fig. 5.7 shows that linear plots are observed with this system and the slopes are in good agreement with the theoretical slope of -1.0 for univalent solute and competing cations. Sevenich and Fritz [16] have tested eqn. (5.63) for divalent and trivalent cations retained on a lowcapacity (0.06 mequiv/g) cation-exchanger using perchloric acid eluents. Linear plots were observed only when activity effects were considered; that is, when log kM' was plotted against log (HClO4). where the parentheses indicate activity. Fig. 5.8 illustrates such a plot for several trivalent rare earth cations and Table 5.5 summarizes some of the slopes obtained for divalent and trivalent cations.

154

Chapter 5

''7

0

Li+

0 = Na*

Slopes -1.13 -1.07

-1.06 -1.07 I

-3.0

-2.9

i

1

-2.8 -2.7 log [H+l

I

I

-2.6

-2.5

Fig. 5.7 Plots of log k M qversus log [H+I for nimc acid eluents used on a Waters IC Pak C surface-sulfonated cation-exchange column. The slope of each plot is given in the Figure. Reprinted from 171 with permission.

TABLE 5.5

SLOPES OF PLOTS OF LOG kM' VERSUS LOG (HC104) USING PERCHLORIC ACID ELUENTS. DATA FROM 116,181

Cation

Slope

Mg2+ Ca2+ Sr2+

- 1.66 -1.87 -1.91 - 1.97 - 1.89 - 1.98 -1.87 - 1.96 - 1.92

B82+

Mn2+ Zn2+ Ni2+ Ua2+ CU2+

Cation

Slope

Fe2+

- 1.79 -1.86 -2.08 -2.00 - 1.92 -2.88 -2.67 -2.99

Co2+ Cd2+

Pb2+ Hg2+ A13+ Bi3+ Fe3+

Cation

Slope

Cation

Slope

h3+

-2.78 -3.01 -2.95 -2.99 -2.95 -2.95 -3.01 -3.01

Tb3+ Gd3+ Eu3+ sm3+ Nd3+

-3.02 -2.99 -3.01 -2.97 -2.97 -2.95 -3.01 -3.04

LU3+ Yb3+ Tm3+ Y3+

Er3+ HO~+

Dy3+

~r3+

Ce3+ ~a3+


155

1.82 log k'

1.32

~03+

Nd3' Eu3+ Tb3+

0.82

HO~+

0.321, -0.55

, -045

,

,

,

,

Lu 3+

-0.35

-0.25

-0.15

-0.05

log (HCIOh activity)

Fig. 5.8 Plots of log kM'versus log (HClO4) for perchloric acid eluents used on a low-capacity surface-sulfonated cation-exchanger. Reprinted from [ 161 with permission.

Agreement between the slopes obtained experimentally and those predicted from eqn. (5.63) is generally good, although there are some exceptions. Lederer 1191 has reviewed a large volume of literature relating to the effective charge on a metal ion (or metal complex ion) when this charge is measured using ion-exchange methods. The conclusions of this study are: Activity effects are of importance, especially for polyvalent cations and when the eluent strength is high. Complete correction for activity effects requires knowledge not only of the activity coefficient of the competing cation in the eluent phase, but also of the activity coefficients for metal ion in the eluent phase and the metal ion and the competing cation in the resin phase. Most of these coefficients are not available. Neglecting activity effects will tend to produce slopes which are smaller than those predicted from eqn. (5.63). (ii) For some cations, the effective charge is reduced due to steric effects which prevent the cation interacting with a stoichiometric number of functional groups on the resin surface. For example, a trivalent cation may not be able to approach closely three functional groups. This effect will tend to be most significant for large, polyvalent cations and for stationary phases of low ionexchange capacity which therefore have a diffuse spread of functional groups.

(i)

156

Chapter5

Reduction in the effective charge on the solute cation by this effect will result in slopes which are smaller than those predicted from eqn. (5.63). (iii) Under conditions of relatively high eluent strength, ion-pair formation may occur between the solute cation and the sulfonic acid groups on the resin surface. The outcome of this is that the interaction between the solute and the resin will be much stronger than that expected from purely electrostatic attraction. The effective charge on the solute is therefore increased and the slope becomes greater than that predicted from eqn. (5.63). Examination of the probable magnitudes of these effects in IC, where dilute eluents are used with low capacity ion-exchange resins, suggests that effects (i) and (ii) can be expected to occur to a significant degree, especially for solute cations with a charge greater than 2. Effect (iii) is likely to be the least significant because the chromatographic conditions employed are not favourable for ion-pair formation. There is some evidence from the data of Table 5.5 to support the occurrence of effect (3). The deviation of the experimentally observed slopes for divalent alkaline earth cations from those predicted by eqn. (5.63) follows the order Mg2+ > Ca2+> Sr2+ > Ba2+,which is the same order for the diameters of the hydrated ions (181. It is clear from the above discussion that, under conditions where the eluent contains a single type of competing cation (which is the normal situation for cationexchange), eqn. (5.63) is followed closely when monovalent cations are used as solutes. Deviations may occur for solutes of higher charge, with these species requiring the use of activity corrections if meaningful estimates of effective charge are to be made. 5.3.2

Complexing eluents

Separation by cation-exchange In Section 4.3.2, we noted that eluents containing a complexing ligand are frequently used for the separation of cations. Fig. 4.16 showed schematically a number of ways in which such a complexing eluent could be employed. We will now extend the above cation-exchangeretention model to include a complexing eluent component. When a solute cation, W+,is eluted by a competing cation, Ey+. and the eluent also contains H2L. where L2- is a ligand capable of reacting with h P ,we have the situation depicted in Fig. 4.16(c). Under these conditions, the equilibrium shown in eqn. (5.59) applies, but we must also consider the further equilibria shown in eqns. (5.66)-(5.69):

H2L f HL;, +

(5.66)

(5.67)

%++ L,2- ej x-2

ML,

x-2

ML,

+ L,2- %

x-4

ML2

(5.68)

(5.69)


157

We can now define a M as the fraction of the total concentration of metal ion in the eluent which is present in the free form, i.e. as W+:

(5.70)

where CM is the total concentration of metal ion in the mobile phase, regardless of the form in which it is present. CMis given by:

Assuming that Mx+is the only form of the metal which is bound to the cationexchange resin ( i x . the complexes formed with L2- are either neutral or anionic), then the distribution coefficient for solute Mx+ can be defined as:

(5.72)

Substituting eqns. (5.70) and (5.71) into eqn. (5.60) gives:

(5.73)

Rewriting eqn. (5.5) for the cation case and employing eqn. (5.8), we obtain:

(5.74)

which can be presented in logarithmic form as:

If we now consider only the case of a divalent metal cation (x = 2) and we assign equilibrium constants of Pal,Ka2, (i.e. acid dissociation constants) to eqns. (5.66)and (5.67), and K1 and K2 (stepwise formation constants) to eqns. (5.68) and (5.69), then aM is given by: a M

1

=

1

+ K l a L C L + K1K2aLC[

(5.76)

Chapter5

158

2.0-

1.5-

tog tk ~a3*

1.0~ e 3 +

~r 3+

0.5

-

Nd3+ Dy3+ Er 3' Tm3. Lu 3+

-2.1

-3.0

-2.5

-2.L -2.3

log [en ~ ~ 2 * ]

Fig. 5.9 Plots of log t

~ versus ' log [enH22+]for ethylenediamine-tartrate eluents, in which the total tartrate concentration is maintained at 2.0 niM and the pH is consrant at 4.5. A low capacity surface-sulfonated cation-exchanger was used. Reprinted from [20] with permission.

where CL is the total concentration of ligand species in the eluent and aL is the fraction of CL which exists as L2-. aL is given by:

Eqn. (5.75) applies to any cation-exchange separation mechanism which uses a complexing eluent. It is therefore applicable to both of the methods depicted in Fig. 4.16(a) and 4.16(c). The only difference between these two methods is that complexation of Mx+exerts an overwhelming effect in Fig. 4.16(a) (where Na+ or Li+ act as very weak competing cations), whilst separation in Fig. 4.16(c) results from a combination of complcxation and cation-exchange because an effective competing cation (enHz2+)is prescnt. Some rctcntion data for cations using ethylcnediamine-tartrate eluents are available for experimental vcrification of cqn. (5.75) [20]. Fig. 5.9 shows plots of the logarithm of the adjusted retention time, t', versus the logarithm of the ethylenediamine Concentration, obtained under conditions where the tartrate concentration in the eluent


159

1.5r

1.0

Log tk

0.5

-

-1.80

-1.50

-1.20 ''9 aMILl

-0.90

Fig. 5.10 Plots of log tM' versus log a M for ethylenediamine-tartrate eluents, in which the ethylenediamine concentration is maintained at 2.0 mM and the pH is constant at 4.5. The column used was identical to that in Fig. 5.9. Reprinted from [20] with permission.

and the pH (and hence CXM) are constant. The adjusted retention time is equal to t,k', where to is the retention time for a non-retained solute. Thus substitution oft' for k' in eqn. (5.75) does not alter the equation except for an additional log(b) term on the right hand side. Under the conditions used, all terms in eqn. (5.75) are constant, except for log[EY+,], so we expect a straight line plot of slope -x/y. The solutes used have a charge of +3, whilst the competing cation in the eluent has a charge of +2, which gives a theoretical slope of 1.5. Fig. 5.9 shows that the predicted linear plots were obtained, with slopes that are all close to -1.0. The discrepancy between theory and experiment may have been due to the fact that no activity corrections were applied in this study. Divalent solute cations showed much better agreement with theory, giving slopes averaging -0.9 1201. If the pH and the concentration of ethylenediamine are now kept constant while the concentration of tartrate is varied, aM will change. It is therefore possible to further versus ) log t'. Since all terms except CLM are test eqn. (5.75) by plotting ~ O ~ ( C X M constant, we expect a linear plot of slope equal to 1.0. The results are presented in Fig. 5.10, which shows that linear plots were obtained, with slopes averaging 1.2. This is in reasonable agreement with the theoretical slope.

Chapter5

160

Separation by anion-exchange Some ligands will form anionic complexes with the solute cations, leading to the possibility of separation by anion-exchange. This process was illustrated in Fig. 4.16(b) and a representative chromatogram was shown in Fig. 4.17(b). A retention model for this situation can be developed using the same approach as that employed for cationexchange with a complexing species present in the eluent. To simplify the derivation, we will assume that the solute ion is divalent. Eqns. (5.66) and (5.67) apply to this case and we can rewrite eqns. (5.68) and (5.69) for a divalent cation to give: 2-

ML,

+ L;

(5.78)

f ML;-

(5.79)

The anion-exchange equilibrium for the solute, MLz2-, and a competing anion, Ey-,

is

YMLi-

m

+

Y-

2-

2E, 6 yML2

r

Y-

+ 2E,

(5.80)

which enables us to write the relevant selectivity coefficient:

Defining a M L Z as the fraction of total metal species in the eluent (CM)existing as MLz2-, we have:

(5.82)

where CMis as defined by eqn. (5.71). We can calculate am2 according to:

where K1 and K2 are the stepwise formation constants for eqns. (5.78) and (5.79), ~ as previously defined (eqn. (5.77)). Since ML22- is the respectively, and CL and c 1 are only form of M which is retained on the column, the distribution coefficient for M is given by:


161

(5.84)

Substituting eqns. (5.82) and (5.84) into eqn. (5.81) gives:

If we rewrite eqn. (5.85) in terms of the capacity factor and make the usual substitution for column capacity, Q,we obtain:

(5.86)

which in logarithmic form becomes:

Turning now to the case where the solute cation is trivalent and forms an anionic bis complex with L2- of the type M L i , eqn. (5.87) can be rewritten:

When the total ligand concentration, CL,and the pH are constant, we expect the same linear relationship between log kM' and log [EY-,,,which ] was found earlier to be valid for anion-exchange (Section 5.2.1)). Under conditions where the concentration of competing anion in the eluent is constant, we also expect a linear relationship between log kM' and log aML2. 5.4 1 2 3

REFERENCES Gjerde D.T., Schmuckler G. and Fritz J.S., J. Chromatogr., 187 (1980) 35. Haddad P.R. and Cowie C.E., J. Chromatogr.,303 (1984) 321. Van 0 s M.J., Slanina J., De Ligny C.L., Hammers W.E. and Agterdenbos J., Anal. Chim. Acza, 144 (1982) 73. Vlacil F. and Vins I., J. Chromatogr., 391 (1987) 133. Rocklin R.D., Pohl C.A. and Schibler J.A., J. Chromatogr.,41 1 (1987) 107. Diop A., Jardy A., Caude M. and Rosset R., Analusis, 14 (1986) 67. Gjerde D.T. and Fritz J.S., J . Chromatogr., 176 (1979) 199. Jardy A., Caude M., Diop A., Curvale C. and Rosset R., J. Chromutogr.,439 (1988) 137.

162 9 10 11 12 13 14 15 16 17 18 19 20

Chapter 5

Diop A., Jardy A., Caude M. and Rosset R., Analusis, 15 (1987) 168. Hoover T.B., Sep. Sci. Technol., 17 (1982) 295. Jenke D.R. and Pagenkopf G.K., Anal. Chem., 56 (1984) 85. Jenke D.R. and Pagenkopf G.K., Anal. Chem., 56 (1984) 88. Jenke D.R. and Pagenkopf G.K., in Jonsson J.A., (Ed.), Chromatographic Theory and Basic Principles, Marcel Dekker, New York, NY, 1987, p. 313. Jenke D.R., Anal. Chem., 56 (1984) 2674. Jenke D.R. and Pagenkopf G.K., J . Chromutogr.,269 (1983) 202. Sevenich G.J. and Fritz J.S.,J. Chromutogr., 371 (1986) 361. Foley R.C.L. and Haddad P.R., J . Chromurogr.,366 (1986) 13. Gjerde D.T., J . Chromutogr.,439 (1988) 49. Lederer M., J . Chromorogr.,452 (1988) 265. Sevenich G.J. and Fritz J.S., Anal. Chem., 55 (1983) 12.

Part II Ion-Interaction, Ion-Exclusion and Miscellaneous Separation Methods

164

Stationary phases

. ION-INTERACTION

Eluents

CHROMATOGRAPHY (Chap 6)

L

-E

Dynamic coating

-€

Permanent coating Anions

Retention model

Cations

,Stationary phases

FURTHER SEPARATION METHODS

ION-EXCLUSION -CHROMATOGRAPHY (Chap 7)

IiDVB

+

Eluents

Microporous

Macroporous

L

Acids Complexing eluents

Retention model Reversed-

Coordination compounds Organometallics

MISCELLANEOUS - SEPARATION METHODS (Chap 8)

Schematic overview of Part II.

t

Carboxylic acids Chemically phases

Micelle exclusion

Crown ethers Anions { Cations

165

Chapter 6 Ion-Interaction Chromatography 6.1

INTRODUCTION

Hydrophilic ionic solutes, such as the inorganic anions and cations of interest in IC, show little or no retention on lipophilic stationary phases when typical reversed-phase eluents are used. However, retention and subsequent separation of such ionic solutes on these stationary phases can be achieved by the addition to the eluent of a lipophilic reagent ion having the opposite charge sign to that of the solute ion. This added reagent ion, and the chromatographic process itself, have been described by a variety of names, some of which are listed in Table 6.1. Most of these names impIy some sort of mechanism for the process and may therefore be misleading. Throughout this book, the terms ion-interaction chromatography and ion-interaction reagent (IIR) will be used, since these are quite general terms. In this Chapter, we will examine some of the mechanisms which have been proposed for ion-interaction chromatography and we will then consider the types of stationary phases and eluents which are used with this technique. Specific applications of ion-interaction chromatography to the separation of inorganic anions and cations will then be discussed.

TABLE6.1 ALTERNATIVE NAMES USED TO DESCRIBE ION-INTERACTION CHROMATOGRAPHY AND THE REAGENT ION ADDED TO THE ELUENT [11 Chromatographic process

Reagent ion

Reference

Ion-pair chromatography Paired-ion chromatography Surfactant chromatography Dynamic ion-exchangechromatography Ion-interactionchromatography Hetaeric chromatography Mobile phase ion chromatography (MPIC)

Pairing ion PIC reagent Surfactantion Ion-pairing reagent Ion-interaction reagent Hetaeron Pairing reagent

2 3 4 5

6 7

7

Chapter 6

166

6.2

MECHANISM

6.2.1

Trends in solute retention in ion-interaction chromatography

A convenient way to highlight the trends in solute retention is to compare the retention of a solute on a chromatographic system comprising a lipophilic stationary phase and an eluent consisting of an IIR dissolved in a mixture of water and one or more organic solvents with the retention of the same solute under the same chromatographic conditions, except using an eluent which does not contain the IIR. When this comparison is made, the following trends are observed: The retention of neutral solutes is not altered significantly when the IIR is added to the eluent. (ii) The retention of solutes having the same charge as the IIR is decreased when the IIR is added to the eluent. (iii) The retention of solutes having the opposite charge to the IIR is increased when the IIR is added to the eluent.

(i)

In addition, the following effects on retention are observed when the composition of the eluent is altered: (iv) The retention of solutes having the opposite charge to the IIR is increased when the concentration of IIR in the eluent is increased. (v) The retention of solutes having the opposite charge to the 11R is increased when the lipophilicity of the IIR is increased. (vi) Retention of all solutes decreases when the percentage of modifier in the eluent is increased, and vice versa. Any mechanism suggested for ion-interaction chromatography must necessarily explain these trends in retention behaviour. A large volume of literature has been devoted to the study of such mechanisms and a detailed discussion of this literature is beyond the scope of this book. Moreover, the most of these studies have been devoted to mechanisms for the retention of orgunic ionic species, such as carboxylic acids and organic bases. A summary of the most commonly suggested mechanisms [l] will be presented below and this will be followed by an evaluation of these mechanisms in terms of their applicability to the retention of inorganic ions. 6.2.2

The ion-pair model

In this model [7- 1 I], an ion-pair is envisaged to form between the solute ion and the IIR. This occurs in the aqueous-organic eluent and the resultant neutral ion-pair can then be adsorbed onto the lipophilic stationary phase in the same manner that any neutral molecule with lipophilic character is retained in reversed-phase chromatography. Retention therefore results solely as a consequence of reactions taking place in the eluent. The degree of retention of the ion-pair is dependent on its lipophilicity, which in turn depends on the lipophilicity of the IIR itself. Neutral solute molecules are unaffected by

Ion-InteractionChromatography

167

Bulk eluent

Fig. 6.2 Schematic illustration of (a) the ion-pair, (b) the dynamic ion-exchange and (c) the ioninteraction models for the retention of anionic solutes in the presence of a lipophilic cationic IIR. The solute and the IIR are labelled on the diagram. The large, hatched box represents the lipophilic stationary phase, the black circle with the negative charge represents the counter-anion of the IIR, whilst the white circle with the positive charge represents the counter-cationof the solute. Adapted from [ 11.

the presence of the IIR in the eluent and interact with the stationary phase in the conventional reversed-phase manner. An increase in the percentage of organic solvent in the eluent decreases the interaction of the ion-pairs with the stationary phase and therefore reduces their retention. The ion-pair model is illustrated schematically in Fig.

168

Chaprer6

6.l(a), using a positively charged IIR and a negatively charged solute as an example. 6.2.3 The dynamic ion-exchange model The dynamic ion-exchange model [12-151 proposes that a dynamic equilibrium is established between IIR in the eluent and IIR adsorbed onto the stationary phase, as follows:

where the subscripts E and S refer to the eluent and stationary phases and the superscript on the IIR indicates that it may carry either a positive or negative charge. The adsorbed 1IR imparts a charge to the stationary phase, causing it to behave as an ion-exchanger. The total concentration of IIR adsorbed onto the stationary phase is dependent on the percentage of organic solvent in the eluent, with higher percentages of solvent giving lower concentrations of IIR on the stationary phase. In addition, the more lipophilic the IIR, or the higher is its concentration, then the greater is its adsorption onto the stationary phase. Thus, for a given eluent composition, the concentration of adsorbed IIR (and hence the "ion-exchange'' capacity of the stationary phase) remains constant. However, constant interchange of IIR occurs between the eluent and stationary phase, so the stationary phase can be considered to be a dynamic ion-exchanger. Introduction of a solute with opposite charge to the IIR results in retention by a conventional ion-exchange mechanism. The competing ion in this ion-exchange process may be the counter-ion of the IIR, or another ionic species deliberately added to the eluent. Since the retention times will be dependent on the ion-exchange capacity of the column, they are also dependent on the lipophilicity of the IIR and the percentage of organic solvent in the eluent. Solutes having the same charge as the IIR are repelled from the charged stationary phase surface and show decreased retention times in comparison to those observed in the absence of IIR, whilst retention times for neutral solutes are unaffected by the IIR. Fig. 6.l(b) gives a schematic representation of the dynamic ion-exchange model, again using a positively charged IIR and a negatively charged solute as an example. 6.2.4 The ion-interaction model

The ion-interaction model [ I . 6, 16-19] can be viewed as intermediate between the two previous models in that it incorporates both the electrostatic effects which are the basis of the ion-pair model and the adsorptive effects which form the basis of the dynamic ion-exchange model. The lipophilic IIR ions are considered to form a dynamic equilibrium between the eluent and stationary phases, as depicted in eqn. (6.1). This results in the formation of an electrical double-layer at the stationary phase surface. The adsorbed IIR ions are expected to be spaced evenly over the stationary phase due to repulsion effects, which leaves much of the stationary phase surface unaltered by the IIR. The adsorbed IIR ions constitute a primary layer of charge, to which is attracted a diffuse, secondary layer of oppositely charged ions. This secondary layer of

Ion-InteractionChromutogrqhy

169

charge consists chiefly of the counter-ions of the IIR. The amount of charge in both the primary and secondary charged layers is dependent on the amount of adsorbed IIR, which in turn depends on the lipophilicity of the IIR, the IIR concentration, and the percentage of organic solvent in the eluent. The double-layer is shown schematically in the top frame of Fig. 6.l(c). Transfer of solutes through the double-layer to the stationary phase surface is a function of electrostatic effects and of the solvophobic effects responsible for retention in reversed-phase chromatography. Neutral solutes can pass unimpeded through the double layer, so their retention is relatively unaffected by the presence of IIR in the eluent. A solute having opposite charge to the IIR can compete for a position in the secondary charged layer, from which it will tend to move into the primary layer as a result of electrostatic attraction and, if applicable, reversed-phase solvophobic effects. The presence of such a solute in the primary layer causes a decrease in the total charge of this layer, so to maintain charge balance, a further IIR ion must enter the primary layer. This means that the adsorption of a solute ion having opposite charge to the IIR will be accompanied by the adsorption of an IIR ion. The overall result is that solute retention involves a pair of ions (that is, the solute and IIR ions), but not necessarily an ion-pair. This process leads to increased retention of the solute compared to the situation in which the IIR is absent from the eluent. The lower frame of Fig. 6.l(c) depicts this process for a positively charged IIR and a negatively charged solute. Solutes having the same charge as the IIR will show decreased retention due to electrostatic repulsion from the primary charged layer. 6.2.5 Evaluation of mechanistic models in retention of inorganic ions

Many studies have examined the applicability of the above models to the retention of organic species [e.g. 20-231, but we will consider here only the case of inorganic ions. Such species are very hydrophilic and, in most cases, are unlikely to form ion-pairs in aqueous-organic solutions. Moreover, conductance measurements would be expected to reveal the formation of ion-pairs and such measurements have failed to provide supporting evidence for significant ion-pair formation [6,241. Furthermore, the ionpair model would require that the neutral ion-pairs formed by different solute ions should have varying degrees of lipophilicity in order for them to be separated. These differences can be expected to be very slight for a series of inorganic ions (e.g. C1-, Br-, N a - , NOS-and S04*-),yet the ensuing discussion in this chapter will show that these species are separated readily by ion-interaction chromatography. Despite these shortcomings, there is a persistent trend in the literature to discuss the retention of inorganic ions in this form of chromatography in terms of interactions occurring between the solute and IIR in the eluentphse. The dynamic ion-exchange model generally provides an accurate prediction of the retention order of solutes, since this usually follows the established ion-exchange selectivity order discussed earlier in Chapter 2. In addition, the role of the counter-ion of the IIR is also predicted correctly if this counter-ion is considered to act as an ionexchange competing ion. Nevertheless, there are some shortcomings to the dynamic ionexchange model. Once such shortcoming can be seen by comparing the elution behaviour of solutes in an ion-interaction system in which a particular competing ion is

170

Chapter6

Ion-Interaction Chromatography

17 1

used to that of a conventional fixed-site ion-exchange system in which the same competing ion is employed. In the latter system, there will be a stoichiometric exchange of solute and competing ions at the ion-exchange site, so that the elution of a solute ion will always be accompanied by a decrease in the concentration of competing ion. This behaviour is not always observed in ion-interaction chromatography, where increases in the concentration of the IJR and its counter-ion often occur [25]. A more detailed discussion of this aspect and its utility for detection purposes will be found in Section 12.3.2. A study of the retention of inorganic anions in ion-interaction chromatography showed results which were in general agreement with the dynamic ion-exchange retention model, but some significant deviations from the predicted dependences indicated that the actual mechanism was more complex [26]. The ion-interaction model, and the formation of an electrical double-layer at the stationary phase surface, gives the most consistent agreement with experimental measurement. Consideration of the double-layer in terms of the Stern-Gouy-Chapman theory enables the effect on solute retention of the ionic strength of the eluent to be predicted accurately for lipophilic organic solute ions [ 161. This approach also permits the effect on solute retention of the concentration of IIR to be predicted. Similarly, electrostatic surface potential calculations, coupled with a Langmuir isotherm for adsorption of the IIR, predicts solute retention behaviour which is in good agreement with experimental results [17, 181. Moreover, studies concerned specifically with the retention of inorganic anions [27] and cations [28] in ion-interaction chromatographic systems have concluded that the ion-interaction model is the most appropriate.

6.3

STATIONARY PHASES AND ELUENTS

6.3.1 Stationary phases Ion-interaction chromatography has been performed successfully on a wide range of stationary phases, including neutral polystyrene divinylbenzene (PS-DVB) polymers 1e.g. 29, 301 and bonded silica materials with c18 [e.g. 311, c8 [32], phenyl [33] and cyano [34] groups as the chemically bound functionality. Each of these stationary phases gives satisfactory retention of ionic solutes, provided the eluent composition is such that an appropriate amount of the IIR is adsorbed. The choice between stationary phases is usually based on such considerations as chromatographic efficiency [35], pH stability [36] and particle size 1371, rather than on differences in chromatographic selectivity. However, it has been noted 1381 that the elution order for solutes can vary when the nature of the stationary phase used to support the IIR is altered. This point is illustrated in Fig. 6.2, which shows chromatograms for inorganic anions, obtained on three different stationary phases. It can be noted that the elution position of sulfate differs markedly between the c18 and PS-DVB stationary phases. Also apparent is the improved chromatographic efficiencies of the silica-based stationary phases compared to that of the PS-DVB material. Further factors to be considered sin the selection of a stationary phase for ioninteraction chromatography are specific interactions existing between the stationary phase and either the IIR or the solutes, and the role of residual silanol groups on silica-

172

Chapter 6

TABLE 6.2 TYPICAL REAGENTS USED AS IIRs IN DYNAMIC COATING ION-INTERACTION CHROMATOGRAPHY OF ANIONS AND CATIONS IIR

Detection mode$

References

Tetramethylammonium Tetrapmpylammonium Tetrabutylammonium Tetrapentylammonium Hexylammonium

spec C, Indirect Spec, Amp C, Spec,Amp spec c,spec, Indirect spec c,spec, Indirect spec spec, ICP C, Spec,Amp spec Spec. Amp Indirect spec Indirect spec Indirect spec Indirect spec Indirect spec Indirect Fluor

38,42 36,43,44 45-47 17, 24, 29 48 49-5 1 41,52-54 26, 31, 53, 55 56 57 58.59 59, 60 59, 60 61 62-64 65

spec RI, ICP spec Spec, PCR. RI PCR Indirect spec Spec,Indirect Amp

66

octylammonium

Hexadecyltrimethy l u m cetyl~thyl~nium Tricaprylylmethylammonium Dodecyltriethylammonium Benzyltributylammonium Naphthylmethy lmbutylammonium Naphthylmethy l t r i p r o p y l a m m o n i u m Methylpyridinium Iron(II)-l,I@phenanthroline Ruthenium(II)-l,lO-phenanthroline on -ations Butanesulfonate Pentanesulfonate Hexanesulfonate Octanesulfonate Dodecylsulfate

Naphthalenesulfonate Diethyldithidamate a

6, 67 66 28, 68,69 70 71 72,73

Spec = spectmphotometry, Amp = axnperomeay, C = conductivity, Fluor = fluorescence, ICP = inductively coupled plasma atomic emission, PCR = post-column reaction, RI = refractive index.

based stationary phases. Some IIRs (e.g. cetylpyridinium ions) and solutes (e.g. iodide) show particularly strong adsorption to PS-DVB stationary phases and this has been attributed to the occurrence of K-x interactions with the aromatic moiety of the polymer [35]. Residual silanol groups on silica-based packings have been shown to act as weak cation-exchange sites and this behaviour exerts an influence on the ion-interaction separation of both anions and cations on these stationary phases [29]. Scrutiny of the literature reveals that the majority of ion-interaction separations are performed on conventional Cis silica-based reversed-phase materials or on neutral PSDVB polymers (such as Hamilton PRP-I, Rohm & Haas XAD-2 and Dionex MPIC columns).

173


TABLE6.3 EFFECT OF THE ALKYL CHAIN LENGTH OF THE IIR ON RETENTION TIMES OF ANIONS [37] Solute

Retention ti&

Chloride Bromide Fluoride Iodide Nitrate

(min)

Hexylamine salicylate

Octylamine salicylate

DeCyl& Salicylae

n.r.b n.r. n.r. n.r. n.r.

2.0 f 0.3 2.0 f 0.3 2.3 f 0.3 2.5 f 0.3 2.5 f 0.3

7.0 f 0.3 7.3 f 0.4 8.0 f 0.4

6.5 f 0.3

8.3 f 0.4

A Lichrosphere RP-18column (5 p n particle size) was used. The eluent concentration was 5 mM and the flow-rate was 2.0 mumin. b n.r. = not retained.

a

6.3.2 Type of ion-interaction reagent

Requirements of the IIR The most important component of the eluent in ion-interaction chromatography is the IIR itself. The prime requirements of the IIR are as follows: (i) (ii) (iii) (iv)

An appropriate charge, which is unaffected by eluent pH. Suitable lipophilicity to permit adsorption onto non-polar stationary phases. Compatibility with other eluent components. Compatibility with the desired detection system.

"Dynamic coating" ion-interaction chromatography Anion separations are normally performed using strong base cations, such as tetraalkylammonium ions, as the IIR whilst cation separations are usually performed using strong acid anions, such as aliphatic sulfonate ions. Table 6.2 lists some IIRs which are used for ion-interaction separations. In each case, the IIR is present at a constant, specified concentration in the eluent in order to maintain a desired concentration of IIR on the stationary phase. That is, the coating of IIR is in dynamic equilibrium, as shown in eqn. (6.1). and the column can be said to be "dynamically coated" with IIR. Table 6.2 shows that the IIRs used for dynamic coating ion-interaction chromatography vary from moderately lipophilic (e.g. tetrabutylammonium ions) to very lipophilic (e.g. hexadecyltrimethylammonium ions). The lipophilicity of the IIR governs the degree of adsorption of the IIR onto the stationary phase, which in turn governs the effective ion-exchange capacity of the column and hence the retention times of solute ions. This point is illustrated in Table 6.3, which lists retention times for

r

f

i

0.032AU

L co

0.01 AU

i

n Pr NI

M

1

I

I

0

2

I

I

4 6 Time (minl (a1

I

I

1

I

I

I

I

1

8

10

11

0

4

8 Time (minl (bl

12

16

Fig. 6.3 Separation of cations by dynamically coated ion-interaction chromatography. (a) A C1g column was used with 10 mM octanesulfonate and 45 mM tartrate at pH 3.4 as eluent. Reprinted from [5] with permission. (b) A 5pm Supelco C1g column was used with an eluent formed from a linear gradient of 0.05-0.40 mM a-hydroxyisobutyric acid at pH 4.2, containing 30 mM octanesulfonate and 7.5% methanol. Reprinted from [68] with permission. Post-column reaction detection was used in each case.

B o\


175

anions obtained using IIRs with differing alkyl chain lengths (and therefore differing lipophilicity). Some of the detection modes available for use with the various IIRs are also listed in Table 6.2 and a discussion of detection for this mode of chromatography will be presented in Part I11 of this book. It is interesting to note that inorganic complexes, such as the 1,lo-phenanthroline complexes of iron(II1) and ruthenium(I1) can be employed as IIRs. Pietrzyk and co-workers [62, 64, 65, 741 have shown that these complexes are adsorbed readily onto PS-DVB stationary phases and their absorbance or fluorescence can be used for indirect detection of anions. Some typical chromatograms obtained for the separation of anions by dynamic coating ion-interaction chromatography were presented in Fig. 6.2, and typical separations of cations are shown in Fig. 6.3. These chromatograms illustrate the excellent chromatographic efficiency which can achieved using ion-interaction as the separation mode. Many of the species listed in Table 6.2 are surfactants which will form micelles if the IIR concentration exceeds the critical micelle concentration for that particular species. These micelles can be adsorbed onto the stationary phase in the same manner as other IIRs. However, it has been shown that the retention times of anionic solutes in micellar eluents decreases as the concentration of the micellar IIR is increased [53]. This behaviour is opposite to that normally observed and has been attributed to interaction of the solute anions with micelles in the eluent, which reduces the electrostatic interaction of these solutes with adsorbed micelles and thereby reduces their retention. The elution order observed for anions in this system is similar to that for conventional anionexchangers, as illustrated in Fig. 6.4.

"Permanent coating" ion-interaction chromatography A quite distinct alternative to the dynamic coating method can also be used. In this approach, a very lipophilic IIR is used to initially equilibrate the stationary phase and is then removed from the eluent in the actual separation step [5, 35, 401. The equilibration process establishes a very strongly bound coating of IIR on the stationary phase and this coating persists for long periods of subsequent usage. For this reason, the method is known as "permanent coating" ion-interaction chromatography. Since the stationary phase has now been converted into an ion-exchanger by virtue of the adsorbed IIR, the eluents used in the separation step are identical to those employed with conventional fixed-site ion-exchange materials (see Chapter 4). Permanent coating ion-interaction chromatography has a number of attractive features when compared with conventional ion-exchange chromatography. These include:

The ion-exchange capacity of the column can be varied over a wide range by altering the composition of the equilibrating solution. Parameters which may be varied are the lipophilicity and concentration of the IIR and the percentage of organic solvent in the equilibrating solution. (ii) The adsorbed layer of IIR can be removed or renewed as desired. Removal of the adsorbed coating can be accomplished by washing the column with an organic solvent, such as methanol or acetonitrile. (iii) The same column can be converted into an anion-exchanger or a cationexchanger through the use of an appropriate IIR.

(i)

176

r~

0

Chapter6

2

I

6

I

I

10 14 Time (min)

1

18

1

22

Fig. 6.4 Separation of anions by dynamic coating ion-interaction chromatography using a micellar eluent. A Spherisorb c18 column was used with 0.01 M cetyltrimethylammonium chloride (buffered at pH 6.8) and acetonitrile (35%) as eluent. Detection was by UV absorption at 205 nm. Reprinted from [53] with permission.

(iv) The high chromatographic efficiency of reversed-phase packing materials is retained when these packings are converted into permanently coated ionexchangers. Table 6.4 lists some of the IIRs used for permanent coating ion-interaction separations of anions and cations. It will be noted that cetyltrimethylammonium is listed as an IIR in Table 6.4 for the permanent coating method and also appears in Table 6.2 for the dynamic coating method. However, when used for permanent coating, cetyltrimethylammonium is not present in the eluent during the separation step. Permanent coating of the column is usually achieved by passing a solution (approximately M)of the IIR in dilute (5%) methanol or acetonitrile through the column for about 20 min. The purpose of the organic solvent is to wet the surface of the lipophilic stationary phase in order to improve binding of the IIR. This coating procedure typically results in the immobilization of about 50 mg of IIR onto a 25 cm column [401, giving an ion-exchange capacity similar to that for fixed-site ionexchangers used in IC. These coated columns have been found to be stable for at least one week [5].


177

TABLE 6.4 REAGENTS USED AS IIRs IN PERMANENT COATING ION-INTERACTION CHROMATOGRAPHYOF INORGANIC ANIONS IIR

structure

References

CH3(m2)15(m3)3N+ ~3(cH2)15N+c5HS ~3t~3(cH2)1113N+ [CHdCH2)714N+ CH3[CH3(m2)713N+

75-79 37,40. 80-84 5, 85 5, 82 5 86.87

Anions Cetyltrimethylammonium Cetylpyridinium Tridodecylmethy l a i m Tetraoctylammonium Trioctylmethylammonium Methyl Green

Cations Eicosanesulfate

As mentioned earlier, the eluents used in permanently coated ion-interaction chromatography are the same as those employed in IC using ion-exchange columns. Thus, aromatic carboxylate ions, such as phthalate [77, 881, hydroxybenzoate [86], salicylate [81] and aimesate [80] are commonly used for anion separations, and tartrate has been used for cations [5]. Fig. 6.5 shows typical chromatograms for the separation of inorganic anions and cations using the permanent coating ion-interaction method. 6.3.3 Role of the counter-ion of the IIR The counter-ion of the IIR fills a very important role in dynamic coating ioninteraction chromatography of anionic solutes. This counter-ion usually acts as an ionexchange competing anion and is responsible for the elution (and in many cases also the detection) of the solute anions. Typical counter-ions are hydroxide [38], fluoride [29], chloride [53], perchlorate [49], bromide [24, 441, phthalate [23, 891, citrate [26] and salicylate [90]. The nature of the counter-ion determines the type of separation which is achieved and the following strengths of counter-ions in reducing the retention of anionic solutes has been reported for a PRP-1 column using a quaternary ammonium salt as the IIR [24,91]:

The counter-ion of the IIR also influences the detection modes which are applicable to a particular separation. This occurs in exactly the same manner as applies in ionexchange chromatography with fixed-site exchangers. Thus, counter ions such as citrate, phthalate and hydroxide are suitable for conductivity detection; hydroxide, fluoride and chloride are suitable for direct spectrophotometric detection; and phthalate is suitable for

CI-

cu

I

02-

k: I

I

I

0

L

8

I

I

12 16 Time (min) (a I

I

I

20

2L

r

0

I

I

50

I

I

I

I

150 100 Time Is) (b)

I

I

200

I

1

250

Fig. 6.5 Separation of (a) inorganic anions and (b) inorganic cations using permanent coating ion-interaction chromatography. (a) A Hamilton PRP1 column coated with cetyltrimethylammonium bromide was used with 24% methanol - 1 mM potassium hydrogen phthalate - 20 mM Tris buffer (pH 9.3) as eluent. Detection was by indirect spectrophotometry at 282 nm. Reprinted from [84] with permission. (b) A C18 column coated with C20H41S04Na was used with 75 mM tartrate at pH 3.4 as eluent. Detection was by spectrophotometry at 530 nm after post-column reaction. Reprinted from [5 ] with permission.

179

Ion-InteractionChromatography Tetrabutylammonium salicylate (0.4 mM) C18

20

Octylamine (5 .mM) salicylate (5 mM) C18

Cetrimide (1 mM) citrate (4 mM) C18

Cetrimide (0.1 C ) phosphate (0.1 M)

CN

Tetrapentylammonium fluoride (1 mM) PRP-1

I s20;-

Fig. 6.6 Typical retention times for anions in dynamic coating ion-interaction chromatography using c18. CN and PRP-1stationary phases. Data taken from [31,34,37,90,91].

indirect spectrophotometric detection. Fig. 6.6 shows some typical retention times for anionic solutes in dynamic coating ion-interaction chromatography. It is not essential that the counter-ion of the IIR serves as the ion-exchange competing anion. An alternative approach is to use a separate eluent component, such as phosphate [92], citrate [26, 661, oxalate 1661 or phthalate [27], for this purpose. This method is sometimes used to assist in the elution of strongly retained ions. The nature of the counter-ion of the IIR is of less importance in ion-interaction chromatography of cations. The reason for this is that the elution of solute cations is usually accomplished with the aid of a complexing ligand, such as a-hydroxyisobutyric acid 1681, which is added to the eluent. 6.3.4 Summary of eluent and stationary phase effects

The discussion thus far has indicated a number of parameters which affect the adsorption of the IIR onto the stationary phase in ion-interaction chromatography. These parameters are summarized below, together with some other factors which influence the retention of solutes:

180

Chapter 6

Fig. 6.7 Dependence of the capacity factor of inorganic anions on the concentration of IIR in the eluent. A Partisil ODs-3 column was used with an eluent containing 1.5 mM phthalate and the indicated concentrationsof tetrabutylanumniurniodide. Reprinted from [27] with permission.

(i) (ii) (iii) (iv) (v) (vi) (vii)

The nature of the stationary phase. The lipophilicity of the IIR. The concentration of the IIR in the eluent. The ionic strength of the eluent. The nature of the competing ion in the eluent. The concentration of the competing ion in the eluent. The eluent pH.

The first four of these factors will determine the surface concentration of the IIR on the stationary phase, and hence the surface charge density and the effective ionexchange capacity. The higher the surface concentration of IIR, the greater is the retention of solutes having a charge sign opposite to that of the IIR. Thus, retention times will increase as the lipophilicity of the IIR is increased and as the percentage of modifier in the eluent is decreased. We can also note that solute retention generally increases with the concentration of IIR in the eluent, but there is a threshold concentration above which solute retention decreases with further increases in the concentration of IIR. This retention pattern is illustrated in Fig. 6.7 and the reasons underlying this behaviour will be explored fully in Section 6.4.1 below. At this stage, it will be sufficient to note that the stationary phase surface becomes saturated with IIR and any further addition of IIR to the eluent results in decreased retention because of the increased concentration of the IIR counter-ion. The nature and concentration of the eluent competing ion (whether this is the counter-ion of the IIR or an ion which is added separately) will determine the retention times and elution order for solute ions. Increases in the concentration of the eluent

Ion-InteractionChromatography

181

competing ion will result in decreased solute retention, in the same manner as observed for ion-exchange separations. Finally, the eluent pH will influence the charges on the competing ion and the solutes, provided that these species are weak acids or bases. An example of this effect is the influence of pH in an ion-interaction chromatographic system using tetrabutylammonium as the IIR and phthalate as the competing anion. Increases in eluent pH over the range 4.0-6.0 cause a decrease in solute retention as a result of increased ionization of phthalate, leading to the formation of a strong, divalent competing anion. 6.3.5

Guidelines for eluent selection in ion-interaction chromatography

It is apparent that selection of the correct eluent composition in ion-interaction chromatography requires consideration of all of the factors discussed in the preceding section. This is especially true of the dynamic coating approach, in which small changes in eluent composition can often result in large variations in retention times. Guidelines for eluent selection in this mode of chromatography are available [8], and are summarized below.

Anion separations The most commonly used IIRs are tetraalkylammonium salts. The more lipophilic IIRs (i.e. those with longer alkyl substituents) are best suited to the separation of NOa-, Br and NO3-, whilst the less lipophilic IIRs are hydrophilic anions, such as F,0, best suited to the separation of hydrophobic anions, such as aromatic sulfonates or sulfates. The counter-anion of the IIR must be selected on the basis of the desired ionexchange competing effects and the detection mode which is to be employed. For example, counter-anions such as phthalate, salicylate or Br- will result in shorter solute retention times than F- and OH, but the use of OH- as the counter-anion is necessary if suppressed conductivity detection is to be employed. The degree of retention which is achieved can then be manipulated by varying the type and amount of organic modifier added to the eluent. Further changes in solute retention times can be accomplished by addition to the eluent of an additional competing anion (e.g. S042-, C032-or Ct), sometimes referred to as an "inorganic modifier", or by varying the eluent pH when the solutes are weak acid anions. Cation separations The major factors to be considered in the separation of cations by ion-interaction chromatography are the type of IIR used, the nature of the eluent ligand, and the eluent pH. Aliphatic sulfonic acids are the most commonly used IIRs and the lipophilicity of the IIR (as determined.by the length of the alkyl chain) exerts a strong effect on solute retention. Elution of solute cations is achieved predominantly by complexation with the eluent ligand, so the conditional formation constants for the solutes are of prime importance. These conditional formation constants are determined by the nature and concentration of the ligand and by the eluent pH. Ligands such as citrate, tartrate, oxalate and a-hydroxyisobutyric acid are suitable, with each of these species showing increased complexation as the eluent pH is raised. This effect occurs only until

Chapter6

182

ionization of the ligand is complete, beyond which point further increases in pH do not significantly alter retention times. 6.4

6.4.1

RETENTION MODELS FOR DYNAMIC COATING IONINTERACTION CHROMATOGRAPHY

Model for anion retention

The following retention model for ion-interaction chromatography under dynamic coating conditions was proposed originally by Iskandarani and Pietrzyk [24] and later modified by Xianren and Baeyens [27]. The model is based on adsorption of the IIR onto the stationary phase. We will consider the situation where a lipophilic stationary phase is equilibrated with an eluent consisting of an IIR (which will be assumed to be a quaternary ammonium salt, designated as Q'C-) and a competing anion, A-:

where A, is the number of free adsorption sites on the stationary phase and the subscripts m and r refer to the mobile (eluent) and stationary phases, respectively. The equilibrium constant for eqn. (6.3) will be denoted by K1 and is given by:

The ion-exchange equilibrium between the competing anion (A') in the eluent and the counter-ion, C-, is given by: (6.5)

and the equilibrium constant, K2,for this equation can be written:

If a solute anion, X-, is now introduced into the chromatographic system, we can write an ion-exchange equilibrium as follows:

and the equilibrium constant, K3, for this equation can be written:

ton-Interaction Chromatography

183 (6.8)

The sorption capacity, KO,for the stationary phase is a measure of the total number of sites that can be occupied in the retention process. We can write a mass balance equation which accounts for all occupied and free sites, as follows:

Now the capacity factor, kx’,for solute X- can be written as: (6.10)

where q is the phase ratio. Combining eqns. (6.3)-(6.9) with elimination of Ar, (QA), and (QB), and substitution of the solution for (QX), into eqn. (6.10) gives:

which can be rearranged to give:

Eqn. (6.12) predicts that the reciprocal of the capacity factor is linearly related to [C-],, [A-lm and [X-lm, and inversely related to [Q+]m.

Effect of [AA‘],,, and [X-1, Fig. 6.8 provides a plot of the reciprocal of capacity factor versus the concentration of competing anion (i.e. [A-lm) in the eluent. In this example, the competing anion was phthalate and each of the parameters [Q+]m, [C-], and [X-], was held constant. The predicted linear relationship between the reciprocal of capacity factor and [A-]m is observed. Fig. 6.9 shows a plot of the reciprocal of capacity factor versus the amount of sample injected (i.e. [X-],), with the parameters [Q+Im,[C-], and [A-]m being held constant. Again, the predicted linear relationship is observed. It is important to note that the amounts of injected sample shown in Fig. 6.9 are relatively high and when smaller amounts of sample are injected, the reciprocal of capacity factor does not alter appreciably when the sample size is varied [24].

184

Chupter 6

30

1

0.0

: 1

I

I

2

I

I

3

I

1

I

[KHP] lmM1 Fig. 6.8 Dependence of the capacity factor of inorganic anions on the concentration of the competing anion bhthalate) added to the eluent. A Partisil ODs-3 column was used with an eluent containing 1 mM tetrabutylammonium iodide and the indicated concentrations of potassium hydrogenphthalate. Reprinted from [27] with permission.

Effect of the concentration of IIR in the eluent Eqn. (6.12)'also allows us to examine the effect of increasing the concentration of IIR in the eluent [27]. Increasing the concentration of the IIR (i.e. Q+C-)will increase simultaneously both [Q+Im and [C-],. Two opposing effects are predicted from eqn. (6.12). The f i s t is an increase in analyte retention due to increased adsorption of IIR onto the stationary phase, so we expect the reciprocal of the solute capacity factor to be linearly related to the reciprocal of [Q+]m. The second is a decrease in analyte retention where the reciprocal of the capacity factor is expected to be linearly related to [C-1,. The observed changes in retention which accompany an increase in the eluent concentration of the IIR were illustrated in Fig. 6.7. We can now use eqn. (6.12) to rationalize this behaviour, but first this equation can be rewritten by using eqn. (6.4) to obtain the following expression for [C-],: (6.13)

which can be substituted into eqn. (6.12) to give:


185

0.180.16

-

Amount injected (mg)

Fig. 6.9 Dependence of the capacity factor of inorganic anions on the concentration of the analyte. A PRP-1 column was used with 1 mM tetrapentylammoniumfluoride in 1:3 acetoniaile-wateras eluent. Reprinted from [91] with permission. When [A-]m and [X-1, are constant, eqn. (6.14) can be simplified to: (6.15)

where [Q+]m can be considered to be the amount of IIR not adsorbed at equilibrium, (QC), is the amount of IIR adsorbed at equilibrium and A, is the number of free sites at equilibrium. We will now consider the effects of increasing the eluent concentration of the IIR. At low IIR concentrations, [Q+]m is low, Ar is high and (QC), is small compared to A,. From eqn. (6.15), this gives:

1 a-

, or kx a

I

kx

[Q'lm

(6.16)

[Q'Im

This relationship is evident in the early part of Fig. 6.7. When half of the available adsorption sites on the stationary phase are occupied, then A, and (QC), are equal, so that: (6.17)

This means that the slope of the plot of kx' versus the concentration of IIR is half that observed at lower eluent concentrations of IIR. We can see that this slope will decrease progressively as more IIR is adsorbed. This process continues until a

186

Chapter 6

maximum in the plot is attained, as in Fig. 6.7. At this point, A, is very small in comparison to (QC),, such that: (6.18)

Considering the relative magnitudes of A, and (QC),, the slope of the plot of capacity factor versus the concentration of IIR will be very small, or even zero. Further increases in the concentration of IIR will not result in further adsorption of Q+ onto the stationary phase since all of the available adsorption sites have been exhausted. However, [C-1, will continue to rise, so that the solute capacity factor will fall in accordance with eqn. (6.12).

6.4.2 Model for cation retention In order to develop a retention model for cations in dynamic coating ioninteraction chromatography, we recognize that the eluent composition used in this case usually differs from that employed for a typical anion separation. Eluents for anion separations contain a cationic IIR and its counter-anion, together with an added competing anion. On the other hand, eluents for cation separations contain an anionic IIR and its counter-cation, together with an added ligand which assists in solute elution by complexation effects. This difference between anion and cation eluents also exists in ion-exchange separations (see Chapter 4) and arises because of the strong ion-exchange affinities of many cations. We can therefore consider the situation where a lipophilic stationary phase is equilibrated with an eluent comprising an IIR (which will be assumed to be an aliphatic sulfonic acid salt, designated as P-C+)and a ligand, L-. Using the same approach adopted above for anions. we can write:

A,

+ P, + C,+

% (PC),

(6.19)

where A, is again the number of free adsorption sites on the stationary phase and the subscripts r and m denote the stationary and eluent phases, respectively. The equilibrium constant for eqn. (6.19) will be denoted by Kq and is given by: (6.20)

When a solute cation, X+, is introduced, the following ion-exchange equilibrium exists:

(PC), +

x;

% (PX), +

c,+

and the equilibrium constant (Ks)for this equation can be written:

(6.21)


187

(6.22)

The solute cation will also participate in complexation reactions with the eluent ligand, L-,so that the solute X will exist as both the free ion (i.e. X+) and as a complex. If we define ax as the fraction of the total concentration of solute cation existing as the free ion, X+, then we have:

ax =

R'Im -

(6.23)

[X'IT

where [X+]T is the total concentration of solute X in the eluent, regardless of whether it is present in the free or complexed forms. We again write a mass balance equation in terms of the sorption capacity, KO, of the stationary phase:

KO = A,

+ (PC), + (PX),

(6.24)

The capacity factor kx' of solute X+ can be written in terms of the phase ratio, q,

as: (6.25)

Combining eqns. (6.19) - (6.25) gives: (6.26)

Eqn. (6.26) represents rhe simplest case in which the solute cation and the countercation of the IIR are both singly charged. The form of this equation will therefore alter when the solute cation has a multiple charge. Nevertheless, from eqn. (6.26) we can predict that the capacity factor for a solute cation will be directly proportional to ax and [P-lm, and inversely proportional to [C'], and [X+]m. Few published data are available to investigate the validity of these relationships for inorganic cations, despite the fact that this separation mode is commonly employed for transition metal and lanthanide ions. However, a study of the retention behaviour of transition metal cations on a reversedphase column using octanesulfonate as the IIR and oxalate as the eluent ligand has been reported [93]. This work shows that there is a linear relationship between log kx' and log ax when all other variables are held constant (in accordance with eqn. (6.26)), and that a plot of log kx' versus log [Pa], (illustrated in Fig. 6.10), again with all other variables held constant, has the same general shape as that observed for anions. That is, an initial proportionality exists between log kx' and log [P-lm, but the curve reaches a

188

Chapter6

-0.2 O.O\

I

-0.4 -2h

I

-2.5

I

-2.6

I

-2.7 log

rw

I

-2.8

1

-29

1

-3.0

Fig. 6.10. Dependence of the capacity factor of inorganic cations on the concentration of IIR in the eluent. A Waters pBondapak Clg column was used with an eluent containing 2.5 m M oxalic acid at pH 3.4 and the indicated concentrations of octanesulfonate. Reprinted from [93] with

permission. maximum due to saturation of the available adsorption sites with P- and the increased concentration of the counter-cation, C+. in the eluent. These results are in general agreement with eqn. (6.26). 6.5

APPLICATIONS

Ion-interaction chromatography has found extensive application in the separation of inorganic anions and cations. The technique offers some advantages over the use of fixed-site ion-exchangers in that there is a wide range of eluent variables which can be used to manipulate the retention of solutes. For this reason, ion-interaction chromatography is often applied to the resolution of difficult mixtures of solutes. Numerous examples of such applications can be found in Part V of this book, however some representative applications are listed in Table 6.5 in order to illustrate the scope of the technique. As a general observation, it can be said that ion-interaction chromatography finds its strongest usage in the separation of transition metal and lanthanide cations, for which it is undoubtedly the method of choice. These species may be separated as simple, hydrated metal ions, or as anionic complexes, using a suitable ligand. The first of these approaches has been developed extensively by Cassidy and co-workers and was illustrated in Fig. 6.3(b), which shows an excellent separation of lanthanides. The

ton-Interaction Chromarography

189

second approach is illustrated in Fig. 6.11, which shows the separation of anionic metal cyan0 complexes. Both of these methods have been applied to a wide variety of complex sample matrices.

TABLE6.5 APPLICATIONS OF ION-INTERACTION SEPARATIONSIN IC Solute(s)

Sample

Stationary phasea

IIRb

Det'n Ref methodc

9 8

TBA sulfate TBAphosphate

DSpec ICP

94 95

ISpec DSpec ISpec C ISpec DSpec Amp DSpec

75

Anions Alkylbenzene sulfonates A s O ~ ~organo-, arsenic compounds Carboxylic acids CI-, NO^-, ~043-, ~ 0 4 2 EDTA, ~2042-, citrate F-, C104~-, BF4-, F,cI-, NO^, ~ 0 4 2 IIIO~-.BIO.~-, NO~-,N%-, I-, SCN, Fe(CN)64N&; Br-, NO3N@-,NO3P043-, Cl-, Bf, NOg',

Detergents Shale oil

c18

Lysimeter solns Plants, soils Reactor water Plating baths Tapwater salt Serum various

CIMA chlorided TBA hydroxide PRP- 1 8chlorided MPIC-NS1 TBA hydroxide TBA phthalate C18 Ouylamine c18 HDTMA chloride ClS PS-DVB CTMA citrate

Foods PRP- 1 Meat,vegetables Ci8 C18 Fruit juices

TPAbromide TBAphosphate TBAsalicylate

DSpec DSpec C

98

Pharmaceuticals Urine Batteries

Amp

47 92

c8

TBA sulfate TBA sulfate OctylamineTSA

DSpec

SCN-

ISpec

51

Cis MPIC-NS1 MPIC-NS1 C18

Octanesulfonate Hexanesulfonate TBA hydroxide TBAhydroxide

PCR C C DSpec

68

c18

Hexanesulfonate

PCR

103

I-,so~~s&-. Iso42-

CIS cl8

CIS c18

96 80 30

97 50 54 55

99 87

Cations

Rare earths Ethanolamines Au(I), Au(III) cyanides CN- complexes of transition metals Transition metals a

Leach liquor Ambient air Plating baths Gold process solutions Brass, urine

MPIC-NS1, PRP-1 and PS-DVB are all styrene-divinylbenzenestationary phases. TBA = tetrabutylammonium, CIMA = cetyltrimethylammonium,CP = cetylpyridinium, HDTMA = hexadecyltrimethylamnium, TPA = tetrapentylammonium, TSA = toluenesulfonate. DSpec = direct spectrophotometry, ISpec =indirect spectrophotometry, C = conductivity, Amp = ampemmeay, PCR = post-column reaction, ICP = inductively coupled plasma. Permanent coating ion-interaction chromatography.

100 101 102

190

Chapter 6

Pd(ll1

k PliIII

0I

10 I

20

Time imin

30

LO

I

Fig. 6.11 Ion-interaction separation of metal-cyano complexes. A Waters Nova Pak C18 column was used with 23:77 acetonitrile-water containing 5 mM Waters Low UV PIC A as eluent. Detection was by direct spectrophotometry at 214 nm. Reprinted from [38] with permission.

6.6 1 2 3 4 5 6

7 8 9 10 11 12 13

14

REFERENCES Bidlingmeyer B.A., J. Chromatogr.Sci., 18 (1980) 525. Billiet H.A.H., Drouen A.C.J.H. and De Galan L., J . Chromatogr., 316 (1984) 231. Paired Ion Chromatography, an Alternative to Ion-Exchange, Waters Bulletin F61, May, ( I 976). Tomlinson E., Jefferies T.M. and Riley C.M., J. Chromatogr., 159 (1978) 315. Cassidy R.M. and Elchuk S., Anal. Chem., 54 (1982) 1558. Bidlingmeyer B.A., Deming S.N.. Price W.P., Jr., Sachok B. and Petrusek M., J . Chromarogr. Sci., 186 (1979) 419. Horvath C., Melander W., Molnar I. and Molnar P., Anal. Chem., 49 (1977) 2295. Dionex Technical Note 12R. Waters Chromatography Division, Milford, MA USA. Wittmer D.P., Nuessle N.O. and Haney W.G., Jr, Anal. Chern., 47 (1975) 1422. Horvath C., Melander W. and Molnar I., J . Chrornurogr., 125 (1976) 129. Kroeff E.P. and Pietrzyk D.J., Anal. Chem., 50 (1978) 502. Kraak J.C., Jonker K.M. and Huber J.F.K., J. Chrornutogr.,142 (1977) 671. Hoffman N.E. and Liao J.C., Anal. Chem., 49 (1977) 2231.

Ion-InteractionChromatography 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57

191

Kissinger P.T., Anal. Chern., 49 (1977) 883. Cantwell F.F. and Puon S., Anal. Chern., 51 (1979) 623. Stahlberg J. and Hagglund I., Anal. Chem., 60 (1988) 1958. Stahlberg J., J. Chromatogr., 356 (1986) 231. Deelder R.S. and van den Berg J.H.M.. J. Chromutogr.,218 (1981) 327. Bartha A. and Vigh Gy., J. Chromatogr.,395 (1983) 503. Bartha A., Billiet H.A.H., De Galan L. and Vigh Gy., J. Chromatogr.,291 (1984) 91. Bartha A., Vigh Gy., Billiet H. and De Galan L., Chromatographia,20 (1985) 587. Barlha A., Vigh Gy., Billiet H.A.H. and De Galan L., J. Chromatogr.,303 (1984) 29. Iskandarani Z. and Pietrzyk D.J., Anal. Cheni., 54 (1982) 1065. Barber W.E. and Carr P.W., J. Chrornatogr..260 (1983) 89. Mingjia W., Pacakova V., Stulik K., Sacchetto G.A., J. Chromatogr.,439 (1988) 363. Xianren Q. and Baeyens W., J. Chromatogr..456 (1988) 267. Smith R.L. and Pietrzyk D.J., Anal. Cheni., 56 (1984) 1572. Smith R.L., Iskandarani Z. and Pietrzyk D.J..J. Liq. Chroniutogr.,7 (1984) 1935. Dionex Application Note 41. Wheals B.B., J. Chronturogr., 262 (1983) 61. Schwedt G., Chromatographia,12 (1979) 613. Crommen J.. Schill G., Westerlund D. and Hackzell L., Chronutographia,24 (1987) 252. Cassidy R.M. and Elchuk S., J. Chromutogr. Sci., 21 (1983) 454. Reeve R.N.,J. Chrornatogr.,177 (1979) 393. Dasgupta P.K.. Anal. Chem.. 56 (1984) 769. Gennaro M.C., J. Chromatogr.,449 (1988) 103. Hilton D.F. and Haddad P.R., J. Chromatogr,,361 (1986) 141. Molnar I., Knauer H. and Wilk D.. J. Chronurogr.,201 (1980) 225. Cassidy R.M. and Elchuk S., J. Chromatogr., 262 (1983) 31 1. De Kleijn J.P., Analyst (London), 107 (1982) 223. Rochester N.E. and Haddad P.R., Proc. 9th. Aust. Symp. Anal. Chern., (1987) p. 329. Andrew B.E.. LC.GC, 4 (1986) 1026. Rabin S.B. and Stanbury D.M., Anal. Chem., 57 (1985) 1130. Weiss J., Mockel H.J.. Muller A., Dicmann E. and Walberg H.-J., J. Chrornatogr.,439 (1988) 93. Rotsch T.D. and Pietrzyk D.J.. J. Chrontarogr. Sci., 19 (1981) 88. Lookabaugh M., Krull I.S. and LaCourse W.R., J. Chrornatogr..387 (1987) 301. Gennaro M.C., J. Liq. Chromatogr.. 10 (1987) 3347. Dreux M., Lafosse M. and Pequignot M., Chronutographia.15 (1982) 653. De Kleijn J.P., Deutsche Leben. Rundsch.,79 (1983) 184. Dreux M., Lafosse M., Pequignot M., Moiin-Allory L. and Douady M., J. HRC & CC,7 (1984) 712. Irgolic K.J., Stockton R.A., Chakraborti D. and Beyer W., Spectrochint.Acto, 38B (1983) 437. Mullins F.G.P. and Kirkbright G.F., Analyst (London), 109 (1984) 1217. Hurst W.J., Evans S.L., White W.W. and'Miller K.L., LC.GC, 6 (1988) 5. Wheals B.B., J. Chronutogr., 402 (1987) 115. Mangia A. and Lugari M.T.,Anal. Chim A m , 159 (1984) 349. Lookabaugh M. and Krull LS., J. Chronmrogr., 452 (1988) 295.

192 58 59 60 61 62 63

Chapter6

Hammers W.E., Aussems C.N.M. and Janssen M., J. Chromatogr., 360 (1986) 1. Barber W.E. and Cam P.W., J . Chromatogr.. 301 (1984) 25. Barber W.E. and Can P.W., J. Chromarogr., 316 (1984) 211. Arvidsson E., Crommen J., Schill G. and Westerlund D., Chromatographia,24 (1987) 460. Rigas P.G. and Pietnyk D.J.. Anal. Chem., 59 (1987) 1388. Rigas P.G. and P i e q k D.J., Poster presented at "11thInternational Symposium on Column Liquid Chromatography",Amsterdam, 1987. 64 Rigas P.G. and Pietnyk D.J., Anal. Chem., 60 (1988) 454. 65 Rigas P.G. and Pietnyk D.J., Anal. Chem., 60 (1988) 1650. 66 Kirk A.D. and Hewavitharana A.K., Anal. Chem., 60 (1988) 797. 67 Krull I.S., Bushee D., Savage R.N., Schleicher R.G. and Smith S.B., Jr., Anal. Left., 15 (1982) 267. 68 Barkley D.J., Blanchette M., Cassidy R.M. and Elchuk S., Anal. Chem.,58 (1986) 2222. 69 Bushee D., Krull I.S., Savage R.N. and Smith S.B., Jr., J. Liq.Chromatogr.. 5 (1982) 463. 70 Lavine B.K., McMillan S., Ward A.J.I. and Donoghue O., in Jandik P. and Cassidy R.M.(Eds.) Advances in Ion Chromarography,Vol. 1, Century International, Inc., Franklin, MA, 1989, p. 195. 71 Hackzell L., Rydberg T. and Schill G., J. Chromatogr.,282 (1983) 179. 72 Smith R.M., Anal. Proc., 21 (1984) 73. 73 Smith R.M. and Yankey L.E., Analyst (London), 107 (1982) 744. 74 Rigas P.G. and Pietnyk D.J., Anal. Chem., 58 (1986) 2226. 75 Barkley D.J., Dahms T.E. and Villeneuve K.N., J. Chromarogr.,395 (1987) 631. 76 Frohlich D.H., J. HRC & CC, 10 (1987) 12. 77 Mullins F.G.P., Analyst (London), 112 (1987) 665. 78 Takeuchi T. and Yeung E.S.,J . Chromatogr.,370 (1986) 83. 79 Takeuchi T., Suzuki E. and Ishii D., J. Chromatogr.,447 (1988) 221. 80 Cassidy R.M. and Elchuk S . , Anal. Chem.,57 (1985) 615. 81 Heffer W.D., Takeuchi T. and Yeung E.S., Chromntographia,24 (1987) 123. 82 DuVal D.L. and Fritz J.S., J. Chromatogr., 295 (1984) 89. 83 Papp E., J . Chromarogr.,402 (1987) 211. 84 Papp E. and Fehervari A., J. Chromatogr.,447 (1988) 315. 85 Al-Omair A.S. and Lyle S.J., Talantu,34 (1987) 361. 86 Golombek R. and Schwedt G., J. Chromatogr.,452 (1988) 283. 87 Schmuckler G., Rossner B. and Schwedt G., J . Chromatogr., 302 (1984) 15. 88 Fuchtner F. and Schmidt W., Z. Chem., 28 (1988) 149. 89 Pilkington A.E. and Waring R.H., Med. Sci. Res., 16 (1988) 35. 90 Bidlingmeyer B.A., Santasania C.T. and Warren F.V., Jr.. Anal. Chem., 59 (1987) 1843. 91 Iskandarani Z and Pietnyk D.J., Anal. Chem..54 (1982) 2427. 92 Kalbasi M. and Tabatabai M.A., Commun Soil Sci. Plant Anal., 16 (1985) 787. 93 Haddad P.R. and Foley R.C., J. Chromarogr., 500 (1990) 301. 94 Bear G.R., J. Chromatogr., 371 (1986) 387. 95 LaFreniere K.E., Fassel V.A., and Eckels D.E., Anal. Chem., 59 (1987) 879. 96 Bossle P.C., Reutter D.J. and Sarver E.W., J . Chromurogr.,407 (1987) 399. 97 Perrone P.A. and Grant J.R., Res. Do..September (1984) 96. 98 Iskandarani Z. and Pietnyk DJ., Anal. Chem.,54 (1982) 2601.

Ion-InteractionChromatography 99 100 101 102 103

Wootton M., Kok S.H.and Buckle K.A., J . Sci. Food Agric.. 36 (1985) 297. Bouyoucos S.A. and Melcher R.G..Am. I d . Hyg.Assoc. J., 47 (1986) 185. Dionex ApplicationNote 4OR. Grigorova B., Wright S.A. and Josephson M., J. Chromutogr.,410 (1987) 419. Schmidt G.J. and Scott R.P.W.,Analyst (London), 109 (1984) 997.

193


195

Chapter 7 Ion-Exclusion Chromatography 7.1

INTRODUCTION

Ion-exclusion chromatography, first introduc d by Wheaton and Bauman in 1953 [l], involves the use of strong anion- or cation-exchange resins for the separation of ionic solutes from weakly ionized or neutral solutes. In this mode of chromatography, the charge sign on the ion-exchange resin used is the same as that of the weakly ionized solutes. That is, solutes with a partial negative charge (such as carboxylic acids) are separated on a cation-exchange resin having anionic sulfonate functional.groups, whereas solutes with a partial positive charge (such as weak bases) are separated on an anionexchange resin having cationic quaternary ammonium functional groups. This is the opposite situation to that occumng in ion-exchange chromatography. As with other IC separation techniques, ion-exclusion chromatography has been described by a variety of names, some of which are listed in Table 7.1. Each of these names implies a mechanism for the separation process and as we will see in the ensuing discussion, the actual mechanism of the process is not clearly defined, but is certainly quite complex. We shall therefore continue to use the term "ion-exclusion chromatography" to describe the technique, whilst recognizing that this title is probably somewhat inaccurate.

7.1.1 Basic principles Ion-exclusion chromatography finds application in the separation of a wide range of small, neutral or partially ionized molecules. Applying the definition of IC used in this text, we will limit discussion of the technique to include its application to certain TABLE 7.1 ALTERNATIVENAMES FOR ION-EXCLUSION CHROMATOGRAPHY Name

Reference

Ion-exclusion chromatography Ion-chromatography exclusion (ICE) Ion-exclusion partition chromatography Donnan exclusion chromatography Ion-moderatedpartition chromatography

1 2 3 4 5

1%

Chapter 7

Fig. 7.2 Schematic representation of ion-exclusionchromatography for (a) acidic solutes, such as acetic acid and HCI, and (b) basic solutes, such as N H 3 and NaOH.

solute types only; namely, carboxylic acids, inorganic weak acid anions, weak organic bases, and water. It may appear that this restricted group of solutes could diminish the importance of ion-exclusion chromatography in comparison to ion-exchange and ioninteraction chromatography, but it will be demonstrated later in this chapter that ionexclusion chromatography is of major importance in IC. The principles of ion-exclusion chromatography can be illustrated in a schematic manner by considering the chromatographic system to be comprised of three distinct phases. The first of these is the flowing eluent, which passes between the beads of ionexchange resin (i.e. through the interstitial volume). The second zone is the polymeric network of the resin material itself, together with its bound ionic functionalities, whilst the third zone is liquid occluded inside the pores of the resin bead. The polymeric resin can be considered as a semi-permeable, ion-exchange membrane which separates the flowing eluent from the stationary occluded liquid inside the resin [ I , 31. The manner in which solutes are separated in ion-exclusion chromatography is illustrated in Fig. 7.1. We fiist consider the behaviour of two solutes, hydrochloric acid and acetic acid, on a cation-exchange resin using water as the eluent. From Fig. 7.l(a), we see that Cl- cannot penetrate into the occluded liquid phase because it is repelled by the anionic functional groups on the resin, in accordance with the Donnan exclusion effect. The C1-ions therefore remain in the flowing eluent phase and are not retained by the column. On the other hand, the acetic acid is only weakly ionized and exists predominantly as neutral acetic acid molecules, with only a small percentage present as acetate anion. The ionized and neutral acetic acid molecules are in dynamic equilibrium with each other, so that the effective negative charge on the acetic acid is therefore determined by the proportions existing in each form. In a water eluent, this effective charge is quite small and because of this, acetic acid can penetrate the negatively charged resin zone and move into the occluded liquid phase. This results in some degree of retention of acetic acid, so that it is eluted somewhat later than hydrochloric acid. As an historical note, this particular separation was that originally reported in the first publication on ion-exclusion chromatography [ 11. In a similar manner, an anion-exchange resin can be used to separate a weak base (ammonia) from a strong base (NaOH), again using water as eluent. This is illustrated in Fig. 7.l(b), which shows that Na+ is repelled by the cationic functional groups on the

Ion-ExclusionChromatography

197

resin phase and is unretained. On the other hand, ammonia, by virtue of its low degree of ionization and hence its low overall charge, can penetrate into the occluded liquid phase and is therefore retained. Fig. 7.1 suggests that retention of solutes in ion-exclusion chromatography is influenced solely by the charge on the solute. That is, all fully ionized solutes can be expected to be unretained and so be eluted together at the void volume of the column, and the retention of partially ionized solutes can be expected to increase as the degree of ionization decreases. These predictions are not fully supported in practice and it can be shown that other factors also play a role in solute retention. These factors will be discussed in Section 7.4, but at this stage we will make the assumption that solute charge is the dominant parameter in determining retention. 7.2

STATIONARY PHASES

Ion-exclusion chromatography was first performed on large particle size, high capacity, fully functionalized polystyrene-divinylbenzenepolymers. Modem stationary phases are essentially the same materials, but differ in some important respects. Some of the stationary phase parameters which can exert an influence on solute retention are: (i) (ii) (iii) (iv)

Particle size. Ion-exchangecapacity. Resin structure. Degree of resin cross-linking.

As with other chromatographic techniques, the separation efficiency is strongly influenced by the particle size of the column packing material. Modem ion-exclusion chromatography is generally performed on 5 or 10 pn particles, however some of the resins used are relatively soft and the use of small diameter particles means that eluent flow-rates must be kept low to avoid compression of the resin bed. The ion-exchange capacity of a typical ion-exclusion packing material is generally greater than that of packings used for ion-exchange separations in IC. These high capacity materials are preferred so that the number of functional groups on the resin is sufficient to exert an appropriate Donnan exclusion effect. The resin structure is also important, with microporous or gel-type resins being most commonly used (for a description of this type of resin, see Section 3.3). However, there has been a recent trend towards macroporous materials of greater rigidity [ 6 ] . Early studies [3] showed that the degree of cross-linking of the resin exerts a considerable effect on solute retention. Highly cross-linked materials (e.g. those with 812% divinylbenzene) show stronger Donnan exclusion effects than resins of lower crosslinking. This means that both strongly and weakly ionized species show more penetration into thk occluded liquid phase on softer resins of low cross-linking (e.g. those with 2% divinylbenzene). Most commercial ion-exclusion resins have approximately 8% cross-linking, with materials of lower cross-linking being used for the separation of those solutes likely to show higher degrees of ionization under the experimental conditions used (e.g. acids in the pKa range 2-4). Ion-exclusion columns

Chapter 7

198

TABLE 7.2 CHARACTERISTICS OF SOME TYPICAL ION-EXCLUSION COLUMNS Column

Aminex A-5

Aminex HPX-87H Brownlee Polypore H anion-exclusion Dionex HPICE-AS1 Dionex HPICE-AS2 Dionex HPICE-AS3 Dionex HPICE-ASS Dowex 50W-X2 Dowex 50W-X4 Dowex 50W-X12 Hamilton PRP-X300 Hitachi 2613 cation-exchanger Hitachi 2632 anion-exchanger Interaction ORH-801 Interaction ION-310 Sarasep WA 1 SP-Sephadex C-25 TSK-gel SCX Waters Fast Fruit Juice Waters Ion-exclusion Wescan 269-006 exclusion Wescan 269-038 exclusion Wescan 269-051 Yokagawa Elecmc YEW SCS 5-252

linking

Particle size

(a>

(vm)

Cross-

8 8

13

-b

5-8 10 10 25 25 7 38-75 75- 180 38-63 7 18 18 8 8 10 50-100

20 15 15 8 8 9 30

7 7 10 10 10 6

.b

8 8 4 6 2 4 12 -b

8 8 8 8 8

5

Ion-exchange capacity (mequiv/g)

Refa

1.7 1.7

7 8,9 10, 11 12-14 15 16 17 18, 19 20-22 3 6, 23 24-27 28 30-33 34 35 36, 37 38-41 42 43-45 46 47,48 49 50,51

-b

3-5 3-5 3-5 1.5

1.1 0.6 2.1 0.17 4.5 -b -b -b

3.0 4.5 4.2 5.0 5.0 1.4 1.4 1.8 4.5

a These references include some publications in which the indicated columns have been utilized.

Data unavailable. are usually large in comparison to conventional IC columns because a considerable volume of resin material is necessary to provide sufficient occluded liquid phase to permit the separation of solutes of similar size and charge. A typical column would be 30 cm in length, with an internal diameter of 7 mm or more. The characteristics of some representative ion-exclusion packing materials are listed in Table 7.2. Cation-exchange resins are generally used in the H+ form, whilst anion-exchangers are used in the OH- form.

lon-Exchion Chromatography

7.3

199

ELUENTS

7.3.1 Water eluents The eluents used in ion-exclusion chromatography are often very simple in composition. Most of the early work was performed using deionized water as eluent [2, 18, 22,521 and the degree of ionization of the solutes (and hence their retention times) is therefore determined by their pKa or pKb values. The limitations of water as an eluent are that stronger acids or bases show too great a degree of ionization to be retained and the peak shape obtained for solutes which are retained is often poor. This is illustrated in Fig. 7.2(a) for carboxylic acid solutes. In view of the problems observed with peak shape, water is rarely used as an eluent in modem ion-exclusion chromatography, except for the separation of very weakly ionized compounds. An example of such a separation is given in Fig. 7.2(b).

7.3.2

Acid eluents

Following from the original suggestion by Turkelson and Richards [22], it is now common for dilute solutions of strong mineral acids to be employed for the elution of anionic solutes, or dilute solutions of strong bases to be employed for the elution of cationic solutes. In this way, ion-exclusion chromatography can be extended to the separation of relatively strong acids and bases by limiting their degree of ionization. The most commonly used eluents are formed from strong acids, such as sulfuric acid [e.g. 5,7, 10.54, 551, hydrochloric acid [e.g. 13,56-591 and aliphatic sulfonic acids [8,43,45, 60,611. When sulfuric acid is used as the eluent, detection of eluted solutes is generally accomplished by monitoring UV absorbance at low wavelength (200-220 nm). Fig. 7.2(c) shows a typical chromatogram obtained with this eluent. On the other hand, hydrochloric acid is most often used with conductivity detection, after the eluent is passed through a suitable suppressor (see Section 9.5). Aliphatic (and aromatic) sulfonic acids can also be employed for conductivity detection, but because of the relatively low background conductance of these eluents, suppression is not necessary. Weak acids may also be utilized as eluents in ion-exclusion chromatography. Examples include phosphoric acid [62-641, tridecafluoroheptanoic acid (perfluorobutyric acid) 1651, carbonic acid [66, 671, n-butyric acid [19] and benzoic acid [53]. It is interesting to note that eluents of the same pH, when used on the same stationary phase, produce virtually identical chromatograms, regardless of the nature of the acid used. The choice between different eluent acids is therefore governed primarily by the detection method which is to be used. We can also note in passing that organic modifiers, such as methanol, acetonitrile or acetone, are sometimes added to the eluents used in ion-exclusion chromatography. The function of these modifiers is related to the participation of solute adsorption effects in the retention process. This factor is discussed more fully in Section 7.4.3. Some typical capacity factors for carboxylic acids, obtained with various eluents and column types, are illustrated in Fig. 7.3. General trends for the retention of carboxylic acids in ion-exclusion chromatography have been reported [3] and are summarized below:

CO32-

P

loxalic

I Formic

,Lactic Propionic Ektyric

Ja

,Fumoric

Valcric

I

0

1

l

10

l

l

l

l

20 30 Time lminl la)

l

l

60

I

1

50

-

0 6 8 12 16 20 Time fminl fb)

I

I

I

0

2

4

I

I

6 8 Time fmin) Icl

I

I

10

12

11

_i!

U

1-

0

6

8121620

T i m (mid Id1

Fig. 72 Ion-exclusion chromatograms with various eluents. (a) A 5 pm TSK cation-exchange resin was used with water as eluent. Reprinted from [53] with permission. (b) A Dionex HPICE-AS1column was used with water as eluent. Reprinted from [12] with permission. (c) An Interaction ORH-801 column was used with 0.01 N H2SO4 as eluent. Chromatogram courtesy of Interaction (d) A Dionex HPICE-AS1 column was used with 50 mM mannitol as eluent. Reprinted from [12] with permission.

P u

Interaction ORH-801 10 mN H2SO4 35 o c oxalic aconitic maleic oxaloacetic a=ketoglutaric citric isocitric pyruvic tartaric. ascorbic malic lactic succjnic formic acetic

-

1

L

0

c

0

m

2-i

4

Loxalic

aconitic oxaloacetic rnaleic a detoalutaric citric isocitric pyruvic tartaric ascorbic rnalic succinic lactic

-

formic acetic Dropionic

kpropionic

Q

4

Interaction ORH-801 10 mN H2SO4 65 OC

kfumaric

'

I

Bio-Rad HPX-87H 4 rnN H2SO4 5OoC

-

oxalic maleic oxaloacetic B.-ke tog Iuta r ic

F Cp:I:U' "v c \tartaric

Benson OA850 1 mN H s O 4 25 OC a-ketoglutaric malgic -L isocitric

i

\;tl$p

-succinic /formic 'lactic -acetic adipic

-

Dionex HPICE-AS5 Perfluorobutyric acid IDH 2.8)

E

pyruvic

-

-0

oxalic

-1

tartaric orrnic

acetic

L

rnalonic ascorbic lactic rnaleic

propionic -adipic

malic

I

Fig. 7.3 Typical capacity factors for organic acids using ion-exclusion chromatography. Data courtesy of Dionex, Benson, Interaction and Bio-Rad.

I4

Chapter 7

202

(i)

Members of a homologous series, such as the aliphatic carboxylic acids, are eluted in order of increasing molecular weight, decreasing acid strength (i.e. increasing pKa) and decreasing water solubility. Thus. the elution order of low molecular weight carboxylic acids is formic c acetic c propionic. (ii) Dibasic acids are generally eluted earlier than monobasic acids of the same carbon number. For example, oxalic acid is eluted earlier than acetic acid, and malonic acid earlier than propionic acid. (iii) Carboxylic acids with branched structures are eluted earlier than the corresponding straight chain isomer. For example, iso-butyric acid is eluted earlier than n-butyric acid. (iv) A double bond serves to increase the retention of an acid. For example, acrylic acid is eluted after propionic acid.

7.3.3

Complexing eluents

The retention and detection properties of some solutes can be enhanced if a complexing agent is added to the eluent. An example of this approach is the use of a mannitol eluent for the determination of boric acid [12], in which the mannitol serves to complex the boric acid to form a species which is more easily detectable by conductivity measurements than is boric acid alone. Fig. 7.2(d) shows a typical chromatogram obtained with this approach. A further application of complexing eluents is the use of a tetraborate eluent in the determination of formaldehyde [68, 691.

7.4 7.4.1

FACTORS INFLUENCING RETENTION IN ION-EXCLUSION CHROMATOGRAPHY Degree of ionization of the solute

The degree to which the solute is ionized is the most significant factor which determines solute retention. As the solute becomes more ionized, the Donnan exclusion effect increases in magnitude and this leads to decreased retention. When only the Donnan exclusion effect is considered. solute retention (expressed as the retention volume, VR)is given by:

where Vo is the interstitial volume of eluent (i.e. the volume of eluent flowing between the particles of stationary phase), Vi is the internal volume of eluent (i.e. the volume of occluded liquid inside the pores of the stationary phase) and DA is the distribution coefficient for the solute between the interstitial eluent and the occluded liquid. The value of DA is dependent on the degree of ionization of the solute. Fully ionized solutes have DA = 0, due to the total exclusion of such solutes in accordance with the Donnan effect. The retention volume of fully ionized solutes is therefore given by:

Ion-Exclusion Chromatography

203

and we expect all solutes of this type to be eluted at the same retention volume. On the other hand, neutral solutes have DA = 1, since these solutes can distribute freely between the interstitial eluent and the occluded liquid, without influence from the Donnan effect. The retention volume for neutral solutes is therefore given by:

and again we expect all neutral solutes to be eluted at the same retention volume. Solutes which are ionized partially will be eluted at retention volumes intermediate between the two extremes given by eqns. (7.2) and (7.3), with the observed retention volume for a particular solute being dependent on the acid or base dissociation constant of that solute. The above equations are identical in nature to those which are used to describe retention in size-exclusion (gel permeation) chromatography, in which retention volumes fall between the two extremes determined by total exclusion of solutes, and total penetration of solutes.

Dependence of solute retention on pK, The effect of solute charge on retention in ion-exclusion chromatography has been examined for acidic solutes on a strong cation-exchanger (8% cross-linked) in the H+ form, using water as the eluent [27]. Under these conditions, the degree of ionization of the solute is determined solely by the acid dissociation constant (pKa) of the solute. A plot of retention volume versus pKa1 for a range of solutes is given in Fig. 7.4. The retention behaviour depicted in Fig 7.4 is in close accordance with the predictions made above in eqns. (7.1) - (7.3). Strong acids, such as HNO3, H2SO4, HCl, etc., are completely excluded from the stationary phase and are eluted at the same retention volume (12.8 ml). This volume corresponds to the volume of interstitial eluent present in the chromatographic column (Vo). Solutes which exist as neutral species in the water eluent, such as methanol, HCN and H2CO3, are eluted together at a retention volume of 28.5 ml (Vo + Vi). Solutes which are ionized partially in water, such as H3B03, HCOOH and CH3COOH, are eluted at retention volumes between 12.8 and 28.5 ml. There is a strong correlation between pKa1 and VR. Substitution of the measured values of VR,Vo and Vi into eqn. (7.1) permits the calculation of values of DA for each solute. These values are listed in Table 7.3. Inspection of the data in Table 7.3 shows that some values of DA exceed the theoretical maximum of 1.0. This behaviour is evident for propionic acid and H2S and it can be seen that these solutes show anomalous retention volumes in Fig. 7.4. The reasons underlying this will be discussed further below. A similar study of solute retention volumes has been undertaken using a highly cross-linked (30%)cation-exchange column, with acidic eluents [50]. The results of this study are presented in Fig. 7.5, which shows some of the same characteristics as Fig. 7.4, in that a fully ionized solute (H2SO4) defines the lower limit of retention volume, with neutral solutes (methanol and H2CO3) defining the theoretical upper limit of retention volume. A straight line is drawn through the points for oxalic acid, HF and H2CO3, as was done for Fig. 7.4, and it can be seen that many of the solutes tested do not conform to the retention behaviour evident from Fig. 7.4. That is, there is poor correlation

204

Ckpter 7

p Methanol

151-

E-

-vo-

0

5c

"25

I

m (Y

0

Y

n

-5

104

I

.*#.I

-10

Retention volume (mll

Fig. 7.4 Relationshipbetween retention volumes and first dissociationconstants (pKal) for acids on a stationary phase with 8% cross-linking, using water as the eluent. Reprinted from 1271 with permission. TABLE 7.3 DISTRIBUTION COEFFICIENTS FOR ACIDS, CALCULATED FROM THE RETENTION DATA SHOWN IN FIG. 7.4 [27] Acid

DA

0 0 0 0 0 0 0.01 0.06 0.08 0.09 0.11

Acid

DA

0.36

0.43 0.65 0.81 1.10 1.00 1.00 1

.oo

0.98 1.02 1.40

Ion-Excluswn Chromatography

-Vo

10

-

__z(t___

I I

205

Vi

Methanol

I I

Monocarboxylic acid

I

5-

-

Dicarboxylic acid

0

Y

a.

0-

I

I

I I I

''W O L

I I I

I

I

I

I

Fig. 7.5 Relationship between retention volume and first dissociation constant (PKal) for carboxylic acids on a stationary phase with 30% cross-linking,using 1 mM H a 0 4 as eluent. Reprinted from [50] with permission.

between pKa1 and retention volume. This behaviour indicates that retention of many solutes is influenced by parameters other than the degree of ionization of the solute.

Dependence of solute retention on eluent pH When changes in the eluent pH produce a change in the degree of ionization of the solute, we can expect this to cause a change in the retention time of that solute. This behaviour is illustrated in Fig. 7.6,for both mono- and dicarboxylic acids on a highly cross-linked cation-exchanger. Each of the solutes shown has at least one fully ionized carboxylate group at pH 6 and is therefore eluted at a retention volume of Vo (i.e. k' = 0) at this pH. For lower pH values, the expected decrease in retention volume (and hence k') with increasing pH is evident. 7.4.2 Molecular size of the solute

The results presented in the preceding Section show that for some solutes, DA (and hence the retention volume) is somewhat less than that predicted by consideration of charge alone. This behaviour is evident in Fig. 7.5 for the C3' and C4' dicarboxylic acids. It has been suggested by a number of authors [4,36,70-741that size-exclusion effects may contribute to the retention process in ion-exclusion chromatography by restricting the

Chapter 7

206

4r

PH (a) Fig. 7.6 Effect of eluent pH on the capacity factors of (a) monocarboxylic acids and (b) dicarboxylic acids. A 30% cross-linked stationary phase was used with 1 m M Na2HP04 (pH adjusted with oxatic acid) as eluent. Reprinted from [50] with permission.

access of larger solute molecules to the occluded liquid in the pores of the stationary phase. Size-exclusion effects should result in the following retention characteristics: Retention volumes for large, partially ionized solutes should be smaller than expected on the basis of solute charge. (ii) Large, neutral molecules can be expected to show DA values which are less than the theoretical value of 1 .O. (iii) Large, neutral solutes which are eluted at retention volumes less than (Vo + Vj) should be eluted in order of decreasing molecular size. (i)

The retention volumes of the C3' and C4' dicarboxylic acids in Fig. 7.5 are in accordance with (i) above and the retention characteristics described in (ii) and (iii) have been confirmed for neutral lactones [70] and oligosaccharides [ 5 ] . From these results, we can conclude that size-exclusion effects make some contribution to the retention of large solutes.

7.4.3 Hydrophobic interactions between the solute and stationary phase The retention behaviour of the C6'- Cg' dicarboxylic acids and the C3 - C5 monocarboxylic acids in Fig. 7.5 cannot be explained on the basis of solute size and charge. All of these solutes show retention volumes which are larger than those


207

predicted on the basis of solute charge and they are eluted in order of increcising molecular weight (which is the opposite of that expected from size exclusion effects). It is clear that the retention of these solutes is influenced by a third factor, in addition to solute charge and size-exclusion effects. The anomalous retention behaviour described above can be attributed to hydrophobic adsorption of the solutes onto the neutral, unfunctionalized regions of the polymeric stationary phase [50, 53, 751. We have noted previously in the discussion of resin-based ion-exchange columns that many organic molecules and ions show strong reversed-phase interactions with styrene-divinylbenzene packing materials. A plot of the logarithm of capacity factor versus the number of carbon atoms is close to linear for many solute types eluted by ion-exclusion chromatography [5]. This behaviour is similar to that observed in reversed-phase HPLC and gives strong support to the proposal that reversed-phase, hydrophobic interactions play a part in the ion-exclusion retention process. Hydrophobic adsorption effects can be expected to increase in magnitude as the alkyl chain length of the solute is increased, leading to larger retention volumes. This behaviour is evident from Fig. 7.5. We can also note that the size-exclusion effect discussed above will be in competition to the hydrophobic adsorption effect. That is, an increase in alkyl chain length of the solute will cause a decrease in retention under the size-exclusion effect and an increase in retention under the hydrophobic adsorption effect (provided the solute charge is constant). This competition can be used to explain the shape of the retention plot for dicarboxylic acids in Fig. 7.5, where size-exclusion is dominant for the C3' and C i acids, whereas hydrophobic adsorption dominates for the Cg' - C{ acids.

Use of organic modifiers in the eluent The existence of hydrophobic adsorption effects creates the possibility for manipulation of solute retention by adding typical reversed-phase organic modifiers, such as methanol or acetonitrile, to the eluent. A decrease in the retention volume for some solutes can be anticipated and this behaviour has been demonstrated by Tanaka and Fritz [53] using benzoic acid as the eluent. Fig. 7.7(a) shows the effect of the addition of methanol to the eluent and it can be seen that small solute molecules, such as CH3COOH and HCOOH, show little or nn change in retention with increasing methanol, whereas larger solutes, such as valeric acid, show decreased retention. It is evident that the chromatographic resolution of a mixture of all the solutes in Fig. 7.7(a) will decrease with increasing modifier content. A further, opposing effect can result from the addition of an organic modifier to the eluent. When the added modifier has a low dielectric constant (e.g. dioxane), the decreased dielectric constant of the eluent causes an increase in the pKa of the solutes [24]. That is, the solutes become weaker acids and their ionization is therefore suppressed, leading to increased retention. This effect is illustrated in Fig. 7.7(b) for some condensed phosphates, which can be separated only when the organic modifier content in the eluent is high. It can be seen that chromatographic resolution of these solutes increases as dioxane is added. This same effect has been reported for the retention of NH4+ on a strong anion-exchange column [28].

LO

-

PZOf

9 0105~,0,3Methanol concn. % ( v l v ) (a1

Dioxane concn. % (vlv)

(b)

Fig. 7.7 Effect of organic modifiers on solute retention in ion-exclusion chromatography. (a) A 5 pm TSK cation-exchange resin (H+ form) was used with an eluent comprising 0.5 mM benzoic acid and the indicated concentrationsof methanol. Reprinted from [53]with permission. (b) An 18 pm Hitachi 2613 cation-exchangeresin (H+form) was used with an eluent comprising water and the indicated concentrationsof dioxane. Reprinted fmm [24] with permission.

n

88 w


209

Many ion-exclusion columns have definite limits to the amount of organic modifier which can be added to the eluent without causing column damage. It is therefore essential that column specifications be consulted before organic modifiers are used to alter solute retention. In conclusion. we note that the ability to manipulate retention in ion-exclusion chromatography using organic modifiers opens up the possibility of gradient elution. This has been achieved using a sulfonated macroporous resin with a methanol gradient [76], and is illustrated in Fig. 7.8.

7.4.4 Ion-exchange capacity of the stationary phase The effect of the ion-exchange capacity of sulfonated resins on the retention of carboxylic acids has been studied by Lee and Lord 1761. They have synthesized a range of sulfonated macroporous PS-DVB resins in which the degree of functionalization ranges from 0-91% and the retention behaviour of carboxylic acids on these resins is shown in Fig. 7.9. It can be seen that there is an increase in retention for most solutes when the ionexchange capacity is increased from 0 (i.e. for the unfunctionalized resin) to 0.20 mequiv/g (i.e. for a partially functionalized resin). This trend is surprising since an increase in the negative charge density due to sulfonate groups would be expected to

Formic

Propionic

I

0

I

I

4

8

Time ( m i d

1

12

Fig. 7.8 Gradient elution in ion-exclusion chromatography. A Hamilton PRP-X300 column was used with an eluent consisting of a linear gradient (over 5 min) of 6-641methanol in 1 mN H2SO4. Detection was by UV absorbance at 210 nrn Reprinted from [76] with permission.

Chapter 7

210

3

k' 2 1

0 0.0 0.2

0.4 0.6 0.8 1.0 1.2 Exchange capacity (mequiv/g)

Fig. 7.9 Effect of stationary phase ion-exchange capacity on retention of carboxylic acids in ionexclusion chromatography. The eluent was 1 m N H2SO4. S = succinic acid, A = acetic acid, L = lactic acid, C = citric acid, M = malic acid, T = tartaric acid. Reprinted from [76] with permission.

cause a decrease in the retention of the partially ionized solute acids. A suggested explanation is that hydrogen bonding between the neutral solute molecules (which are present in far grcatcr numbers than the ionized solute molecules) and the sulfonic acid groups on the resin may contribute towards solute retention [76]. The presence of some sulfonate groups on the resin could then lead to the observed increase in retention. However, further increases in ion-exchange capacity (beyond 0.20 mequiv/g) show predictable behaviour in that solute retention decreases as the surface charge on the resin increases. It can be concludcd from these results that there is an optimal ion-exchange capacity for each resin type, so that the use of fully functionalized materials is thereforc not always advantageous. 7.4.5

Ionic form of the ion-exchange resin

In the discussion thus far, all of the parameters affecting retention have been related to ion-exclusion chromatography performed either on cation-exchange resins in the Ii+ form, or on anion-exchange resins in the OH- form. We now turn to the effects which arise when the ionic form of the resin is varied. It has been demonstrated [50J that the retention of carboxylic acids on cation-exchangers decreases in the following sequence of ionic forms:


211

This sequence is the reverse of that found for the hydrated radii of these cations, except for NH4+, which has the same radius as K+. It has been suggested that the presence of a bound cation of large radius serves to decrease the available hydrophobic surface area of the resin and to alter the values of both Vo and Vi for the column, all of which can result in reduced solute retention [50]. Some ion-exclusion separations are possible only if the column is in a particular form. This is especially true of the separation of monosaccharides on a calcium-form cation-exchanger, for which ligand-exchange involving interaction between calcium ion and the non-bonding orbitals of the sugar oxygen is thought to occur [5].

7.4.6

Temperature

Temperature can affect retention in ion-exclusion chromatography either by alteration of the chromatographic efficiency in the same manner as observed in most forms of chromatography, or by influencing the degree of ionization of the solute. The first of these effects is evidenced by somewhat reduced retention volumes, improved peak shapes and better separations at elevated temperature due to faster mass-transfer characteristics. In addition, the lower solvent viscosity at higher temperatures permits the use of faster flow-rates on gel-type stationary phases which are subject to pressure limitations. The effects of temperature on solute ionization often vary from solute to solute, as can be seen in the first two columns of Fig. 7.3, which differ only in the temperature used. Some solutes show changes in form at elevated temperature and may therefore be eluted at different retention volumes at different temperatures. An example of this behaviour is the increased retention of partially ionized aldonic acids at higher temperature. due to their conversion into neutral lactones [70]. Increased temperature can also cause a change in the dielectric constant of the eluent, especially when an organic modifier is present. This, in turn, will affect the PKa of the solute and hence its retention. An example of the effects of temperature on retention is illustrated in Fig. 7.10, for phosphate, phosphite and hypophosphite ions eluted with an aqueous acetone eluent [26].

7.4.7

Summary

From the discussion thus far in Section 7.4, we can appreciate that numerous factors play a part in the retention process in ion-exclusion chromatography. These factors are listed below in approximate order of importance. The relative influences of these factors have been determined by examining retention data for carboxylic acids, and may therefore differ for other solutes. The degree of ionization of the solute (which is determined by the pKa of the solute, the eluent pH and the organic modifier content of the eluent). (ii) Hydrophobic (reversed-phase) interactions between the solute and the stationary phase (which are dktermined by the nature of the solute and the organic modifier content of the eluent).

(i)

212

Chapter 7

"0.5 Or

i tu H PO32-

o*o10

0

10 20 30 40 50 60 70 80 Column temp. ('C)

Fig. 7.10 Effect of temperature on the retention of inorganic phosphates using ion-exclusion chromatography. A Hitachi 2613 stationary phase was used with 4050 (v/v) acetone-water as eluent. Reprinted from [26] with permission.

(iii) (iv) (v) (vi) (vii)

The molecular size of the solute. The degree of cross-linking of the stationary phase. The temperature at which the separation is performed. The ion-exchange capacity of the stationary phase. Hydrogen-bonding (normal-phase) interactions between the solute and the stationary phase. (viii) The ionic form of the stationary phase.

7.5

RETENTION MODEL FOR ION-EXCLUSION CHROMATOGRAPHY

The fact that many parameters influence retention in ion-exclusion chromatography makes it difficult, if not impossible, to develop a retention model unless certain simplifying assumptions are made. The most important of these assumptions is that the retention process is dominated by a Donnan exclusion equilibrium mechanism. That is, none of the additional retention processes discussed in Section 7.4 above plays any significant role in solute retention. Using this assumption, Glod and Kemula [77] have reported the following derivation of a retention model. We consider a weak acid, HA, which dissociates according to:

Zon-Exclusion Chromatography

HA % H+ + A-

213 (7.4)

Both HA and A- may exist in both the mobile phase and the stationary (resin) phases (designated by the subscripts m and r, respectively). In a thermodynamic Donnan equilibrium, the chemical potentials of the acid on both sides of the membrane are equal, and if activity effects are neglected, the equilibriumcondition assumes the form:

The dissociation constant of the acid, HA, in both phases is given by: (7.6) The electroneutrality conditions in both phases can be written: (7.7) (7.8) where the Concentration of dissociated functional groups in the stationary phase is given by [S03-lr. Now the concentration, c. of the sample at the peak maximum can be written in terms of the total concentration of all forms of the acid, to give:

The distributioncoefficient, DA.is given by: (7.10) From eqns. (7.6), (7.7) and (7.9) we can write: (7.1 1)

The value of [A-Im can be obtained from eqns. (7.6)-(7.9). if we make the following assumption: c cc

[SO;],

This enables us to write the following expression for DA:

(7.12)

Chapter 7

214

1.0

-

- 2.8 -

0

-

-

- 2.L -

0.6DA 0.L

- 2.0

0.8

-

-

"R

- 1.6 -

c

- 1.2

0.2-

-

-

- 0.8

0l

l

l

l

l

t

l

l

l

l

l

l

l

l

l

Fig. 7.11 Plot of the distribution coefficient, DA,as function of log (C/Ka). The solid line is calculated from eqn. (7.14), whilst the points represent experimental values obtained for 33 solutes. LiChrosorb KAT was used as the stationary phase, with water as eluent. Reprinted from [77]with

permission.

DA

=

2c

+ K, - JKZ + 8K,c 2~ - 2Ka

(7.13)

which can be rewritten as:

(7.14)

2-

G

-2

Ka

Eqn. (7.14) shows that DA (and hence VR) depends only on one experimental parameter, the ratio C/Ka. A plot of the theoretical relationship between DA and log (C/Ka) is shown in Fig. 7.11, together with experimental points obtained for 33 solutes (mineral acids, carboxylic acids and nitrophenols). Good agreement is obtained between theory and experiment. The above retention model has been recently extended to remove the requirement to calculate the sample concentration at the peak maximum (c), and to consider the case where a buffer is added to the eluent [78]. An iterative, numerical procedure is required in order to calculate DA for different solute and eluent conditions. Since this is a lengthy and complex process, the extended model will not be considered here.


215

n5

1

I

I

0

5

I

1

15 ‘firnc (min)

I

I

20

25

Fig. 7.12 Analysis of human urine using ion-exclusion chromatography. An Interaction ORH801 column was used with an eluent comprising 10 mN H2SO4 containing 10% methanol. Detection was by spectrophotometryat 254 nm. Solute identities: 1 = oxalic acid, 2 = oxaloacetic acid, 3 = a-ketoisovaleric acid, 4 = ascorbic acid and a-keto-P-methyl-n-valeric acid, 5 = pphenylpyruvic acid, 6 = uric acid, 7 = a-ketobutyric acid, 8 = homoprotocatechuic acid, 9 = unknown, 10 = unknown, 11 = hydroxypheylacetic acid, 12 = p-hydroxyphenyllactic acid, 13 = homovanillic acid. Reprinted from 1321 with permission.

7.6

APPLICATIONS OF ION-EXCLUSION CHROMATOGRAPHY

Ion-exclusion chromatography has many applications in IC, but for the purposes of illustration, we will consider here only three of the more important applications. These are the determination of carboxylic acids, weakly ionized inorganic compounds, and water. Further applications may be found in the Tables comprising Part V of this book. 7.6.1

Carboxylic acids

The separation of carboxylic acids is the most common application of ion-exclusion chromatography. This mode of chromatography is undoubtedly the method of choice for these solutes. When coupled with direct spectrophotometric detection at low wavelength, ion-exclusion chromatography yields excellent separations and relatively

216

Chqpter 7

TABLE 7.4 SOME APPLICATIONS OF THE DETERMINATION OF CARBOXYLIC ACIDS BY IONEXCLUSION CHROMATOGRAPHY Sample Acid rain Antarcticice Blood coffee Fhit juice Milk

Warrnaceuticals Plasma Plating baths Ringers solution Solder fluxes Sugar cane juice Urine Urine Wine Wine a

COlUmn

Dionex HPICE-AS2 Aminex HPX-87H Dionex HPICE-AS1 Dionex HPICE-AS1 Bio-Rad AG5OW-X2 Aminex HPX-87H Dionex HPICE-AS1 Dionex HPICE-AS1 Wescan 269-006 Dionex HPICE-AS1 Waters Ion-Exclusion Aminex HPX-87H Interaction ORH-801 Aminex HPX-87H Intaaction ORH-801 Dionex HPICE-AS1

EluenP 2.0 mM HCl 5.0 mM MSA 10 mM HCl 10 mM HCl 0.75 mv1n-BA 10 mM H2SO4 25 mM 0.5 mMH2CO3 3.2mMHN03 5.4 mM H2CO-J 1.O mM OSA 5 mM H2SO4 10 mM H2SO4 25 mM H2SO4 10 mM H2SO4 2.0 mM OSA

Detection

Detection Ref

methdb

limit

C

0.03 ppm 15 8 79 lppm 56 0.5 ppm 83 1 ppm 5 1 ng 84 2ppm 67 0.5 ppm 89 0.1 ppm 16 50ppb 90 5PPm 9 0.3 ppm 32 0.5 ppm 54 0.5 ppm 30 1PPm 60

Spec (200nm) 6-9 ppb

C C C Spec (210 nm)

RI C C C C

RI Spec (254 nm) Spec (200nm)

Spec (210 nm) C

MSA = methane sulfonic acid, n-BA = n-butyric acid, OSA = octane sulfoNc acid. C = conductivity. RI = refractive index, Spec = spectrophotometry.

clean chromatograms for a wide variety of very complex sample matrices. These samples include biological materials, such as urine [54, 791, tissue [13], blood 1801, plasma [67, 80,811, serum [82] and bile [82]; foods and beverages, such as wine [31,60, 76, 831, coffee [56], milk [5] and cane juice 191; and pharmaceuticals, such as Ringers solution [16, 661, tablets [67] and intravenous solutions [84, 851. Fig. 7.12 shows a chromatogram for a urine sample, obtained without sample pretreatment, and illustrates the relatively clean chromatograms which can be achieved for the above complex samples. Industrial and environmental applications of carboxylic acid determinations are also common and include samples such as acid rain [ 151, diesel exhaust [86], plating baths [87-891 and sewage [Sl]. Table 7.4 lists some of the chromatographic conditions employed in these separations.

7.6.2 Weak inorganic acids and bases Ion-exclusion chromatography has found increasing usage for the determination of weakly ionized inorganic species. It is especially attractive as an adjunct to ion-exchange chromatography since the selectivities obtained by these two techniques are quite different (see Sections 2.1.4 and 7.4). Solutes such as fluoride [43, 911, carbonate [39],

Ion-ErcluswnChromarography

217

A s (V)

32-

[O.Ol AU

-

Anions

L rill

0

1

2

Time (min)

3

0

6

12 18

Time (min) (b)

Fig. 7.13 Determination of inorganic species by ion-exclusion chromatography. (a) A Brownlee Polypore high-speed anionexclusion column was used with 6 mM H2SO4 as eluent. Detection was by amperometry using a Pt electrode at +0.4V versus AdAgCl. The sample was peppers in vinegar. Reprinted from [l 11 with permission. (b) An Aminex HPX-87H column was used with 10 mM H3PO4 as eluent. Detection was by spectrophotomeay at 200 nm. Reprinted from [62]

with permission. cyanide [58], borate [42]. sulfite [47], phosphates [24], nitrite 1921, arsenite 1621. arsenate [62] and ammonium [92] have been determined using this approach. Interference from strongly ionized species is minimal because these solutes are unretained and appear at the column void volume. Ion-exclusion chromatography can therefore readily separate weakly ionized solutes in samples containing high concentrations of ionic species, e.g. seawater and wastewater. Table 7.5 lists some of the applications of this technique in inorganic analysis. Fig 7.13 shows typical chromatograms obtained in two important applications, namely the determination of sulfite (Fig. 7.13(a)) and inorganic arsenic ions (Fig. 7.13(b)). The fact that all strongly ionized solutes are eluted at the void volume in ionexclusion chromatography opens up the possibility of a two-dimensional chromatographic system in which these solutes are collected and then separated on an ion-exchange system. This type of chromatographic system will be discussed in Section 15.4.

218

Chapter 7

TABLE7.5 SOME EXAMPLES OF THE DETERMINATION OF INORGANIC SPECIES BY IONEXCLUSION CHROMATOGRAPHY

Solute(s)a

Sample

column

Eluentb

Demc Ref

Plating baths Mineral water Seawater Fluxes Mouthwash Wastewater Biological fluids Water Water Disinfectant Effluents

Dionex HPICE-AS 1 Aminex HPX-87H Aminex HPX-87H Waters ion-exclusion Wescan 269-006 Cation-exchange resin Dionex HPICE-AS 1 TSK-gel SCX Waters Fruit Juice Waters Fruit juice Dionex HPICE-AS 1 TSK-gel SCX

1 mMH2SO4 lOmMH3P04 5mMH3po4 1.0 mM OSA 3mMH2SO4 40% M e o w 2 0 Water 1 mM benzoic acid 1.25 mM H2SO4 1 mMH2SO4 50mMmannitol 0.1 M fructose

Amp 87 Amp 62 spec 63 C 43 93 C Coul 91 82 C C 39 42 RI 94 RI 12 C C 41

Water Foods Process water Process water Process water Process water

Bio-Rad AG50W-X8 Wescan ion-exclusion Hitachi 2632 Anion-exchange resin Cation-exchange resin Hitachi 3613

C 58 Amp 47 C 28 Coul 92 Spec 92 Spec 95

Hitachi 2613

1 mM HCl 5mMH2SO4 Water 10%MeOH/H20 10%MeOH/H20 0.1 mMH2SO45% MeOH Dioxane-water

Coul

24

Wescan 269-05 1 Dionex HPICE-AS I

5mMH2SO4 5.4mMH2C03

Amp

49 16

C

a DMSO = dimethylsulfoxide.

OSA = mane sulfonic acid. Amp = amperornetry, Spec = spectrophotomeay,C = conductivity, Caul= coulomeay, RI = refractive index

7.6.3

Water

One of the more significant recent developments in ion-exclusion chromatography is the application of the technique to the determination of water. This determination is a very important and frequently encountered analytical problem. Water, being a small, neutral molecule, can be expected to show retention on an ion-exclusion column, provided a suitable non-aqueous eluent is employed. Stevens et al. [20] showed that water can be separated from other sample components by ion-exclusion chromatography on a short column packed with Aminex 50W-X4resin (H+) form, using an eluent comprising methanol and a small amount of HCI, H2SO4 or p-toluenesulfonic acid. Detection was achieved by conductivity measurements, with the water showing decreased conductance relative to that of the eluent. Fig. 7.14(a) shows the chromatogram

219


(0)

lime lminl 0 2 1 8

Fig. 7.14 Determination of water using ion-exclusion chromatography. (a) A short (9 x 21 mm) column packed with Aminex 50W-X4 (H+form) resin was used, with a methanolic eluent containing 1.2 mM HCI. Conductivity detection was used. Reprinted from [20] with permission. (b) A 150 x 2.1 mm I.D. column packed with Aminex Q-150s resin in the H+ form was used with 1.0 mM cinnamaldehyde in methanol as eluent. Detection was by spectrophotometry at 300 nm. The sample was 0.184% H20 in dichloroethane. Reprinted from [97] with permission.

obtained. The main drawback with this method was variability in the detector response as the concentration of water in the sample was altered. Fritz and co-workers [96, 971 have reported an alternative ion-exclusion method in which the water is separated using a cation-exchange column in either the Li+ or H+ form, with cinnamaldehyde in methanol as the eluent. Detection is accomplished by spectrophotometric monitoring of the equilibrium existing between cinnamaldehyde and cinnamaldehyde dimethylacetal:

2CH30H + cinnamaldehyde % H 2 0

+ cinnamaldehyde dimethylacetal

(7.15)

This reaction does not occur to any appreciable extent until an acid catalyst is present. The catalyst may be the hydrogen-form cation-exchange resin in the column, or

Chaprer 7

220

an acid can be added to the eluent. After catalysis, the above equilibrium lies well to the right. When water is injected, there is a small shift in the equilibrium towards the formation of cinnamaldehyde. This change can be detected spectrophotometrically at 300 nm. Excellent sensitivity is achieved by this method and the analysis can be completed in less than 2 min. Fig. 7.14(b) shows a typical chromatogram obtained using this approach. Both of the methods depicted in Fig. 7.14 have been applied to a wide range of sample types, with good results. 7.7 1 2 3 4 5

6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32

REFERENCES Wheaton R.M. and Bauman W.C., ind. Eng. Chem., 45 (1953) 228. Rich W., Smith F.,Jr., McNeil L. and Sidebottom T..in Sawicki E. and Mulik J.D.(Eds.), lon Chromatographic Analysis of Environmental Pollurants, Vol. 11. AM Arbor Sci. Publ., Ann Arbor, MI, 1979, p. 17. Harlow G.A. and Moman D.H., Anal. Chem., 36 (1964) 2438. Waki H. and Tokunaga Y..J. Chromatogr., 201 (1980) 259. Jupille T., Gray M.,Black B. and Gould M., Am. Lab., 13 (1981) 80. Walser P., J. Chromarogr., 439 (1988) 71. Albman G. and Collins C.H., J. Chromarogr., 395 (1987) 623. Haddad P.R. and Jackson P.E.,J. Chromarogr., 447 (1988) 155. Blake J.D., Clarke M.L. and Richards G.N..J. Chromafogr., 398 (1987) 265. Ianniello R.M., Anal. Lerr., 21 (1988) 87. Kim H.-J.. Park G.Y. and Kim Y.-K., Food Technol., 41 (1987) 85. Cameron A. and Pohlandt-Watson C., Mintek Repon No. M284 (1986). Dahl A.R.. Miller S.C. and Pemdou-Fischer J., Toxicol. Lerr., 36 (1987) 129. Okada T. and Dasgupta P.K., Anal. Chem., 61 (1989) 548. Bachman S.R. and Peden M.E., Wafer Air Soil Pollut., 33 (1987) 191. Itoh H. and Shinbori Y.,Chem. Lett., (1982) 2001. Dionex HPICE-AS5Product Information'Bulletin. Lehotay J. and Traiter M.,J. Chromarogr., 91 (1974) 261. Monk P.R. and Iland P.G., Food T e c h f . Aust.. 36 (1984) 16. Stevens T.S., Chritz K.M.and Small H., Anal. Chem., 59 (1987) 1716. Richards M.,J. Chromafogr., 115 (1975) 259. Turkelson V.T. and Richards M., Anal. Chem., 50 (1978) 1420. Lee D.P., Bunker M.T. and Lord A.D.,in Jandik P. and Cassidy R.M. (Eds.). Advances in Ion Chromarography, Vol. 1, Century international, Inc., Franklin, MA, 1989, p. 451. Tanaka K. and Ishizuka T.,J. Chromarogr., 190 (1980) 77. Tanaka K. and Ishizuka T., Wafer Res., 16 (1982) 719. Tanaka K. and Sat0 H., Bunseki Kagaku, 27 (1978) 95. Tanaka K.,Ishizuka T. and Sunaham H.,J. Chromatogr., 174 (1979) 153. Tanaka K., Ishizuka T. and Sunahara H., J. Chromarogr., 177 (1979) 227. Benson J.R. and Woo D.J., J. Chromarogr. Sci., 22 (1984) 386. Woo D.J. and Benson J.R. Organic Acid Analysis Column ORH-801. Interaction Product Bulletin (1982). Woo DJ. and Benson J.R.. LC. 1 (1983) 238. Woo DJ.and Benson J.R.. Am. Clin. Prod. Rev., January (1984) 20.

Ion-Exclusion Chromatography 33 34 35 36 37 38 39

40 41 42 43 44

45 46 47 48 49 50 51 52 53 54 55 56 57 58 59

22 1

Woo D.J. and Benson J.R., Am. Lab., January (1984) 50. Interaction Separations#1, March (1989). Gjerde D.T. and Mehra H., in Jandik P. and Cassidy R.M. (Eds.), Advances in Ion Chromatography,Vol. 1, Century International, Inc., Franklin, MA, 1989, p. 139. Waki H. and Tokunaga Y..J. Liq. Chromutogr.,5 (1982) 105. Tokunaga Y., Waki H. and Ohashi S., J. Liq. Chromatogr.,5 (1982) 1855. Tanaka K. and Fritz J.S., J. Chromatogr.,409 (1987) 271. Tanaka K. and Fritz J.S., Anal. Chem., 59 (1987) 708. Okada T., Anal. Chem.,60 (1988) 1336. Okada T. and Kuwamoto T., Fres. Z . Anal. Chem., 325 (1986) 683. Jones W.R., Heckenberg A.L. and Jandik P., J. Chromutogr., 366 (1986) 225. Dunn M.H., LC.GC, 7 (1989) 138. Heckenberg A.L., Jones W.R., Wildman WJ., Krol J.A. and Alden P., in Jandik P. and Cassidy R.M. (Eds.), Advances in Ion Chromatography,Vol. 1, Century International, Inc., Franklin, MA, 1989, p. 333. Jones W.R., Jandik P. and Swartz M.T., J. Chromatogr.,473 (1989) 171. Jupille T., Burge D. and Togami D., Chromutographia, 16 (1982) 312. Kim H.-J. and Kim Y.-K.,J. FoodSci., 51 (1986) 1360. Kim H.-J. and Kim Y.-K., in Jandik P. and Cassidy R.M. (Eds.). Advances in Ion Chromatography,Vol. 1, Century International, Inc.. Franklin, MA, 1989, p. 391. Nguyen J.H., Kim H.-J. and Gjerde D.T., Am. Lab., May (1988) 122. Kihara K., Rokushika S. and Hatano H., J. Chromutogr., 410 (1987) 103. Murayama T., Kubota T., Hanaoka Y., Rokushika S., Kihara K. and Hatano H., J . Chromatogr.,435 (1988) 417. Manning D.L.and Maskarinec M.P., J. Liq. Chromatogr..6 (1983) 705. Tanaka K. and Fritz J.S., J. Chromutogr., 361 (1986) 151. Buchanan D.N. and Thoene J.G.,Anal. Biochem., 124 (1982) 108. Haginaka J., Wakai J., Yasuda H. and Nomura T., J. Chromatogr., 447 (1988) 373. Dionex Application Note 19. Muller H., Nielinger W. and Horbach A., Angew. Makromol. Chem.,108 (1982) 1. Pohlandt C., NIM Report No. 2107 (1981). Rossiter W.J., Jr., Godette M., Brown P.W. and Galuk K.G., Sol. Energy Mat., 11 (1985) 455.

Dionex Application Note 21. Waters IC Lab. Report No. 241A. Butler E.C.V., J. Chromatogr..450 (1988) 353. Ivey J.P. and Haddad P.R., J. Chromatogr.,391 (1987) 309. 64 Hanai T. and Hubert J.,J. Chromutogr., 316 (1984) 261. 65 Dionex Technical Note 17. 66 Itoh H. and Shinbori Y., Bull. Chem. SOC.Jap., 58 (1985) 3244. 67 Itoh H., Shinbori Y.and Tamura N., Bull. Chem.SOC.Jap., 59 (1986) 997. 68 Kim W.S., Geraci C.L. and Kupel R.E., Am. Ind. Hyg. Assoc. J., 41 (1980) 334. 69 Kim W.S., Geraci C.L., Jr. and Kupel R.E., in Sawicki E. and Mulik J.D.(Eds.), Ion ChromatographicAnulysis of Environmental Pollutants, Vol. II, Ann Arbor Sci. Publ., Ann Arbor, MI, 1979, p. 171. 70 Hicks K.B., Lim P.C. and Haas M.J., J. Chromatogr., 319 (1985) 159. 71 Wheaton R.M. and Bauman W.C., Ann. N.Y. Acid. Sci., 45 (1953) 228 60 61 62 63

222 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97

Chapter 7 Asher D.R. and Sirnpson D.W.. J. Phys. Chem.. 60 (1956) 518. Reichenberg D.,Chem. Ind (London), (1956) 958. Reichenberg D. and Wall W.F.. J. Chem. SOC..(1956) 3364. Waki H., Tsuruta K. and Tokunaga Y . , J. Liq. Chromarogr.. 8 (1985) 2105. Lcc D.P. and Lord A.D., LC.GC. 5 (1987) 261. Glod B.K. and Kernula W.. J. Chromurogr.. 366 (1986) 39. Glod B.K., Piasecki A. and Slafiej I., J. Chromurogr., 457 (1988) 43. Rich W.E.. Jr. and Johnson E.L., Eur. Par. Appl., EP. 38720 (1981). Rich W.E., Johnson E.. Lois L.,Stafford B.E., Kabra P.M. and Marton L.J., in Kabra P.M. and Marton L.J. (Eds.), Liquid Chromurographyin Clinical Analysis, Hurnana: Clifton, NJ, 1981, p. 393. Rich W., Johnson E., Lois L., Kabn P., Stafford B. and Marton L.. Clin. Chem, 26 (1980) 1492. Krcling J.R. and DeZwaan J., Anal. Chem., 58 (1986) 3028. Monk P.R. and Hand P.G., Food Technol. Ausr., 36 (1984) 18. Iwinski G. and Jenke D.R., J. Chromutogr., 392 (1987) 397. Waters IC Lab. Report No. 292. Bodek 1. and Menzies K.T.. Chemical. Hour& in the Workplace, ACS Symp. Ser. 149 (1981) 599. Dionex Application Update 110. Haak K.K.. Plat Su$ Finish. 70 (1983) 34. Wescan Application #136a. Waters IC Lab. Report No. 299. Tanaka K., Bunseki Kagaku, 32 (1983) 439. Tanaka K., Bunseki Kagaku, 31 (1982) T106. Wescan Application #90. Waters IC Lab. Report No. 257. Tanaka K. and Ishihara Y . ,Mizu Shod Gijrrrsu. 23 (1982) 855. Fortier N.E. and Fritz J.S., J. Chromarogr.. 462 (1989) 323. Chen J. and Fritz J.S., J. Chromutogr., 482 (1989) 279.

223

Chapter 8 Miscellaneous Separation Methods 8.1

INTRODUCTION

In addition to the mainstream separation methods discussed thus far, we will also consider a number of alternative approaches which can be used for the separation of inorganic ions and carboxylic acids. In the strict sense, none of these methods can be defined accurately as IC, yet the fact that they can be employed for the same solutes as those normally encountered in IC suggests that a brief discussion is merited. The purpose of this discussion will be to indicate the operating principles of each approach and to provide some representative chromatograms in order to facilitate comparison with the ion-exchange, ion-interaction and ion-exclusion methods discussed in Chapters 2-7. Fig. 8.1 provides an overview of the separation methods which will be considered.

Coordination compounds

Reversed-

- phase

HPLC

i -r

Organometallics Carboxylic acids (Ion-suppression)

Miscellaneous separation methods

-

Chelating stationary phases

Chemically bound ligands

L Crown ether stationary phases Anions

chromatography

Cations

Fig. 8.2 Overview of miscellaneous separation methods.

Chapter 8

224

8.2

REVERSED-PHASE LIQUID CHROMATOGRAPHY

Ionic or partially ionized solutes are generally retained only weakly on conventional c18 HPLC stationary phases. We have already seen in Chapter 6 that retention of these solutes can be increased through the use of an ion-interaction reagent added to the eluent. In this Section, we now turn to the use of non-polar stationary phases for the separation of coordination compounds, organometallics and carboxylic acids.

Coordination compounds

8.2.1

A convenient and frequently used method for the determination of metal ions is to first complex the metal ions with a suitable ligand, and to then separate the resultant coordination compounds by conventional reversed-phase or normal-phase HPLC. The complexes formed are often uncharged and this permits separation to be achieved on Cis or silica stationary phases. Several comprehensive reviews on HPLC of coordination compounds are available 11-61, from which it can be seen that there are a number of desirable properties of both the ligand and the chelate. These include [3-51:

(i) (ii)

(iii)

(iv)

(v)

The ligand should form neutral complexes with a large number of metals, using relatively simple preparation methods. The complexes formed should be coordination saturated, since this gives the greatest probability of separation of complexes formed from different metals. Moreover, the ligand should not be too large, so that specific properties of the central metal atom are retained in the coordination complex. The donor atoms in the ligand should have low total electronegativity to minimize adsorption effects on silica-based reversed-phases. Preferred donor atoms are N-. 0- and S-. Separation selectivity increases when ligand substituents do not have large induction or steric effects, and also when electronegative atoms exist in close proximity to the chelate ring. The complexes should have high stability, good detectability and high solubility in non-polar organic solvents.

Many ligands have found application in HPLC separations of metal chelates. These ligands include dithiocarbamates (71, 8-hydroxyquinoline [8, 91, P-diketones [lo]. 4-(2pyridylaz0)-naphthol [ 11],4-(2-pyridylazo)-resinol [ 121, dialkyldithiophosphates[ 13, 141, xanthates [ 151, 2.3-diaminonaphthalene [16], pyrazolones [17] and hydrazones [18. 191. No single ligand is suitable for all metal ions and typically only a few metals are determined in a single chromatographic separation. In most cases, water-insoluble chelates are formed and these must be extracted into a suitable organic solvent, prior to the chromatographic separation step. This sometimes involves extraction with solvents which cannot be injected directly into a reversed-phase HPLC system, so that evaporation and redissolution become necessary. Alternatively, complexes can be formed in-situ by injecting metal ions into a mobile phase which contains the ligand and

Miscellaneous Separation Methods

225

an appropriate buffer [20] or through the use of solid-phase reaction on a suitable precolumn 1141. The stability of the metal complexes is also of great importance because these complexes are generally injected at very low concentrations and are therefore prone to dissociation as they traverse the chromatographic system. This is particularly true of complexes which may undergo ligand-exchange reactions at the surfaces of metallic chromatographic components, such as the injector, interconnecting tubing and the inlet and outlet frits in the column. Kinetic stability is of more importance than thermodynamic stability, since kinetically inert complexes are more likely to pass intact through the chromatographic system. The large volume of literature on HPLC of metal chelates precludes a comprehensive discussion of this topic. Table 8.1 provides a selected listing of some applications of HPLC of metal chelates. To illustrate the utility of this technique, we will focus attention only on the use of dithiocarbamate ligands, since these reflect most of the trends which exist for other ligands.

Dithiocarbamate complexes Alkyldithiocarbamate ligands form complexes with a very wide range of metal ions and therefore offer the potential for separation of a larger number of metal ions than any other ligand. Diethyldithiocarbamate (DEDTC) complexes have been studied extensively and Fig. 8.2 shows a typical separation of metal-DEDTC complexes achieved on a reversed-phase column with a ternary eluent comprising water, methanol and acetonitrile [7]. Two important characteristics are evident from this chromatogram. First, there is a peak due to the ligand oxidation product, bis(diethy1thio-carbamy1)disulfide (usually referred to as disulfiram), which results either from excess ligand in the extracting solution, or from ligand produced by dissociation of labile complexes. Second, the peak shape for Pb(I1) is very poor, due to the low kinetic stability of this complex and the resultant likelihood of dissociation or ligand-exchange reactions. This behaviour occurs with other unstable complexes, such as those of Cd(II), Fe(II1) and Zn(II), and is especially evident when columns with stainless-steel frits are used. The porous nature and high surface areas of these frits provide ideal sites for ligand-exchange reactions to occur between the injected metal complexes and metal ions produced from the oxidation of stainless steel components, especially nickel. These reactions can be minimized either by using columns without porous metallic frits, such as radial compression columns [7], by deactivating the frits with an organosilane [25],or by addition of EDTA to the mobile phase 171. Formation of the chelate It should be stressed that it is the metal chelates which are separated in the above example, rather than the metal ions themselves. Therefore, these chelates must be formed before the sample reaches the analytical column. Chelate formation is usually achieved by buffering the sample and then extracting with a solution of the ligand in a suitable organic solvent. This process can be automated using the apparatus shown in Fig. 8.3, wherein the sample is added to a solution stream of ligand (diethyldithiocarbamate) in acetonitrile [26]. The mixture is then passed, in turn, to a heated

TABLE 8.1 TYPICAL DETERMINATIONS OF METAL CHELATES BY REVERSED-PHASEHPLC Solute(s)

Liganda

Stationary phase

Al(III), In(1LI) As(III), Sb(III), Bi(II1) Cd(II), PWII), Ni(II), Co(III), Hg(I1). Cr(III), Se(IV), Cu(II), Te(W Cd(II), Co(II), Pb(II), Ni(II), Wrr)

PMBP DEDTP DEDTC

BHEDC

BHEDC Oxine HFAA Oxine DEDTC sew) Ti(IV), FeOII), U(VI), V(V)

DAN DAPMP, DAPMT

Detection modec

Detection limitd

Ref

Shim-Pack CLC-ODs ACN-MeOH 10 mM DEDTP in ACN Hypersil ODS Waters C18 Rad-Pak 40:35:25 MeOH-ACNwater

S (290 nm) S (280 nm) S (254 nm)

21-121 ng 2 ng 0.5

17 14 7

O.lmM HEDC in 40:a MeOH-water with 0.1 m~ Zn2+ SUPelCO cl8 25 mM TEA acetate Cg Reversed-phase 1 mM oxine in borate buffer (pH 9) 100% cHZC12 Silica c18 reversed-phase 10 mM oxine, ACN an^ acetate buffer (pH 6) 0.05% DEDTC in waterHypersil ODs MeOH-cHC13 60:40MeOH-water pBondapak C18 Polymer Labs PLRP-S 10%-40% ACN-water gradients

S (300 nm)

7-53 ppb

21

S (255 nm) S (254 nm)

5 PPb

22 23

DCP S (400 nm)

n.s. 100 PI--

24 8

S (350 nm)

0.5 ppm

20

S (254 nm), Fluor S (340 nm)

10 ppb n.s.

16

pBondapak cl8

Mobile phaseb

n.s.

19

PMBP = l-phenyl-3-methyl-4-benzoyl-5-pyrazolone, DEDTP = diethyldithiophosphate, DEDTC = diethyldithiocarbamate, BHEDC = bis(2hydroxyethy1)dithiocarbamate. Oxine = 8-hydroxyquinoline, HFAA = hexafluoroacetylacetone, DAN = 2,3 diaminonaphthalene, DAPMP = 2,6diacetylpyridine bis (N-methylenepyridiniohydrazone),DAPMT = 2,kliacetylpyridine bis(N-methylene-N,N,N,-trimethylammoniohydnuone). MeOH = methanol, ACN = acetonitrile, TEA = tetraethylarmnonium. S = spectrophotometry, Fluor = fluorhetry, DCP = direct current plasma atomic emission spectrometry. n.s = not stated. a

5

00


227

I

0.005 Ahsorhancc

Fig. 8.2 Separation of a mixture of diethyldithiocarbamate complexes by reversed-phase HPLC. The mobile phase comprised 40:35:25 methanol-acetonitrile-waterand a Waters c18 Rad-Pak was used as the column. The flow-rate was 2.0 rnl/rnin and the detection wavelength was 254 nm. Peak identities: A-disulfiram, B-Cd(II), C-Pb(II), D-Ni(II), E-Co(III), F-Cr(III), G-Se(IV), HCu(II), I-Hg(II), J-Te(1V). Reprinted from [7] with permission.

reaction coil, a bubble capture device and to the sampling loop of an auto-injector. Excess ligand in the solution is removed by an anion-exchange guard column placed before the c18 analytical column. An alternative method for the formation of the dithiocarbamate complexes is to inject the metal ions into an eluent which contains the ligand. On-column complex formation provides a potentially quick and easy method for multi-element identification and determination. When DEDTC is added to the mobile phase, Cd(II), Pb(II), Co(II), Hg(I1) and Cu(I1) can be separated [20, 271. The chief problem encountered with oncolumn complexation is the high background detector signal produced by the presence of the ligand in the mobile phase. This requires that a selective wavelength be used in the case of spectrophotometric detection, or alternatively, amperometric detection must be used [28]. A further problem is the poor solubility of many dithiocarbamate complexes in typical mobile phases for reversed-phase HPLC, but this may be overcome either by the addition of a small amount of chloroform to the mobile phase [20], or through the use of a ligand which forms water-soluble complexes [21]. An example of the latter approach is the use of bis(2-hydroxyethy1)dithiocarbamate (BHEDTC), in which the

Ckpter 8

228

DEDTC in acetonitrile prewure Sample

Fig. 8.3 Schematic diagram of apparatus for automated pre-column formation of diethyldithiocarbamate (DEDTC)complexes, prior to separation by reversed-phase HPLC. Reprinted from [26] with permission.

hydroxy groups on the ligand cause metal complexes to be water-soluble at low concentrations [22]. Fig. 8.4 compares chromatograms obtained using pre-column (Fig. 8 4 a ) ) and on-column (Fig. 8 4 b ) ) complex formation with BHEDTC. 8.2.2 Organometallic compounds

One of the factors which limits the applicability of HPLC analysis of metal chelates is the necessity to form the chelate itself. This limitation does not exist for many of the organometallic species which are amenable to chromatographic analysis, since these species often occur in a wide range of samples. The more important organometallic species which can be analyzed by HPLC include alkyllead, alkylmercury, alkylarsenic and alkyltin compounds. Of these, the organoarsenic species are the most widely studied. Monomethylarsonate, dimethylarsinate and phenylarsonate (as well as the inorganic ions arsenate and arsenite) are formed by the action of many common yeasts, fungi and bacteria on arsenic present in soils. The high toxicities of these compounds necessitate their accurate determination, especially in water samples. Separation can be accomplished by reved-phase chromatography [29], as well as by anion-exchange [30, 311 or ion-interaction 132, 331 methods. Organomercury compounds, such as methylmercury and ethylmercury, have also been separated by reversed-phase HPLC [34]. In most of the above examples, atomic spectroscopic detection methods have been employed Table 8.2 lists some further applications of the determination of organometallic species by reversed-phase HPLC, and Fig 8.5 shows chromatograms obtained for organomercury and organotin compounds.

% c;.

TABLE 8.2 TYPICALDETERMINATIONS OF ORGANOMETMC SPECIES BY REVERSED-PHASE HPLC Solute(s)

Sample

Alkyl Hg compounds Alkyl Pb compounds Ethyl Sn compounds Fe, Mo carbonyl complexes Methylmucury, ethylmacury Methyl Sn compounds Organo As compounds Organo Hg compounds

n.s. PeIml

Tetraphenyl Pb Transition metal cluster complexes

Stationary phase

Eluent

Detection modes

Detection limit

ICP ICP

Reactionmixtures

HyperSd c18 H m f i c18 Spherisorb S5W ODs ZarbaXCg

ICP

n.s. 35 llppb 35 50-100pg 36 1 PPb 35

Tuna

Waters pic0 Tag

1:2 EtOH-0.05 M NaBr 75% EtOH-water 7030 acetone-pentane 7030 EtOH-water or J3OH-water gradients 6omManrmoniumacetate, 0.005% 2-mer~aptOethan01 6040 acctone-pentane 100% MeOH 4050 MeOH-water + 0.06 M ~ O A + C 0.01% 2-mexaptOethanol 90:lO MeOH-water 7525 MeOH-water

ICP-MS

1 PPb

34

Hydride AAS GFAAS

2-2opg 5 ng 2 PPb

37 38 39

480pg n.s.

40 41

Water

n.s. n.s. Fish

n.s. n.s.

FtI

DP Amp ZeemanAAS

spec

I B = inductively coupled plasma atomic emission spe!cmehy, Hydxide AAS = hydxide generation atomic absorption spectrometry, GFAAS = graphite furnace atomic absorption specaometry,RI = refractive index, MS = mass spectrometry,DP Amp = differential pulse. ampemmefry, Spec = specfrophotomefry. n.s. = not stated.

a

Ref

is

E

8e

I

230

Chapter8

CO2'

0

L

8

l i m e (minl (a)

12

-

0

5 10 lime (minl (bl

15

Fig. 8.4 Comparison of chromatograms obtained using (a) pre-column and (b) on-column formation of bis(2-hydroxyethy1)dithiocarbamate (BHEDTC) complexes. (a) The pre-formed BHEDTC complexes were injected onto a Supelcosil c18 column using an eluent comprising 4060 methanol-water containing 25 mM triethylammonium acetate. Detection was by spectrophotometry at 255 nm. Reprinted from [22]with permission. (b) Metal ions were injected onto a Waters WBondapak cl8 column using an eluent comprising 4050 methanol-water containing 0.1 mM Detection was by spectrophotometry at 300 nm. Reprinted from [21] BHEDTC and 0.1 mM a*+. with permission.

8.2.3

Carboxylic acids (Ion-suppression)

A further strategy which can be employed in the determination of ionizable solutes by reversed-phase HPLC is to suppress the ionization of these solutes by adding a buffer of appropriate pH to the eluent. Retention of the solutes on non-polar stationary phases is therefore increased and separation can then be accomplished. Acidic buffers are used for the separation of weak acids, whilst alkaline buffers are used for the separation of weak bases. This method, often described as "ion-suppression",is generally considered to be applicable only to those weak acids and bases for which the ionization can be suppressed using buffers having pH values in the range 3 - 8 [42]. The reason for this is that C1g stationary phases are unstable outside this pH range. Although this is undoubtedly a major limitation, many weak acids may separated on C18 columns, as demonstrated by Skelly [43]. Restrictions in eluent pH do not apply to the use of non-

23 1


MeSn( EtSnCI: Me2SnCI; Me3SnCI MebSr

#Hg+

I I

1

1

1

1

.L 1

1

1

1

0 2 4 6 8 10 12 14 16 Time (min) (a)

1 1 1 T

-

0 1 2 3 Time (min)

0 1 2 3 1 5 Time (min)

(bl

(C 1

Fig. 8.5 Separation of (a) organomercury, (b) methyltin and (c) ethyltin compounds by reversedphase HPLC. (a) A Spherisorb ODS column was used with an eluent comprising 40% methanolwater, 0.06 M NH4OAc (pH 5.5) and 0.01% 2-mercaptoethanol. Detection was by differential pulse amperometry. MeHg+ = methylmercury, EtHg+ = ethylmercury, @Hg+= phenylmercury. Reprinted from [39] with permission. (b) A Spherisorb ODs column was used with 60:40 acetonepentane as the eluent and hydride generation atomic absorption spectrometricdetection. Reprinted from [37] with permission. (c) Conditions as for (b) except that a 70:30 acetone-pentane eluent was used. Reprinted from [37] with permission.

polar polymeric stationary phases, so these materials can therefore be employed for the separation of a wider range of solutes using the ion-suppression technique than is possible with c18 stationary phases. The utility of ion-suppression on polymeric stationary phases can be appreciated by considering the separation of the homologous series of aliphatic carboxylic acids. Neither ion-exchange nor ion-exclusion chromatography yields a complete separation of these species. However, ion-suppression coupled with gradient elution and suppressed conductivity detection enables the separation of butyric through to stearic acid, as illustrated in Fig. 8.6. The gradient used involved an increase in the percentage of organic modifier in the eluent and a decrease in eluent pH. Carboxylic acids more hydrophilic than butyric acid were eluted as a single peak at the column void volume.

232

Chupter8

ityric

Lauric

Capric

I I

0

I

5

I

10

I

15

I

I

20 25 lime (min)

I

30

I

35

1

LO

I

45

Fig. 8.6 Gradient elution ion-suppression chromatogram of carboxylic acids, obtained on a polymeric reversed-phasecolumn. A Dionex MPIC-NS1 column was used with a gradient of 100% eluent A ( t 4 ) to 100% eluent B (t=20 min), with maintenance of eluent B after this time. Eluent A is 24% acetonitrile and 6%methanol in 0.03 mM HC1. Eluent B is 60% acetonitrile and 24% methanol in 0.05 mM HCl. Detection was by suppressed conductivity. The baseline conductance for a blank w e n t has been subtracted in the chromatogram shown. Reprinted from I441 with permission.

8.3

CHELATING STATIONARY PHASES

8.3.1 Chemically-bound ligands Metal ions may be separated on a stationary phase in which a suitable ligand is immobilized onto the stationary phase. Numerous chelating stationary phases have been synthesized using stryene-divinylbenzenepolymers or silica as the support material. In each case, the ligand is chemically bound to the support using an appropriate reaction, such as silylation reactions with silica. Some examples of the ligands which can be bound in this way include iminodiacetate (Chelex 100, Dow Chemical Company), propylene-diaminetetraacetate 1451. P-diketones (e.g. trifluoroacetoacetate [461), 8hydroxyquinoline [47], isothiuronium (481, hydroxamic acids [49], dithiocarbamates (501, phenylhydrazones [51] and dithizone [52]. Solute retention can be manipulated by varying the eluent pH or through the addition of a competing ligand to the eluent. The key factor in the success of the above materials as chromatographic stationary phases is the rate at which the metal-ligand complex is formed and dissociated. Slow rates will lead to poor peak shape in the chromatogram. An evaluation of the literature suggests that most ligands give unacceptably slow rates of reaction, so that chromatograms are typically characterized by very broad peaks. A recent study has compared 9 different chelating stationary phases and has shown that some useful separations can be achieved on dithizone silica gels, as illustrated in Fig. 8.7. It is interesting to note that the same metal ions shown in Fig. 8.7 can be well separated using either ion-exchange or ion-interaction chromatography (e.g. see Figs. 4.17 and 6.3).

Miscellaneous Separation Methotlr

233

Pb2

1 1 1 1 1

0

10 20 Time (mid

Separation of metal ions on a stationary phase formed by binding dithizone functionalities to silica gel. The eluent was 15 mM tartrate at pH 4.0. Detection was by specaophotometry after post-column reaction with 4-(2-pyridylazo)-resinol. Reprinted from [52] with permission.

Fig. 8.7

8.3.2 Crown ether stationary phases

Considerable effort has been expended over recent years on the development of stationary phases in which a crown ether is chemically bound to a suitable support, such as silica or an appropriate polymer. Crown ethers (or cyclic polyethers) are cyclic compounds which possess an inner cavity, generally consisting of oxygen atoms linked by ethylene bridges. Fig. 8.8 shows two examples of these compounds. Crown ethers are non-systematically named according to the total number of atoms in the ring, the number of oxygen atoms and any substituents on the ring. Thus the fiist crown ether in Fig. 8.8 is called 18-crown-6 (total of 18 atoms in the ring, with 6 oxygen atoms), whilst the second is called benzo-18-crown-6 (to indicate the benzene ring substituent). Cryptands are related compounds having two interconnected rings which produce a three dimensional cavity. The final compound in Fig. 8.8 is one such cryptand.

Synthesis of stationary phases Crown ether stationary phases may be synthesized in three ways. The simplest approach is to impregnate a silica particle with a solution of a suitable crown ether in formic acid, followed by cross-linking with formaldehyde [53], and is illustrated schematically for dibenzo-18-crown-6 in Fig. 8.9(a). The resultant material is

234

Chapter 8

18-crown-6

Benzo- 18-crow n-6

O

d

Cryptand-n-decyl-2.2.2 Fig. 8.8 Structures of some cyclic polyethers

Fig. 8.9 Structures of some crown ether stationary phases. (a) The crown ether is coated onto silica and then polymerized. (b) Typical bonding arrangement of a crown ether onto silica. (c) Typical bonding arrangement of a crown ether onto a resin. Reprinted from [53, 581 with permission.

Miscellaneouy Separation Methods

235

stable mechanically and is resistant to hydrolysis. Alternatively, the crown ether can be chemically bound to silica by silyl ether linkages using conventional silylation reactions. An example of the resultant stationary phase structure is shown in Fig. 8.9(b) for benzo18-crown-6. Finally, similar bonding reactions can be performed using resins as the support and a typical stationary phase structure is depicted in Fig. 8.9(c), again using benzo- 18-crown-6 as the ligand. Chromatographic properties The chromatographic utility of crown ether stationary phases rests in their ability to complex cations of a specified size. As the size of the cavity in the crown ether is altered, so too does the selectivity of the stationary phase. For example, Li+, Na+ and K+ are bound preferentially to 12-crown-4, 15-crown-5 and 18-crown-6 stationary phases, respectively, by virtue of the increasing cavity size. However, it is not essential for the solute cation to reside within the cavity since layered structures in which the solute is located between two cavities can also be formed [54]. The bound metal ion imparts a positive charge to the crown ether and this is balanced by a suitable counter anion. The binding of metal ions on crown ether stationary phases is .dependent on the following factors: (i) The size of the cation. (ii) The nature of the associated anion. (iii) The organic modifier content of the eluent. The influence of cation size has been discussed above. The most common elution trend for alkali metal ions on benzo-18-crown-6 stationary phases (which show preference for K+) is for K+ to be eluted last, with the remainder being eluted in order of size. That is, retention usually follows the sequence:

The nature of the associated anion can sometimes exert an influence on retention which dominates all other effects. "Soft" (polarizable) anions such I- and SCN- are more strongly associated with the bound cation than are "hard" anions, such as S04*-and C1-. It has been demonstrated that, when a mixture of anions and cations is injected, the most strongly retained species will be the preferred cation associated with the softest anion [55]. For example, injection of a mixture of Li+, K+,C1-, Br- onto a dibenzo-18-crown6 stationary phase gives four peaks corresponding to LiCI, KCI, LiBr and KBr [56]. The first eluted peak (LiCI) contains the least preferred cation (Li+) and the hardest anion (Cl-), whilst the last eluted peak (KBr) contains the preferred cation (K+) and the softest anion (Br-). Water alone can be used as an eluent in this form of chromatography, but the separations which are obtained are often relatively poor. The reason for this is that water complexes the solute cations and competes effectively with the crown ether. Addition of an organic modifier, such as methanol, to the eluent lowers its polarity and

236

Chapter8

5Br

KBI

Liar

KBr

1

0

I

5

1

I

1

10 15 20 Time (min) (a 1

I

25

-

0 10 20 30 LO Time (min) (b)

Fig. 8.10 Separation of (a) cations and (b) anions on crown ether stationary phases. (a) A paly(benzo-l5-crown-5)-modifiedsilica was used as stationary phase with water as eluent. Detection was by conductivity. Reprinted from [64]with permission. (b) A benzo-18-crown-6 modified silica stationary phase was used with water as eluent. Conductivity detection was used. Reprinted from [65] with permission.

decreases complexation with the solutes. This, in turn, results in increased solute retention and therefore improved separation. However, detection is more simple with a water eluent than with aqueous-organic mixtures, so the former eluent is preferable.

Applications Crown ether stationary phases can be used in two ways. First, a series of alkali metal salts with a common anion can be separated. These salts will be eluted with the cations following the sequence determined by the preference of the particular crown ether used 153, 57-62]. An example of such a separation on silica coated with poly(benzo-t5-crown-5) is shown in Fig. 8.lqa). We note that the cations must be present as their Br- salts for the separation to be reproducible. An alternative application is the separation of anions, using the cation bound to the crown ether as a site for selective retention of anions. The salts will be eluted in the sequence of reducing hardness of the anions [53,56, 57, 59, 60,62, 631. A typical separation by this method is given in Fig. g.lO(b). We again note that the separation is dependent on each anion being present as the K+ salt. If other cations are present, the elution order could vary.

MiscellaneousSeparation Methods

237

The separations shown in Fig. 8.10 are of limited practical applicability because of the need to specify a particular counter-anion or counter-cation for the solute ions. It has been suggested that a sample could be converted to the correct form by passage through a suitable ion-exchange column prior to analysis [54], but no results using this approach have been reported. A more attractive alternative is to use a sufficiently high concentration of the desired counter-cation or anion in the eluent so that the sample counter-ions do not influence solute retention. This approach has been reported for anion determinations using a stationary phase comprising the macrocycle cryptand ndecyl 2.2.2, which has the structure shown in Fig. 8.8, coated onto reversed-phase supports with alkali metal hydroxides as eluents [66, 671. The cryptand is hydrophobically bound to the support in the same manner as used in "permanentcoating" ion-interaction chromatography (see Section 6.3.2). Excellent chromatographic efficiencies were obtained and solute retention was not dependent on the cations present in the sample.

Gradient elution on macrocyclic stationary phases The above-mentioned studies with the cryptand n-decyl 2.2.2 have opened a new possibility for gradient elution of anions. The cation bound to the macrocycle acts as an anion-exchange site for solute anions. The number of bound cations, and hence the anion-exchange capacity of the column, is dependent on the identity of the alkali metal hydroxide used as eluent. The highest anion-exchange capacity is produced with KOH eluents (because of the preferential binding of K+ to the macrocycle), whereas the smallest capacity results when LiOH is used as the eluent. Thus, a gradient in which the eluent is changed from, for example, 30 mM NaOH to 30 mM LiOH will therefore result in a progressive decrease in column ion-exchange capacity. The change in baseline conductivity in such a system will be minimal, especially if a suppressor is employed. Fig. 8.1 1 shows a chromatogram obtained using this method, which has been called "gradient capacity IC" [67]. This method has considerable promise, especially in view of the ease with which the stationary phase can be prepared. 8.4 8.4.1

MICELLE EXCLUSION CHROMATOGRAPHY

Introduction

Many surfactant molecules form micelles in concentrated solution. Micellar chromatography [68, 691 utilizes micellar eluents as a means of improvement of the chromatographic selectivity. We have seen earlier (Section 6.3.2, Fig. 6.4) that micellar eluents can be employed,in ion-interaction chromatography of inorganic anions. The chromatograms obtained show good chromatographic efficiency, but the elution order of solutes (and hence the selectivity of the method) is the same as that for conventional ionexchange chromatography. This can be explained by the fact that in this particular case, the surfactant was strongly adsorbed onto the reversed-phase column which was used, so that ion-exchange was the dominant retention mechanism. On this basis, use of a different type of stationary phase, such as a size-exclusion material, could lead to changes in chromatographic selectivity. Okada [70,711 has investigated this approach

238

Chapter 8

F-

p-

Acct, SC N-

I r

0

I

10

1

20 Time (min)

30 1

LO I

Fig. 8.11 Gradient elution on a macrocyclic stationary phase using variation of the ion-exchange

capacity of the column. A Dionex MPIC-NS1 column was used after conditioning with cryptand-ndecyl-2.2.2. The eluent was a linear gradient (over 20 min) from 30 mM NaOH to 30 mM LiOH. Detection was by suppressed conductivity. Chromatogram courtesy of John D. Lamb. for the separation of inorganic anions and cations. The eluent consists of an aqueousorganic mixture containing a micellar solution of a surfactant of appropriate charge. The micelles in the eluent are excluded from part of the stationary phase, whilst the solute ions partition between the micelle and the bulk solvent in the eluent, and also between the stationary phase and the eluent, This method has been entitled "micelle exclusion chromatography" on the basis of this separation mechanism. 8.4.2

Micelle exclusion chromatography of anions

When anionic solutes are to be separated, the surfactant in the eluent should be positively charged. Hexadecyltrimethylammonium chloride (HTAC) and dodecyltrimethylammonium chloride (DTAC) are therefore suitable surfactants [71]. Use of a size-exclusion stationary phase means that the aggregated surfactant forming the micelle

Miscellaneous Separation Methodr

239 Mnll

L I I'

0

30

Time (rnin) (01

I

LO

r

0

I

10

I

20 lime (rnin)

I

30

1

LO

(bl

Fig. 8.12 Determination of (a) inorganic anions and (b) inorganic cations by micelle exclusion chromatography. (a) An Asahipak GC910H (poly(viny1 alcohol)) size-exclusion column was used with 0.01 M hexadecyltrimethylammonium chloride as eluent. Detection was by spectrophotometry at 210 nm. Reprinted from [70]with permission. (b) The same column as for (a) was used, with a 10 min gradient from 100%eluent A (commencing at t = 10 min) to 100%eluent B. Eluent A was 25 mM sodium dodecyl sulfate (SDS) and 80 m M a-hydroxyisobutyric acid at pH 4.0. Eluent B was 25 mM SDS and 60 mM tartaric acid at pH 4.0.Detection was by spectrophotometry after post-column reaction with 4-(2-pyridylazo)-resorcinol.Reprinted from [71] with permission.

can penetrate only partially into the pores of the stationary phase, whereas the monomeric surfactant can penetrate fully by virtue of its smaller size. The small solute anions can also penetrate into the pores and are therefore retained in this region of the stationary phase by ion-interaction chromatography. Solute anions will also partition between the bulk eluent and the surfactant micelles, and between the bulk eluent and the liquid inside the pores of the column. Retention is therefore based on three distinct processes. Fig. 8.1 2(a) shows a chromatogram obtained using HTAC as the surfactant. The elution order in Fig. 8.12(a) is different from that exhibited by conventional ionexchange or ion-interaction systems, particularly for I- and Br-. The retention order and

Chaprer 8

240

retention times observed in this system are dependent on a number of parameters, including: (i) (ii) (iii) (iv) (v)

The concentration of surfactant The nature of the surfactant used The nature of the surfactant counter-anion The Concentration of added salt The concentration of organic solvent in the eluent

All of these parameters also affect retention in ion-interaction chromatography, which supports the hypothesis that this process plays a significant role in the mechanism of micelle exclusion chromatography. However, the observed chromatographic selectivity shows that there are additional conmbutions to solute retention which are attributable to the partitioning processes described above.

8.4.3 Micelle exclusion chromatography of cations Micelle exclusion chromatography of cations requires an anionic surfactant, such as sodium dodecylsulfate (SDS),and a size-exclusion stationary phase [71]. Again, the surfactant micelles are partially excluded from the pores of the stationary phase, whereas monomeric surfactant and solute cations can penetrate fully. Retention occurs by the same mechanisms outlined above for anions and the retention times are dependent on the same parameters as listed earlier. As with ion-exchange and ion-interaction methods, further control over solute retention can be accomplished through the addition of a complexing agent, such as citrate, to the eluent. Moreover, gradient elution can be achieved by changing the nature of the complexing agent whilst keeping other eluent parameters constant. Fig. 8.12(b) shows a chromatogram obtained using this approach.

8.5

REFERENCES

1 2

Nickless G., J . Chromurogr.. 313 (1985) 129. Krull I.S., in Bemhard M.,BMkman F.E. and S d e r P.J. (Eds.), The fmponance of Chemical "Speciation" in Environmenral Processes, Springer-VerlagBerlin, Heidelberg,

3 4 5 6

Timerbaev A.R., Petrukhin O.M.and Zolotov Yu.A., Fres. 2. Anal. Chem., 327 (1987) 87. Steinbrech B., J. Liq. Chromurogr., 10 (1987) 1. OLaughlin J.W.. J . Liq. Chromurogr.,7 (1984) 127. Machnald J.C., in MacDonald J.C. (Ed.) Inorganic Chromatographic Analysis, WileyInterscience, 1985, p. 285. Hutchins S.R., Haddad P.R. and Dilli S., J. Chromufogr.,252 (1982) 185. Mooney J.P., Meaney M., Smyth M.R., Leonard R.G. and Wallace G.G., Analyst

1986, p. 579.

7 8

(London), 112 (1987) 1555.

9 10 11

12

Heisz O., GfT Fuchz. Lob., 29 (1985) 113. Gierira R.C. and Can P.W., J. Chromurogr.Sci., 20 (1982) 461. Schwedt G. and Rudde R.,Chromographia, 15 (1982) 527. Roston D.A., Anal. Chem., 56 (1984) 241.

Miscellaneous Separation Methodr

13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50

51 52 53

241

Cardwell T.J. and Caridi D.. J. Chromatogr., 193 (1980) 53. Irth H., Brouwer E.. De Jong G.J., Brinkman U.A.T. and Frei R.W., J. Chromatogr.,439 (1988) 63. Eggers H. and Russel H.A.. Chromatographia, 17 (1983) 486. Wheeler G.L. and Lott P.F., Microchem. J., 19 (1974) 390. Tong A., Akarna Y. and Tanaka S., J. Chromatogr.478 (1989) 408. Casoli A., Mangia A. and Predieri G., Anal. Chem., 57 (1985) 561.. Main M.V. and Fritz J.S., Anal. Chem., 61 (1989) 1272. Smith R.M., Butt A.M. and Thakur A.. Analyst (London), 110 (1985) 35. Ge H. and Wallace G.G.. Anal. Chem., 60 (1988) 830. King J.N. and Fritz J.S.. Anal. Chem., 59 (1987) 703. Hoffman B.W. and Schwedt G., J. HRC & CC, 5 (1982) 439. Mazzo D.J..Elliott W.G., Uden P.C. and Barnes R.M., Appl. Spectrosc., 38 (1984) 585. Shih Y-T. and Carr P.W., Talanta, 28 (1981) 41 1. Bond A.M., Garrard W.N.C.. Heritage I.G., Majewski T.P.. Wallace G.G., McBumey M.J.P., Crosher E.T. and McLachlan L.S., Anal. Chem., 60 (1988) 1357. Smith R.M. and Yankey L.E., Analyst (London), 107 (1982) 744. Bond A.M., Knight R.W., Reust J.B.. Tucker D.J. and Wallace G.G., Anal. Chim Acta, 182 (1986) 47. Irgolic K.J., Stockton R.A.. Chakraborti D. and Beyer W., Spectrochim Acta, 38B (1983) 437. Fish R.H., Brinkman F.E. and Jewett K.L., Environ. Sci. Technol., 16 (1982) 174. Haswell S.J.. ONiell P. and Bancroft K.C.C.. Talanta, 32 (1985) 69. Lawrence K.L., Rice G.W. and Fassel V.A., Anal. Chem.. 56 (1984) 289. LaFreniere K.E., FasseI V.A., and Eckels D.E., Anal. Chem., 59 (1987) 879. Bushee D.S.. Analyst (London), 113 (1988) 1167. Cast C.H., Kraak J.C., Poppe H. and Maessen F.J.M.. J. Chromatogr.. 185 (1979) 549. Burns D.T., Glockling F. and Harriott M., J. Chromatogr.. 200 (1980) 305. Burns D.T.. Clockling F. and Haniott M..Analyst (London), 106 (1981) 921. Brinkman, F.E., Blair W.R., Jewett K.L. and Iverson W.P.. J. Chromatogr. Sci., 15 (1977) 493. MacCrehan W.A. and Durst R.A., Anal. Chem.. 50 (1978) 2108. Vickrey T.M.. Howell H.E. and Paradise M.T., Anal. Chem., 51 (1979) 1880. Enos C.T., Geoffroy G.L. and Risby T.H., J. Chromatogr. Sci.. 15 (1977) 83. Bidlingmeyer B.A.. J. Chromatogr. Sci., 18 (1980) 525. Skelly N.E., J. Chromatogr.,250 (1982) 134. Slingsby R.W., J. Chromatogr.. 371 (1986) 373. Moyers E.M. and Fritz J.S., AnaL Chem., 49 (1977) 418. DenBleyker K.T., Arbogast J.K. and Sweet T.R., Chromatographia,8 (1983) 449. Jezorek J.R and Freiser J.R., Anal. Chem., 5 1 (1979) 366. Koster G. and Schmuckler G., Anal. Chim.Acta, 38 (1967) 179. Phillips R.J. and Fritz J.S.. Anal. Chim Acta. 121 (1980) 225. Raja R.. Am. Lab., 14 (1982) 35. Simonzadeh N. and Schilt A.A., Talunta, 35 (1988) 187. Faltynski K.H. and Jezorek J.R.. Chromatographia,22 (1986) 5. Blasius E. and Janzen K.P., Israel J. Chem. 26 (1985) 25.

242 54

55

56 57 58 59 60 61 62 63

64 65 66 67 68 69 70 71

Chapter 8 Dasgupta P.K., in Tarter J.G. (Ed.), Ion Chromurogruphy, Marcel Dekker, Inc.. New York, NY, 1987, p. 191. Blasius E., Janzen K-P., Klein W., Nguyen V.B., Nguyen-Tien T., Pfeiffer R., Scholten G.,Simon H., Stockemer H. and Touissant A.. J . Chromutogr.. 201 (1980) 147. Igawa M., Saito K., Tsukamoto J. and Tanaka M., A d . Chem., 53 (1981) 1942. Lauth M. and Gramain P.,J. Liq. Chromutogr., 8 (1985) 2403. Iwachido T., Naito H., Samukawa F., Ishimaru K. and Toei K., Bull. Chem. SOC.Jap., 59 (1986) 1475. Igawa M., Ito I. and Tanaka M., Bumeki Kuguku, 29 (1980) 580. Nakajima M., Kimura K. and Shono T., Bull. Chem. SOC.Jup., 56 (1983) 3052. Nakajima M., Kimura K., Hayata E. and Shono T., J. Liq. Chromatogr., 7 (1984) 21 15. Blasius E., Janzen K-P.. Simon H. and Zender J., Fres. 2.A d . Chem., 320 (1985) 435. Igawa M., Saito K., Tanaka M. and Yamabe T., Bumeki Kuguku, 32 (1983) E137. Nakajima M., Kimura K. and Shono T., Anal. Chem., 55 (1983) 463. Lauth M. and Gramain P.,J. Chromutogr., 395 (1987) 153. Lamb J.D. and Drake P.A., J. Chromatogr.,482 (1989) 367. Lamb J.D., Drake P.A. and Wooley K.E., 2nd Int. lon Chromutogr. Forum, Boston, September (1989). Armstrong D.W. and Nome F., Anal. Chem., 53 (1981) 1662. Armstrong D.W. and Fendler J.H., Biochim. Biophys. Acta, 478 (1977) 74. Okada T., Anal. Chem., 60 (1988) 1511. Okada T., Anal. Chem., 60 (1988) 21 16.

Part III Detection Methods

244

Principles

AC bridge Bipolar pulse 4-electrode cell Differential detection Packed column Hollow-fibre Packed-fibre Micromembrane Electrochemical Post-suppressors

and :ONDUCTIVITY $Cellscircuitry Chap 9 ) Suppressors

Principles

E-

Modes

Amperometry Coulometry Polarography Direct F n p t

Electrodes

$ :a Reference i;

Types

2LECTROCHEMICAL Chap 10)

z-. z-, z-

Cells

DETECTION METHODS

Thin-layer

Fl?l;:tpugh

-

Principles

Ion-selective electrodes Coated-wire Metallic copper Direct Indirect

Indicator electrodes POTENTIOMETRY (Chap 11)

Modes Cells

Spectrophotometry

UV visualization

?:z$.t

Refractive index SPECTROSCOPY (Chap 12)

.POST-COLUMN REACTION (Chap 13)

Atomic spectroscopy -C

fluorescence Indirect fluorescence Indirect phosphorescence Atomic absorption Atomic emission

Types

Solution PCR Packed-bed PCR

Photoluminescence

Hardware

'M"iz:ichambers

Anions

Phosphorus oxoanions Ferric perchlorate

Cations

L Organics Schematic overview of Pan 111.

+Direct

- = -

~ ~ , n a z o

245

Chapter 9 Co n d u c tiv ity D e tec tion 9.1

INTRODUCTION

The first report on the use of conductivity as a method for the detection of ions in the eluate from a chromatographic column can be credited to James et al. in 1951 [l]. The technique was used sporadically over the next 25 years or so, being seen as too insensitive for all but preparative scale chromatography. Conductivity detection gained popularity only after the introduction of specialized IC columns and post-column signal enhancement methods, the former of which have been described in Chapter 3 and the latter of which will be discussed later in this Chapter. Conductivity detection has two major advantages for inorganic ion analysis. The first of these advantages is that all ions are electrically conducting, so that conductivity detection should be universal in response, and the second is that conductivity detectors are relatively simple to construct and operate. Conductivity detection is very widely employed in IC and applications are therefore abundant. A survey of the literature (Appendix A) shows that this form of detection is employed in approximately 55% of publications dealing with IC and is utilized with ion-exchange, ion-interaction and ionexclusion separation methods. Conductivity detection will be discussed here in terms of the principles of its operation, the modes of detection employed, cell designs, postcolumn signal enhancement (suppression), performance characteristics of conductivity detectors, and applications.

9.2 9.2.1

PRINCIPLES OF CONDUCTIVITY DETECTION Nature of electrical conductivity of electrolyte solutions

A solution of an electrolyte will conduct an electrical current if two electrodes are inserted into the solution and a potential is applied across the electrodes. The more current conducted by the solution, the higher is its electrical conductivity. Ohm's Law applies, so that:

V = IR

(9.1)

where V is the applied potential (volts), I is the current (amps) and R is the resistance (ohms), The reesislance of the solution will be determined by several factors, including the concentration and type of ionic species in the solution, and the temperature. The

Chapter 9

246

conductance of the solution, G, (having the units of Siemens, which are represented by the symbol S) is given by the reciprocal of resistance. Note that the older units for conductance are reciprocal ohms (mhos).

G = -1

R

(9.2)

The resistiviry (p). in units of ohms.cm, is given by:

p=- A R

L

(9.3)

where A is the cross-sectional area (cm2) of the electrodes inserted into the solution, and L (cm) is the distance between them, The inverse of resistivity is the conductivity, k, which has units of S.cm-', and is also often referred to as the specific conductance.

The specific conductance of an electrolyte will vary with concentration, so it is necessary to introduce a further term to facilitate the comparison of electrolytes. This term is the equivalent conductance (A) and has the units S.cm2equiv-'. A is given by:

where C is the concentration of the electrolyte, expressed as equivalents per lo00 em3 of solution. We can now combine the geometrical characteristics of the cell (i.e. A and L) into a single term, called the cell constant, K, which has units of cm-'. K is given by:

K = -L

A

(9.6)

This enables us to rewrite eqn. (9.4) as:

k=GK

(9.7)

so that the conductance can now be defined as:

G=- AC 1000 K If G is expressed in pS, eqn. (9.8) becomes:

(9.8)


G =

247

1OOOAC -- A C K 10-~K

(9.9)

The conductance of a solution can be seen to be proportional to the equivalent conductance of the electrolyte and its concentration. In addition, the lower the cell constant, the higher the conductance. Looking at the factors which influence K (eqn. (9.6)), we see that conductance is increased for cells with large surface area electrodes which are close together. The equivalent conductance is subject to activity effects (particularly ion-ion interactions) and therefore shows variation with increasing concentration of the electrolyte solution. lon-pairs may form as a result of ion-ion interactions, leading to a decrease in the effective conductance of the solution. The relationship between G and C therefore becomes non-linear at high ionic strength. If the value of equivalent conductance for an infinitely dilute solution of the electrolyte is given by ho, then the decrease in equivalent conductance observed at higher concentrations is given by the Debye-Huckel-Onsager equation: 1

A = 120 - (A + BAJ (C)?

(9.10)

where A is a constant dependent on the dielectric constant, temperature and viscosity of the solvent, and B accounts for the relaxation effect resulting from asymmetrical charge distribution arising from movement of the solvated ions under the influence of the applied potential. This latter effect serves to retard the movement of ions and causes a decrease in the effective conductance. Eqn. (9.10) applies only when the electrolyte concentration is significantly higher than that commonly encountered in IC. Thus, conductances in IC can be calculated within acceptable error using A0 (limiting equivalent Conductance) values by substitution in eqn. (9.9), which is more correctly written as: (9.11)

It must be appreciated that the conductance of the solution results from bofh the anions and cations of the electrolyte. We must therefore calculate conductance using values for the limiting equivalent ionic conductances (h) of the individual anions and cations in solution. Eqn. (9.1 1) can now be rewritten as:

G =

(h, + hJ

c

(9.12)

K where A+ and Ic are the limiting equivalent ionic conductances of the cationic and anionic components of the electrolyte, respectively. Typical limiting equivalent ionic conductances for common solute and eluent species used in IC are listed in Table 9.1. It

Chapter 9

248 TABLE9.1

LIMITING EQUIVALENT IONIC CONDUCTANCES OF SOME IONS IN AQUEOUS SOLUTION AT 25 OC [2]

OH Fe(CN)64Fe(CN)63m42-

CN' so42-

BrI-

a' Cz042'

m2NQm430 4 -

SCN-

clo3ciaate3-

HCOO F HWcH3cooPhthalate? c2HscooBenzoate'

198 111 101 85 82 80 78 77 76 74 72 71 69 67 66 65 56 55 54 45

41 38 36 32

H30+

Rb+ cs+

K+ NH4+ Pb2+ Fe3+ Ba2'

A ? ca2+

Sr*+ CH3NH3'

350 78 77 74 73 71 68

64 61 60 59 58

cu2+ cd2+

55

Fe2+ Mg2+

54

co2+

7h2+

Na+ Phenylethylammonium+ Li+ N(c2H5>4+ Benzyhmmonium+ Methylpyridinium+

54 53 53 53 50 40

39 33 32 30

can be noted that H30+ is a very strongly conducting cation, showing a 5 value about 5 times higher than other cations. Similarly, OH- is the most strongly conducting anion, having a A value which is 2-3 times greater than most other anions. As an example of the practical application of eqn. (9.12), we can calculate the conductance of an eluent comprising 5 mM LiOH, measured in a cell with a cell constant of 10 cm-l. From Table 9.1 we note that the limiting equivalent ionic conductances of Li+ and OH- are 39 and 198 S.cm2 equiv-', respectively. Substitution into eqn. (9.12) gives: (9.13)

ConductiviryDetection

249

9.2.2 Factors influencing limiting equivalent ionic conductance

The limiting equivalent ionic conductance of an ion is given by eqn. (9.14) [3]: (9.14)

where e is the electronic charge, F is the Faraday constant, Z j is the number of charges on the ion, q is the viscosity, and r is the ionic radius. An increase in the number of charges on an ion does not necessarily lead to an increase in Xi, since an inverse proportionality exists between and the ionic radius, r. Thus, large ions with multiple charges can exhibit the same Li value as smaller, singly charged ions. This has important ramifications for IC in that eluent ions of multiple charge are powerful competing ions, but may have inappropriately high background conductances unless the ionic radius is large. When the combined attributes of high eluotropic strength and low background conductance are required, large ions with multiple charge, such as phthalate, citrate and trimesate, are therefore suitable. 9.2.3

Theory of conductivity response

The operating principles of conductivity detection in IC can best be illustrated by reference to ion-exchange as the separation mode, and by considering the conductance of a typical eluent prior to and during the elution of a solute ion. For simplicity, we will consider only anion-exchange at the present time, but the detector response equations which will be developed are equally applicable to cation exchange, provided the obvious amendments are made. The response equations developed here below follow the method of Fritz et al. [4-61. We first calculate the background conductance of the eluent; i.e. the conductance which results when onZy the eluent is flowing through a fully equilibrated ion-exchange column and thence to the cell of a conductivity detector. It is assumed that the eluent contains a single species which may be only partially dissociated into the ions E+and E-. Eqn. (9.12) can be modified slightly to give: (9.15)

where CE is the total concentration of the eluent and IE is the fraction of eluent species which is present in the ionic form, i.e. as E-.We note in passing that the cell constant can be readily determined in practice by measuring the conductance of a solution containing species with known limiting equivalent ionic conductances ( e . g . KCl, for which 5 = 74 and A- = 76 S.cm2.equiv-l at 25OC). The values of conductance calculated from eqn. (9.15) are, of course, only approximate because values of limiting ionic equivalent conductance are used and the cell constant determined by the above-mentioned method is of limited accuracy. We must now examine the concentration changes which occur when a solute anion is eluted from the column through which the above eluent flows. In the anion-exchange

Chapter 9

250

C

.-0

C

Eluent

E! C 3 C

C

Solute 1 Eluent flow

-

0

0

(a 1 C

.-0

Eluent

C

E

C

C

Eluent f Low

-

s C 0

0

(b)

Fig. 9.1 Schematic representation of ion concentrations on the column and in the eluent in ion-

exchange chromatography. (a) shows concentrations immediately after injection, whilst (b) shows concentrations during peak elution. Reprinted From [7] with permission. system under consideration, we can write the following simple equilibrium:

Si

+ E;

f S;

+ Em

(9.16)

where the subscripts m and r refer to the mobile and resin phases, respectively, and S- is the solute ion. When the solute is injected at the head of the equilibrated column, S - ions are bound to the column and E- ions are released. The concentration of E- in the sample volume therefore depends on the concentration of S - in the sample. A band of E- of concentration greater or less (depending on the sample concentration) than that of the bulk eluent is produced and, to a first approximation, moves through the column to produce the characteristic solvent peak observed in IC. This solvent peak can therefore be positive or negative (Fig. 9.1(a)). Eluent ions, which continue to be pumped through the column, compete with S- for the ion-exchange sites, causing the sample ions to move down the column at a speed determined by their selectivity coefficients. The total ion concentration in the column and eluent remains constant throughout this elution process, as illustrated schematically in Fig. 9.l(b). The eluted S - ions therefore displace an equivalent number of E- ions from the eluent. If the concentration of solute passing through the detector is given by Cs and the degree of ionization of the solute is Is. then the eluent concentration in the detector cell during sample elution is given by: (9.17)


25 1

The conductance measured in the cell at .this time originates from the eluent and solute anions, together with the eluent cations which are required to maintain electroneutrality. The solute cations need not be considered since these are unretained on the anion-exchange column used in this example. The conductance during solute elution is therefore given by: (9.18) The change in conductance (AG) which accompanies the elution of the solute can be obtained by subtracting the background conductance (eqn. (9.15)) from the conductance during solute elution (eqn. (9.18)) to give: (9.19) The solute concentration during elution will follow an approximately Gaussian profile and the detector response will therefore change in a similar manner. Eqn. (9.19) is applicable to all forms of ion-exchange TC and shows that the detector response depends on solute concentration, the limiting equivalent ionic conductances of the eluent and solute anions, and the degree of ionization of solute. The last of these parameters is governed by the eluent pH (since the eluent is generally well buffered) and eqn. (9.19) suggests that weak acid anions such as acetate, fluoride, formate, phosphate, oxalate, etc., should show appreciable decreases in sensitivity as the eluent pH is lowered; this behaviour has been observed in practice [ 5 ] . However, many of the solutes determined by IC are fully ionized in aqueous solution, so eqn. (9.19) generally takes the form:

AG =

(As- - AE-)cs

(9.20)

10-~K This equation shows that if a conductivity detector is used to monitor the effluent from an ion-exchange column, the signal observed for an eluted solute is proportional to the solute concentration and to the difference in limiting equivalent ionic conductances between the eluent and solute ions. A similar relationship (eqn. (9.21)) can be derived for the conductimetric detection of cations after ion-exchange separation [8]. (9.21) where S+ and E+ are the solute and eluent cations, respectively. Eqns. (9.20) and (9.21) are fundamental to the understanding of the function of conductivity detection and can be used as a basis for discussing the various modes of conductivity detection employed in IC. as outlined in the next Section.

Chapter 9

252

-

x

GC 0

CI

u

3 '0

c

0

0 c

C

0,

Solute

d

0

.-> 3

(A,)

CT P,

0)

.-cC

z

Eluent 2

3

(AEZ)

Schematic representation of indirect (negative peak) and direct (positive peak) conductivity detection using an eluent competing ion with high (eluent 1) or low (eluent 2) limiting equivalent ionic conductance. Fig. 9.2

9.3

MODES OF CONDUCTIVITY DETECTION

9.3.1 Direct and indirect conductivity detection From eqns. (9.20) and (9.21), it is clear that sensitive detection can result as long as there is a considerable difference in the limiting equivalent ionic conductances of the solute and eluent ions. This difference can be positive or negative, depending on whether the eluent ion is strongly or weakly conducting. It should be remembered here that the response equations developed are applicable to any ion-exchange system in that they describe the conductance changes occurring at the column outlet. When no further chemical change is imposed prior to passage of the eluent-solute mixture to the conductivity detector, then eqns. (9.20) and (9.21) give an accurate description of the conductivity detector signal. This situation exists for non-suppressed IC,but not for suppressed IC. It will therefore be necessary to extend the above response equations to cover the case of suppressed IC and this extension is provided in Section 9.5. If the limiting equivalent ionic conductance of the eluent ion is low, then an increase in conductance occurs when the solute enters the detection cell. In general, we can define this detection mode as direct, where the solute has a higher value of the measured property than does the eluent ion. Alternatively, an eluent ion with a high limiting equivalent ionic conductance can be employed and a decrease in conductance would occur when the solute enters the detection cell. Once again, we can generally define this type of detection as indirect, where the solute has a lower value of the measured property than does the eluent ion. These detection modes are illustrated schematically in Fig. 9.2, which shows that positive peaks occur with direct detection and negative peaks result with indirect detection.

253

ConductivityDetection

K4

r 0

,

5

1

10 l i m e (min)

I

1

15

20

1

0

1

1

4

1

1

1

I

0 12 Time (min)

I

r'6

1

Fig. 9.3 Direct conductivity detection of (a) anions and (b) cations. (a) A TSK-GELEX-620 anion-exchange column was used with 1.3 mM Na2Bq07 - 5.8 mM H3B03 - 1.4 mM potassium gluconate (pH 8.5) in water-acetonitrile (88:12). Reprinted from [12] with permission. (b) A Waters IC Pak C column was used with 0.15 mM benzylamine (pH 7.14) as eluent. Reprinted from [1I] with permission. Direct conductivity detection is used for most IC methods involving the separation of anions. Eluents for non-suppressed IC, formed from salts such as potassium hydrogen phthalate [9] or sodium benzoate [lo] contain competing anions with moderately low limiting equivalent ionic conductances (see Table 9.1). Similarly, direct conductivity detection of cations is possible with eluents formed from organic bases [8, 111. Chromatograms showing direct conductivity detection of anions and cations are given in Fig. 9.3. Indirect conductivity detection can be applied to anions using hydroxide eluents [ 131, and to cations using mineral acid eluents [ 111. Examples of these separations are given in Fig. 9.4. Each of the eluents used in these examples provides excellent detector response because of the very high limiting equivalent ionic conductances of the hydroxide and hydrogen ions, which act as the competing ions.

254

Chapter 9

(b) Timelmin)

0

L

8

12

16 I

I

Fig. 9.4 Indirect conductivity detection of (a) anions and (b) cations. (a) A TSK-GEL 620 SA column was used with 2 mM KOH as eluent. Reprinted from [ 131 with permission. (b) A Waters IC Pak C column was used with 2 mM HNO3 as eluent. Reprinted from [l 11 with permission.

Simultaneous direct and indirect conductivity detection

The possibility also exists for two eluent ions to contribute simultaneously to the detector response. This may occur when the eluent contains two competing ions, provided that the concentrations and selectivity coefficients of the two ions are not such that only one dominates analyte elution. When both eluent ions participate in solute elution and both have similar valucs of limiting equivalent ionic conductance, there is little change in detection sensitivity compared to the use of each eluent alone. However, when one eluent ion produces direct conductivity dctection and the other produces indirect detection, then the overall detector response will be much lower than for the individual eluents. This behaviour has been reported [81 for cation elution with aromatic base eluents operated at acidic pH. Solute elution is accomplished jointly by the aromatic base cation (low h+, direct detection) and H3O+ (high A+, indirect detection). The net outcome is that solutes give negative or positive peaks (or even no peak at all), depending on the relative influences of the two eluent ions. Similar behaviour can be expected for anion separations using aromatic acid elucnts at high pH. 9.3.2

Magnitude of conductance change on sample elution

An estimate can be made of the conductance change occurring on elution of a typical solute in IC. If we assume that a 10 ppm CI- solution is injected and that a fivefold dilution of the sample occurs during passage of the solute through the

255


chromatographic column, then the concentration of C1- in the detection cell is 2 ppm (5.63 x M). Assuming a cell constant of 10 cm-' for the conductivity detector, then the conductance change on sample elution, together with the background conductance of the eluent can be calculated. These parameters are shown in Table 9.2, for typical anionexchange eluents. It is seen readily that the conductance changes which occur are very small indeed and this places stringent requirements on the conductivity detectors which are applicable to IC. TABLE 9.2

CONDUCTANCE CHANGE ON ELUTION OF 10PPM CHLORIDE WITH VARIOUS ELUENTS

Eluent

Background conductance (PS)

Conductance change (AG)

1 mM KBza, pH 7 1 mM K2Pb, pH 7 1 mM KOH

10.6 18.4 27.2

0.25

Direct Direct

1.7 rnM NaHC03, 1.8 mM Nap203

48.1

0.13 -0.69 O.OlC, 0.09d

Detection mode

(ClS 1

Indirect Direct

a Bz = benzoate anion.

P = phthalate di-anion. Calculated assuming that CI' is eluted only by COs2-. Calculated assuming that C1' is eluted equally by HCO3- and C032-.

9.4 9.4.1

ELECTRONIC CIRCUITRY AND CELL DESIGN FOR CONDUCTIVITY DETECTION Introduction

The measurement of conductivity in liquids is performed by the application of an electric potential between two electrodes. Under the influence of this field, anions move towards the anode, whilst cations move towards the cathode. The current which results is dependent on the applied potential and also on the nature and concentration of ionic species present in the solution. The limiting equivalent ionic conductances listed in Table 9.1 show that different ionic species have different ionic mobilities in solution. I: is usual for the potential to be applied in a pulsed or sinusoidal manner, i.e. as an alternating current. The amplitude of this potential must be such that thermal effects or chemical reactions at the electrode surfaces (generally Faradaic oxidation or reduction) do not occur to any significant extent. At the same time, the amplitude should be as high as possible, since the detection signal i s directly related to the applied potential. The suitable range of frequencies for the applied potential is from approximately 50 Hz to 10,000 Hz [3]. As the frequency of the applied potential increases, ions gradually become unable to transfer all the energy imposed on them by the field into translational

Chapter 9

256

Fig. 9.5 AC conductance bridge (a) and equivalent circuit of the conductance cell in full (b) and simplified (c) forms. R, is the solution resistance, GJis the double layer capacitance at the electrode surface, ZFis the Faradaic impedance, Rc is the contact and lead resistance, Cc is the contact and lead capacitance, Ci is the inter-electrode capacitance, C, is the combined parallel capacitance, and C, is the combined series capacitance. Reprinted from [161 with permission.

motion. Other phenomena such as distortion of the counterionic atmosphere or the formation of dipoles also become predominant [14].

9.4.2 AC conductance bridge The simplest alternating current conductance circuit is based on the Wheatstone bridge design. Fig. 9.5(a) shows the circuit used. There are several problems involved in the balancing of such a bridge circuit and these arise from phenomena occumng within the cell itself. The more important of these phenomena are [15]:

A double-layer of ions can arise due to the attraction of a thin layer of ions to the electrode surface, which in turn causes a more diffuse layer of ions of opposite charge to collect in the adjacent solution. That is, local ordering of ions at the electrode surfaces occurs. This double-layer acts as a capacitor (at each electrode) capable of storing charge and its structure is greatly influenced by interactions of the solvent with the solute ions, and by the nature of the electrode surface. Faradaic (or electrolytic) processes may occur at the electrodes. These processes act to partially short-circuit the double-layer and behave as impedances. Concentration polarization may occur if Faradaic removal of ions at the electrodes occurs at a faster rate than they can be supplied by diffusion from the bulk electrolyte.


out

257

In

Fig. 9.6 Simple two-electrode, flow-through conductance cell using annular electrodes. El and E2 are the measuring electrodes, which are separated by FTFE isolators (designated by Ins in the Figure). The modulated voltage is applied to electrode El. The internal cell volume is approximately 3 p1. Reprinted from [17] with permission. (iii) The sample liquid in the conductance cell itself imposes a resistance to the motion of ions because they must overcome frictional forces. (iv) Whilst imposition of an alternating field can largely eliminate the Faradaic effects discussed in (ii) above, this can itself introduce further complications. The electrical components themselves show frequency dependence which must be considered. The above factors can be incorporated into an equivalent circuit for the conductance cell, such as that shown in Fig. 9.5(b). This circuit includes the resistance of the solution (Rx),the double layer capacitance at each electrode ( c d ) , the Faradaic impedance at each electrode (ZF), the resistance and capacitance of the contacts and leads in the circuit (& and Cc), and the inter-electrode capacitance (Cj). It is convenient to simplify the equivalent circuit by combining the series capacitances and the parallel capacitances, as shown in Fig. 9.5(c). This type of conductance cell has found very widespread use and is employed in many conductivity detectors used for IC. A simple, low-volume, two-electrode, flowthrough cell design for this type of detector can be made using annular stainless steel electrodes. Fig. 9.6 shows a typical cell of this type. Conductivity cells with internal volumes of less than 0.5 pl have been reported for use with microcolumn IC [lS, 191 and a combined conductivity-amperometry cell of 15 nl internal volume has also been described [20].

258

Chopter 9

BPD

I I'

II

Cell A H S u D p r e s s o H Cell

B

b

Waste

Fig. 9.7 Experimental arrangement for a comparison of the sensitivities of bipolar-pulse (BPD) and conventional AC conductance bridge (ACCB) conductivity detectors. Cell A was taken from a Wescan 219-900 detector, cell B is from a Dionex model 10 [23].

9.4.3

Bipolar-pulse circuitry

The drawbacks of the conventional conductance bridge include slow response to rapidly changing conductances, inapplicability to solutions of high conductance, and the existence of the series and parallel capacitances. One approach to overcoming these drawbacks is the bipolar-pulse technique, in which two consecutive voltage pulses of equal amplitude and pulse width, but of opposite polarity, are applied to the cell. The current is measured precisely at the end of the second pulse [21, 221. The advantage of this approach is that parallel and series capacitances of the equivalent circuit of the conductance cell no longer influence the cell current. Thus, measurement of this current cnables calculation of the electrolytic conductance, free of Faradaic and other distortions. The first application of bipolar-pulse conductivity measurement in IC was reported by Keller [23], who used a home-made bipolar-pulse detector prior to the suppressor in a suppressed IC system and a commercial AC conductance bridge-type of detector after the suppressor. The arrangement of chromatographic components in this experiment is shown in Fig. 9.7. The sensitivity of the bipolar-pulse detector in the non-suppressed mode exceeded that of the conventional detector monitoring the suppressed eluent. Bipolar-pulse conductivity detectors for IC are available commercially from a number of manufacturers. 9.4.4

Four-electrode conductance measurement

The simplest practical arrangement for measurement of the resistance of the sample solution (and hence its conductance) is through the use of two electrodes, as discussed above. However, i t has also been noted that this measurement is complicated by the presence of contact resistances and other extraneous phenomena. An alternative experimental arrangement is the four-contact mode, where leads and contacts supplying the current are separated from those probing the voltage drop across the sample solution. This arrangement is illustrated in Fig. 9.8. Since only an infinitesimal current (12) now flows through the probing circuit, the potential drop corresponding to the contact resistances R,I and Rc2 can, for all practical purposes, be neglected. This gives a more accurate measurement of the unknown (solution) resistance, Xu. Pig. 9.9 shows a simplified experimental set-up for fourcontact measurement of resistance and conductance in static solutions (24).


259

I

Voltmeter

Source of current

Fig. 9.8 Circuitry for four-contact measurement of resistance (and hence conductance). Ru is the unknown resistance, &l and &2 are contact resistances. Note that I p - 1 2 .

The same approach can be applied to flowing solutions using a series of annular electrodes. Fig. 9.10 shows a four-electrode, flow-through conductivity cell of low internal volume (4 pl) [25]. A variable AC generator (ACG) controlled by a differential amplifier (DA1) supplies a sinusoidal current to the outer two electrodes of the cell (El, E4). The two inner electrodes (E2, E3) are connected to the input terminals of DA1, which forces the current generator to maintain a constant potential drop between E2 and E3. The current flow through E2 and E3 is negligible, so Faradaic impedances, contact resistances, etc., are largely eliminated. As DA1 forces the ACG to vary the current flow in response to changes in conductance between E2 and E3, the potential across the range resistor (RR) will also vary. These changes are converted by the differential amplifier (DA2) to an appropriate analog signal, which is measured by the voltmeter V1.

Fig. 9.9 Apparatus for four-contact measurement of resistance and conductance. Reprinted from [24] with permission.

260

Chapter 9 ACG

out

CH

CH

Fig. 9.20 Four-electrode conductimetric cell. See text for an explanation of the symbols.

Reprinted from [25] with permission. In this manner, the conductivity of the liquid flowing through the cell is measured. The cell holder (CH)is connected to ground and also through a resistor (R)to the output of DA2. The cell holder thus serves as a guard electrode against capacitance effects to ground [26].

9.4.5

Differential conductivity detection

A dual-cell configuration, in which the column effluent passes through a sample cell and eluent is passed through a reference cell (Fig. 9.11(a)), has been suggested for differential conductivity detection [27, 281. Subtraction of the signal arising in the reference cell from that in the sample cell should permit cancellation of the background conductance of the eluent itself and should also provide a means to compensate for temperature fluctuations. When applied to eluents with conductance of 30 pS or less, this approach permits full-scale recording of a conductance difference between the eluent and column effluent of 0.1 pS, with low baseline drift. This, in turn, permits the detection of common anions at ppb levels. Differential conductivity detection has also

Column effluent

te

Refercnc eluent fl

Column eMuent

Waste Reference cell (a)

Reference cell (b)

Fig. 9.22 (a) Two-cell configuration for differential conductivity detection. (b) Cell configuration for first derivative conductivity detection. Reprinted from [30]with pennission.


261

I

--

0

2

L

lime (min) (aI

6

0

2

4

lime (mint

6

(bl

Fig. 9.12 Use of first derivative conductivity detection for identification of co-eluted peaks. Chromatogram (a) shows the direct conductivity signal for two co-eluted solutes, whilst chromatogram (b) shows the first derivative signal for the same injection. Reprinted from [30]with

permission. been applied to the simultaneous detection of anions and cations when eluents of relatively high conductance are used [29]. If a dual-cell conductivity detector is modified so that the column effluent passes through the sample cell, a delay loop, and finally returns to the reference cell, a first derivative conductivity signal results [30, 311. Fig. 9.11(b) shows the flow-path used. The first derivative signal can be used to identify coelution of peaks (Fig. 9.12) or to improve the apparent separation of poorly resolved peaks. 9.5

SUPPRESSORS IN IC

9.5.1 Function of the suppressor Table 9.2 shows that direct conductivity detection of anions in HC03-/C032-eluents is quite insensitive because of the limiting equivalent ionic conductances of these ions. However, this situation can be improved greatly by exchanging hydrogen ions for the cations in the eluent, prior to the measurement of conductance. The HCO3- and C032ions are thereby converted into weakly conducting HzCO3, and the background

262

Chapter 9

conductance of the eluent is said to be suppressed. The most simple means of accomplishing eluent suppression is to pass the eluent through a cation-exchange column in the hydrogen form. As an example of the reactions which take place in such a suppressor column, consider the case of CI-ions as solute and an eluent composed of NaHC03. The eluent reaction in the suppressor is given by eqn. (9.22), whilst the reaction of the solute is given by eqn. (9.23). Resin-H++ Na'HCO; Resin-H+

% Resin-Na+

+ Na+ + C1'

+ 3C0,

% Resin-Na+

+ H+ + C1-

(9.22) (9.23)

The combined result of these processes is that the eluent conductance is decreased greatly, whilst the conductance of the sample is increased by virtue of the replacement of sodium ions (A+= 50 S.cm*.equiv-') with hydrogen ions (k+= 350 S.cm2.equiv-'). The detectability of the solute is therefore enhanced. A similar procedure can be applied to cation-exchange, where the suppressor is now an anion-exchange column in the OH- form and operates by the addition of OH- ions to the eluent. As an example, consider the suppressor reactions of Na+ ions eluted with a HCI eluent, as shown in eqns. (9.24) and (9.25).

+ Resin-C1' + H 2 0

Resin-OH-

+ H+ + CI-

Resin-OH-

+ Na+ + C1- %

Resin-Cl-

+ Na' + OH

(9.24) (9.25)

The eluent is converted into water, whilst the conductance of the sample band is increased due to replacement of CI- ion (h. = 76 S.cm2.equiv-') by OH- ions (h. = 198 S.cm2.equiv-1). It is important to note that suppression reactions are not limited to acid-base reactions, such as those shown in the above examples. Indeed, any post-column reaction which results in a reduction of the background conductance of the eluent can be classified as a suppression reaction. We have seen earlier in Table 4.6 some other suitable suppressor reactions. However, the ensuing discussion of suppressor design and performance will be restricted to those which employ acid-base reactions, since these are the most widely used.

9.5.2

Packed-column suppressors

The original suppressor device [32] was an ion-exchange column in the H+ or OHform, which operated according to the mechanism discussed above. To enable the suppressor to be used for as long a time as possible, high capacity ion-exchange materials were used. The suppressor was regenerated periodically by passing an appropriate solution (such as 0.25 N H2SO4 for the H+ form suppressor) through the column to displace the accumulated eluent cations. Packed-column suppressors suffer from a

263

ConductivityDetection HNOl

I

'a

so;

+

v

HNO,

Fig. 9.13 Ion-exclusion processes occumng in the resin bead of a hydrogen form packed-column suppressor. It can be noted that H N e , produced in the suppressor by protonation of the solute N02-, is retained on the suppressor resin by ion-exclusion. This process will occur for any solute which is the conjugate base of a weak acid. Ionized solutes (e.g. NO33 are not retained.

number of disadvantages which include: (i)

The limited total ion-exchange capacity of the packing material in the suppressor means that periodical regeneration is necessary. (ii) Significant broadening of the solute band occurs in the suppressor, resulting in loss of chromatographic efficiency. (iii) Solute ions which are protonated easily may show variable retention in the suppressor column due to ion-exclusion effects. This process is illustrated in Fig. 9.13 for nitrite. The nitrite will not become protonated until it reaches an active zone of the suppressor (i.e. where unexchanged H+ ions exist), so penetration of HNO2 into the resin will not commence until this time. Clearly, the length of active suppressor (which is a variable quantity) will determine the elution time for this species. (iv) Some ions undergo chemical reaction in the suppressor. For example, nitrite has been shown to undergo oxidation reactions in the suppressor [33]. The "water dip" resulting from elution of the sample solvent (usually water) (v) often hampers trace analysis of those solute ions which are eluted early in the chromatogram.

Chapter9

264 Silicone rubber

Centre tube

Eluent + inlet

+ Eluent

outlet

+

fibres

Regenerant outlet

4

Regenerant inlet

Fig. 9.14 Schematic drawing of a hollow-fibre suppressor. Reprinted from [34] with permission.

Despite these disadvantages, packed column suppressors provided the foundation on which suppressed IC was built. These suppressors were in use from 1975-1981,at which time the first of the membrane-based suppressors was introduced.

9.5.3

Hollow-fibre membrane suppressors

Hollow fibres constructed from polymeric ion-exchange material provided an alternative means for eluent suppression. These fibres may be prepared by introducing a liquid monomer into the walls of a porous fibre, after which the monomer is polymerized to give a cross-linked ion-exchange polymer. The eluent is passed through the interior of the fibre, whilst a suitable regenerant (or scavenger) solution passes over the exterior of the fibre, usually in a countercurrent direction.

Operating principle The first hollow-fibre suppressor was reported by Stevens et al. [34] and consisted of a collection of sulfonated cation-exchange fibres, with which sulfuric acid was used as the regenerant. The physical design of this suppressor is shown in Fig. 9.14 and its mode of operation with a HC03-/C032- eluent is illustrated schematically in Fig. 9.15(a). The operation of a cation suppressor, with which HCI is used as eluent and barium hydroxide as regenerant, is shown in Fig. 9.15(b). In the suppression of anion-exchange eluents (Fig 9.15(a)), there is a transfer of sodium ions out of the eluent stream, with concomitant transfer of hydrogen ions into the eluent stream. Cation-exchange eluents (Fig. 9.15(b)) are suppressed by transfer of CI- out of the eluent and OH- into the eluent. The overall results of these processes are identical to those achieved by the column suppressor, but the hollow-fibre design has the chief advantages of greatly reduced bandbroadening and continuous regeneration [35,36]. It has also been noted that suppression efficiency is increased at elevated temperatures because of improved diffusion of ions both in solution and through the membrane [35,371. The operation of the hollow-fibre suppressor with a typical eluent (e.g. tetrabutylammonium hydroxide, TBA+OH-)used for ion-interaction separation of anions is illustrated in Fig. 9.15(c), in which sulfuric acid is employed as the regenerant. It can be seen that the suppression process converts the eluent species to water. Similarly, suppression of a typical eluent (HCI) for ion-exclusion chromatography of carboxylic acids is illusaated in Fig. 9.15(d), in which TBA+OH-is used as the regenerant. In this case, the eluent species are converted in the suppressor to the weakly conducting TBA+CI-salt.

ELUENT

ELUENT

Ha

TBA+OH-

TBA)zSO,

'4

I ~

+

TBA+OIT ~

Anion-exchange hollow-fibre

TBA+CI- TBA+OA' Catton-exchange

llpa hollow-fibre

Fig. 9.15 Schematic operation of a hollow-fibre suppressor for eluents used with (a) anion-exchange (b) cation-exchange, (c) ion-interaction and (d) ion-exclusion separation modes.

h,

E

Chapter 9

266

Fig. 9.16 Zig-zag packing of beads inside a packed-fibre membrane. Reprinted from [40] with

permission.

Regen era n ts The regenerant used with hollow-fibre suppressors must be selected with care. The regenerant must supply the ion required for effective eluent suppression (e.g. H+ or OH-), but must not contaminate the eluent with any other ion. The chief potential contaminant is the regenerant ion having the same charge sign as that of the solute. This ion is theoretically prevented from entering the eluent stream as a result of Donnan exclusion by the ion-exchange functionality on the hollow-fibre. However, this repulsive effect may not totally prevent penetration of the forbidden ion, especially when the regenerant concentration is high. It has been demonstrated that large ions have a smaller penetration rate through the membrane [35,361. Thus, dodecylbenzenesulfonic acid is a useful regenerant for an anion-exchange suppressor [38] because of the low penetration rate by dodecylbenzenesulfonate ions through the anionic fibre. Packed-fibre suppressors The hollow-fibre must have a very narrow bore to permit a sufficient rate of transfer of ions. A bore diameter of less than 400 pm is desirable, but tubing of this type is not readily available. For this reason, some type of inert packing is usually employed inside the tube. This may be a nylon filament (such as fishing line) [3Y] or polystyrene beads [40,41]. in the first case, insertion of a nylon filament permits coiling the hollow-fibre into a helix so as to increase suppression efficiency. In the second case, the beads used have a slightly larger diameter than the bore radius of the hollow-fibre,

n

Packed-f ib re

n

Regenerant outlet

Eluent outlet

Eluent inlet

Regenerant inlet Rod (28 x 100 rnm)

Cylinder (34 ID x 100 rnrn)

Fig. 9.17 Design of a packed-fibresuppressor. Reprinted from [41] with permission.


267

p so&

I

0

1

1

2

1

1

1

1

L 6 Time (min)

1

1

8

1

1

1

10

(01

1

0

1

1

2

1

1

1

1

i 6 lime (min)

1

1

8

1

1

1

10

fb)

Fig 9.18 Chromatograms obtained with (a) a hollow-fibre and (b) a packed-fibre suppressor. Note the improved resolution and sharper peaks for early eluted solutes. Reprinted from [41] with

permission.

so that the beads become arranged in a zig-zag pattern as shown in Fig. 9.16. This configuration reduces the dead volume inside the suppressor and provides turbulent flow, which improves the transfer of ions across the membrane. The packed-fibre is then wound around a central support, as shown in Fig. 9.17. The improvement in chromatographic efficiency resulting from the use of an inert packing inside the hollowfibre is evident from Fig. 9.18. The packed-bead fibre suppressor has been the subject of many innovative suggestions designed to increase its suppression efficiency. These have included replacing the inert beads with ion-exchange resin beads, packing beads around the exterior of the fibre to provide mechanical support, altering the shape of the packing beads and application of an ultrasonic field to the system. The effects of such approaches have been discussed in some detail by Dasgupta [42], who in the same article also provides an excellent treatment of practical aspects of the preparation of fibre suppressors, such as packing procedures and methods for joining fibres to conventional PTFE tubing. 9.5.4

Micromembrane suppressor

Hollow-fibre suppressors had provided a solution to the problem of frequent regeneration which existed for packed suppressors, but introduced a new limitation relating to the ion-exchange capacity of the fibre. The small internal diameters of the fibres employed in these suppressors meant that the surface area of the fibre available

268

Chqpter 9

Eluent Regenerant

t I

out

in

Regenerant Eluent

+I

out

I

in

\ \

I

1

Regeneran t

Regenera n t out

in

Ion-exchange membrane Ion-exchange screen Gasket material

n

Fig. 9.19 Design of a micromembrane suppressor. Adapted from [43].

for exchange between eluent and regenerant ions was low, and this, in turn, led to low ionexchange capacity. The eluent concentrations suitable for use with such a suppressor were therefore restricted. In addition, the hollow-fibres were intolerant towards some organic solvents and ion-interaction reagents. These factors provided stimulus for the development of an improved suppressor which was capable of continuous regeneration, possessed high ion-exchange capacity, and showed minimal band-broadening effects.

Design of the micromembrane suppressor The micromembrane suppressor is able to meet the above requirements by replacing the ion-exchange tubing used in the hollow-fibre suppressor design with flat sheets of membrane [43].The surface area available for exchange between eluent and regenerant ions is thereby increased greatly in comparison to the hollow-fibre suppressor, and so is the ion-exchange capacity. The design of a micromembrane suppressor is shown in Fig. 9.19. The eluent passes through a central chamber which has ion-exchange membrane sheets as the upper and lower surfaces. Regenerant flows in a countercurrent direction over the outer surfaces of both of these membranes. Mesh screens constructed from a polymeric ion-exchange material are inserted into the eluent cavity and also into the cavities which house the flowing regenerant solution. The entire device is constructed in


269

a sandwich layer configuration with gaskets being used to define the desired flow-paths. The volume of the eluent chamber is very small (40pl [43]), so band-broadening is minimal.

Operating principles The micromembrane suppressor operates on the same principles as the hollow-fibre suppressor, as illustrated in Fig. 9.15. That is, regenerant ions (H+ or OH-) are transferred into the eluent across the membrane, whilst eluent counter-ions pass simultaneously into the regenerant. However, unlike the hollow-fibre suppressor, which relies on diffusion to transport the appropriate ions to the membrane surface, the micromembrane suppressor utilizes high-capacity ion-exchange screens to perform this task. These screens promote ion transport in two ways. First, the three-dimensional over-and-under square weave pattern of the screens causes a disruption to laminar eluent flow and directs eluent flow to the membrane surface. Second, the ion-exchange sites on the screen enable site-to-site transport of the desired ions to the membrane. The latter mechanism of ion transport plays an increasingly important role as the eluent passes from the suppressor inlet towards the outlet. During this passage, the suppressor reaction advances towards completion and there are relatively fewer eluent ions remaining to react. The transport of these residual eluent ions to the membrane surface is greatly facilitated by the ion-exchange screens. It is useful at this stage to describe suppressor performance in terms of dynamic capacity. That is, the number of microequivalents of eluent which can be suppressed per unit time. Clearly, there must be a limit to the eluent concentration which can be suppressed by a membrane of a specified surface area. It has been shown that the dynamic capacity of the micromembrane suppressor increases with increasing ionexchange capacity of the screen material [43].Dynamic capacity also increases with the concentration of the regenerant solution, but practical limits exist above which there is some penetration of the forbidden regenerant ion through the membrane into the eluent. It is often convenient to use a regenerant concentration considerably less than that at which penetration of the forbidden ion occurs, but to increase the flow of the regenerant in order to increase dynamic capacity. When 12.5 mM sulfuric acid is used as regenerant at a flow-rate of 10 ml/min, the dynamic capacity of the micromembrane suppressor exceeds 100 pequiv/min [a]. This means that an eluent of 100 mM (i.e. 100 pequiv/ml) NaOH, flowing at 1 ml/min, can be suppressed effectively. We can note that in this example, the flux of H+ions in the regenerant is 250 pequiv/min, which exceeds the flux of OH- ions in the eluent by a factor of 2.5. This factor is close to the minimum required for effective operation of the micromembrane suppressor. Advantages of the micromembrane suppressor The micromembrane suppressor combines the advantages of other suppression devices and at the same time, eliminates their drawbacks. These advantages can be summarized as: (i)

Small internal volume, leading to minimal band-broadening effects and hence low detection limits.

Chapter 9

270

(ii) Continuous regeneration. (iii) High dynamic suppression capacity which can be varied readily by changing the nature, concentration and flow-rate of the regenerant. (iv) Suitable for gradient elution with appropriate eluents. (v) Resistant to many organic solvents and ion-interaction reagents. (vi) A wider choice of eluent types is possible because of the high dynamic suppression capacities which can be achieved.

9.5.5

Post-suppressors

Post-suppressors are devices inserted between the suppressor and the detector for the purpose of further lowering of the background conductance of the eluent in suppressed anion-exchange IC systems. When a carbonate buffer is used as eluent, the fully suppressed eluent contains HzCO3, which is dissociated partially in aqueous solution (to form H+ and HCO3-) and so contributes to the background conductance. Moreover, the HCO3- present in the suppressed eluent also causes a reduction in the conductance of a sample peak by reaction with the H+ ions which accompany the elution of the anion of a strong acid. This effect is discussed in more detail in Section 9.6.1. A further disadvantage of the formation of H2CO3 in the suppressed eluent is that this species diffuses through the suppressor membrane at a rate which is dependent on pressure, and this may lead to the formation of pump pulsations in the detector baseline [42]. It is therefore desirable if H2CO3 can be removed from the suppressed eluent.

Design of post-suppressors A convenient way to remove H2CO3 is to pass the suppressed eluent through a length of tubing which is permeable to carbon dioxide [45]. Quantitative removal of dissolved C02 from the suppressed eluent will result in the background conductance of the eluent approaching that of pure water. Post-suppressors constructed of porous PTFE tubing have been shown to remove 90% of dissolved COz. This removal was attained only with the aid of some mechanical means of improving mass transport to the membrane wall, such as inserting a knotted nylon fishing line or a twisted stainless steel wire into the PTFE tubing. An air current around the outside of the post-suppressor tubing is used to cany the C02 away after diffusion through the tubing. Leaking of the eluent through the tubing does not occur at moderate pressure because of the hydrophobic nature of the tubing material. The design of this post-suppressor is illustrated in Fig. 9.20. Improved post-suppression can be achieved through the use of silicone rubber tubing (which is highly permeable to C02. but is non-porous) and by passing a heated solution of KOH over the outside of the tubing in order to remove the C02 as it diffuses from the eluent 1461. Shintani and Dasgupta 1471 have developed a specialized postsuppressor membrane tubing consisting of porous polypropylene coated with silicone rubber, with a nylon monofilament inserted into the interior of the tubing. Transport of C 0 2 to the silicone rubber layer is very rapid because of the porous nature of the underlying polypropylene membrane. This post-suppressor tubing was shown to be superior to the use of silicone rubber tubing or uncoated porous polypropylene tubing.

Conductiviry Detection

From

+

suppressor

27 1

To detector

4

L--=+;r Ascarite Fig. 9.20 Schematic design of a post-suppressor. Reprinted from [45] with permission.

Advantages of post-suppression The advantages which may be attained through the use of post-suppressors include: Decreased baseline noise resulting from pump pulsations. Decreased baseline conductance. Virtual elimination of the water and carbonate dips from the final chromatogram. Enhanced detectability of eluted anions since there is no loss of conductance signal due to reaction of H+ ions in the analyte band with H2CO3 in the suppressed eluent. Gradient elution using carbonate buffers as eluents is possible with postsuppressors of suitable efficiency. 9.5.6

Other post-column signal enhancing devices

Electrochemical suppressor A type of micromembrane suppressor in which transfer of ions across the membrane is enhanced with an electric field has been suggested as a more efficient design in comparison to those suppressors in which ion transport is accomplished by diffusion alone [48]. This suppressor is illustrated in Fig. 9.21(a). The eluent flow compartment consists of a chamber packed with cation-exchange resin, with cationexchange membranes forming the chamber walls. Two chambers containing 0.1 M sulfuric acid are located on the outer surfaces of these membranes and platinum-plated titanium electrodes are housed in each of these chambers. Application of a suitable voltage (about 4 V) to give a cell current of 50 mA creates an electric field, under the influence of which H+ from the regenerant moves rapidly into the eluent compartment

Chqter 9

272 ELUENT: NazCOf, NaHC(?I Anodc

SAMPLE: Na* +

cT

:atbode

Cation-excbange bollow-fibre Cation-exchange

0 resin

t

Steel tubing (4 Cation-exebange membrane Cation-excbange membrane Pt wire (+)

Regenerant flow-paths Eluent flow-path

(b) Fig. 9.21 Schematic designs of electrochemical suppressors. Adapted from [48,49].

and Na+ from the eluent migrates towards the cathode. Using this process, the complete suppression of 5 mM Na2C03 eluent at a flow-rate of 2 ml/min can be accomplished. It should be noted that this suppression efficiency is achieved with a static regenerant solution, resulting in simplified operation and reduced consumption of regenerant. A flow-through type of electrochemical membrane suppressor has also been reported [49]. In one of several possible configurations, this device uses a concentric arrangement of the following components, in sequence: a platinum wire, a cationexchange membrane fibre (400 pn ID, 50 pm wall thickness), a second cation-exchange membrane fibre (625 p ID. 125 pm wall thickness), and a length of stainless steel tubing (Fig. 9.21(b)). The eluent flows through the annular cavity between the two membranes, whilst regenerant flows in a countercurrent direction through the two remaining annular cavities (i.e. between the Pt wire and the inner membrane, and between the outer membrane and the steel tubing). A dc potential of 3-8 V is applied between the PI wire and the steel tubing, with the Pt wire being positively polarized. The above study provides an interesting comparison between chemical suppression and electrochemical suppression. When an acid (e.g. dodecylbenzenesulfonic acid) is


273

used as the regenerant with a NaOH eluent, suppression efficiency is roughly the same (about 95%), regardless of whether a potential is applied across the electrodes. The reason for this is that in the absence of potential, both membranes contribute towards the removal of Na+ from the eluent. However, when a potential is applied, Na+ is removed from the eluent onfy through the outer (or cathodic) membrane, albeit at a faster rate than without the potential. Nevertheless, a significant finding of this work is that electrochemical suppression can function effectively even when water is used as the regenerant. The same operating principles apply, but there is now no longer any requirement for specialized regenerant solutions.

Signal enhancement devices for ion-exclusion chromatography Organic species (such as carboxylic acids) eluted from an ion-exclusion column by an acidic eluent are difficult to detect using conductivity measurements. The reason for this is that these solutes are retained on the column only in their neutral or weakly ionized forms, which show little conductance. It therefore becomes necessary to modify the eluted solutes if conductivity detection is to be employed. Cation-exchange membrane suppressors have been applied successfully when sulfuric or octanesulfonic acids are used as eluents [50-531. A suitable alkaline regenerant (such as tetrabutylammmonium hydroxide or dilute NaOH) leads to a reduction in the conductance of the acidic eluent and at the same time, ionization of the solute acids occurs. This mechanism is depicted in Fig. 9.22(a). An interesting effect occurs if the concentration of the NaOH regenerant is increased (e.g. from 10 mM to 500 mM). Under these conditions, the peak direction changes from positive (increasing conductance) for the 10 mM regenerant to negative (decreasing conductance) for the 500 mM regenerant [52]. The reason for this is that the more concentrated regenerant overcomes the Donnan exclusion effect of the cation-exchange membrane, so OH- ions move into the eluent, giving a high background conductance. This process is shown in Fig. 9.22(b). The eluted carboxylic acids react quantitatively with OH-, causing a decrease in conductance and hence indirect conductivity detection. This detection mode is more sensitive than direct conductivity because of the high limiting equivalent ionic conductance of OH- (see eqn. (9.20)). Finally, it can be noted that signal enhancement in ion-exclusion chromatography of organic acids can be achieved using a regenerant comprising a solution of a neutral salt (e.g. KzSO.4). The exchange of H+ ions from the eluent and the solute acids with K+ from the regenerant, as shown in Fig. 9.22(c), leads to a reduction in the eluent conductance and an enhancement of the solute conductance 1531. Post-column enhancement of the conductivity signal has also been applied to the detection of HzCO3 after ion-exclusion separation [54]. The effluent from the ionexclusion column is first passed through a cation-exchange column in the K+ form, and then through an anion-exchange column in the O H form. The first of these columns converts HzCO3 into KHC03, whilst the second converts KHCO3 into KOH, which is the species ultimately detected.

From column

From column

H2SQ

10 mM NaOH

Na2S04 RCOO'Na' To detector

From column

H2 SQ

H2S04

10 mM NaOH

500 mM

NaOH

Na2S04 500 mM RCOO"a+ NaOH NaOH To detector

K2SO4

K ~ S O ~ K2SO4 RCOOK+ To detector

(b)

Fig.9.22 Signal-enhancing devices for conductivity detection of carboxylic acids after ion-exclusion separation. A cation-exchange fibre is used in each case, with (a) 10 m M NaOH, (b) 500 mM NaOH or (c) K2SO4 as the regenerant solution.

B

Y)

275


9.5.7

Response equation for suppressed conductivity detection

Development of a response equation for conductivity detection in suppressed IC systems can be approached by considering the factors which contribute to the detector signal during elution of a sample band [55]. Because of the diversity of suppression reactions, we will confine the present discussion to the case of a suppressed anionexchange IC system in which the eluent consists of an ionic salt, NaA, present at concentration CE, whilst the regenerant consists of a solution of a suitable acid. A fully ionized sample anion, S-, is eluted from the column at a concentration of Cs. This solute is accompanied by an equivalent concentration of H+ as a result of the suppression reaction. For suppression to be effective, the eluent anion, A-, must be the conjugate of a weak acid, HA. Passage of the eluent through the suppressor results in the formation of HA, which ionizes according to:

HA % H+ + A-

(9.26)

For which we can write: (9.27)

When the eluent is converted fully to HA in the suppressor, the conductance of the suppressed eluent and the suppressed sample results from H+, A- and S-ions. However, when the suppression reaction is not quantitative (i.e. not all of the Na+ ions from the eluent are removed), we can expect the suppressed eluent to contain some residual Na+ ions which will also contribute to the conductance. Finally, we must consider any OHwhich may be present in the suppressed eluent, so that:

Kw = [H+l[OHl

(9.28)

and we can write the following charge balance and mass balance equations:

[H+l + "a+] = [A-I + [OH-] [HA] + [A-I = CE - Cs

+ [S-I

(9.29) (9.30)

Eqns. (9.27)-(9.30) can be combined to give an expression for [H+], as follows:

Solution of this equation permits the value of [H+]to be calculated for given values of the parameters KHA, Kw, CE, Cs and "a+]. This value can then be used to calculate

276

Chapter 9

the remaining parameters using eqns. (9.32)-(9.34). That is: (9.32)

(9.33)

(9.34)

The conductance measured during sample elution is given by:

In the ideal case, the suppression reaction will be quantitative (i.e. "a+] = 0) and the product of the suppressor reaction (HA) will not dissociate to any appreciable extent (i.e. [A-1 = 0). Under these conditions, the detector signal arises entirely from S-and an equivalent amount of H+. That is: (9.36)

Similar derivations can be used to formulate detector response equations for suppressed conductivity detection in cation-exchange, ion-interaction and ion-exclusion separation modes. 9.5.8

Suppression based on precipitation or chelation reactions

Thus far, we have considered only those suppressors in which an acid-base reaction forms the basis of the suppression mechanism. It is important to note that other types of chemical reactions can also be employed as a means of reducing the conductance of the eluent. This topic has already been discussed in Section 4.4, but it is pertinent to reiterate briefly the major reactions used, as part of the present discussion of suppressors. One simple example is the use of precipitation reactions to physically remove an eluent component. This approach has been used for the suppression of NaI eluents using a suppressor in the Ag+ form (resulting in the formation of a precipitate of AgI) [56] and for the suppression of Ba(N03)2 eluents using a suppressor in the S042- form (resulting in the formation of a precipitate of BaS04) [57]. Other examples of suppressors using precipitation reactions were presented in Table 4.6. A further interesting possibility is the use of complexation reactions to achieve suppression of the eluent conductance. When an eluent containing a complexing agent


277

(such as dipotassium ethylenediamine-NN-diacetate, K2EDDA) passes into a suppressor containing a cation-exchange resin in the Cu2+form, the K+ from the eluent binds to the suppressor, releasing Cu2+ ions which then complex with the eluent anion (EDDA2-), forming the neutral complex, Cu-EDDA. This effectively lowers the eluent conductance [58]. An example of a chromatogram obtained using this approach was presented in Fig. 4.20.

9.6

PERFORMANCE CHARACTERISTICS OF CONDUCTIVITY DETECTORS

9.6.1 Non-linearity of calibration plots in suppressed IC Effect of hydrogen ions in the sample band From eqn. (9.36), we see that the conductance change accompanying the elution of a solute ion from a suppressor is proportional to the solute concentration in the detector cell (Cs).Linear plots of the conductance signal versus Cs can therefore be expected from an ideal system, provided that care is taken not to overload the low-capacity ionexchange columns used. That is, provided the calibration plot is prepared over a concentration range in which the adsorption isotherm for the solute is linear. However, we must consider an additional complication which arises when a suppressor is used. In the case used for the derivation of a response equation in Section 9.5.7 above, we have noted that the sample passes through the detector as a dissociated, strong acid (ix.H+ and S-). The hydrogen ions in the sample band will exert an influence on the dissociation equilibrium for HA (eqn. (9.26)) by forcing this reaction to the left. That is, the conductance of the baseline during sample elution will be lower than the background level existing when eluent alone passes through the suppressor. This effect is summarized graphically in Fig. 9.23. The background conductance of the eluent (in the absence of sample) is given by GIII. GI shows the Gaussian change of conductance due only to the solute anion and its accompanying hydrogen ions. The decrease in background conductance caused by the influence of sample on the eluent equilibrium is given by GII. This latter effect gives a non-Gaussian profile. The overall conductance change during sample elution (AG) is given by: (9.37)

Detailed mathematical expressions for each of the above terms have been derived by Doury-Berthod et al. [59] and these permit the conductance changes depicted in Fig. 9.23 to be calculated. The contribution GI describes the rise and fall of the conductance signal during the lifetime of the peak, whilst GI[ shows the change in eluent background conductance during development of the same peak. As the concentration of the sample (and associated hydrogen ions) reaches a maximum, GII is depressed to its minimum value. This minimum value will depend on the sample concentration, since this determines the [H+] in the sample band. The final peak shape, given by eqn. (9.27), is non-Gaussian. The end result of this effect is that the slope of the peak height calibration curve varies with the concentration of injected sample. As the sample concentration

Chapter 9

27 8

AG

GI

t

G I I (non goussion)

G I , ( n o n goussion)

Volume Fig. 9.23 Three contributions to the shape of the eluted peak in suppressed IC with conductimemc detection. See text for an explanation of the origins of GI, Gl1 and G ~ I Reprinted . from [591 with permission.

Conduetonce ( o r b i t r o r y units)

Fig. 9.24 Non-linear peak height calibration plots in suppressed IC resulting from the influence of hydrogen ions in the sample band on the eluent conductance. The curve has two limiting slopes, one at low sample concentration (hli,) and one at high sample concentration (h'lim). Reprinted from [591 with permission.

279


decreases, the slope approaches a limiting value (hlim), whilst at higher concentrations of solute, the slope approaches a different limiting value (h’lim). The final calibration curve is similar to that depicted in Fig. 9.24. Similar studies on the non-linearity of calibration curves in suppressed IC have been conducted by other workers [55, 60-631. In one of these studies 1601,an equation is presented which permits correction for the influence of sample elution on the background conductance. Using this correction equation, linear calibration plots were obtained using peak areas for solute concentrations in the range 0-40ppm.

Practical consideration of the hydrogen ion effect The degree to which the above effect exerts an influence on the linearity of calibration curves in practical situations depends on a number of factors [631. First, the higher the PKa of the eluent acid, the less pronounced will be the effect of hydrogen ions in the sample band on background conductance. The reason for this is that the equilibrium shown in eqn. (9.26) already lies well to the left for very weak acids. This behaviour is illustrated in Fig. 9.25, which shows that the linearity of calibration plots improves as the PKa of the eluent acid increases. Thus, hydroxide eluents can be expected to give calibration plots which are very close to linear since the eluent acid produced in the suppressor i s H20. Similarly, the strong acid eluents (HCl, HNO3) used for cation separations should give linear calibrations. Second, lower eluent concentrations should minimize the sample acid effect, resulting in a more linear calibration curve. Finally, the effect of hydrogen ions in the sample band will be dependent on the type of suppressor used. Micromembrane

Sample Concn. 1 0 5 M

Fig. 9.25 Peak height calibration plots in suppressed IC for eluent acids of differing strengths. The PKa of the eluent acid is (1) 12, (2) 10,(3) 8 and (4) 6. The peak heights were calculated by assuming that the solute ion was C1-. Reprinted from [55] with permission.

Chuprer 9

280

'5

1

l2

1

12

E,

E

' ? 9

9

f

i

,u

F

P

6

I

r

X

-.L

c

0

k

3

0

0 Sample Concn.

loJ M

(4

6

12

18

24

30

Sample Concn. lo-' M

(b)

Fig. 9.26 The effect of residual "a+] in the suppressed eluent on peak height calibration plots for suppressed IC. The p b of the eluent acid is 10 for (a) and 6.36 for (b). The residual "a+] in each Figure is (1) 0, (2) 50 pM, (3) 100 pM and (4) 150 pM. Reprinted from [55] with

permission. suppressors give a lower background conductance than packed suppressors due to diffusion of carbon dioxide through the membrane, so the effect of hydrogen ions in the sample band is decreased. Similarly, penetration of the HzSO4regenerant in the micromembrane suppressor into the eluent will further decrease the effect of hydrogen ions in the sample band. A study of linearity in suppressed IC has demonstrated that calibration curves for anions and cations show excellent linearity from the detection limit, measured in partsper-billion, to above 0.196, when dilute eluents are used in conjunction with a micromembrane suppressor [63]. This concentration range covers almost five orders of magnitude.

Effect of suppression efficiency on linearity of calibration A further factor which may cause non-linearity of calibration curves in suppressed IC is the efficiency of the suppressor. If we again consider a chromatographic system in which the eluent is an aqueous solution of NaA (where A is an anion such as HCOj) and the injected solute is C1-, then the efficiency of suppression can be judged by the concentration of sodium ions present in the suppressed eluent. A perfectly efficient suppressor will give a residual "a+] of zero (seeeqns. (9.35) and (9.36)). The effect of the residual "a+] on the shape of calibration plots has been studied by Tian et al. (551. Calibration curves calculated for eluent acids of pK, 10 and 6.36 are shown in Figs. 9.26(a) and 9.26(b), respectively. for various values of residual "a+]. From Fig. 9.26(a), it can be seen that the calibration plot is linear provided that the

Conductiviry Detection

281

sample concentration, [S], exceeds the residual “a+]. When [S]< residual “a+], the peak height is severely depressed, due chiefly to replacement of H+in the sample band by the weakly conducting Na+ ion. This reduction in [H+]causes the equilibrium depicted in eqn. (9.26) to move to the right, which partially compensates the decreased sample conductance due to reduced [H+]in the sample band. The magnitude of this latter effect will be greater for eluents acids which are more ionized (i.e. lower p k ) , as illustrated in Fig. 9.26(b). This shows that an eluent acid with pK, of 6.36 (i.e. carbonic acid) is less influenced by residual “a+] than is a weaker eluent acid. Thus, the linearity of calibration plots prepared using carbonate eluents show decreased dependence on suppression efficiency in comparison to those prepared in hydroxide eluents. However, in both cases, the sensitivity, linearity and reproducibility of conductivity detection of low concentration samples are seriously affected by the residual “a+], especially at low sample concentrations. 9.6.2 Temperature effects in conductivity detection Temperature fluctuations have been shown to affect both the sensitivity and reproducibility of conductivity detection in IC [64-671. Improvements in detection limits of up to 20-fold have been achieved by insulating the separation column and other exposed components of the IC [65]. The precision of analytical results was also improved considerably. The four most common equations describing the temperature dependence of electrolytic conductance measurements have been evaluated [67]. These relationships are given below: (9.38)

(9.39)

(9.40) (9.41)

where G25 and GT are the conductances at 25OC and P C , respectively, TT is the viscosity of water at temperature P C , and K1-K4 are constants. Literature values for conductances of KCl were used to assess the validity of each of the above equations. Fig. 9.27 shows the difference between the measured and calculated values of the parameter (G~~/GT at)several temperatures, for each of the equations. The best agreement between calculated and measured conductances was obtained for eqn. (9.38). It can be seen from Fig. 9.27 that the popular rule-of-thumb calculation (as described by eqn. (9.39)) [68], which estimates a 2% conductance change for each degree of temperature change, holds well only in a very narrow temperature range of 10-25OC. Outside this range, this calculation may be in error by as much as 25%.

Chapter 9

282

'"I

O' 0.05

.

\

'

'0

\ 0

\

.-0 4

\

\

-0.15

\

0 CK -0.20

\ \

-0.25

0

1

1

1

1

10

20

30

40

Temperature,

b

I

50

OC

Fig. 9.27 Difference between calculated and literature values of conductance ratios for KCI at various temperatures. The calculated values were determined by eqns. (9.28) [o],(9.29) [a], (9.30) [o] and (9.31) [A]. Reprinted from [67] with permission.

9.6.3 Conductivity detection without the use of standards Non-suppressed IC can be applied quantitatively to samples in which the identities of the sample component ions are unknown [69]. The conductivity detector response is measured using first an eluent competing ion of low limiting equivalent ionic conductance and then an eluent competing ion of high limiting equivalent ionic conductance. Two equations of the same form as eqn. (9.20) result and these can be solved simultaneously for the two unknowns Cx (the solute concentration) and h, (the limiting equivalent ionic conductance of the solute ion). In this way, the identity and concentration of each solute ion can be determined. The equations derived for this method are:

c, =

x, =

(F") +

s,

(9.42)

(9.43)

where C represents molar concentrations, S represents peak areas, and h is limiting


283

Sample concentration (mM1

Fig.9.28 Dependence of the conductance of the injection peak on the concentration of an injected monovalent anion. Calculated and experimental (0)values are compared. The eluent is 2 mM benzoic acid at pH 4.37. Reprinted from 1701 with permission.

equivalent ionic conductance. The subscripts have the following meanings: 1 refers to the first eluent; 2 refers to the second eluent; a denotes the peak area of the anion of eluent 1 when eluted by eluent 2; b denotes the peak area of the anion of eluent 2 when eluted by eluent 1. Provided the values of Sa, Sb, S1, S2, C1, C2, hl and h2 are known (or have been determined from previous experiments), it becomes possible to solve the above equations for Cx and Ax. For similar determinations without the use of standards in suppressed IC, the authors recommend the use of a third column, which in the case of anion separations could be an anion-exchanger in the chloride form. All anions eluting from the suppressor after their separation on the separator column would thus be exchanged for C1- in the third column. The number of equivalents of Cl- indicated by a detector calibrated with a standard solution of CI- would thus correspond to an equal number of equivalents of the analyzed anion. 9.6.4

Utilization of the injection peak in conductivity detection

Chromatograms obtained with conductivity detection show an injection peak corresponding to elution of the sample co-ions, displaced eluent ions, and the sample solvent. This peak is analogous to the void volume peak encountered in other liquid chromatographic methods, and is sometimes called the solvent peak or the water dip. In a non-suppressed IC system, the injection peak may be positive or negative and its conductance is related quantitatively to the total ionic content of the injected sample.

284

Chapter 9

KC I

NaCl M

6.0 x lomL M

r-121737

4=9L25&

6.05 Y

A=5&6&0

A-53861

I

CaClz 3.0 x

M

\=91053

A-5333:

I

Time Pig. 9.29 Injection peaks and sarnpie peaks for three different chloride salts. The area of each peak is indicated in the Figure. The eluent was 1.5 mM phthalate at pH 4.3. Reprinted from [71] with permission.

Fig. 9.28 illustrates this relationship, which may be used to assist in the quantification of a sample mixture. For example, a solute mixture comprising a well-retained ion (such as S042-) and several ions of short retention time (such as Cl-, NOy, Br) can be quantitated by measuring the peak areas of the injection peak, together with those of all solutes except the well-retained ion. Since the injection peak contains the sample cations and the eluent ions displaced by initial adsorption of all solute ions injected onto the column, the area of the well-retained ion can be deduced 1701. If samples contain eluent ions at the same concentration and pH as those present in the eluent itself, the injection peak in a non-suppressed anion-exchange IC system can be used to obtain quantitative data on the cation concentration of the injected salts [71]. Under the above experimental conditions, the area of the injection peak is linearly proportional to the cation concentration in the sample. Moreover, the ratio of the areas of the solute anion peak and the injection peak is specific for a given combination of solute anion and cation. This is illustrated in Fig. 9.29, which shows chromatograms for solutions of three chloride salts, each of which has the same [Cl-I. The C1- peaks are therefore of almost equal area, but the areas of the injection peaks differ as the cation is varied. This characteristic ratio can therefore be used to identify the components of an injected sample, because the solute anion is readily identifiable by its retention time, and the area ratio determines the identity of the solute cation. Binary salt mixtures, or mixtures of salts with acids and bases, have been analyzed using this approach [71].


9.7

285

APPLICATIONS OF CONDUCTIVITY DETECTION IN IC

Conductivity detection has always been the mainstay of IC and a brief perusal of the numerous applications listed in Part V of this book will reveal the frequency with which this detection mode is employed. Moreover, most of the chromatograms included thus far in this text have provided ample illustration of the utility of conductivity detection. It is therefore difficult to offer a comprehensive summary of the areas in which conductivity detection is employed. Instead, the current status of conductivity detection will be described by reference to its application to a few selected classes of ions.

9.7.1

Anions

The analysis of anions of strong inorganic acids has been, and continues to be, the most common application of IC. Detection limits for direct injections of these species vary from anion to anion, and with the nature of the conductimetric method used. Generally, it can be said that detection limits between 50 and 100 ppb can be achieved in both suppressed and non-suppressed conductivity methods without the need for sample preconcentration. With suitable preconcentration methods (see Section 14.6). detection limits in the lower half of the parts-per-trillion range result. The real limits in this case are imposed more by sample handling considerations than by the sensitivity of the conductivity detection used. One of the strengths of conductivity detection, arising from its character as a bulk property technique, is its utility for speciation after separation. That is, the identification of different forms of the same element. Fig. 9.30 shows the speciation of anions of sulfur, chlorine and phosphorus using conductivity detection.

9.7.2 Cations Conductivity detection of alkali metal cations and ammonium is extremely sensitive with both suppressed and non-suppressed conductivity methods. Detection limits for direct injection are often a factor of 5-10 times lower than those for anions, so preconcentration is rarely necessary for these solutes. Low molecular weight bases can also be detected readily. Divalent cations are amenable to conductivity detection, although not with the sensitivity attainable with post-column colour reactions (see Chapter 13). However, conductivity detection represents a simple and often preferable approach, as is illustrated in Fig. 9.31.

9.7.3 Weakly ionized species Detection of partially ionized inorganic and organic species has always presented a challenge for conductivity detection. In the suppressed mode, the ionization of weak acids is suppressed, whilst in the non-suppressed mode, the sensitivity of detection is impaired for any ion which is not fully dissociated. The use of high pH eluents (such as hydroxide), coupled with indirect conductivity detection, gives optimal results for these species in non-suppressed IC. On the other hand, the sensitivity of conductivity detection of weak acids can be improved with signal enhancing devices such as those shown earlier in Fig. 9.22. This method is illustrated in Fig. 9.32 for borate as the solute.

CIOi

203-

SCN-

CI'

I

I

0

10

I

I

20 30 Time (min) (a1

I

1

40

50

Y

0

I

f

1

I

5

10

15

0

mL

(b)

I

I

5 10 Time (min)

1

15

(C)

Fig. 9.30 Speciation of (a) sulfur, (b) chlorine and (c) phosphorus using conductivity detection. (a) Lichrosphere c18 column with an eluent containing 1 mM tetrabutylammonium hydroxide, 7.5 mM H3B03 and 50% acetonitrile. Solute concentrations 1 ppm. Detection by suppressed conductivity. Reprinted from [72] with permission. (b) Spherisorb A5Y column with 0.5 mM H3P04 as eluent. Detection by non-suppressed conductivity. Reproduced from [73] with permission. (c) Waters IC Pak Anion SW column with 2 mM tartaric acid as eluent. Solute concentrations 20 ppm. Reprinted from [3] with permission.

B i7 E:

v,

287


15 (min.)

I

Fig. 9.31 Conductimetric detection of divalent metals. Waters IC Pak C column with 3.5 mM ethylenediamine and 10 mM cimc acid as eluent. Solute concentrations: 5 ppm. Reprinted fiom [31 with permission.

Carbonate

I 0 Borate J

0

2

4

8 1 0 1 2 1 4 Minutes

6

Fig. 9.32 Detection of borate using a membrane suppressor. Dionex HPIC-AS1 column with 1 mM octanesulfonic acid as eluent. The regenerant used in the micromembrane suppressor was 10 mM m0H. The borate concentration was 10 ppm. Reprinted from [74] with permission.

Chapter 9

288

9.8 1 2 3 4 5 6 7 8 9 10 11 12 13 14

15 16 17 18 19 20 21 22 23 24

25 26 21 28 29 30 31 32 33 34 35 36 37 38

REFERENCES James A.T., Martin A.J.P. and Randall S.S., Biochem. J.. 49 (1951) 293. Shpigun O.A. and Zolotov Yu.A.. Ion Chromatographyin Water Analysis, Ellis Horwood, Chicester, 1988, p. 85. Jandik P., Haddad P.R. and S m k P.E.. CRC Crit.Rev. Anal. Chem., 20 (1988) 1. Fritz J.S., Gjerde D.T. and Pohlandt C., Ion Chromatography,Huthig, Heidelberg, 1982. p. 119. Gjerde D.T. and Fritz J.S., A d . Chem., 53 (1981) 2324. Fritz IS., Gjerde D.T. and Baker R.M., Anal. Chem., 52 (1980) 1519. Fritz J.S., Anal. Chem., 59 (1987) 335A. Foley R.C.L. and Haddad P.R., J . Chromatogr., 366 (1986) 13. Gjerde D.T., Fritz J.S. and Schmuckler G., J. Chromatogr., 186 (1979) 509. Glatz J.A. and Guard JE., J. Chromatogr. Sci., 20 (1982) 266. Haddad P.R. and Foley R.C., Anal. Chem., 61 (1989) 1435. Haddad P.R. and Heckenberg A.L., J. Chromarogr..300 (1984) 357. Okada T. and Kuwamoto T., Anal. Chem:,55 (1983) 1001. Rock A. and McConkey G.,Topics in Chemical Physics, Elsevier, Amsterdam (1962) p. 246. Braunstein J. and Robbins G.D., J. Chem. Educ., 48 (1971) 52. Daum P.H. and Nelson D.F., Anal. Chem., 45 (1973) 463. Svoboda V. and Maesal J., J . Chromatogr., 148 (1978) 111. Tesarik K. and Kalab P.,J. Chromafogr.,78 (1973) 357. Kourilova D., Slais K. and Krejci M., Collect. Czech. Chem. Commun., 48 (1983) 1129. Slais K., J. Chromarogr., 436 (1988) 413. Dasgupta P.K., Soroka K.and Vithanage R.S., J . Liq. Chromarogr., 10 (1987) 3287. Johnson D.E. and Enke C.G.,Anal. Chem., 42 (1970) 329. Keller J.M., AnaZ. Chem., 53 (1981) 344. Holler J.F. and Enke C.G., in Kissinger P.T. and Heineman W.R. (Eds.) Laboratory Techniques in ElectroanalyticalChemistry,Marcel Dekker, New York. 1984, p. 235. Baba N. and Housako K., US.Patenr 4,462,962 (1984). Mysels E.K., Sholten P.C. and Mysels K.J., J. Phys. Chem., 74 (1970) 1174. Popovich D.J., Dixon J.B. and Ehrlich B.J.. J. Chromatogr. Sci., 17 (1979) 643. Sat0 H., Bunseki Kagaku, 31 (1982) T23. Tarter J.G., J . Chromatogr., 367 (1986) 191. Jen J.-F., Daugherty K.E. and Tarter J.G., J . Chromatogr., 436 (1988) 86. Scott R.P.W ., Liquid Chromatography Detectors {Journal of ChromatographyLibrary, Vol. 1 I), Elsevier, Amsterdam, 1977, pp. 209-215. Small H., Stevens T.S. and Bauman W.C., Anal. Chem., 47 (1975) 1801. Koch W.F., Anal. Chem., 51 (1979) 1571. Stevens T.S., Davis J.C. and Small H., Anal. Chem., 53 (1981) 1488. Hanaoka Y., Murayama T., Muramoto S., Matsuura T. and Nanba A., J. Chromatogr., 239 (1982) 537. Rokushika S., Qiu Z.Y. and Hatano H., J. Chromatogr., 260 (1983) 81. Rokushika S., Qiu Z.Y., Sun Z.L. and Hatano H., J. Chromatogr., 280 (1983) 69. Dionex Technical Note 15.

ConductivityDetection 39

40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57

289

Dasgupta P.K.. Anal. Chem.. 56 (1984) 103. Stevens T.S., Res. Dev., September (1983) 96. Stevens T.S.. Jewett G.L. and Bredeweg R.A., Anal. Chem.. 54 (1982) 1206. Dasgupta P.K.. in Tarter J.G., (Ed.) Ion Chromatography. Marcel Dekker, New York, 1987, p. 191. Stillian J., LC,3 (1985) 802. Rocklin R.D., Pohl C.A. and Schibler J.A.. J. Chromatogr., 411 (1987) 107. Sunden T., Cedergren A. and Siemer D.D., Anal. Chem., 56 (1984) 1085. Siemer D.D. and Johnson V.J., Anal. Chem., 56 (1984) 1033. Shintani H. and Dasgupta P.K., Anal. Chem, 59 (1987) 802. Tian ZW., Hu R.Z. Lin H.S. and Hu J.T., J. Chromatogr., 439 (1988) 159. Strong D.L. and Dasgupta P.K., Anal. Chem., 61 (1989) 939. Rocklin R.D.. Slingsby R.W. and Pohl C.A., J. Liq. Chromatogr.. 9 (1986) 757. Slingsby R.W., J. Chromutogr., 371 (1986) 373. Haginaka J., Wakai J., Yasuda H. and Nomura T., J. Chromatogr., 447 (1988) 373. Murayama T., Kubota T., Hanaoka Y., Rokushika S., Kihara K. and Hatano H., J. Chromatogr.. 435 (1988) 417. Tanaka K. and Fritz J.S.. Anal. Chem.. 59 (1987) 708. Tian Z.W., Hu R.Z.,Lin H.S. and Hu W.L., J. Chromatogr., 439 (1988) 151. Pohl C.A. and Johnson E.L., J. Chromatogr. Sci., 18 (1980) 442. Nordmeyer F.R., Hansen L.D., Eatough D.J.. Rollins D.K. and Lamb J.D.. Anal. Chem, 52 (1980) 852.

58 59

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60

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61 62 63 64 65 66 67 68 69 70 71

Gob1 M., GIT Fachz. Lab., 27 (1983) 261. Gob1 M., GIT Fachz. Lab., 27 (1983) 373. Polite L.N., McNair H.M. and Rocklin R.D., J. Liq. Chromutogr., 10 (1987) 829. Cassidy R.M. and Elchuk S., J. Chromutogr. Sci., 21 (1983) 454. Jenke D.R. and Pagenkopf G.K., Anal. Chem.. 54 (1982) 2608. Fortier N.E. and Fritz J.S.. Talanta, 34 (1987) 415. Sorensen J.A. and Glass G.E.,Anal. Chem.. 59 (1985) 1954. Dionex Technical Note 9R. Wilson S.A., Yeung E.S. and Bobbit D.R., Anal. Chem, 56 (1984) 1457. Hershcovitz H.. Yarnitzky C. and Schmuckler G.,J. Chromutogr., 244 (1982) 217. Strassburg R., Fritz J.S.. Berkowitz J. and Schmuckler G..J. Chromatogr., 482 (1989)

72 73 74

Weidenauer M., Hoffmann P. and Lieser K.H., Fres. Z Anal. Chem., 331 (1988) 372. Schmitt G.L.,PhD Thesis, University of Iowa, Iowa City (1985). Weiss J., Handbook of Ion Chromatography, Dionex Corporation, Sunnyvale, CA, 1986, p. 102.

(1985) 2257. 169.

343.


29 1

Chapter 10 Electrochemical Detection (Amperometry, Voltammetry and C o u lo me try) 10.1 INTRODUCTION

10.1.1

Definitions

The term "electrochemical detection" is applied loosely to describe a range of detection techniques involving the application of an electric potential (via suitable electrodes) to a sample solution, followed by measurement of the resultant current. Conductivity detection, which was discussed in full in Chapter 9, quite properly falls into this category, but is usually treated as a distinct detection method. In a similar manner, potentiometric detection, which is discussed in Chapter 11, can also be validly regarded as an electrochemical technique, but has some distinguishing features which render it more suitable for separate consideration. For the purposes of the discussion in this Chapter, we will interpret electrochemical detection to embrace the techniques of voltammetry, amperometry and coulometry. The common characteristic of these techniques is that a chemical reaction (e.g. a Faradaic oxidation or reduction) occurs during the measurement. It will be remembered that steps to avoid such reactions were taken in conductivity detection, and the same is true of potentiometry.

Voltammetry and polarography Voltammetry is a well-established technique which involves the application of a changing potential (measured with respect to a reference electrode) to a working electrode, followed by measurement of the current resulting from the reaction of analyzed species at the working electrode. Within the general field of voltammetry, we can identify polarographic techniques as those which measure current-voltage relationships using mercury as the working electrode. The key factor in voltammetry (and polarography) is that the applied potential is varied over the course of the measurement. Amperometry and coulometry The term amperometry describes the technique in which a fixed potential (again measured with respect to a reference electrode) is applied to a working electrode and the current resulting from oxidation or reduction reactions occurring at the working

292

Chapter 10

electrode is measured. In the case of chromatographic detection. the working electrode is located in a suitable flow-cell, through which the eluent stream passes. The analyte to be detected undergoes a Faradaic reaction if the applied potential has appropriate polarity and magnitude. However, the surface area of the working electrode in amperometry is generally quite small (0.5 cm2 or less), so the Faradaic reaction of the analyte is incomplete, causing only a fraction of the total analyte to react. In fact, less than 10% of the analyte is reacted in a typical amperometric flow-cell at flow-rates around 1 ml/min. The use of working electrodes of larger surface area can lead to quantitative reaction of the analyte at the electrode, and when this occurs, the technique used is described as high-efficiency amperometry, or coulometry. Thus, amperometry and coulomeny can be distinguished by the extent to which the analyte undergoes a Faradaic reaction at the working electrode. 10.1.2

Interrelationships between voltammetry, amperometry, and eoulometry

Potential window Consider the situation where a working electrode is inserted into a solution containing an electrolyte (e.g. KCl), but with no electrochemically-active(i.e. oxidizable or reducible) solute present. The potential of the working electrode is now varied with time from a negative value (E2) through to a positive value (El). This potential-time profile is shown in Fig. lO.l(a). The experimental configuration will require that a reference electrode be in electrolytic contact (but not necessarily inserted into) the sample solution. Current flow will be measured between the working electrode and an inert, auxiliary electrode, which is also inserted into the sample solution. Three electrodes are therefore present; the working electrode to which the potential is applied, the auxiliary electrode which measures the current flowing, and the reference electrode. The current which flows in the above experiment varies with the applied potential and follows the general shape depicted in Fig. lO.l(b), which is called a voltammogram. The current is negligible over most of the potential range because no electroactive solute is present, and appreciable current flow is observed only at the extremes of the potential range. At the most positive potential (El), the observed current is due to oxidation of the electrolyte in the solution, or of the working electrode itself, and is therefore represented as an oxidative current. At the most negative potential (E2). the observed current is due to reduction of the electrolyte or of the working electrode, and is therefore represented as a reductive current. Clearly, the actual potentials at which these oxidative and reductive currents occur will be dependent on the nature of the working electrode and the type of electrolyte used. Thus, for a given combination of electrode material and supporting electrolyte, there exists a range of potentials over which current flow is minimal when no electroactive solute is present. This range of potentials is known as the potential window for that system. The potential window in Fig. lO.l(b) is indicated by Ew. Most IC applications of electrochemical detection are performed at positive (oxidative) potentials because the reduction of oxygen occurs at small, negative potentials and this severely limits the reductive potentials which can be used in solutions which have not been

Elecbochemical Derection

1

293

Ired

Fig. 20.2 Voltammograms obtained by applying the potential profile illustrated in (a) to a solution of supportingelectrolyte alone (b) and to the same electrolyte containing an oxidizable species (c) or

a reducible species (d). deoxygenated.

Addition of an electroaetive solute When an electroactive solute (i.e. one capable of Faradaic oxidation or reduction at the working electrode) is added to the electrolyte solution, and the potential is again varied from E2 to El, a different potential-current relationship results. Fig. lO.l(c) shows the potential-current relationship when the added solute is oxidizable. An S shaped step (i.e. a wave) appears in the voltammogram. The height of the wave (given by the anodic current, Ia) is proportional to the concentration of the oxidizable solute (Co), whilst the position of the middle point of the wave on the potential axis (called the half-wave potential, El@) is characteristic of the oxidizable solute. Fig. lO.l(d) shows the voltammogram which results for a reducible solute and it can be seen that a cathodic, reduction current IC results, which is proportional to the concentration of the reducible species (CR).Again, the half-wave potential is characteristic of the reducible solute involved.

Chapter 10

294

Fig. 10.1 shows that voltammetry can be used to identify a solute and to determine its concentration. Provided only one electroactive species is present, the same result could be achieved by monitoring the current flow at a fixed potential, that is, by amperometry. The potential used should be one at which the analyte gives maximal current flow, but the residual current due to the electrolyte, working electrode. etc., is minimal. Thus, E3 would be a suitable potential for amperometric measurement of the oxidizable solute in Fig. lO.l(c), and E4 would be suitable for the reducible solute in Fig. lO.l(d).

Hydrodynamic voltammograms Successful amperomemc or coulometric detection can result only if the applied potential is chosen correctly. Extensive compilations of half-wave potentials for many solutes, measured with various working electrodes and under differing experimental conditions, are available in the literature (e.g. [ 11). These compilations are valuable, but sometimes do not contain information which is specific to the actual experimental conditions being employed. In these circumstances, voltammograms such as those shown in Fig. 10.1 must be obtained. This is a relatively simple matter when voltammetric instrumentation is available which permits the potential to be varied continuously whilst the current is monitored. However, amperometric instruments are designed to operate at a fixed applied potential, so true voltammograms cannot be generated. In these cases, a voltammogram can be obtained by measuring the current output for a fixed concentration of analyte, using a series of discrete potentials covering the desired range. When the amperometric instrumentation used for this procedure consists of a flow-through cell, the voltammogram .is obtained under flowing conditions and is referred to as a hydrodynamic voltummogram [2]. Hydrodynamic voltammograms find extensive use in chromatographic applications of amperometry and coulometry and are especially useful for the comparison of the performance of different electrochemical detectors. Analysis of mixtures of electroactive species Mixtures of electroactive species can be analyzed by simple voltammetry, provided that.the half-wave potentials and relative concentrations of the components of the mixture are such that the separate waves for each component can be discerned. Amperometry is not suitable for such mixtures because the measured current will often be due to more than one solute. However, when amperometry is coupled with chromatographic separation, a powerful combination results. The sample components are separated in the chromatographic system and pass sequentially over the working electrode, where they are detected amperometrically. Moreover, this form of chromatographic detection is specific for electroactive solutes and in IC therefore serves as a valuable adjunct to universal detectors, such as conductivity detectors. 10.1.3

Basic instrumentation for electrochemical detection in IC

An electrochemical detector can be formed from a potential supply, appropriate circuitry for the measurement of current, and a suitable sample cell. In the case of IC,

Electrochemical Detection

Reference electrode

Auxiliary electrode

295

Buffer

= I

Voltage

Current-

Eluent stream

Fig. 10.2 Basic configuration of an electrochemical detector

a flow-through type of sample cell is used and details of such cells are provided later in Section 10.4. As discussed above, the cell should accommodate three electrodes. Fig. 10.2 shows a schematic representation of the basic configuration of an electrochemical detector for flowing sample streams. The recorded analog signal is commonly generated by conversion of amplified oxidative or reductive currents generated in the cell to voltages, using the current-voltage converter shown in Fig. 10.2. More detailed descriptions of the electronic circuitry for voltammetric and amperometric measurements can be found elsewhere [2-71. It should be noted that considerable versatility of detection can be achieved by varying the manner in which the potential is applied to the working electrode. It is not necessary that a simple DC voltage be used for this purpose. Indeed, it is often very advantageous to use potential pulses, or sequences of pulses, and to measure the resultant current at specific times. Pulsed detection will be discussed further in Section 10.2.2. Table 10.1 provides a summary of the more important characteristics of voltammetric, amperometric and coulometric detection.

10.1.4 Usage patterns for electrochemical detection in IC Voltanimetry (including polarography [8-lo]), amperometry and coulometry have all found application as detection methods for IC. Other electrochemical methods, such as electrocatalytic detection [1 11, double-layer capacitance electrosorptive detection [ 12, 131 and indirect electrosorptive detection [ 141 have also been successfully used. However, it is fair to say that amperometry and coulometry are the most widely applicable methods, and of these, amperometry predominates. It is interesting to speculate on why this usage pattern exists. The difficulty in constructing a low-volume cell containing the dropping mercury electrode, coupled with the limited range of oxidative potentials available with a mercury electrode, combine to restrict the application of polarographic detection in IC. Voltammetry at solid electrodes does not offer significant advantages over amperometry using the same

s:

TABLE 10.1 BASIC PRINCIPLES OF VOLTAMMETRIC, AMPEROMETRIC A N D COULOMETRIC DETECTION

Q,

Method

Controlled quantity

Measured quantity

Cell design

Electronic unit

Remarks

Ampemmetry

Potential is held constant

Current

Three e l e c d e s

Potentiostat

Less than 10% conversion of the analyte at the electrode. Electrode materials: carbon, silver, gold, mercury. Wellestablished technique. Large choice of commercialinstrumentation.

Coulornetry

Potential is held constant

Current, charge

Working electrodes larger than used for

Potentiostat

A p p x . 10% conversion of rhe analye. WelleStabllished technique. Few commercial instruments available

Voltammetric instnunentation providing modulated potentials and various current sampling modes

This technique can be used by combining ampemmetric flow-cells with voltammetric instrumentation.

See voltammetry. MicrOproCeSSOr instrumentation is necessary

Reproducibility of ampemmetric detection on some solid electrodes is improved with this technique.

~pemmetry

Voltammetry

Potential is changed as a function of time

Current is evaluated either continuously or by sampling

Pulsedampemmetry

Potential is applied as pulses to clean the electrodebetween measurements

Current is evaluated between two cleaning pulses

electrodes

Three electrcdes

3


297

TABLE 10.2 SOME ELECI’ROACITVESOLUTES SUITABLE FOR DIRECT AMPEROMETRIC DETECTION IN IC ArseNte

Mde

Bromate Bromide Carbohydrates Chlorate Chloride chlorite

Chromate

Cyanide Hexathionate Hypochlorite

Iodate Iodide Nitrate Nitrite Oxalate Pentathionate

Sulfide sullite Sufite adducts

Teaathionate Thiocyanate Thiosulfate Transition metals

Transition metal complexes

electrode types, and requires more sophisticated instrumentation. Amperometry and coulometry therefore remain as the electrochemical methods of choice. The relative merits of these two techniques has been the subject of some debate [15171. At first sight, it might appear that coulometry would offer better sensitivity than amperometry because of the greater extent of reaction of the electroactive species which occurs in coulometric detection. However, as the electrode surface area is increased in order to improve the efficiency of the Faradaic reaction, the background current due to breakdown of the solvent electrolyte also increases. Little, if any, gain in sensitivity therefore results. Coulometric cells are often awkward in design and difficult to dismantle and maintain. In addition, these cells are sometimes expensive and can be used with only a limited range of working electrode materials. Considered together, these factors provide some cogent reasons why amperometry is used more frequently than coulometry. In view of the above discussion, the ensuing treatment of electrochemicaldetection techniques in this Chapter will focus on amperometry, with passing reference to coulometry and polarography.

10.2 MODES OF OPERATION OF ELECTROCHEMICAL DETECTORS 10.2.1 Direct and indirect detection

Direct electrochemical detection The majority of amperometric and coulometric applications in IC involve the direct detection of electroactive solutes. That is, the recorded current arises from a Faradaic reaction involving the particular solute ion under consideration. Table 10.2 gives a partial listing of the inorganic and organic ions which show Faradaic reactions in the potential windows accessible to modem electrode materials. This listing includes only a small fraction of the ions detectable using conductivity measurements, so direct electrochemical detection can be considered to be quite selective, as well as offering extreme sensitivity. This selectivity is illustrated in Fig. 10.3, which shows peaks for S2-

298

Chapler 10 1

Slan d a i d

Saniple 1

.-. u

2

E

C -

I

18 niin

18 min

Fig. 10.3 Direct amperometric detection of (1) sulfide and (2) thiosulfate in standard solutions and in Kraft White liquor. Solute concentrations in the standard solution were 1 pprn and the sample was diluted 1:1O,OOO. A Waters M460 amperometric detector was used with a silver electrode. The eluent was 5 mM Na2HP04 at pH 6.5. Chromatograms courtesy of Waters.

and Sz0j2- obtained in a Kraft White liquor produced in pulp and paper processing. None of the electroinactive anions (such as C1- and OH-) present in the sample appear in the final chromatogram. Direct electrochemical detection will be discussed at some length in the remainder of this Chapter.

indirect electrochemical detection In recent years, some indirect electrochemical detection methods have also been developed to enable the detection of solutes which are not electroactive. Indirect detection methods can be defined as those in which the measured current is not due to a Faradaic reaction involving the solute ion. One approach to indirect detection is to use an eluent competing ion which is itself electroactive and to monitor the changes i n concentration of this species as solute ions are eluted [18]. Salicylate or 2,Sdihydroxybenzoate have been suggested as suitable eluent ions for anion-exchange IC and Fig. 10.4 shows indirect amperometric detection of CI- and NO3- after their separation on an anion-exchange column, It is noteworthy that the electroactive eluent anion need be present at very low concentrations only and may therefore be used in conjunction with a second eluent anion which has the prime function of separating the desired

Electrochemical Deitction

299

-

0 1 2 3 Time Imin)

Fig. 10.4 Chromatogram obtained with indirect amperometric detection. A Biotronik BT I1 AN anion-exchange column was used with 0.1 mM salicylic acid-0.9 mh4 sodium salicylate as eluent. Amperometric detection using a carbon-paste electrode at +950 mV was used. Sample concentrations were 5 ppm for Cr and 10 ppm for NOg. Reprinted from [18] with permission.

solutes. The eluent ions responsible for separation and detection of the solute anions can therefore be different, giving more flexibility in eluent choice than when a single eluent anion serves both functions. The latter situation, of course, exists in all other forms of indirect detection. Indirect amperometric detection has been applied to suppressed IC by utilizing the pH change which accompanies the elution of strong acid anions or metal ions. In the simplest case, the hydrogen ions eluting together with strong acid anions after their passage through the suppressor, have been found to give a change in the baseline current of an amperometric detector [19, 201. This permits detection of electroinactive species, as illustrated in Fig. 10.5(a), which also shows the pH change occurring during elution of the same sample peaks. This pH change may be used in yet another way by virtue of its effect on the protolytic equilibrium of a suitable electroactive species. For example, p-benzoquinone undergoes a reversible reduction to p-hydroquinone, as shown below:

p-benzoquinone + 2H++ 2e- 5 p-hydroquinone

(10.1)

n NO3

.i

[

100 nA

_-

[10

nA

SOL*-

I,

iL

f

L I

,

0

2

i

,

6

1

1

1

1

e 1 0 1 2 l i

Time (min) (a)

r

0

. I

5

JQ* N"L+

L Rb* Cs+

I

I

1

I

I

1

10

15

20

0

30

60

Time (min) (b)

Time (min) (C 1

Fig. 10.5 Indirect electrochemical detection in suppressed IC using the pH change which accompanies sample elution. (a) A Dionex HPIC-AS3 column was used with a Dionex AFS fibre suppressor. The eluent was 3 mM NaHCO3 and 2.4 mM Na2C03 and the sample concentrations were 4 ppm for F-and C1-, and 20 ppm for the remaining ions. The amperometric detector was operated at 0.3 V versw AglAgCl. Reprinted from [191with permission. (b) A Dionex column was used. The effluent from the suppressor was mixed with 10 mMp-benzoquinone, 1 mM hydroquinone and 0.1 M KCl. Coulomemc detection using a carbon cloth working electrode at 0.45 V versus Ag/AgCl was employed. Solute concentrations were 5 ppm and 1 ml of sample was injected. Reprinted from [21] with permission. (c) A Dionex cation column was used with 3 mM HNo3 as eluent, and with a cation fibre suppressor. Hydrcquinone was added to the suppressor effluent. Coulometric detection at 0.45 V versus AglAgI was used. Solute concentrations: 20 ppm of each ion. Reprinted from [26] with permission.

z

301

Elecrochem'cal Detection

This reaction is pH dependent and will therefore be influenced by H+or OH- ions eluted in the sample band. Indirect amperometric or coulometric detection has been accomplished in suppressed IC using this approach for anions (by detecting the co-eluted H+ ions) [21-241 and for cations (by detecting the co-eluted OH-ions) [21,25,26]. Both ion-exchange and ion-exclusion separation methods have been used. Fig. 10.5(b) shows the detection of anions using indirect coulometry, whilst Fig. 10.5(c) shows this approach applied to the detection of cations. Indirect electrochemical detection of metal ions after ion chromatographic separation can also be achieved by adding an electroactive ligand (or an electroactive metal complex) to the column effluent. Complexation or ligand exchange reactions occurring with the eluted metal ions lead to a reduction in the detector signal due to the added component. This method has been used with dithiocarbamate ligands [27, 281, dithiocarbamate complexes [29] or diethylenetriaminepentaacetic acid (DTPA) 1211 as the post-column reagent. In the latter case, the ligand is added as a labile complex of Cu(II), which reacts with eluted metal ions to give the following electrode reaction: [Cu-DTPAI3-

+ M2+ + 2e-

% [M-DTPAI3- + Cuo

(10.2)

where I$+ represents an eluted metal ion. Fig. 10.6 shows a chromatogram obtained with this approach.

10.2.2 Amperometric detection with pulsed potential

Problems with conventional VC amperometry The discussion thus far has concentrated on single potential (dc) amperometry. In this mode, the selected potential is applied continuously between the working electrode and the reference electrode. The auxiliary electrode (also called the counter electrode) prevents a deleterious current flow through the reference electrode. The sample is in direct contact with the working electrode. A serious problem encountered with this mode of amperometric detection is a gradual loss of detection sensitivity. To understand the nature of this problem, it must be appreciated that the amperometric detector utilizes a heterogeneous electrochemical reaction which occurs at the interface between the working electrode and the sample solution. In some cases, reaction products can accumulate and adhere to the electrode surface, thereby blocking the surface and hindering further reaction. In other cases, the electrode surface itself can show deterioration. The outcome of either of these processes is a decline in the efficiency of the Faradaic reaction occumng at the working electrode. This leads to a decrease in the current produced and is manifested in a flowing system as a decrease in the peak height recorded for the particular solute under study. In addition, baseline noise and drift may also increase as the electrode becomes coated (or "poisoned"). The sensitivity must therefore be monitored carefully by frequent injections of a standard and, when the detector response becomes unacceptable, the cell must be reconditioned by replacing or polishing the working electrode.

Chapter I0

302

I

0

I

15 Time (min)

$0

Fig. 10.6 Indirect coulometric detection of metal ions using post-column ligand exchange reactions. An Aminex A-4 column was used with 0.18 M sodium tartrate, 0.04 M tartaric acid and 0.04 M sodium chloride as eluent. The post-column reagent was 10 mM Cu-diethylene-

triaminepentaacetate, 1.0 M NhOH, 0.1 M NH4NO3. The coulometric detector was operated at 0.75 V. Solute concentrations 0.5 ppm. Reprinted from [21] with permission.

Pulsed amperometric detection (PAD) One elegant method for overcoming the problem of adsorption of reaction products on certain electrode surfaces is to use potential pulses instead of a continuously applied dc potential [30-321. In the pulsed mode, the detector measures current only during a short sampling interval, so there is less likelihood of electrode fouling. In addition, the potential can be stepped to values more negative or positive than the measuring potential as a means of cleaning the electrode surface or activating it to improve detection response. Fig. 10.7 shows a typical pulsed potential waveform which might be applied to a working electrode. The measuring potential (El) is applied and the current is determined over a suitable time period. Some reaction products become deposited on the electrode during this process, but if the potential El is applied for only a short time, then these deposits can be expected to occur in only very small amounts. After measurement, the potential is raised for a short time (E2) and then lowered (E3) for a further short interval. Note that E2 and E3 are more positive and more negative, respectively, than El. These steps permit the desorption from the working electrode of reaction products which are oxidizable or reducible. At the conclusion of this cycle, the measuring potential can then be applied to a clean electrode surface.

Elecnochemical Detection

E

303

Cleaning

Cleaning

E2 Measuring

El

El Activation E3

E3

)t Fig. 10.7 Typical triple-pulse potential waveform used for pulsed amperometric detection.

A further advantage of pulsed amperometric detection is that activation of the working electrode may occur due to the cycling of potentials. For example, at the potential E3, activation may result from reduction of surface sites on the electrode or possibly adsorption of the analyte [30, 331. An important example of this effect is the catalytic oxidative detection of carbohydrates at a platinum or gold electrode, as illustrated in Fig. 10.8 [34]. Using this approach, Johnson and co-workers have reported the detection of several classes of compounds which were thought previously to be undetectable by amperometry [30, 351. A summary of the applications of pulsed amperometric detection in IC is available [36].

Reverse-pulse methods We have noted previously that electrochemical detection is generally performed at positive potentials, since the presence of dissolved oxygen limits the accessible range of reductive potentials. This means that direct detection of metal ions is not possible unless oxygen is removed. Oxygen can be eliminated using a variety of procedures (e.g. [37]), and further steps must also be taken to prevent redissolution of oxygen. For example, the entire chromatograph can be blanketed with nitrogen. Reverse-pulse amperometry provides a means to detect metal ions through their reduction reactions at a dropping mercury electrode, without interference from dissolved oxygen 1381. This is achieved by use of a potential waveform in which a large, negative initial potential (Ei) is applied during the majority of the lifetime of the mercury drop. Transport-limited reduction of the analyte ions occurs and the analyte is deposited onto the elecuode surface, forming an amalgam. The potential is then stepped to a positive, final value (Ef), near the anodic limit for mercury. An oxidative current results as the deposited analyte is oxidized from the electrode. Dissolved oxygen does not react. Hence, cathodically active (reducible) analytes are detected by way of an anodic reaction. Fig. 10.9 shows the detection of some transition metal ions using this method.

Chapter 10

304

6 8

10

1

3

'1516

1 I

I

I

I

I

0

5

10

15

20

I

25.

I

I

30

35

I LO

Time (min)

Fig. 10.8 Pulsed amperometric detection of carbohydrates following separation by gradient elution. A Dionex HPIC-AS6A column was used with a gradient of 0-50 mM NaOH. NaOH (0.3 M) was added as a post-column reagent. Peak identities (and concentrationsin ppm): 1 - inositol (15), 2 - sorbitol (40),3 - fucose (25), 4 - deoxyribose, (25) 5 - deoxyglucose (20), 6 - arabinose (25), 7 - rhamnose (25), 8 - galactose, 9 - glucose, 10 - xylose, 11 - mannose, 12 - fructose, 13 melibiose, 14 - isomaltose. 15 - gentiobiose, 16 - cellobiose, 17 - turanose, 18 - maltose. Reprinted from [34] with permission.

10.3 ELECTRODES FOR AMPEROMETRIC DETECTION 10.3.1

Reference and auxiliary electrodes

Early cell designs included only two electrodes. that is, the working and reference electrodes. We have already noted that this configuration is undesirable because the reference electrode must c a m a current and hence its potential does not remain constant. The auxiliary electrode is therefore included to carry the cell current, so that the reference electrode is maintained under conditions of zero current flow. The most widely utilized reference electrodes are the Ag/AgCl electrode and the saturated calomel electrode (SCE). A palladium-hydrogen electrode and quasi-reference electrodes based on platinum and other metals have also been employed. Potentials of

ElectrochemicalDetection

305

-

0

5 10 Time (min)

15

Fig. 10.9 Detection of reducible metal ions by reverse-pulse ampemmetry, without removal of dissolved oxygen. A column packed with Dunum DC-4A cation-exchange resin was used with 0.10 M sodium hydrogentartrate and 60 mM Mg2+at pH 4.0 as eluent. The potential values used were Ej = -1.30 V,Ey = -1-0.20V. Reprinted from [38] with permission.

reference electrodes are available in the literature [39] and should be considered when an attempt is made to reproduce a detection method developed originally with a different reference electrode. Auxiliary electrodes should be constructed of inert material and should ideally be situated as close as possible to the working electrode. This minimizes potential drop due to the resistance of the sample solution. Typical auxiliary electrode materials are platinum and glassy carbon. In some detector cells, the stainless steel capillary tubing used to connect the chromatographic column to the cell may serve as the auxiliary electrode. This function may also be filled by the cell body itself, provided that it is constructed of a suitably conducting, inert material.

10.3.2

Working electrode materials

Many different materials have been used for the constmction of working electrodes for amperometric detection. Several comprehensive reviews on electrochemistry include discussion on this subject [5-7,40,41]. In this Section, the significant characteristics of the more important electrode materials will be listed.

306

Chapter 10

Mercury For reduction reactions, the electrode material of choice has been, and remains, mercury. The primary reasons for this choice are the high overpotential for reduction of the hydrogen ion, the formation of amalgams with many metals, and the ease of replacement of mercury-drop electrodes. The high hydrogen overpotential on mercury means that mercury electrodes can be used in acid solution without interference by the reduction of hydrogen ions. Mercury electrodes have the widest negative potential range of any electrode material. On the other hand, mercury is quite easily oxidized (at about 0.4 V versus SCE) and this prevents its use for the study of most oxidative processes. This limitation can be overcome when analytes forming complexes or precipitates with mercury ions are involved. Representative reactions of this type are shown below, where the analyte is represented as Ln- [7]: Hg

+ 2Ln-

2Hg

2(1-n)+ % HgL, + 2e-

+ 2Ln-

% Hg,L,

2( 1-n)+

+ 2e-

(10.3)

(10.4)

Since these electrode processes involve oxidation of mercury in the presence of complex-forming species, they occur at potentials less positive than for oxidation of the mercury itself. Removal of oxygen is unlikely to be necessary and only a moderate positive potential is required. This can be contrasted to the case where the oxidation of the same species is conducted on other electrode materials. Here, more positive potentials would be required and the likelihood of interference is therefore increased. Examples of the inorganic anions which react with mercury include S2-and CN-, which form a precipitate and a complex, respectively, as shown below:

Hg

+ S2- f

Hg

+ xCN- %

HgSl

+ 2e-

[Hg(CN),]

(10.5)

(2-x)+

+ 2e-

(10.6)

Both of these ions have been detected by amperometry with a mercury electrode [421.

Mercury electrodes can be formed from mercury drops (either flowing or static) or from thin films of mercury coated onto a suitable substrate. Mercury adheres well to silver and platinum, but since it slowly dissolves these materials, the thickness of the mercury film decreases with time. Carbon Carbon can be used as an electrode material in a number of forms, including carbon paste, carbon impregnated into a suitable binder, glassy carbon, pyrolytic graphite, carbon fibres, etc. Carbon paste electrodes are manufactured from particles of carbon suspended in an oil (such as Nujol) or a wax which is immiscible with the

307


solution phase. These electrodes are relatively simple to manufacture and replace, and give high detection sensitivity due to the very low residual currents produced. Disadvantages of carbon paste electrodes include some variability in electrode performance which results even when successive electrodes are prepared from the same constituent materials, and the long equilibration times required for sensitive operation [40]. Composite electrodes, in which the carbon is impregnated into a suitable binder such as polyvinylchloride, neoprene rubber or Kel-F, give more consistent performance but are more difficult to prepare. Glassy carbon is a very popular electrode material because it can be formed into a variety of shapes and can be easily polished. Glassy carbon is a gas impermeable material formed by the heating of phenol-formaldehyde resins in an inert atmosphere. Electrodes of this type give higher residual currents than carbon-paste electrodes, but can be used over a 2 V working potential range which covers both positive and negative potentials. In addition, carbon is resistant to the formation of oxides at the electrode surface, so the electrode maintains integrity over prolonged periods of usage. Glassy carbon is sometimes also used for the construction of auxiliary electrodes as well as working electrodes.

Silver, platinum and gold Working electrodes may also be constructed of pure metals, which are usually inert materials so that the available potential window is not unduly restricted. It can be noted in passing that some reactive metals, such as copper [43],nickel [44]and copperized cadmium [45], have also found limited application as working electrodes in amperometry. Platinum and gold have wide potential windows which extend above +1.0 V in the oxidative region. Silver has a much smaller potential window, but is a valuable electrode material because of the reactions it can undergo with solutes during the detection process. It has been shown that amperometric detection of inorganic anions can be improved through the use of electrode materials that mediate or participate directly in the electrode reaction. This participation may be in the form of complexation or precipitation reactions, along the same lines discussed earlier for mercury electrodes. Silver has special importance in this regard and is a suitable electrode material for the amperometric determination of halides, CN-, S2-, S03*-, SCN- and S ~ 0 3 ~ As - . an example, we will consider the detection of CN-at a silver electrode. Cyanide can be oxidized to cyanate at a glassy carbon electrode, but only at a moderately high positive potential. The possibility of interference by other oxidizable anions therefore exists. With a silver electrode, the silver itself is oxidized in the presence of CN-to form a silver cyan0 complex, as shown in the following reaction [46]:

Ag

+ 2CN-

% Ag(CN)i

+ e-

(10.7)

This oxidation occurs readily at 0.0 V versus AgIAgCl, at which potential interferences are considerably less than encountered with a glassy carbon electrode operated at the higher potential necessary with this electrode material [7]. The ability of silver, gold and platinum electrodes to form complexes with halide ions restricts the

308

Chapter 10

potential range accessible to these electrodes in eluents which contain halides. Metal electrodes constructed from inert materials, such as gold and platinum, can show pronounced changes in performance with use. This behaviour arises from the adsorption of electrode products or of components from the sample itself. An example of this effect is the response of a platinum electrode used for the amperometric (oxidative) detection of I- [47]. A new electrode gave very low response to I-, whereas a used electrode which had just been polished gave very high sensitivity, but the signal produced was unstable and the baseline drifted severely. Reproducible electrode performance was obtained only when the platinum was conditioned by immersing it in a saturated solution of KI for 30 minutes, in order to form a uniform layer of chemisohed 1- on the electrode surface. It was necessary to repeat this procedure after 5 working days. Similar adsorption effects have been noted for S2- on silver working electrodes [48] and in the detection of CrOq2- with an iodized platinum working electrode [49].

Criteria for selection of the working electrode material The preceding discussion has revealed a wide range of materials which are available for the construction of working electrodes in amperometric detection. Some guidelines for selection of the correct material for a given application are therefore required. The following aspects should be considered when making this selection 1411: The potential window for the working electrode in the chromatographic eluent to be used. (ii) The involvement of the electrode itself in the electrochemicalreaction. (iii) The kinetics of the electron-transfer reaction. (i)

The potential window will vary with the pH of the eluent. Fig. 10.10 shows the approximate potential windows accessible to five common electrode materials when used in acidic and basic solution. As the applied potential approaches the potential limit, background current increases. This increase is much more rapid for metal electrodes than for carbon. Glassy carbon and platinum have the highest accessible positive potentials and are therefore preferred for oxidations. Mercury, glassy carbon and silver are preferred for reductions. Involvement of the electrode material itself in the electrochemical reaction can often allow a particular material to find use in a potential range in which other electrode materials are normally considered to be preferable. This behaviour has been illustrated above for the oxidative reaction of CN- with a silver electrode. Rocklin [41] has suggested that a formation constant of loio for reaction of the analyte ion with the electrode material indicates a high probability that sensitive detection will be achieved for that ion. Electrode reactions with fast kinetics require a working electrode potential slightly (about 100 mV) more positive (for an oxidation reaction) than the standard reduction potential (EO)for that reaction. At this potential, all of the oxidizable species reaching the electrode surface will react and further increases in potential will not lead to larger currents (and hence larger peaks). However, electrode reactions with slow kinetics do

ElectrochemicalDerecrion -1.5

- 2 .O

309 -1.0

I

I

I

I I

I I

1.0

I

I

I

I

I

I

Gold

I

I

I I

I

I

I

I I

I

I

I I

I

I

I

I

I

I

II t)

I

Silver

I

I

I I

I

I I

I I I I

I

-1.0

I

I

I I

II

I

Mercury

-1.5

I

I

Platinum

I

1 I

!

I

I '

I

I -2.0

0.5

I

I

I

I

0

I

Glassy carbon

I

I

-0.5

I

-0.5

I

I

!

!

I

0 0.5 VOLTS (wrt Ag/ApCl)

I

! I

1.0

1.5

Fig. 10.10 Approximate potential windows accessible with various electrode materials in acidic solution (white boxes) and in alkaline solution (shaded boxes). Data from [6,41].

not follow this rule and require substantially more positive potentials to achieve quantitative reaction of the analyte at the electrode surface. The reaction kinetics therefore play a prominent role in determining the optimal applied potential and this, in turn, affects the choice of the working electrode material. The eluent used for the particular IC separation under consideration also exerts a considerable influence on the selection of the working electrode. For example, when basic eluents are used for the ion-exchange separation of anions of weak acids, different electrode materials may be required in comparison to the detection of the same solutes under acidic conditions, such as after elution from an ion-exclusion column. 10.4 FLOW-CELL DESIGN AND RESPONSE EQUATIONS

Electrochemical cells for use with flowing streams may be classified as flow-by, in which the eluent flows parallel to the surface of the working electrode;flow-through, in which the eluent follows a tortuous path between surfaces of the working electrode; and flow-at, in which the eluent i,mpinges perpendicularly onto the surface of the working electrode.

Chapter I0

310 Outlet (to reference electrode)

t

eiectrpde

n

WoriLlng electrode

Fig. 10.11 Thin-layer amperometric flow-cell in (a) schematic form and (b) in commercial form. a - cell inlet, b - holder for working electrode, c - contact for working electrode, d - gasket, e FTFE holder for reference electrode, f - flow-through diaphragm, g - outlet. Courtesy of Waters. 10.4.1

Thin-layer cells

The most common type of amperometric cell is the flow-by, which is illustrated schematically in Fig. 10.1l(a). A thin spacer (in the form of a gasket) held between two rigid blocks defines the thickness, width and length of the flow-channel, and thereby the cell volume. In early versions of this cell, the working electrode was housed in one of the blocks comprising the cell, whilst the auxiliary and reference electrodes were mounted downstream in another compartment. Newer designs have the auxiliary electrode positioned close to the working electrode, and in some cases, even the reference electrode is positioned in the same cell compartment. A commercial cell design of this type is shown in Fig. 10.1l(b). These changes result in greatly improved

Electrocktnikal Detecrion

311

cell performance because the electrical resistance of the cell is lower and potential control is improved. In addition, the cell is simple to dismantle and the working electrode can be polished easily. The cell current for a thin-layer amperometric cell is given by [35]:

(10.8)

where i is the cell current in amperes, n is the number of electrons in the electrode halfcell reaction, F is the Faraday constant, A is the electrode area in cm2, D is the diffusion coefficient in cm2s-', h is the flow-channel thickness and d is the flow-channel width in cm, Uv is the volume flow-rate in cm3s-l, and C is the analyte concentration in mol ~ m - ~ . Eqn. (10.8) predicts that there is a linear relationship between cell current and analyte concentration, under conditions of constant flow-rate. It is also predicted that the cell current will increase with flow-rate, but this has been shown to be invalid for real cells [17]. Thin-layer cells attain detection limits in the sub-nanogram range, have wide linear dynamic ranges (three to five orders of magnitude) and exhibit good measuring reproducibility. Cell volumes of less than 1 pl can be achieved by decreasing the thickness of the spacer, but the baseline noise increases rapidly with decreasing cell thickness. The quality of polishing of the faces of the two cell blocks is also important, since the better the polishing, the thinner the spacer can be without the baseline noise becoming too great. 10.4.2

Flow-through, high-efficiency cells

The electrochemical efficiency of a cell can be increased by using electrodes of large surface area. This can be achieved in a thin-layer cell configuration by using a long, planar working electrode, but the design can be made more compact through the use of a flow-through electrode made from a packed bed of porous particles or from a porous plug of electrode material (such a.; reticulated vitreous carbon). Reaction of the analyte in this type of cell approaches 100%; that is, the cell operates in the coulometric mode. The response of such high-efficiency cells is given by [17]:

i = nFCUv

(10.9)

where the terms have the same meanings as for eqn. (10.8). If the current is integrated during the passage of the analyte through the cell, we obtain:

Q = nFM

(10.10)

where Q is the integrated current (i.c. the coulombs of electricity from the analyte) and M is the moles of analyte injected. Eqns. (10.9) and (10.10) show that the cell current for a high efficiency flow-through cell is larger and much more dependent on flow-rate than is the thin-layer cell, but the integrated current, Q, is independent of flow-rate.

312

Chaprer 10

Inlet Inlet

(a)

Working elect rode (b)

Fig. 20.12 Wall-jet cells in (a) constrained and (b) unconstrained formats.

Whilst the relative merits of amperometry and coulometry (and hence those of high-efficiency flow-through cells and thin-layer cells) may be debated, certain attributes of the high-efficiency cell are clear. These cells are well-suited for screening and sample clean-up applications where electroactive interferences in the sample or eluent can be reacted before the analyte band reaches the detector. On the other hand, these cells are not well-suited to applications in which the potential is swept or pulsed because the large surface area has an associated large double-layer capacitance effect. 10.4.3

Wall-jet cells

The wall-jet cell (shown schematically in Fig. 10.12(a)) is the most common example of the flow-at type of electrochemical cell. The sample flow is directed perpendicularly onto the surface of the working electrode and may exit from the cell through a narrow flow-tube (as shown in Fig. 10.12(a)), or may pass into a larger volume compartment surrounding the working electrode (Fig. 10.12(b)). The theoretical response equation for this type of cell is given by [50]: (10.11)

where v is the kinematic viscosity, R is the radius of the working electrode in cm, and a is the inside diameter of the jet in cm. The other terms in eqn. (10.1 1) are as previously defined. The theoretical flow-rate dependency of the wall-jet cell differs from that for the thin-layer cell, but in practical studies, cells such as that shown in Fig. 10.12(a) exhibit the same flow-rate dependency as a thin-layer cell. These are called constrained wall-jet cells, since the design causes the reflected flow to move parallel to the working electrode surface, so the cell is effectively a thin-layer type with radial flow patterns and therefore responds according to eqn. (10.10). A true wall-jet cell, which responds in accordance with eqn. (lO.ll), has a jet diameter, a, which is smaller than the radius of the working electrode, R. A sufficient


313

gap between the jet and the working electrode is also required, so that the tip of the jet does not interfere with the reflected flow from the electrode surface. Such cells are called unconstrained wall-jet cells, or large-volume wall-jet cells, and are depicted schematically in Fig. 10.12(b). Some important characteristics of wall-jet cells deserve mention. The effective cell volume is much smaller than the volume of the electrode compartment, since the effective volume actually consists only of the thin layer of liquid at the electrode surface. After detection, the sample band disperses rapidly in the relatively large electrode compartment, so wall-jet detectors can be used in tandem with other detectors only when placed last in line. (ii) Wall-jet cells show a greater flow-rate dependence than thin-layer types and are therefore more susceptible to flow-pulsations in the pumping system. (iii) Wall-jet cells are suitable for miniaturization without loss of concentration detection limits because the ratio of electrode radius to jet diameter is more important than electrode radius alone. (iv) The relatively small size of the working electrode results in small doublelayer capacitance effects and therefore a shorter time constant for the cell. Thus, this cell is suitable for those applications in which the electrode potential is changed rapidly, such as pulsed-amperometric detection. (i)

10.4.4

Polarographic cells

Polarographic cells require that a dropping mercury electrode (DME)or a hanging mercury drop electrode (HMDE)is used as the working electrode. The most difficult problem encountered with the DME is the attainment of a small cell volume without impairing the function of the DME. Two typical cell constructions are shown in Figs. 10.13(a) and 10.13(b), in which the DME is arranged vertically and horizontally, respectively. Cell volumes as low as 1 pl can be achieved, but current oscillations occur as a result of the physical disturbance of the solution as the drop falls. These current oscillations may be suppressed using an RC filter, but this increases the time constant of the cell and renders it unsuitable for rapid potential changes. Alternatively, a very short drop time may be employed, and this is best achieved with a horizontal capillary (Fig. 10.13(b)). A wall-jet type polarographic cell can also be used in which the eluent jet is directed vertically onto a macroscopic DME placed in a vessel containing a base electrolyte (Fig. 10.13(c)). The effective volume of such a cell is around 1 pl. The construction of detectors with a HMDE is more simple than with a DME,but the advantage of periodical renewal of the electrode surface is lost in these cells. 10.4.5

Multi-functional cells

More than one mode of electrocheqical detection can be combined into the same cell as long as the correct electrodes for each detection mode are supplied. For example, amperometry (at a glassy carbon working electrode) and polarography (at a DME)can be performed simultaneously using a bi-electrode cell [53]. The flowing sample solution

Chapter 10

314

1

Fig. 10.13 Polarographic detector flow-cells. (a) Thin-layer type with vertical DME. 1 - DME, 2 - inlet, 3 - reference electrode, 4 - auxiliary electrode. (b) Thin-layer type with horizontal DME. 1 - inlet, 2 - DME. 3 - mercury pool auxiliary electrode, 4 - tube to mercury reservoir. (c) Wall-jet type. 1 - DME, 2 - jet, 3 - base electrolyte, 4 - reference electrode, 5 - auxiliary electrode. Reprinted from [Sl, 521 with permission.

contacts both electrodes and the combination of electrode materials which is employed permits reductive and oxidative chromatograms to be recorded simultaneously on the electrode material most suited to each. It is also possible for conductivity and amperometric detection in IC to be used in the same detector cell [53]. The electrode arrangement (Fig. 10.14(a)) utilized in this case consists of a platinum working electrode inserted concentrically into a stainless steel capillary tube, with suitable insulation between the platinum and steel. Eluent flow is directed onto the working electrode in a wall-jet configuration and the cell volume is only 15 nl. Chromatograms with simultaneous conductivity and amperomeuic detection were obtained on a microcolumn for common anions, using an ion-interaction separation with phthalate as eluent (Fig. 10.14(b)). It is noteworthy that the amperometric and conductivity signals from the cell are quite independent. 10.4.6

Dual-electrode amperometric cells

Thin-layer cells, such as those shown in Fig. 10.11, can be modified by inclusion of a second working electrode. This electrode can be arranged in parallel or in series with the first working electrode. These configurations are illustrated in Fig. 10.15, together with typical operating modes of the dual-electrode cell. It is should be noted that the dual, parallel arrangement can also be produced in the cell shown in Fig. 10.11(a) by mounting a second working electrode in the position of the auxiliary electrode. In the dual, series electrode configuration, the first electrode (set at a positive potential) can be used to oxidize solutes, whilst the second electrode (set at a negative potential) can be used for reduction. Solutes which undergo reversible or quasireversible reactions will be oxidized at the first electrode, and then reduced at the second electrode, if the potentials are correctly chosen. On the other hand, solutes which are irreversibly oxidized will not be detected at the second electrode (Fig. 10.15(a)).


3 15

c I-

i,: NOf

4\

c

1 nA

/l

Hydroquinone

1 mm

(a)

I

0

I

I

10 20 Time (min)

I

30

(b)

Fig. 10.14 Simultaneous conductivity and amperomemc detection in IC using the same flowcell. (a) Design of the flow-cell. 1 - platinum working electrode, 2 - stainless steel capillary tube, 3 - insulation, 4 - stainless steel body of the cell. Arrows indicate the flow-path of the sample. (b) Chromatograms obtained with conductivity (upper trace) and amperomemc (lower trace) detection. A clfjmicrobore column was used with 0.75 mM teaabutylammonium phosphate and 0.25 mM phthalate at pH 6.3 as eluent. Solute concentrations: (21-, NOz-, Br- and NO3- all 1 mM; hydroquinone 0.1 mM. A polarizing voltage of +1.1 V was used. Reprinted from [54] with permission.

Interferences can therefore be eliminated. This approach has been utilized for the - , I- alone is reduced at the second electrode detection of I- in the presence of S Z O ~ ~since without interference from S 2 0 ~[55]. ~In the dual, parallel electrode cell configuration, different potentials (El and E2) are applied to each working electrode and the resulting currents are ratioed or subtracted (Fig. 10.15(b)). The two solutes (A and B) depicted in the Figure have different half-wave potentials. Solute B will give the same current at both electrodes, whereas solute A will give a higher current at El than at E2. The ratio of these currents can be used for identification of the solutes and as a measure of peak homogeneity, whilst the difference between the currents can be used to eliminate interferences. This

2 ch

TABLE 10.3 SOME TYPICAL APPLICATIONS OF ELECrrZOCHEMICAL DETECTION IN IC Solutes

Aldehydes Aldehydes (as adducts with S032') As(II1) As(IW, AsW) Bf Bi c2042-

CNCN-, HS-, I', BrCu2+, Ni2+ F ,ci-, PO^, NO^HS-, CNHS-, CN-

r, s2032-

II' IInorganic anions Inorganic anions

Sample

M

Standards Wines

PAD

Mineral water Standards Waters Plasma Urine Dust, particulates Standards Urine Standards Standards Drinking water Pharmaceuticals Iodized salt, KF Sedimentary rocks Seawater Standards Standards

Amp

d

Amp

Cod Amp Amp Amp Amp Amp Amp

Indirect Amp Amp

Pol Amp, DPE Amp Amp Amp Amp

Indirect cod

Working elecrmcleb Au GC

Reference electrodec

Potential

Sensitivity

Ref

Pulsed 0.75

1-3 pprn 5 PPm

57 58

1.oo 0.16 0.10 0.03 1.25 0.02 0-0.3 0.8 0-0.45 0.06 0.00 1.05, 1.15 0.8 0.17 1.o 1.1 v 0.16

0.012 pM n.s 10 PPb 10 PPb 100 PPb 1 PPb 2-30 ppb < 80 ppb c 0.5 pprn 20 PPb 1-2.5 ppm 10-50 ppb 10 PPb 10 PPb 5 PPb 0.05-19 ng c 0.1 ppm

59

W)

60 61 62 63 64 46 65 19 48 42 55

47 66 67 68 21

TABLE 10.3 (Continued)

Lactose Maltose NHs+ No3N@-,NOy Organic acids Po43-

Polythionates, ~ 2 0 3 ~ ~032s2e2S2s2-, s,2-, so32-, s2032-

s2-.s2032S2-, Br, CN-

Transition metals Transition metals Transition metals UWI)

Au

Pulsed pulsed

Potato chips Corn syrup Sewage Natural waters Smked meat, fish Urine Wastewaters Standards Foods Urine, plasma Turbid waters Coal desulfinizaaon samples Kmft White liquor Sour waters

PAD PAD Indirect cod Amp Amp, DPE

Amp Amp

n.s.

Standards

Indirect coul F s Pol IndirectAmp Pol

0.75 -1.30,0.20

Standards Seawater Urine, waters

Au C-cloth

cu-cd

Indirect cod DP Pol

GC GC n.s. HMDE

Amp

Pt

Amp, DPE

GC Ag DME

Amp

Amp

Indirect Pol

0.45 -1.15

1.10, 1.00 1.10 0.45 0-1.6 0.4 0.7,0.9 0.00 0.08

1.0

0.65 -0.65

n.s. 50 PPb 3.3 ng 6.2 pp b 50 n.s. 27 PPb n.s. 0.1 ppm 10pmol 1.9 ppb n.s.

32 32

n.s. n.s. < 0.5 ppm 13-64ppb 0.2-1.0 ppm 80 ng

73 74 21 38

25 45 69

70 23 8

71 56

72 10

29 9

Amp = ampemmetry, Coul = coulometry, Pol = polarography, PAD = pulsed ampemmetry, DPE = dual, parallel electrodes. RP = reverse-pulse, DP = differential pulse. GC = glassy carbon, DME = dropping mercury electrode, HMDE = hanging mercury drop elemode, WAG = wax-impregnated graphite, I-Pt = iodized Pt, C = carton, Cu-Cd = coppenzed cadmium. Fe(1I)FeQII) = fem/femcyanide, SCE = saturated calomel electrode. n c. = not specified. a

I

I

A

B

A'

A -EAl " A "E2

8

-+ El

,E2

8

No reaction

(a)

(bl

Fig. 10.15 Configurations for dual-electrode amperomemc detection in IC. (a) Dual, series electrodes. Irreversibly reacted solute B is excluded from interfering with the detection of reversibly reacted solute A at the second working electrode. (b) Dual, parallel electrodes, which may be operated separately and independently to yield difference and ratio chromatograms for solutes A and B. Reprinted from [7] with permission.

3

$ r, 0


319

approach can again be illustrated using I- and S Z O ~as~ solutes, ’ with El = +1.05 V and V [55]. The current ratios were found to be reproducible and were characteristic of each solute. A further application of dual, parallel electrode detection ~ -urine, without interference from uric acid in amperometry is the detection of S ~ 0 3 in or other electroactive components of urine [56]. E2 = +1.15

10.5 APPLICATIONS OF ELECTROCHEMICAL DETECTION Electrochemical detection has been applied in IC in situations where extreme sensitivity or special selectivity is required. Most commonly, the electrochemical detector has been operated in tandem with a universal, non-selective (e.g. conductivity) detector so that a more general sample analysis can be obtained than is possible with the electrochemical detector alone. Table 10.3 gives a representative listing of some typical applications of electrochemical detection in 1C and includes the approximate detection limits which can be attained. The chromatograms given throughout this Chapter provide further illustration of other applications.

10.6 REFERENCES 1 2 3 4 5 6 7

8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Meites L., Zuman P. and co-workers,CRC Handbook Series in Inorganic Electrochemistry, Vols. I-V, CRC Press Inc., Boca Raton, FL, 1980-1985. Kissinger P.T. and Heineman W.R., (Eds.),Laboratory Techniques in Electroanalytical Chemisrry, Marcel Dekker Inc.. New York, 1984. Sawyer D.T. and Roberts J.L., Experimental Electrochemistryfor Chemisrs, John Wiley, New York. 1974. Rveki R., Talanta, 27 (1980) 147. Stulik K. and Pacakova V., J . Electroanal. Chem., 129 (1981) 1. Palmisano F. and Zambonin P.G., Ann. Chimia, 74 (1984) 633. Johnson D.C., Weber S.G., Bond A.M., Wightman R.M., Shoup R.E. .and Krull I.S., Anal. Chim. Acta, 180 (1986) 187. Takano B.. McKibben M.A. and Barnes H.L., Anal. Chem.. 56 (1984) 1594. Cassidy R.M. and Elchuk S., Inter. J. Environ. Anal. Chem., 10 (1981) 287. Uddin Z., Markuszewski R. and Johnson D.C., Anal. Chim. Acta, 200 (1988) 115. Heuser J.R. and Girard J.E., Anal. Chem., 57 (1985) 2847. Ramstad T. and Weaver M.J., Anal. Chim. Acta, 204 (1988) 95. Ramstad T. and Weaver M.J., J. Chromatogr., 456 (1988) 307. Ramstad T., Anal. Leu., 21 (1988) 331. Kissinger P.T., Anal. Chem., 49 (1977) 487A. Weber S.G. and Purcjue W.C., Anal. Chim. Acta, 100 (1978) 531. Roe D.K., Anal. Lerr., 16 (1983) 613. Horvai G., Fekete J.. Niegreisz Z., Toth K. and Pungor E., J . Chromatogr., 385 (1987) 25. Tarter J.G., J. Liq. Chromatogr., 7 (1984) 1559. Jones V.K. and Tarter J.G., J. Chromurogr., 312 (1984) 465. Girard J.E., Anal. Chem., 51 (1979) 836. Tanaka K. and Ishizuka T., J. Chromatogr., 190 (1980) 77.

320 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39

40 41 42 43 44 45 46 47 48 49 50 51 52

53 54 55 56 57 58 59

60 61 62 63

chapter 10 Tanaka K. and Ishizuka T., Water Res., 16 (1982) 719. Tanaka K., Bunseki Kaguku, 32 (1983) 439. Tanaka K., Ishizuka T. and Sunahara H., J. Chromatogr., 177 (1979) 21. Tanaka K., Ishihara Y.and Nakajima K . , Bunreki Kagnku, 32 (1983) 626. Hojabri H., Lavin A.G.. Wallace G.G. and Riviello J.M.. Anal. Proc., 23 (1986) 26. Barisci J.N., Wallace G.G. and Riviello J.M., Chromarogruphia, 25 (1988) 162. Hojabri H.. Lavin A.G., Wallace G.G. and Riviello J.M., Anal. Chem., 59 (1987) 54. Hughes S. and Johnson D.C., Anal. Chim. Acta, 149 (1983) 1. Anderson J.E.. Bond A.M., Heritage I.D., Jones R.D. and Wallace G.G., Anal. Chem., 54 (1982) 1702. Edwards P. and Haak K.K., Amer. Lob., April (1983) 78. Berger T.A., U.S.Patent 4,496,454. January 29 (1985). Dionex Technical Note 20. Weber S.G., J. Elecrrounul. Chem., 145 (1983) 1. Dionex Technical Note 11R. Bratin K. and Kissinger P.T., J. Liq. Chromutogr.,4 (1982) 321. Hsi T. and Johnson D.C., Anal. Chim. Acta, 175 (1985) 23. Sejeant E.P., Potentiometry and Potentiomenic Tinutiom, John Wiley & Sons, New York, 1984. White P.C., Analyst (London), 109 (1984) 677. Rocklin R.D., LC, 2 (1984) 588. Bond A.M., Heritage I.D., Wallace G.G. and McCormick M.J., Anal. Chem., 54 (1982) 582. Kok W.Th., Brinkman U.A.Th. and Frei R.W., J . Chromatogr., 256 (1983) 17. Buchberger W., Winsauer K. and Breitwieser Ch., Fres. Z. Anal. Chem., 31 1 (1982) 517. Sherwood G.A. and Johnson D.C., Anal. Chim. Acta. 129 (1981) 101. Rocklin R.D. and Johnson E.L.. Anal. Chem., 55 (1983) 4. Han K., Koch W.F. and Pratt K.W., Anal. Chem., 59 (1987) 731. Han K. and Koch W.F., A w l . Chem., 59 (1987) 1016. Lamhelle J.H. and Johnson D.C., Anal. Chem., 50 (1978) 240. Gunasingham H. and Fleet B., Anal. Chem., 55 (1983) 1409. Stulik K. and Pacakova V., CRC Crit. Rev. Anal. Chem., 14 (1982) 297. Stulik K. and Pacakova V., Die Nahrwtg, 5 (1985) 501. Kunter W. and Kemula W., Chromurographia, 17 (1983) 322. Slais K., J . Chromutogr., 436 (1988) 413. Lookabaugh M., Krull I.S. and LaCourse W.R., J. Chromurogr.,387 (1987) 301. Kawanishi T., Togawa T., Ishigami A., Tanabe S.and Imanari T., Bunseki Kagaku, 33 (1984) E295. Rocklin R.D., in Turnski V. (Ed.), Formaldehyde: Analytical Chemistry and Toxicology, American Chemical Society Advances in Chemistry Series, 1985, p. 210. Lawrence J.F. and Charbonneau C.F., J . Chromatogr., 403 (1987) 379. Butler E.C.V., J. Chromatogr., 450 (1988) 353. Hu Z. and Tang Y., Analyst (London), 113 (1988) 179. Pyen G.S. and Erdmann D.E., Anal. Chim. Acta, 149 (1983) 355. Waters Ion Brief No. 88106. Mayer W.J., McCarthy J.P. and Greenberg M.S., J. Chromtogr. Sci., 17 (1979) 656.

Electrochemical Detection 64 65 66 67 68 69 70 71 72 73 74

32 1

Koch W.F., J . Res. Nur. Bur. Srd., 88 (1983) 157. Bond A.M., Knight R.W., Reust J.B., Tucker D.J. and Wallace G.G., Anal. Chim. Actu, 182 (1986) 47. Yang X.-H. and Zhang H., J . Chromutogr.,436 (1988) 107. Itoh K. and Sunahara H., Bunseki Kuguku, 37 (1988) 292. Mingjia W., Pacakova V., Stulik K. and Sacchetto G.A., J. Chromurogr., 439 (1988) 363. Lookabaugh M. and h l l IS., J . Chromurogr.,452 (1988) 295. Woo D.J.and Benson J.R., LC, 1 (1983) 238. Kim H.-J. and Kim Y.-K., J . Food Sci., 51 (1986) 1360. Goodwin L.R., Francom D.,Urso A. and Dieken F.P., Anal. Chem,. 60 (1988) 216. Cox D.. Jandik P. and Jones W., Pulp Pap. Conado, 88 (1987) "3 18. Nadkami R.A. and Brewer J.M., Am. Lob., 19 (1987) 50.


323

Chapter 11 Potentiometric Detection 11.1 INTRODUCTION

Potentiometry is the process in which potential changes at an indicator electrode are measured with respect to a reference electrode, under conditions of constant current (usually zero) flow. The potential of the indicator electrode varies with the concentration of a particular ion (or ions) in the solution contacting the electrode and thereby provides a means for determining ion concentrations. Potentiometric detection in IC is attractive because it has the following characteristics: Simplicity of the cell design, the measurement technique used, and the signal handling necessary. Measurements can be made without interference from dissolved oxygen. When certain types of indicator electrodes are used, the electrode potential is not greatly influenced by the eluent flow-rate. Potentiometric detection functions best when the eluent has a moderate and constant ionic strength, so it is particularly well-suited to IC applications. The indicator electrode does not participate in electrolysis reactions which could result in surface contamination of the electrode. The ohmic resistance existing between the indicator and reference electrodes is not critical. The latter factor permits the reference electrode to be placed remotely from the indicator electrode, provided that electrical contact between the two is maintained via the flowing solution. Potentiometry has been applied widely to the determination of ionic species (particularly inorganic anions) in aqueous solution, generally through the use of ionselective electrodes (ISEs), and for a number of years represented the most attractive approach to this type of analysis. This period saw intensive development of ISEs, which resulted in improvements to their detection sensitivity and also to their selectivity. Whilst high selectivity is essential for the analysis of solutions containing interfering species, and also for potentiometric detection in flow-injection analysis applications, the resulting response to only a limited number of solutes can often represent a disadvantage when potentiometric detection is coupled with a chromatographic separation technique. In such cases, it is desirable that the detector show more general response so that it can be applied to the detection of a wider range of solutes.

Chapter I I

324

Despite the advantages listed above, the usage of potentiometric detection in IC has been somewhat limited. This can be attributed to the moderate sensitivity of the technique and to the problem of high detection selectivity discussed above. Further drawbacks existing with some indicator electrodes in flowing solutions are slow response and poor baseline stability. This instability arises because the electrode potential is poorly defined under conditions where the concentration of the active solute ion is zero; that is, when the eluent alone is in contact with the indicator electrode. It is therefore sometimes necessary to stabilize the baseline electrode potential by addition to the mobile phase of a low, constant concentration of the active solute ion.

11.2 PRINCIPLES OF POTENTIOMETRIC DETECTION 11.2.1 General response equation The response of an indicator electrode is given by the Nernst equation:

E = Const +

(

'e3:FT)

log x

(11.1)

where E is the potential of the indicator electrode in volts, R, T and F have their usual meanings and X is the activity of the solute ion sensed by the electrode. The symbol n refers to the equivalents of electrons per mole of reaction (when a redox couple is involved), or the equivalents of charge per mole of analyte for a membrane electrode. When an inert indicator electrode is used to monitor a redox reaction, the same equation applies except that X becomes the ratio of chemical activities of the oxidized to reduced forms of the redox couple. The potential change (AE) accompanying a change in the activity of the solute ion from XI to Xp is given by: (11.2)

Eqn. (1 1.2) implies that a logarithmic electrode response profile results as the activity of the solute ion is increased. This is sometimes seen as a disadvantage of potentiometric detection, but as will be shown in the next Section, linear response can also occur in the low concentration ranges typically encountered in chromatographic methods of analysis.

11.2.2 Instrumental considerations The instrumentation required for potentiometry consists of a measurement cell comprising the indicator (variable potential) and reference (constant potential) electrodes, together with suitable electronics for accurate determination of the potential of the cell. Fig. 1 1. I shows a schematic representation of this arrangement. The prime requirement is that the current flow through the indicator electrode is kept close to zero,

Fi$

Porenriometric Detection

325

Power supply

1-'

R e f e r e nce electrode

Indicator electrode

Fig. 11.1 Potentiometry using a high impedance voltmeter for potential measurement. Reprinted from [ 13 with permission.

since any current flow will alter the electrode potential. For this reason, high impedance voltmeters are used so that the current flow is very small (typically amp) and the measured voltage is within the experimental error of the true potential of the system.

11.3 INDICATOR ELECTRODES AND RESPONSE PROFILES 11.3.1

Ion-selective indicator electrodes

Both solid-state and membrane type ion-selective electrodes have been employed as indicator electrodes in IC. The solid-state type has the advantage of rapid response in swiftly flowing streams in which the width of the diffusion layer at the electrode surface is minimal, but the selectivity is often too high for chromatographic applications. For this reason, potentiometric detection with solid-state electrodes finds most use when it is used in conjunction with a more general detection method, such as conductivity. On the other hand, membrane electrodes are less selective and are therefore more suited to chromatographic detection, but give slower response and may show marked dependence on eluent flow-rate. The response time of a membrane electrode has been shown to be improved by coating the membrane surface with a Nafion ion-exchange membrane, but this process adversely affected the sensitivity and linearity of response 121. Calibration plots of electrode potential versus analyte concentration show either a linear or logarithmic relationship, depending on the concentration range studied. Under conditions where a background concentration of analyte is added to the eluent stream in order to stabilize the baseline potential, the detector response is usually Nernstian and follows eqn. (1 1.2). In this case, X1 would be equal to the background concentration of analyte in the eluent. However, when theltotal analyte concentration at the electrode surface is very low, a linear relationship dependence on electrode potential may be observed [3-61. That is, the detector is operating in the sub-Nernstian region.

Chapter I I

326

p H electrodes

Glass electrcdes for the measurement of pH can be applied directly to the detection of organic acids 171 but are also useful for the indirect detection of common anions. We have already noted (Section 9.5) that most solute anions in suppressed IC are eluted as fully dissociated strong acids. This phenomenon was utilized in conductivity detection as a means of enhancing the detection signal for such ions. It is also apparent that a decrease in the pH of the eluent will occur as a result of the hydrogen ions introduced into the sample band by the suppressor. This pH change can be utilized for indirect detection of anions using potentiometric detection with a pH indicator electrode [8-10]. A chromatogram of common anions obtained using pH detection was shown in Fig, 10.5(a). The magnitude of the pH change is dependent on the eluent used. A typical carbonate-based eluent comprising 2.4 mM Na2C03 and 3 mM NaHC03 gives 5.4 mM H2CO3 when fully suppressed, resulting in a background eluent pH of 4.28. An analyte band composed of 1.5 ppm (42 pM) CI- gives 42 pM HCI after suppression, which results in a pH of 4.11 at the elution peak [9]: This gives a pH change of 0.17 pH units. On the other hand, a KOH eluent which is converted to water i n the suppressor, gives a pH change of 2.62 pH units for 1.5 ppm of C1-. Some representative limits of detection using pH measurements are 2.8 ppb for CI- and 128 ppb for NO3-, and it has been demonstrated that this form of detection generally gives detection limits which are superior to conductivity detection when a polymer pH electrode is used [lo]. The response equation for pH detection can be derived as follows [9]. If the background pH of the eluent is x, where the acidity is contributed solely by penetration of a strong acid regenerant through the suppressor, the background [H+] is M. An analyte band containing y M of a monovalent anion at the peak apex will have [H+]equal to + y) M, giving a pH of -[log(lOx + y)]. The pH change during sample elution is given by the difference between the background pH and the pH of the analyte band.

Eqn. ( 1 1.3) can be rearranged to give:

loz = 1 + 10Xy

(11.4)

Fig. 11.2 shows some calibration plots for pH detection of common anions, obtained using KOH or Na2C03 eluents. A plot of 10” versus y for the chloride data in Fig. 11.2 is linear and has an intercept of 1.023, which is in agreement with eqn. (1 1.4). We can also note that pH detection in suppressed IC is applicable to the detection of cations which are eluted with an acidic eluent and passed through a suppressor in the OHform. In this case, the suppressor contributes OH- to both the eluent and the sample, so that an increase in pH accompanies the elution of solute cations. The principles of this form of detection are identical to those described above for anions and once again, the detection limits obtained with a polymer pH electrode are generally superior to those for conductivity detection [lo].

PotentiometricDetection

321

x. .1

0.3 7 5

-

h

u)

.-

L

C

0.300 -

Anion concentration,p.M

Fig. 12.2 Calibration plots for anions in suppressed IC using pH detection. The solid lines are for a 30 mM KOH eluent, whilst the broken lines are for a 5 mM Na2C03 eluent. (1) C1-, (2) so42-,(3) NOj. Reprinted from [9] with permission.

Solid-state ion-selective electrodes Solid-state, or crystalline. ISEs use a conducting solid material as the ion-selective barrier between an internal filling solution and the external, sample solution. The most common examples of solid-state electrodes are the fluoride electrode (which uses a LaF3 crystal as the ion-selective barrier) and halide electrodes based on mixtures of AgzS and silver halides. Solid-state ISEs have been employed in IC for the detection of F- [11- 141, Bf 113, 151, I- 13, 151, C1- [15] and SCN- 1151. A solid-state copper ISE has been used for the detection of non-UV absorbing amino acids after post-column addition of 0.1 mM Cu(I1) in acetate buffer 1161. Here, reaction with the eluted amino acids caused a reduction in the concentration of Cu(I1) reaching the electrode, leading to a change in potential and hence indirect detection. At low values of amino acid concentration and for high values of the stability constant for the copper-amino acid reaction, it can be shown that the theoretical potential change of the electrode is linearly proportional to the total amino acid concentration. This relationship was found to be valid in practice, provided formation of the 1:2 copperamino acid complex was negligible. Liquid membrane electrodes ISEs of the liquid membrane type have a membrane separating the internal filling solution from the external analyte solution. The membrane consists of an organic liquid (immiscible with water) held on a porous plastic support. The organic liquid is often an

328

Chapter 11

ion-exchange material. As discussed earlier, these electrodes are generally less selective than the solid-state type and can therefore be used for the chromatographicdetection of a wider range of analyte ions. Commercially available liquid membrane ISEs have been used for the detection of N02- and NO3- [17] and CN- [2] after separation by IC. Home-made electrodes containing valinomycin. benzo- 15-crown-5, nonactin or tetranactin as neutral carrier ligands in a PVC membrane have been used for the detection of alkali metal cations and NH4+1181. 11.3.2.

Coated-wire indicator electrodes

Some very useful potentiomemc detectors have been developed using coated-wires as indicator electrodes. A simple example is a silver-silver chloride electrode prepared by treating a length of silver wire with hydrochloric acid. However, any insoluble silver salt can be used as the coating material. The potential of such an indicator electrode is given by [4,191:

where Ks is the solubility product of the salt, AgX, used to coat the electrode and a,- is the activity of the analyte ion, X-. When the electrode is immersed in a flowing stream, a certain amount of the AgX salt dissolves from the surface, giving rise to a steady state concentration of Ag+ and hence a stable background potential. A new potential is established if X - is added to the flowing stream (e.g. by elution from the chromatographic column), since the concentration of free silver ions will change. The difference between the new electrode potential and the baseline value is governed by the added concentration of X-. By analogy, low concentrations of other anions which form insoluble silver salts should also produce potential changes proportional to the concentrations in which they are added. Thus, a potentiometric detector of this type should be capable of detecting anions such as halides, SCN-and S2032-.

Coating materials A range of coating materials, including AgCI, AgBr, AgI, Ag3P04, Ag2S and AgSCN, have been examined for the preparation of coated silver wire electrodes [20]. When an eluent of sodium perchlorate was used, the AgCl and AgSCN coated electrodes gave optimum response and reproducibility. The coatings were produced by electrical oxidation of a silver wire in an aqueous solution of the selected anion for a period of 3-7 min. Electron microscopy of the coating showed a wide variation in particle size for the precipitated layer and with the exception of silver sulfide, coatings of smaller particle sizes were observed to have faster response kinetics in flowing solution. It was also found that newly prepared electrodes required conditioning by repeated immersion (and rinsing with water) in a solution containing 1 mM concentrations of each of the ions to be detected. When this conditioning process had been performed, the electrode surface was a composite of many silver salts covering the underlying layer of silver chloride precipitate.

Potentiometric Detection

329

Calibration and response characteristics The response of coated-wire indicator electrodes will again be dependent on the concentration of analyte injected and will also be influenced by the nature of the eluent used. A silver-silver chloride electrode used with an acetate eluent has been found to give a linear relationship between electrode potential and analyte concentration for chloride in the sub-ppm range [4]. Hershcovitz et al. [ 191 observed Nemstian response with a silver-silver salicylate electrode for several anions in the 0.1-1 mM concentration range, after separation with a salicylate eluent. On the other hand, Lockridge et al. [20] found the response of a silver-silver chloride electrode in a perchlorate eluent to be neither linear nor logarithmic. However, curvature of the logarithmic plot was sufficiently slight that the calibration was still useful for analytical purposes. Calibration plots obtained with coated-wire electrodes are shown in Fig. 11.3. Potentiometric detectors based on coated-wire electrodes show peak shape characteristics which are dependent on the re-equilibration kinetics of the precipitate film existing at the electrode surface. The response profiles observed for these electrodes are also influenced by consumption of a small, but significant, proportion of the analyte as a necessary part of the detection process. This effect may be an explanation for the fact that response with these electrodes can vary from linear to logarithmic, with unexplained effects occumng most often at low concentration levels. Mass-transfer processes and surface kinetics therefore assume an important role for indicator electrodes which are based on insoluble silver salts, particularly when largearea electrodes are used in conjunction with microbore columns, and these factors merit further investigation.

11.3.3.

Metallic copper indicator electrode

Metallic copper in the form of a wire or tube has been employed as an indicator electrode for IC and flow-injection methods [21-321. When a metallic copper electrode is placed in a flowing stream of an oxygenated, buffered eluent, copper ions (both cuprous and cupric) are formed at the electrode surface. Considering only the cupric ions and assuming that the eluent buffer contains a species B which complexes copper (11), the potential of the electrode may be described by the equation [231: (11.6)

where CCu11 is the total cupric ion concentration at the electrode surface and q u ~ ~ ( ~ , is the side-reaction coefficient for the binding of cupric ions by component B and hydroxyl ion. If a copper complexing ligand, L, now passes the electrode as an eluted peak, the new electrode potential is given by: (11.7)

where

330

L.

70

e

0

e

5

Chapter I I

Potentiometric Detection

331

and the a values describe the side reaction coefficients for Cu(1I) with the indicated components. Under conditions where the complexation of cupric ions by the added ligand is much stronger than by the buffer component and hydroxyl ions, and a 1:l complex is assumed between Cu(I1) and L, then for constant pH and constant composition of the eluent it can be shown that the difference between El and 5 (i.e. the peak height, H, in volts) is given by:

E2 - El = H = Const,

+RT log 2F

(-)PlCL D~L(H)

(11.9)

where is the stability constant for the Cu(1I)L complex, CL is the total concentration of ligand L, D is the dispersion factor and CXL(H) is the side-reaction coefficient for ligand protonation. A similar equation may be derived for complexation of the ligand with cuprous ions. Thus, if a single, stable compound (either a soluble complex or a precipitate) is formed by the solute ligand L with cuprous or cupric ions, a logarithmic relationship with Nernstian slope can be expected between the peak height H and CL (or N, the total number of moles of solute injected). When several complexes are formed in comparable amounts, then a more complicated relationship can be predicted. Similarly, the above Nernstian relationship will not be followed for very small concentrations of injected ligand, and it can be shown that under these conditions the peak height is given by [301:

Regardless of the response relationship applicable, the electrode potential is dependent on the concentration of copper ions which is, in turn, governed by a number of factors which are either constant over the period of the analysis or show variation. Factors of the former type include the oxygen content of the eluent and the flow-rate, whilst the chief variable factor is the changing concentration of the solute species.

Modes of operation of a metallic copper indicator electrode A metallic copper indicator electrode can operate in a variety of detection modes and these are summarized below: If the eluted solute forms a more stable complex with copper ions than does the eluent ion, a local decrease in copper ion concentration will occur at the electrode surface, leading to a decrease in the electrode potential. (ii) If the eluted solute forms a less stable complex with copper ions than does the eluent ion, a local increase in the copper ion concentration will occur at the

(i)

Chapter I1

332 N , nmol 6;

1

-

--

2

0.2

0.4

Succinate

10

Y

m

m

0

1

EI

5

0.5

10

Fig. Z2.4 Calibration plots obtained with a metallic copper indicator electrode operated in (a) direct and (b) indirect detection modes. Reprinted from [33] with permission.

electrode surface, leading to an increase in the electrode potential. This process assumes that an ion-exchange separation is being employed, so that solute ions replace an equivalent number of eluent ions in the mobile phase at the time of elution. (iii) If the eluted solutes are strong oxidants and are able to oxidize the surface of the metallic copper electrode, a local increase in copper ion concentration will occur and the electrode potential will increase. Reducing solutes can also be detected by the reduction of cupric ions to cuprous and this will also result in an increase of the electrode potential due to the higher standard electrode potential of the Cu+/Cuocouple (4.520 V) compared to the Cu2+/Cu0couple (4.337 V). A potentiometric detector employing a metallic copper indicator electrode can therefore function i n a direct mode, wherein only copper complexing ions are detected, or in an indirect mode in which only the eluent needs to show copper complexation properties. Fig. 11.4 shows typical calibration plots for direct and indirect detection. A further consideration is that the direction of the measured peak can be used as a further means of identification of the eluted solute. This type of detector is the most widely applicable potentiometric detector in IC and is influenced by analyte consumption or surface kinetic effects to a lesser degree than other indicator electrodes, such as those based on insoluble silver salts.

Potentiometnk Detection

333

+

Eluent in

n

Eluent in Eluent out (a 1 OUT t"FLAlRFIT" 1

f

IN ...

+J

Eluent out (b)

-

LKrn$!!

Fig. ZZ.5 Typical flow-cell designs for cylindrical ISEs. (a) flow-cap; (b) and (c) electrode housing types. Reprinted from [3, 11, 171 with permission.

11.4 FLOW-CELLS FOR POTENTIOMETRIC DETECTION To enable successful potentiometric detection to be achieved, it is necessary that the indicator electrode (and often the reference electrode as well) be incorporated into a suitable flow-cell. For the maintenance of chromatographic efficiency, it is essential to minimize dispersion of the solute by placing the flow-cell close to the column outlet, by reducing flow turbulence in the cell, and by ensuring that the internal volume of the cell is as low as possible.

11.4.1

Flow-cells for cylindrical ISEs

When cylindrical ISEs are used as indicator electrodes, it is possible to obtain reliable potentiometric readiGgs merely by allowing the column effluent to flow under gravity over the sensing membrane of an inclined electrode [9]. However, it is generally more convenient to use a flow-cell of some kind. The simplest flow-cell configuration is a flow-cap designed to fit over the end of the electrode. A typical flow-cap is illustrated in Fig. 11.5(a) and in the design shown, a small cavity of 5 pl volume, drilled into the

Chapter I 1

334

Eluent

Teflon spacer Fig. 11.6 Typical flow-cell designs for wire indicator electrodes. (a) copper wire indicator electrode (l), silver-silver chloride reference electrode (2), agar gel (3); (b) cell for silver-silver chloride coated wire indicator electrode. Reprinted from [20,2l]with permission.

inside bottom of the cap, serves as the detection chamber [ 171. Most efficient operation of the electrode was achieved when the inlet stream was directed vertically onto the electrode surface and the entrapment of air bubbles was prevented when the outlet tube was orientated at 45O. When flow-caps are used, the distance between the indicator and reference electrodes is generally large and electrical contact between the electrodes can be enhanced by inserting platinum wire into the connecting tubing 1161. Cylindrical indicator electrodes can also be housed in a variety of alternative flow-cells and two suitable designs are shown in Figs. 1 1 S(b) and (c). Both cells are of very low volume (6-10 pi), provide for high linear flow velocities over the electrode surface to promote rapid electrode response [3], and permit the indicator and reference electrodes to be placed in close proximity to each other. 11.4.2

Other flow-cells

Wire indicator electrodes are more easily accommodated into a flow-cell than cylindrical electrodes by virtue of their smaller size, and the design of the flow-cell does


335

Iodide feed solution out

Eluent inlet

+

4

Iodide feed

solution in

Fig. 11.7 Concentric-flow diffusion cell for post-column addition of an electroactive species to the mobile phase. (1) external feed solution; (2) collars from heat-shrinkable plastic; (3) 1.6 mm OD polyethylene tubing transporting the eluent; (4) hollow-fibre dialysis tubing. Reprinted from [3] with permission. not appear to exert a major influence on electrode performance. Fig. 11.6 shows two flow-cells which are representative of published designs. The simple design in Fig. 11.6(a) has low volume (4 pl) and permits easy removal and cleaning of the metallic copper wire indicator electrode [21], whilst the flow-cell shown in Fig. 11.6(b) [20] is more suited to indicator electrodes which do not require frequent treatment. A membrane cell has been reported for potentiometric detection in IC [34]. This cell consists of an ion-exchange membrane which separates two chambers, each of which contains a reference electrode. The column effluent passes through one chamber, whilst eluent is pumped through the second chamber. When a solute ion is eluted into one chamber of the cell, and provided that the concentration of the solute is very small compared to that of the eluent, then the potential of the cell changes linearly with solute concentration. As discussed previously, it is often advantageous if a low background level of an electroactive solute is added to the eluent in order to give a stable baseline electrode potential. Whilst this can be achieved by adding the solute ion to the eluent, such an approach has a number of disadvantages. Samples with levels of the solute less than that present in the eluent will produce negative detector signals, whilst those with concentrations similar to this level cannot be detected at all. An alternative method is to add the desired ion to the eluent after the separation column. A concentric flow diffusion cell has been described 131 for this purpose, wherein addition of I- is required. This cell contains a length of hollow-fibre dialysis tubing, through which the column effluent is passed, whilst eluent containing 0.1 mM I- is circulated around the exterior of the tubing with the aid of a peristaltic pump (Fig. 11.7). Provided pressure fluctuations from the eluent pump are eliminated with a suitable pulse dampener, the rate of diffusion of I- into the column effluent is constant.

Chapter I I

I'

0

2

4

6

Time (min)

Fig. 11.8 Gradient elution with ptentiometric detection using a silver-silver chloride coated wire indicator eleclrode. Eluent: 3.5-10.0 mM sodium perchlorate at 1.6 ml/min; injection volume, 20 pl; analyte concentrations, 1.0 mM. Reprinted from [u)]with permission.

11.5 APPLICATIONS OF POTENTIOMETRIC DETECTION IN IC 11.5.1

Halides and pseudohalides

One of the more commonly encountered applications of potentiometric detection in IC involves the use of a solid state electrode for the detection of F-. This species is eluted early in a chromatogram under eluent conditions suitable for the separation of common anions, such as halides, N O i . NO3-, Pod3- and S042-. In many cases, it is difficult to distinguish the F peak from the solvent front eluted at the column void volume and this provides strong justification for coupling potentiometric detection for Fwith a general purpose detection mode, such as conductivity [ l l , 131. Use of an iodide ISE for the analysis of I- in seawater has also been reported [3]. Silver wire electrodes coated with insoluble silver salts have been applied to the detection of halide and pseudohalide (SCN-and S2O3*-) ions. A silver salicylate coating

Potentiomem*cDetection

337

coupled with the use of a salicylate eluent for the ion-exchange separation has been suggested to provide a stable baseline potential [ 191, whereas other authors [4,201 have found that a silver chloride coating is suitable when the eluent used does not form an insoluble silver salt. Under the latter conditions, the baseline potential of the electrode is not dependent on eluent concentration and this leads to the possibility of gradient elution with potentiometric detection. Fig. 11.8 shows a chromatogram obtained using a silversilver chloride indicator electrode with a linear concentration gradient of sodium perchlorate in the eluent. 11.5.2

Weak acid anions

Problems are often encountered with the conductivity detection of anions of weak acids because these species are weakly retained by anion-exchangeunless the eluent pH is high, and they become partially or fully protonated after passage through a suppressor. For these reasons, potentiometric detection is attractive and the use of a silver sulfide membrane electrode has been reported for the detection of CN- and S2- [2]. No interference from Br', I- and SCN- was observed, provided that chromatographic conditions were chosen so that these ions did not co-elute with the analyte ions. A glass membrane electrode has been employed for the detection of carboxylic acids after ionexchange separation [7] and a liquid-membrane electrode of low selectivity has been successfully used for the determination of NO2- and NO3-, as well as phthalate isomers [171. 11.5.3

Cations

A homemade PVC matrix membrane electrode has been reported for the detection of monovalent cations [18]. The polymer coating contained small quantities of active ligands, such as valinomycin. A calcium liquid-membrane ISE has been applied to the determination of calcium using measurements of conductance rather than potential [35]. An instrument using the bipolar pulse method was employed for conductance measurements and whilst the sensitivity of the electrode under these conditions was similar to that obtained when the electrode was operated in the potentiometric mode, the electrode response time was dramatically shorter for the conductance mode. The times required for the electrode to reach 90% of total response were 10 ms and 4-5 s for the conductance and potentiometric modes, respectively. It is therefore likely that conductance measurements using ISEs could find wider application to IC analysis. 11.5.4

Applications of the metallic copper indicator electrode

Direct detection A metallic copper indicator electrode housed in the flow-cell shown in Fig. 11.6(a) has been widely applied in IC,. Direct detection of copper complexing solutes eluted with weakly complexing eluent ions has been reported for carboxylic acids, halides and amino acids using eluents such as tartrate, phosphate and phthalate, in both ion-exchange and ion-exclusion separation systems [33]. Fig. 1 1.9 shows chromatograms for direct potentiometric detection after ion-exchange separation. Similarly, direct detection of

338

Chaprer 11

EC"

I

CI

1

1

-

1

I

I

Time, min Fig. 11.9 Chromatogram of species exhibiting direct response with a metallic copper indicator electrode. Column: Vydac 302 IC 4.6. Eluent: (a) 1 mM sodium tartrate at pH 3.2, (b) 1 mM potassium orthophosphate at pH 7.0. Injected amounts: (a) 0.5-50 nmol. (b) 5 nmol of each species. Reprinted from [33] with permission.

oxidizing anions (IO3-, BrO3- and ClO3-) and reducing species (ascorbic acid, hydrazine and hydroxylamine) has been achieved [32, 331. Fig. 11.10 illustrates the direct potentiometric detection of oxidizing solutes. The high degree of selectivity offered by direct detection has proven to be advantageous in the determination of oxalate in urine, where differentiation of oxalate from closely eluting excess sulfate is required. Use of the potentiometric detector enabled sample preparation to be confined to simple dilution and no interference from sulfate was observed [31].

Indirect detection Indirect detection with a metallic copper indicator electrode can be applied to inorganic anions and some weakly complexing ciirboxylic acids, as well as alkaline earth and transition metal ions. provided that the eluent used for the ion-exchange separation shows appreciable copper complexation ability. Thus NOz-, NO3- and Sod2- are detectable in a phthalate eluent [29], and alkaline earth and transition metal ions are detectable in an eluent comprising ethylenediamine and a complexing agent such as tartrate, citrate or oxalate 1301. Fig. 11 .I 1 shows chromatograms obtained with indirect potentiometric detection. Post-column addition of Cu(11) has been employed for the indirect potentiometric detection of reducing carbohydrates, using a metallic copper electrode and an ion-exchange column 1251. The ubiquitous complexation characteristics


339

I

EC"

I

4 I

I

0

5

Time, min Fig. 22.20 Chromatogram of oxidizing anions detected at a metallic copper indicator electrode. Column: Vydac 302 IC 4.6. Eluent: 20 mM sodium tartrate at pH 3.2. Injected amounts: 1-100 nmol. Reprinted from 1331with permission.

ECU

1

NO;

I

I

I

I

I

L

0

5

10

0

5

10

Timc,min

Fig. 11.11 Chromatogranis of inorganic and organic anions exhibiting indirect potentiometric response with a metallic copper electrode. Column: Vydac 302 IC 4.6. Eluent: 2 m M potassium phthalate at pH 4.6 (a) or 4.0 (b). Injected amounts: (a) 60-120 nmol, (b) 250 nmol of each species. Reprinted from [33] with permission.

Chapter I1

340

of copper enable a wide range of eluents to be used with the indirect detection mode, which suggests that this form of detection could be applied to almost any ion which can be separated by IC. Table 1 1.1 summarizes some of the published applications of potentiometric detection in IC. TABLE 11.1 APPLICATIONS OF POTENTIOMETRIC DETECTION IN IC Species separated

Indicator electrode

Separation mode

Detection limitsa

Ref.

Alkaline eanh ions Amino acids Amino acids, diamines Ascorbic acid, N2H.4, NH2OH BrCarboxylic acids Carboxylic acids Carboxylic acids CN-, S2-

Metallic copper Metallicqpr Copper - ISE Metallic copper

Ioncxchange RWd-PhaSe Reversed-phase Ionexchange

1 PPm 75-300 ng 10-100 ng 2-10 nmol

27 36 16 32

Hg~Br2- ISE Metallic copper Metalliccopper H+ - glass AgzS - ISE Fluoride - ISE Ag - AgCl AgzS - ISE Ag - Ag salicylate Ag - A@ Iodide - ISE Lead phosphate glasses H+- glass H+ - polymer Metallic copper

Ionexchange Ionexchange Ion-exclusion Ionexc hange Ion-exchange Ionexchange Ionexc hange Ioncxchange Ionexchange Ionexchange Ion-exchange Ionexchange

20 PPb 20-50 nmol 0.2-5.3 C(g 1 pquiv 10 PPb 10-20 ppb 0.5 ppm 10 pM 0.2 ppm

0.05 mM 0.015 pM 50 PPb

13 28 31 7 2 11-14 4 15 19 20.37 3 38

Ion-exchange Ionexchange Ionexchange

0.08-2.0 pM 1.1-6.5 pM 0.02- 10 nmol

9 10 29

PVC membrane H+ - polymer Non-selective membrane cell Nitrate - ISE Metallic copper Metalliccopper Metallic *per

Ionexchange Ionexchange Ion-exchange, ion-interaction Ionexchange Ioncxchange Ionexchange Ion-exchange

1 w 4.7-7.8 pM 10 nmol

18 10 34

0.1-0.3 nmol 1 PPm 1-5 nmol 1-10 nmol

17 26,31 25 24.30

F Halides Halides, SCNHalides, SCNHalides, SCN', Sfl3'IIInorganic anions Inorganic anions Inorganic anions, IO3', B e - , ClO3Monovalent cations Monovalent cations Na+, Li+, F,P043-, CH3COO', Sod2N@-,NO3oxalate

Reducing carbohydrates Transition metal ions a

Detectionlimits are expressedik the same units as used in the original publication.


341

11.6 REFERENCES 1

2 3 4 5 6 7 8 9 10 11

Kennedy J.H.. Analytical Chenhtry Principles,Harcourt Brace Jovanovich, San Diego, 1984, p. 582. Wang W., Chen Y. and Wu M.X, Analyst (London), 109 (1984) 281. Butler E.C.V. and Gershey R. M., Anal. Chim.Acta, 164 (1984) 153. Franks M.C. and Pullen D.L., Analyst (London), 99 (1974) 503. Roehse W., Roewer G.,Boran R. and Hellmig R.. Z. Chem., 22 (1982) 226. Trojanowicz M. and Matuszewski W., Anal. Chim.Acta, 151 (1983) 77. Egashira S., J. Chromatogr., 202 (1980) 37. Tarter J.G., J. Liq. Chromatogr.,7 (1984) 1559. Shintani H. and Dasgupta P.K., Anal. Chem., 59 (1987) 802. Meyerhoff M.E. and Trojanowicz M., Anal. Chem., 61 (1989) 787. Keuken M.P., Slanina J., Jongejan P.A.C. and Bakker F.P., J. Chromarogr.,439 (1988) 13.

12

13

Speitel L.C., Spurgeon J.C. and Filipczak R.A.. in Sawicki E. and Mulik J.D. (Eds.), Zon ChromatographicAnalysis of Environmental Pollutants,Vol. II, Ann Arbor Sci. Publishers, Ann Arbor, MI, 1979, p. 75. Slanina J., Bakker F.P., Jongejan P.A.C., Van Laxnoen L. and Mols J.J., Anal. Chim. Acta, 130 (1981) 1.

14 15 16 17 18 19 20

Talasek R.T., J . Chromatogr.,465 (1989) 1. Muller H. and Scholz R., Ion-SelectiveElectrodes,ACS Anal. Chem. Symp. Series, Vol. 22, 1984, p. 4. Loscombe C.R.. Cox G.B. and Dalziel J.A.W., J. Chromatogr., 166 (1978) 403. Schultz F.A. and Mathis D.E., Anal. Chem., 46 (1974) 2253. Suzuki K.,Aruga H. and Shirai T.. Anal. Chem., 55 (1983) 2011. Hershcovitz, H., Yarnitzky C. and SchmucklerG.,J. Chromatogr.,252 (1982) 113. Lockridge J.E., Fortier N.E., Schmuckler G. and Fritz J.S., Anal. Chim.Acta. 192 (1987) 41.

27 28 29 30 31

Alexander P.W., Trojanowicz M. and Haddad P.R., Anal. Lett., 17 (1984) 309. Alexander P.W., Haddad P.R. and Tmjanowicz M., Anal. Chim. Acta, 177 (1985) 151. Alexander P.W., Haddad P.R. and Trojanowicz M., Anal. Chem., 56 (1984) 2417. Alexander P.W.,Haddad P.R. and Tmjanowicz M., Anal. Chim.Acta, 177 (1985) 183. Cowie C.E., Haddad P.R. and Alexander P.W., Chromatographia,21 (1986) 417. Croft M.Y. and Haddad P.R., Australian Association of Clinical Biochemists Monograph Series, 1986, p. 138. Haddad P.R., Alexander P.W. and Trojanowicz M.. J. Chromatogr.,294 (1984) 397. Haddad P.R., AIexander P.W. and Trojanowicz M., J. Chromatogr.,315 (1984) 261. Haddad P.R.. Alexander P.W.and Trojanowicz M., J. Chromatogr.,321 (1985) 363. Haddad P.R., Alexander P.W. and Trojanowicz M., J . Chromatogr.,324 (1985) 319. Haddad P.R., Alexander P.W., Croft M.Y. and Hilton D.F., Chromatographia,24 (1987)

32 33 34

Haddad P.R., Alexander P.W. and Trojanowicz M., J. Liq. Chromatogr.,9 (1986) 777. Alexander P.W., Haddad P.R. and Trojanowicz M.. Chromatographia,20 (1985) 179. Deelder R.S.. Linssen H.A.J., Koen J.G. and Beeren A.J.B., J. Chromatogr., 203 (1981)

35

Powley C.R., Geiger R.F., Jr. and Nieman T.A., Anal. Chem.,52 (1980) 705.

21 22 23 24 25 26

487.

153.

342 36 37

38

Chapter 11

Alexander P.W., Haddad P.R.,Low G.K.-C. and Maitra C., J . Chromutogr.. 209 (1981) 29. Suzuki K., Aruga H., Ishiwada H., Oshirna T., Inoue H. and Shirai T., Bunseki Kaguku, 32 (1983) 585. Nornura T., Hikichi Y. and Nakagawa G., Bull. Chem. Soc. Jup., 61 (1988) 2993.

343

Chapter 12 Spectroscopic Detection Methods 12.1 INTRODUCTION 12.1.1 Types of spectroscopic detection in IC Spectroscopic methods of detection are very commonly employed in IC and are second only to conductivity detection in their frequency of usage. These spectroscopic methods are listed schematically in Fig. 12.1, from which it can be seen that they can be divided broadly into the categories of molecular spectroscopic techniques and atomic spectroscopic techniques. Molecular spectroscopy includes UV-Visible spectrophotometry, refractive index measurements and photoluminescence techniques (fluorescence and phosphorescence). Atomic spectroscopy includes atomic emission spectroscopy (using various excitation sources) and atomic absorption spectroscopy.

12.1.2

Direct and indirect spectroscopic detection

It will become apparent throughout the discussion in this Chapter that many of the spectroscopic detection methods can operate in a direct or indirect mode. The definitions of these terms are the same as those used to describe the electrochemical detcction modes discussed in previous chapters. That is, direct spectroscopic detection results when the solute ion has a greater value of the measured detection parameter than does the eluent ion. Indirect detection results when the reverse is true.

SPECTROSCOPIC DETECTION IN IC I

Molecular spectroscopic methods Spect rop h otomet ry Refractive index Photoluminescertce

I

Atomic spectroscopic methods

t

Atomic absorption Atomic emission

Fig. 12.1 Schematic representation of spectroscopic detection modes in IC.

Chapter 12

344

12.2 UV-VISIBLE SPECTROPHOTOMETRIC DETECTION IN IC METHODS USING ION-EXCHANGE SEPARATIONS 12.2.1

Detection response equations for ion-exchange

We will begin by illustrating the operating principles of UV-Vis spectrophotometric detection using anion-exchange chromatography as a typical example. Equations will be developed for the absorbance of an eluent prior to and during the elution of a solute ion and from these, a detection response equation will be derived. The general approach used for this derivation has been described previously [l-31. Consider an anion-exchange system in which the eluent consists of the species HE, which is dissociated partially into H+and E-.If the degree of dissociation of the eluent is IE and the total eluent concentration is CE,then:

(12.2)

The background absorbance of an eluent flowing through a fully equilibrated column will arise from both HE and E . In this example, the eluent cation (H+) does not absorb and we will assume this to be true in all cases. The background absorbance can be calculated from Beer’s Law to be:

where EE-is the molar absorptivity of the eluent anion, &HE is the molar absorptivity of the undissociated eluent, and m is the path-length of the detector cell. The elution of a solute S, which is dissociated to a degree Is to form S-,will result in the displacement of an equimolar amount of E-from the eluent (see Fig. 9.1 for a schematic representation of this process). This solute will be eluted as a Gaussian peak, with a concentration of Cs- at the peak maximum. The concentration of E-at the peak maximum is therefore given by:

The absorbance measured at the peak maximum is then given by:

where ES- and ENS are the molar absorptivities of the dissociated and undissociated forms of the solute, respectively. The change in absorbance (AA) accompanying the elution of the solute can now be obtained by subtracting the background absorbance (eqn. (12.3)) from the absorbance during sample elution (eqn. (12.5)) to give:

Spectroscopic DetectionMethods

345

Solute

(ES)

Eluent 2 ( E E 2 ) Fig. I22 Schematic representation of direct and indirect spectrophotomeaicdetection in IC.

Under conditions where the solute is fully dissociated (i.e. Is = 1). which is the usual case for ion-exchange separation, eqn. (12.6) can be simplified to give:

AA =

(ES-

- EE-)CS~

(12.7)

We see that the absorbance change measured by the detector on elution of a sample is proportional to the solute concentration, the cell path-length and to the difference in molar absorptivities between the solute and eluent anions. Eqn. (12.7) applies to the case where both the eluent and solute anions have a single charge. This equation may be rewritten for the general case in which the solute is represented by Sx- and the eluent by Ey-. We therefore obtain [4]:

Y

(12.8)

Eqn. (12.7) shows that direct detection will result when the molar absorptivity of the solute anion exceeds that of the eluent anion, leading to a positive value for AA. Conversely, indirect detection results when the molar absorptivity of the solute anion is less than that of the eluent anion, leading to a negative value for AA. These detection modes are illustrated schematically in Fig. 12.2. Eqn. (12.8) can be rewrigen for the cation case (i.e. for a solute Sx+ and an eluent EY+), as follows: (12.9)

346

Chapter 12

TABLE 12.1

SOLUTES AND TYPICAL OPERATING CONDITIONS FOR DIRECT SPECTROPHOTOMETRIC DETEXTION IN 1C AFTER ION-EXCHANGESEPARATION Solute(s)

Eluent

Wavelength (nm)

Detection limit

Ref

HCl K2m4 N aC1 CH3S03H NaC104 Na2COflaHCO3 TridSO4’-

200

1.5 ppm 0.6 ng 50 PPb 50 PPb 50 PPb 160 PPb 50 ng 300 ng 30 PPb 3 PPb 0.2 ppm 4 PPm Nickel@) Thorium(IV)

Uranium(VI) Vanadium(IV) zinc(II> zirconiumov)

.

0

PAR

.

.

a

.. .. . 0

. .. 8

8

Indicates a positive colour reaction.

13.4.2 PCR detection of transition metals using PAR

A s indicated from Table 13.3, PAR forms complexes with a wide range of metal ions. The colour-forming reaction is rapid and so can be carried out by direct mixing of the PCR reagent and eluent in a simple mixing tee-piece, without the requirement for a delay coil. The PCR reagent consists of dilute PAR solution in an appropriate alkaline buffer (for example, 0.2 mM PAR in 2 M ammonia and 1 M ammonium acetate), and a detection wavelength in the range 500-540 nm is generally used. Some typical detection limits which can be attained under these conditions are listed in Table 13.4, which shows that sensitive detection is possible for lanthanides and transition metals, but not for calcium and magnesium 1311. The detection limits for PCR with PAR are dependent on the reaction conditions, especially the concentration of PAR used, since this affects the baseline noise. The lowest possible concentration of PAR should be employed, consistent with the concentration of metal ions in the sample. Cassidy and Elchuk [31-331 have demonstrated that trace enrichment procedures (see Section 14.6) enable the detection limits shown in Table 13.4 to be reduced substantially. The use of PAR for PCR detection of transition metals has proven successful after separation of these species by either ion-exchange or ion-interaction chromatography. A chromatogram illustrating ion-exchange separation and PCR detection was given earlier in Fig. 4.17(b), whilst Fig. 13.9 illustrates an ion-interaction separation with PCR detection using PAR. Both of these separations are quite rugged and can be applied to complicated samples without the requirement for extensive cleanup procedures. This can be illustrated by the determination of selected transition metal ions in nuclear materials [34), process liquors [35], urine [36], serum [37,381, whole blood [38J,soils


r 0

l

I

I

I

I

I

401

I

I

I

I

I

I

I

10

5

I

1

15

Time (min) Fig. 23.9 PCR detection of transition metals with PAR, after separation by ion-interaction chromatography. A Waters pBondapak c18 column was used with 50 mM tartaric acid and 2 mM

sodium octanesulfonate at pH 3.4 as eluent. The detection wavelength was 546 nm. Solute concentrations: Co (5 ppm); Ni, Cd (2 ppm); remainder (1 ppm). Chromatogram courtesy of Waters Chromatography Division. TABLE 13.4 DETECTION LIMITS ATTAINABLE FOR SOME METAL IONS USING POST-COLUMN REACI'ION DETECTION WITH PAR DATA FROM [31] Metal ion

Bismuth(II1) Cadmium(II) Calcium(Il) Cobalt(ll) COPPem Iron(II)

Detection limiP (ng)

Metal ion

5 25 2500 0.5 1 25

Lanthanides(1Il) Magnesium0 Mmgane=(II) Nickel(ll) M(II) =c(Jn

Detection limP

(4.9 1-5 8300 5 1 5 3

The PCR reagent was 0.2 mM PAR and the detection wavelength was 540 nm. a Measured at a signal-to-noise ratio of 2. [39], etching solutions [40], wastewaters [41] and electroplating solutions [42]. In many of these applications, acid digestion of the sample is required, but the resulting acidity of the sample digest does not interfere in the final chromatographic analysis. The detection sensitivity of PCR with PAR can be enhanced if an equimolar solution of PAR and Zn(I1)-EDTA complex is used as the PCR reagent [43-481. When a

402

Chapter 13 Tb

cu

BLANK GRADIENT I

0

I

1

I

2

I

3

I

L

I

5 Time (min)

I

I

7

6

I

8

I

9

Fig. 13.10 Gradient separation of the lanthanides with PCR detection using Arsenazo 111. A Supelco LC18 column was used. The eluent was formed using a linear gradient at pH 4.6 from 0.05 M HlBA to 0.40 M HIBA over 10 min at 2.0 ml/min, with a constant concentration of 10 mM 1-octanesulfonate in the eluent. The detection wavelength was 635 nm. Reprinted from [54] with permission.

metal ion is eluted, the following reaction occurs:

M(I1)

+

Zn[EDTA]

+

PAR % M(II)[EDTA]

+

Zn[PAR]

(13.3)

Detection is therefore based on the absorbance of the Zn[PAR] complex at 490 nm. This procedure is especially suitable for the detection of alkaline earth metal ions, which do not form strongly absorbing complexes with PAR.

13.4.3 PCR detection of lanthanides using Arsenazo dyes Extensive studies by Cassidy and co-workcrs [49-541have shown that Arsenazo I and Arsenazo I11 are ideal PCR reagents for the detection of lanthanide elements. Reaction of Arsenazo dyes with these ions is very rapid at room temperature and detection sensitivity is relatively uniform for the entire lanthanide series. A typical detection limit attained by this method is 80 ppb [49]. PCR detection is applicable after either ion-exchange or ion-interaction separation. The latter method often gives superior chromatographic resolution and octanesulfonic acid is commonly used as the ion-interaction reagent. Moreover, gradient elution can be applied in this detection mode. A chromatogram obtained using a concentration gradient of a complexing ligand in the eluent, with ion-exchange separation, was shown in Fig.4.22(a), whilst that for ion-interaction separation is given in Fig. 13.10. In the first example, 2-methyllactic acid was used as the ligand and Arsenazo I as the PCR reagent, whereas in the second example, a-hydroxyisobutyric acid (HIBA) is used as the ligand and Arsenazo IT1 as the PCR reagent. With a slight modification of the eluent composition used in Fig. 13.10,


403

TABLE 13.5 ANALYSIS OF STANDARD SOLUTIONS OF LANTHANIDES BY ION-INTERACTION CHROMATOGRAPHY WITH POST-COLUMN REACTION DETECTION USING ARSENAZO n1 [SO] Element L a Pr Sm

Gd

DY Ho Tm

Lu

Injected (ng)

Found (ng)

Deviation

60.4 22.3 34.4 46.9 57.7 12.1 6.16 5.71

61.0 22.5 34.7 46.9 58.6 12.1 6.24 5.69

1.o 0.9 0.9 0.0 1.5 0.0 1.3 0.3

Element

Injected (ng)

Found (ng)

Deviation

ce

146.0 108.9 13.9 8.64 195.8 37.7 39.2

146.8 109.1 14.1 8.72 193.1 37.7 39.9

0.5

(%I Nd Eu

Tb Y Er Yb

0.1 1.4 1.0 1.0 0.0 1.8

yttrium, thorium and uranium can be separated in the same chromatographic run, together with the elements shown in the chromatogram [49]. PCR detection of lanthanides using Arsenazo I11 has proven to be an outstanding method for the analysis of complex samples, such as uranium dioxide fuels [52, 541 and rocks [SO]. Table 13.5 shows the excellent agreement between the true and observed concentrations of lanthanides in standard solutions, as determined by IC. 13.4.4 PCR detection of other metal ions A wide range of other PCR reagents has been reported for the selective detection of particular metal ions using a variety of detection modes. These methods are normally developed for a specific analysis and therefore do not have the general applicability of PCR procedures which use PAR or Arsenazo dyes. Table 13.6 lists some of the pertinent details of these methods. It can be noted from Table 13.6 that several of the methods listed involve indirect detection. For example, monitoring at the absorption wavelength of dithizone or Eriochrome Black T permits the indirect detection of metal ions which form complexes with these species [68]. Similarly, use of an electroactive dithiocarbamate ligand as a PCR reagent permits indirect amperometric detection of metal ions which form electroinactive complexes with the ligand 166,671.

13.5 PCR DETECTION OF ORGANIC SPECIES Some less commonly used applications of PCR detection involve organic species as the analytes. Organic acids eluted from an ion-exclusion column with nitric acid as eluent may be detected by PCR with an acid-base indicator (4-nitrophenol) after passage through a hydrogen form suppressor [701. The pH changes which accompany the sample

Chapter I3

404 TABLE 13.6

SOME EXAMPLES OF FOST-COLUMN REACTION DEIECTION OF CATIONS IN IC Solutes

Postcolumn reagenta

Detection modeb

Detection limits

Ref

Al3+ Alh

Oxine-5-sulfonate Tiron Arsenam I Neo-thorin Luminol DPC PAR BPDSA Oxine

Fluor (512 nm) Spec (310 nm) spec (590 m) Spec (570 nm) Fluor Spec (520 nm) Spec (520 nm) Spec (530 nm) Fluor (530 nm)

1

Pa

55

2-6ng 50ppb 0.5 ng/l 1. 100ppb 1.5pM 10ppb 1-84ng

59 4, 60 61 62 63 64,65

Amper (+0.65V)c Spec (590 nm)c Spec (610 Fluor (534 nm)

0.2ppm 2-long 2-10 ng 16ngSn

66,67 68 68 69

Ca2+,Mg2+ Ca2+,Mg2+

co2+

wm,Cr(VI)

Fe2+,Fe3+ Fe2+,Fe3+ Zr(IV), Ga3+,Sc3+,Y3+, h 3 + , AQ+,h 3 + , ~2+, Cd2+,Ca2+,Mg2+ Transition metals SPDC Transition metals Dithime Transition metals Eriochrome Black T Tributyl tin Morin a

56.57 58

DPC = diphenylcarbohydnzide, BPDSA = bathophenanthroline disulfonic acid, SPDC = sodium pyrrolidinedithiocarbamate. Spec = spectrophotometry,Fluor = fluorescence, A m p = ampenmetry. Indirect detection method.

bands as they leave the suppressor are visualized by changes in the absorbance of the acid-base indicator at 400 nm. This detection system is applicable to gradient elution. PCR has also been used for the detection of formaldehyde after separation by ionexclusion chromatography [7 11. Reaction of the eluted formaldehyde with acetylacetone gives a detection limit of 10 ppb using absorbance at 420 nm. Post-column addition of Cu(1I) has been employed for the detection of amino acids [72] and reducing carbohydrates [73]. In each of these examples, the change in concentration of free copper ions caused by reaction with the eluted analyte species is detected by potentiometry, either with a copper ion-selective electrode or a metallic copper indicator electrode.

13.6 REFERENCES 1 2

3

Schlabach T.D. and Weinberger R., in Krull I.S. (Ed.), Reaction Detection in Liquid Chromatography,Chromatographic Science Series, Vol. 34, Marcel Dekker, New York, 1986, Ch. 2. Colgan S.T.and Krull I.S., in Krull I.S. (Ed.), Reaction Detection in Liquid Chromatography,Chromatographic Science Series, Vol. 34, Marcel Dekker. New York, 1986, Ch. 5. Cassidy R.M., Elchuk S. and Dasgupta P.K., Anal. Chem., 59 (1987) 85.

Detection by Post-Column Reuction

405

Boyle E.A., Handy B. and Van Geen A., Anal. Chem., 59 (1987) 1499. Elchuk S. and Cassidy R.M., Anal. Chem., 51 (1979) 1434. Schmidt G.J.and Scott R.P.W., Anulyst (London), 109 (1984) 997. Lillig B. and Engelhardt, H., in Krull LS. (Ed.), Reaction Detection in Liquid Chromutogruphy, Chromatographic Science Series, Vol. 34, Marcel Dekker, New York. 1986, Ch. 1. Davis J.C. and Peterson D.P.. Anal. Chem., 57 (1985) 768. 8 9 Hirai Y., Yoza N. and Ohashi S.. J . Chromutogr.,206 (1981) 501. 10 Yoza N., It0 K.. Hirai Y. and Ohashi S., J. Chromutogr., 196 (1980) 47 1. 11 Hirayama N. and Kuwamoto T., J. Chromutogr.,447 (1988) 323. 12 Yoza N., Miyaji T., Hiria Y. and Ohashi S., J. Chromutogr.,283 (1984) 89. 13 Nakae A.. Furuya K., Mikata T. and Yamanaka M., Nippon Kaguku Kaishi, (1977) 1655. 14 Nakae A., Furuya K., Mikata T. and Yamanaka M., Nippon Kaguku Kaishi, (1976) 1426. 15 Goguel R., Anal. Chem., 41 (1969) 1034. 16 Imanari T., Tanabe S., Toida T. and Kawanishi T., J. Chromutogr.,2% (1982) 55. 17 Tschabunin G., Fischer P. and Schwedt G., Fres. Z. Anal. Chem., 333 (1989) 117. 18 Dionex Application Note 44R. 19 Waters IC Lab. Report No. 309. 20 Lee S.H. and Field L.R., Anal. Chem., 56 (1984) 2647. 21 Weiss J. and Hagele G., Fres. 2.Anal. Chem., 328 (1987) 46. 22 Toida T., Togawa T., Tanabe S. and Imanari T., J. Chromutogr., 308 (1984) 133. 23 Buchberger W., J. Chromutogr.,439 (1988) 129. 24 Buchberger W. and Winsauer K., Mikrochim. Acta, 1985 III (1986) 347. 25 Lookabaugh M. and Krull I.S., J. Chromutogr.,452 (1988) 295. 26 Meek S.E. and Pietrzyk D.J., Anal. Chem., 60 (1988) 1397. 27 Vaeth E., Sladek P. and Kenar K., Fres. Z. Anal. Chem., 329 (1987) 584. 28 Dionex Application Update 113. 29 Brunt K., Anal. Chem., 57 (1985) 1338. 30 Fritz J.S. and Story J.N.. Anal. Chem., 46 (1974) 825. 31 Cassidy R.M. and Elchuk S., J. Chromutogr.Sci., 18 (1980) 217. 32 Cassidy R.M. and Elchuk S., J. Chromutogr.Sci., 19 (1981) 503. 33 Cassidy R.M., Elchuk S. and McHugh J.O., Anal. Chem., 54 (1982) 727. 34 Cassidy R.M. and Elchuk S., J. Liq. Chromatogr.,4 (1981) 379. 35 Byerley J.J., Scharer J.M. and Atkinson G.F., Analyst (London). 112 (1987) 41. 36 Blaszkewicz M., Baumhoer G., Neidhart B., Ohlendorf R. and Linscheid M., J. Chromatogr.,439 (1988) 109. 37 Takayanagi M. and Yashm T., J. Chromutogr..374 (1986) 378. 38 Ong C.N., Ong H.Y. and Chua L.H., Anal. Biochem., 173 (1988) 64. 39 Waters IC Lab. Report No. 272. 40 Waters IC Lab. Report No. 273. 41 Waters IC Lab. Report No. 276. 42 Wescan Application #283. 43 Arguello M.D. and Fritz J.S., Anal. Chem., 49 (1977) 1595. 44 Yan D. and Schwedt G., Fres. Z. Anal. Chem., 320 (1985) 325. 45 Yan D. and Schwedt G., Anal. Chim. Acta, 178 (1985) 347. 46 Yan D., Stumpp E. and Schwedt G.. Fres. Z. Anal. Chem., 322 (1985) 474. 4 5 6 7

406 47 48 49

50 51 52 53 54 55 56 57 58 59

60 61 62 63

64 65

66 67 68 69 70 71 72 73

Chapter I3 Yan D., Zhang J. and Schwedt G., Fres. 2.Anal. Chem., 331 (1988) 601. Schwedt G., GIT Fachz. L.ub., 7 (1985) 697. Barkley D.J., Blanchette M.. Cassidy R.M. and Elchuk S.. Anal. Chem., 58 (1986) 2222. Cassidy R.M., Chem. Geol., 67 (1988) 185. Cassidy R.M. and Fraser M., Chromurographia,18 (1984) 369. Cassidy R.M., Elchuk S., Elliot N.L., Green L.W.. Knight C.H., Recoskie B.M., Anal. Chem., 58 (1986) 1181. Cassidy R.M., Miller F.C., Knight C.H., Roddick J.C. and Sullivan R.W., Anal. Chem., 58 (1986) 1389. Knight C.H., Cassidy R.M., Recoskie B.M. and Green L.W., Anal. Chem., 56 (1984) 474. Jones P., Ebdon L. and Williams T., Analyst (London), 113 (1988) 641. Bertsch P.M. and Anderson M.A., Anal. Chem., 61 (1989) 535. Dionex Application Note 42. Smith D.L. and Fritz J.S., Anal. Chim. Acta, 204 (1988) 87. Nagashima H., Bunseki Kagaku, 35 (1985) 7. Jones P., Williams T. and Ebdon L., Anal. Chim. Acta, 217 (1989) 157. Dionex Application Note 26. Moses C.O., Herlihy A.T., Herman J.S. and Mills A.L., Talanra, 35 (1988) 15. Saitoh H. and Oikawa K., J. Chromutogr., 329 (1985) 247. Karcher B.D. and Krull I S . , J. Chromutogr. Sci., 25 (1987) 472. Karcher B.D., Krull IS., Schleicher R.G.and Smith S.B., Jr., Chromatographia, 24 (1987) 705. Hojabri H., Lavin A.G., Wallace G.G. and Riviello J.M., Anal. Proc., 23 (1986) 26. Hojabri H., Lavin A.G., Wallace G.G. and Riviello J.M., Anal. Chem., 59 (1987) 54. Hobbs P.J., Jones P. and Ebdon L., Anal. Proc., 20 (1983) 613. Ebdon L. and Alonso J.I.G., Analyst (London), 112 (1987) 1551. Okada T. and Dasgupta P.K., Anal. Chem., 61 (1989) 548. McClure J.E., Anal. Lett.. 21 (1988) 253. Loscombe C.R., Cox G.B. and Dalziel J.A.W., J . Chromutogr., 166 (1978) 403. Cowie C.E., Haddad P.R. and Alexander P.W., Chromutographia,21 (1986) 417.

Part IV Practical Aspects

408

r

1

Extraction Acid di estion Alkali &ion L Combustion Filtration Ion-exchange resins Dialytic methods Cartridge columns

Extraction

Cleanup

SAMPLE HANDLING (Chap 14)

-€

Contamination

-+

Physical handling Filtration devices Cartridge columns Hardware components

Ultra-trace analysis PRACTICAL ASPECTS

Filters Impin ers Adsorgents Denuders

Sample collection

I

Large injection volumes Concentrator columns Dialytic methods

On-column L Matrix elimination <post-column

E-,

Selection of chromatographic parameters

-

METHODS DEVELOPMENT (Chap 15)

Optimization Multi-dimensional methods Automation

Schematic overview of Part IV.

Separation mode

E;;4;i;pyde

Empirical methods Computer methods

fi'

VI P

TABLE 17.1 (CONTINUED) ANALYSIS OF INDUSTRIAL WASTEWATERS AND EFFLUENTS USING IC -

~~

~

__

~

~

Solutes (min)

Sample Prep.

Column

Eluent 5.0 mM octanesulfonic acid,

Ammeline (9.19, melamine (10.03)

adjusted to pH 3.0 with acetic acid (28% MeOH) 1.5 d m i n 0.25 mM rn-phenyleneDionex 30831 Guanidinium (5.1) cation separator diamine dihydrochloride, 0.25 mM hydrochloric acid 2.5 d m i n Dionex cation 1.3 mM o-phenylenediamine Sodium (1.9). magnesium (3.3, calcium hydrochloride separator (5.5) 250 x 6.0 mm ID 138 ml/hr 2.0 mM octanesulfonate, Iron (111) (2.681, copper (4.18). lead (5.38), Nitric acid digestion, Waters c18 50 mM tartaric acid, pH 3.4 pondapak zinc (9.35). nickel (10.35), cobalt (14.72), dilution 300 x 3.9 mm ID 1.O d m i n cadmium (19.43, iron (11) (20.32), manganese (26.62) 2mMsodium Nitric acid and Waters c18 Iron (III) (2.4), copper (4.3), lead (5.4), octanesulfonate, 50 mM peroxide digest, pondapak zinc (9.8), nickel (10.5), cobalt (15.4), 300 x 3.9 mm ID tartaric acid, pH 3.4 resuspension in cadmium (17.0), manganese (28) 1.0 d m i n 0.5% nitric acid 0.35 M to 0.5 M tartrate o n Arninex A5 Preconcentration Copper (2.1), zinc (8.2), lead (10.4), nickel 100 x 4.0 mm ID gradient, pH 3.5 Aminex A5 (12.11, cobalt (15.4) 1.0 ml/min Iron (III) (0.99), copper (4.27), nickel (7.22), zinc (11.33)

Dilution in eluent

TSK silica cation 1.07 gA citric acid, 7.5 gA 50 x 4.6 mm ID tartaric acid, 120 pUl ethylenediamine 1.O d m i n

Detection

Ref

Direct spectrophot. at 40 235 nm conductivity

40

Conductivity

28.41

Direct speccrophot at 520 nm after postcolumn reaction with PAR Direct spectrophot. at 520 nm after postcolumn reaction with PAR Direct spectrophot. at 530 nm afta postcolumn reaction with PAR Direct spectrophot. at 546 ~nafter postcolumn reaction with PAR

42,43

Seep. 487 for notes on the organization of this Table. Seep. 585 for References. Abbreviations are listed in Appendix B (p. 745).

44

45

46

85 v

Ir

TABLE 17.1 (CONTINUED)ANALYSIS OF INDUSTRIALWASTEWATERS AND EFFLUENTS USING IC Solutes (min)

Sample Prep.

Column

Tungstate (9.9). molybdate (15.0), arsenate Preconcentration (21.0), chromate (24.9)

Chmium (In) (0.9). chromium (VI) (4.0)

chromium @I) Chromium 0, Chromium (VI)(3.5)

Chromate (6.0)

Dionex anion brine 500x3.0mmID Addition of cyanide TSKgel ICto prevent chromium Anion-PW 100 x 4.0 mm ID hydroxide ppt

oncolumn preconcenmtion, filtration Neutralization, dilution in eluent

Dionex CS-2 250 x 4 mm ID

Preconcentration (10 ml at 4.0

Dionex AS-2

Wmin)

Dionix CS-5

Eluent 3.0 mM carbonate, 5.0 mM hydroxide 230ml/hr 0.5 mM phthalate, pH 6.5 1.2 d m i n

& E Detection Conductivity

Inductively coupled plasma atomic emission spectrometry Water with multiple injections 3 e l d p l a s m a of 1.O M hydrochloric acid atomic emission 2.0 d m i n spectromeay 10 mM p y r i d i n e d i x y l i c Direct spectrophot. at acid, 148 mM durn 520 ~I after I I postcolumn reaction with hydroxide methanolic diphenyl1.0 Wmin carbohydrazide 20 mM nitric acid, Direct specaophot. at 30 mM hydroxide 365 nm 1.5 mVmin


Ref 47 48

5

B

&

z 2

49

50

51

VI P

W

v,

TABLE 17.2

iz: v

ANALYSIS OF OTHER INDUSTRIAL WATERS USING IC ~~

~

Solutes (min)

~~

~

~

Sample

Sample Prep.

Column

Fluoride (2.1), chloride (3.5). Industrial waters phosphate (7.0), bromide (9.0), nitrate (1 l.O), sulfate (15.5) Chloride (4.3), nitrite (5.0), Indusmal nitrate (12.0). sulfate (18) process waters, water-formed deposits Fluoride (3.66), chloride Industrial run-off (8.64). ninate (19.77) waters

Samples diluted in bicarbonate buffer Dilution, filtration

Dionex anion separator 250 x 4.0 mm ID Dionex anion separator 500 x 3.0 mm ID

Filtration

Waters IC Pak A 50 x 4.6 mm ID

Magnesium (3.9),calcium (5.0) Industrial waters

-

Toyo Soda TSKgel SP-NPR 35 x 4.6 mm ID

Chloride (4.59, sulfate (21.14) Indusmal brine

Dilution, filtration

Waters IC Pak Anion HC 150 x 4.6 mm ID

Lithium (3.16), sodium (4.1I), potassium (6.74)

Indusmal brine


Waters IC Pak C 50 x 4.6 mm ID

Magnesium (2.73), calcium (3.68)

Indusmal brine


Waters IC Pak C 50 x 4.6 mm ID

Eluent

Detection

Ref

3.0 mM bicarbonate, 2.4 mM carbonate 161 ml/hr 3.0 mM bicarbonate, 2.4 mM carbonate 138 mVhr

Conductivity

52

Conductivity

53

10 mM acetate, pH 7.6 (10%ACN) 1.2 mumin 0.05 M phosphate buffer, 0.075 M potassium chloride, 1.0 mM Arsenazo 111, pH 6.0 0.7 mumin

Conductivity

54

1.42 mM gluconate, 5.82 mM boric acid, 0.25% glycerine, pH 8.5 (12% ACN) 1.2 ml/min 2.0 mM nitric acid, 0.05 mM disodium EDTA 1.5 ml/min 0.5 mM ethylenediamine, nimc acid, pH 6.0 1.2 ml/min

Direct 55 spectrophot. at 600 nm after in situ formation of Arsenazo complexes Conductivity 56, 57

Indirect conductivity

56


56

Seep. 487 for notes on the organization of this Table. Seep. 585 for References. Abbreviations are listed in Appendix B ( p . 745).

TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIAL WATERS USING IC Solutes (min)

Sample

Magnesium (6.8), calcium (10.4)

Chlor-alkali brine Dilution, Dionex CS-2 preconcentration on Dionex MFC-1 Processreclaim Wescan 269001 water anion 250 x 4.6 mm ID Processbath Wescan 269001 anion 250 x 4.6 mm ID Process water Vydac anion 250 x 4.6 mm ID

Chloride (2.5), nitrate (4), system peak (7), sulfate (12) Phosphate (lo), chloride (13), chlorate (16), sulfate (17) Chloride (4.4), nimte (5.0), nitrate (10)

Sample Prep.

Chloride (2.9), chlorate (4), sulfate (7)

Processliquor

-

Sulfite (2.5), sulfate (2.8), thiosulfate (8.6)

Process stream solution

-

Phosphate

Processing water Dilution, aqueous extraction

Magnesium (7), calcium (9)

Process water

-

Column

Wescan 269001 anion 250 x 4.6 mm ID Dionex AG-3 (x2) in series 50 x 4.0 mm ID Waters Resolve 150 x 4.6 mm ID Dionex cation separator 250 x 6.0 mm ID

Eluent

Detection

Ref

25 mM hydrochloric acid, 2.0 mM hystidine 1.0 d m i n

Conductivity

58

4.5 mM phthalate, 0.5 mM phthalic acid 3.7 d m i n 5 mMphthalic acid 2.5 d m i n

Conductivity

59

Conductivity

60

1mM phthalate 2.0 d m i n

8.7 mM phthalate, 1.3 mM phthalic acid 2.5 mumin 6.0 mM carbonate 4.6 d m i n 10 mM tetrabutylammonium phosphate, 20 mM formic acid (20% MeOH) 1.O d m i n 2.5 mM nitric acid, 2.5 mM phenylenediamine dihydrochloride 1.15 Wmin

-9 a

$

3

2.

2

Indirect 61 spectrophot. at 280 nm Conductivity 62 Conductivity

63

Direct RI

64

Coulometry, 65 +0.45V, after reaction with hydroquinone v,


v, c

VI

TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIAL WATERS USING IC Sample

Zinc (4.0), samarium (10.0). gadolinium (13.4), terbium (15.7). dysprosium (17.5)

Process stream (lanthanide manufacturing)

Dionex CS-5 cation 2.0 mM pyromellitic acid, separator

50 mM oxalic acid, adjusted to pH 3.5 with lithium hydroxide 1.o d m i n

Succinate (16), formate (17). acetate (201, propionate (211, butyrate (32) Oxalate (2.4),EDTA (6.3), citrate (8.1)

Synfuels plant stream water

Dionex AS-2 ion exclusion

10 mM hydrochloric acid 2.0 d m i n

Reactor decontamination water

sUplc4d c18 coated with cetylpy-ridinium chloride 75 x 4.0 mm ID Dionex anion separator 500 x 3.0 mm ID Dionex anion separator 500 x 3.0 mm ID

1.5 mM phthalic acid, Indirea 68 1.8 mM rris(hydmxymethy1) specmphot. at 254 nm aminomethane (7% ACN) 2.0 d m i n

UraniUm processing solution Fuel reprocessing solutions

Tetrafluomborate (10.8)

Nuclear fuel dissolvent solutions

Column

Eluent

N

Solutes (min)

Dibutylphosphate (2.3, monobutylphosphate (4.7), phophate (14.1) Dibutylphosphoric acid (3.3, monobutylphosphoricacid (5.8)

Sample Prep.

VI

3.0 mM bicarbonate, 2.4 mM carbonate 138 mvhr 1.5 mM carbonate, 0.5 mM hydroxide 138 mvhr

Barium ppt of oxalate and carbonate, C18 Sep-Pak Reconcentration Dionex brine anion 3.0 mM bicarbonate, separator 4.0 mM carbonate on Amberlite XE-243, 500 x 3.0 mm ID 138 mvhr acidification, hydroxide elution

Detection

Ref

Direct 66 specmphot. at 520 nm after post-column reaction with PAR Conductivity 67

Conductivity

69

Conductivity

70

Conductivity

71,72


TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIALWATERS USING IC Solutes (min)

Sample

Sample Prep.

Column

Dibutylphosphate (6.4)

Nuclear fuel reprocessing

Hydroxide extraction, centrifugation Preconcentration on Amberlite XE-243, acidification, hydroxide elution Reconcentration on Dionex TCC- 1

Dionex anion separator 500 x 3.0 mm ID Dionex anion separator 150 x 3.0 mm ID

Dionex CS-3

2.3 diaminopmpionicacid Conductivity monohydrochloride,histidine monohydrochloride 8 mM phthalate. conductivity 2 mMphthalic acid 2.5 ml/min 5.0 mM phthalate. pH 3.8 Conductivity 4.0 ml/min

streams

Terrafluomborate(13)

Nuclear fuel dissolvent solutions

Calcium (8.6), strontium (12.6) Nuclear reprocessing solutions Chloride (2), system peak (4). Refinery accumulated sulfate U),thiosulfate (9) water Chloride (0.9), sulfite (l.l), Refinery sulfate (4.7), thiosulfate (7.0) accumulated Water Sulfide (2.2) Refinery waters

Wescan 269001 anion 250 x 4.6 mm ID Wescan anion/HS

-

Dionex AS-3

Eluent

Detection

Ref

3.0 mM hydroxide 92 ml/hr

conductivity

73-75, 69

3.0 mM bicarbonate, 2.4 mM carbonate 138 ml/hr

conductivity

71,72

1.0mM carbonate, 10 mM sodium dihydrogen borate, 15 mM ethylenediamine 2.0 ml/min 3.0 mM tetraborate 2.0 ml/min

Phenol (4.40)

Refinerywaters

-

Dionex AS-3

Ammonia (5.15). monoethylamine (6.41), diethylamine (8.45), methylamine (10.74)

Refinery waters

-

Dionex MPIC NS-1 5.0 mM hexanesulfonicacid Separator 1.0 ml/min

76 77 30

Ampmmetry 67 at Ag electrode,

0.oov 67 at Pt electrode, +1.ov Conductivity 67 Amperomeay


v1

TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIAL WATERS USING 1C Solutes (min)

Sample

Sample Prep.

Column

Monoethanolamine (20), diethanolamine (29), methyldiethanolamine (36) Chloride (2), nitrate (2.7), sulfate (7), hydroxylamine disulfonate (lo), dithionate (19) Chloride (3.0). sulfide (5.8). sulfate (12), sulfite (12)

Refmerywater

-

Dionex MPIC NS-1 5.0 mM octanesulfonic acid separator 1.O mumin

Flue gas desulfurization liquors Scrubber liquor

Dilution

Eluent

vl

Detection

Ref

Conductivity

41

Wescan 269013 anion/HS 100 x 4.6 mm ID Dionex anion separator 500 x 3.0 mm ID

4 mM phthalate, pH 3.8 2.7 d m i n

Conductivity

78

1.5 mM bicarbonate, 1.5 mM carbonate 2.2 mumin

Conductivity

79

Fluoride, chloride, nitrate, arsenate. sulfate

Flue gas, extract Adsorption, liquor from Cu, dilution Al, Fe refineries

Dionex anion separator 500 x 3.0 mm ID

0.25 @ bicarbonate, I 0.25 @ carbonate I 1.5 d m i n

Conductivity

80

Chloride (4), adipate (14), sulfite (19), sulfate (31)

Scrubber liquors Filtration

Dionex anion separator 500x3.0mmID Dionex anion separator 500 x 3.0 mm ID

1.5 mM bicarbonate, 1.2 mM carbonate

Conductivity

81

138mMr 3.0 mM bicarbonate, 2.4 mM carbonate 2.3 mumin

Conductivity

82

Dionex anion separator 500x3.0mmID Dionex anion separator 500 x 3.0 mm ID

9.0 mM bicarbonate, 7.2 mM carbonate 184mvhr 3.0 mM bicarbonate, 2.4 mM carbonate 138 mvhr

Conductivity

83

Conductivity

84

Chloride (1.8), adipate (3.8), sulfate (4.9)

Flue gas from desulfurization systems

Chloride (2.5), sulfate (4.4), thiosulfate (17)

Scrubber liquors, processing waters Flue gas Dilution scrubber solution

Chloride (3.8), sulfite (1 l.O), sulfate (17.3)

Filtration, peroxide oxidation


PB

c,

u

TABLE 17.2 (CONTINUED). ANALYSIS OF OT

R INDUSTRIALWATERS USING IC

Solutes (min)

Sample

ample Prep.

Chloride (1.3), sulfite (4.3, sulfate (7.0)

Lime, limestone- :&ration, sulfite based sulfur stabilized with dioxide scrubber formaldehyde solutions Flue gas Dilution scrubbing liquor

Chloride (2.7), nitrate (7.4), sulfate (15.0) Sulfate (9.0), oxalate (13.1)

Scrubber liquor, pitch

Nitrate (8.6), sulfate (11.0)

Stack gas trap solution

Sulfite (23), sulfate (45)

Scrubberliquor from flue gas coal desulfurization Flue gas Dilution scrubbing liquor Flue gas Dilution desulfurization liquor Flue gas Dilution scrubber solution

Imidodisulfate (7.3, hydroxyimidodisulfate (16.5) Bicarbonate (6) Sodium (13), ammonium (20), potassium (26) Sodium (2), potassium (7)

Flue gas desulfurization liquors

Column

Detection

Ref

Dionex anion separator 500 x 3.0 mm ID


Conductivity

85, 83

Dionex AS-4

5 mM bicarbonate, 5 mM carbonate 2.0 d m i n

Conductivity

86


Conductivity

87

5.0 mMp-hydroxybenzoic acid, pH 8.6 2.0 d m i n 1.5 mM bicarbonate, 1.2 mM carbonate 1.5 d m i n

Conductivity

30

Conductivity

88

12 mM carbonate 1.5 d m i n Deionized water 1.Od m i n

Conductivity

86

Conductivity

89

5.0 mbl nitric acid 184 mvhr

Conductivity

84

3 mM nitric acid 1.5 ml/min


90

Combustion, Dionex AS-3 sorption in alkaline peroxide Wescan anion R 250 mm

Dilution

Eluent

Dionex anion separator 500 x 3.0 mm ID Dionex AG-4 500 x 3.0 mm ID Wescan 269038 exclusion 250 x 7.1 mm ID Dionex cation separator 250 x 6.0 mm ID Wescan 269024 cation/HS 50 x 3.0 mm ID


VI

TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIAL, WATERS USING IC

x "I

Solutes (min)

Sample

Sample Rep.

Lime, limestone- Filtration based sulfur dioxide scrubber solutions Dilution Magnesium (6.6).calcium (9.9) Flue gas scrubber solution

Sodium (1.6),magnesium (3.4), calcium (5.2)

Fluoride (2.6).chloride (3.2), phosphate (4.4), nitrate ( 5 . 3 , sulfate (7.0)

Water-insoluble wastewater precipitate

Fluoride (5), chloride (6). Sewage phosphate (1 l), nitrate (14), sulfate (22) Chloride (4). nitrate (6), sulfate Sewage sludge (lo), tungstate (12)

Fluoride, chloride, phosphate, sulfate

Sewage sludge

Chloride (7.0),bromide (9.0), nitrate (1 1.5)

Sewage water

Sodium carbonate fusion, SCX in hydrogen form

Column

Detection

Ref

Dionex cation separator 500 x 6.0 mm ID

2.0 mM p-phenylenediamine dihydrochloride

Conductivity

85

Dionex alkaline earth separator 250 x 6.0 mm ID Dionex M30170 anion separator 500 x 3.0 mm ID

1.O mM p-phenylenediamine dihydrochloride 138 mvhr 2.1 mM bicarbonate, 1.7 mM carbonate 2.0 mumin

Conductivity

84

Conductivity

91

9.0 mM bicarbonate, 3.0 mM carbonate 1.38 Wmin 5.0 mM p-hydmxybenzoic acid, pH 8.5 2.0 d m i n

Conductivity

92

Conductivity

93

3.0 mM bicarbonate, 2.4 mM carbonate 4dmh 0.01 M phosphate, pH 3.8 2.0 d m i n

Conductivity

94

Home-packed anion-exchanger 500 x 3.0 mm ID Ashing, aqueous Wescan 269-069 extraction, anion centrifugation, 2 5 0 ~ 4 . 1mmID filtration, barium ppt of sulfate, silica column clean-up, concentration Dionex anion Oxygen bomb separator Dilution

Eluent

Vydac 302 IC 250 x 4.6 mm ID

Direct 95 specmphot. at 190 nm

See p. 487 for notes on the organizarion of this Table. Seep. 585 for References. Abbreviations are listed in Appendix B (p. 745).

2

TABLE 17.2 (CONTINUED). ANALYSIS OF OTHER INDUSTRIAL WATERS USING IC Solutes (min)

Sample

Sample Prep.

Chloride (1.9), bromide (4.0), sulfate (6.9)

Pollutant water

Column

Nimte (7.71, nitrate (9.5)

Sewage sludge

Nitrite (3.3, nitrate (4.8)

Silage extract

Adsorption onto Dionex A S 4 carbon, combustion, all

Ion Chromatography (Journal of Chromatography Library)

Bioaffinity Chromatography (Journal of Chromatography Library)

Affinity Chromatography (Journal of chromatography library)

Instrumental Liquid Chromatography (Journal of Chromatography Library)

Chromatography of Antibiotics: Journal of Chromatography Library

Chromatography of Antibiotics: Journal of Chromatography Library

Chromatography of Antibiotics (Journal of Chromatography Library)

Extraction Chromatography (Journal of chromatography library)