Ion Chromatography Book

Preface 13

Acknowledgments 15

1 Introduction and Overview 1

1.1 Introduction 1

1.2 Historical Development 2

1.2.1 Early Ion-Exchange Separations 2

1.2.2 Cation Separations 3

1.2.3 Separation of Anions 6

1.2.4 On-line Detection 8

1.2.5 The Birth of Modern Ion Chromatography 8

1.2.6 Non-Suppressed-Ion Chromatography 10

1.3 Principles of Ion Chromatographic Separation and Detection 13

1.3.1 Requirements for Separation 13

1.3.2 Experimental Setup 13

1.3.3 Performing a Separation 14

1.3.4 Migration of Sample Ions 15

1.3.5 Detection 17

1.3.6 Basis for Separation 17

2 Instrumentation 21

2.1 Components of an Ion Chromatography (IC) Instrument 21

2.2 General Considerations 23

2.3 Eluent 24

2.4 Pump 26

2.4.1 Gradient Formation 29

2.5 Sample Injector 30

2.6 Columns 31

2.6.1 Column Hardware 31

2.6.2 Column Protection 32

2.6.3 Column Oven 33

2.6.4 Two-dimensional IC 33

2.7 Suppressor 33

2.8 Detector 34

2.9 Data Acquisition and Calculation of Results 35

3 Resins and Columns 37

3.1 Introduction 37

3.2 Polymeric Resins 38

3.2.1 Substrate and Cross Linking 38

3.2.2 Microporous Resins 39

3.2.3 Macroporous Resins 40

3.2.4 Chemical Functionalization 41

3.2.5 Resin Capacity 42

3.3 Resins and Columns for Ion Chromatography 43

3.3.1 Monolith Columns 43

3.4 Anion Exchangers 45

3.4.1 Porous Anion Exchangers 45

3.4.2 Effect of Functional Group on Selectivity 47

3.4.3 Effect of Spacer Arm Length 52

3.4.4 Latex Agglomerated Ion Exchangers 54

3.4.4.1 Effect of Latex Functional Group on Selectivity 56

3.5 Cation Exchangers 57

3.5.1 Sulfonated Resins 57

3.5.2 Weak-acid Cation Exchangers 61

3.5.3 Other Types 63

3.6Other Resins 63

3.6.1 Chelating Ion-exchange Resins 63

3.6.2 Metal Oxides 64

3.6.3 Multi-purpose Resins 64

3.6.4 Ion-exchange Disks 65

4 Detectors 69

4.1 Introduction 69

4.2 Conductivity Detectors 70

4.2.1 Conductivity Definitions and Equations 73

4.2.2 Principles of Cell Operation 74

4.2.3 Conductance Measurement 75

4.2.4 Conductivity Hardware and Detector Operation 75

4.2.5 Contactless Conductivity Detection 76

4.3 Ultraviolet-Visible (UV-Vis) Detectors 77

4.3.1 UV-Vis measurement 77

4.3.2 Direct Spectrophotometric Measurement 78

4.3.3 Post-column Derivatization 81

4.3.4 UV-Vis Hardware and Detector Operation 82

4.4 Fluorescence Detector 83

4.5 Electrochemical Detectors 85

4.5.1 Potentiometric Detection 86

4.5.2 Conductometric Detectors 86

4.5.3 Amperometric/Coulometric Detection 87

4.5.4 Pulsed Electrochemical Detection (PED) 89

4.5.4.1 Pulsed Amperometric Detection (PAD) 91

4.5.4.2 Integrated Pulsed Amperometric Detection (IPAD) 93

4.5.4.3 IC-PED 94

4.5.5 Post-column Derivatization 95

4.5.6 Electrochemical Hardware and Detector Operation 95

4.6 Refractive Index Detector 97

4.7 Evaporative Light Scattering Detector (ELSD) 97

4.7.1 Nebulizer 98

4.7.2 Evaporation Chamber 99

4.7.3 Detection Cell 99

4.8 Other Detectors 100

5 Principles of Ion Chromatographic Separations 105

5.1 General Considerations 105

5.2 Chromatographic Terms 105

5.2.1 Retention Factor 106

5.3 Selectivity 109

5.3.1 Selectivity Coefficients 110

5.3.2 Other Ion-exchange Interactions 112

5.3.3 Selectivity of Sulfonated Cation-exchange Resin for Metal Cations 113

5.3.4 Factors Affecting Selectivity 120

5.3.4.1 Polymeric Matrix Effect 121

5.3.4.2 Resin Functional Group 122

5.3.4.3 Solvation Effects 123

5.4 Chromatographic Efficiency 124

6 Anion Chromatography 131

6.1 Scope and Conditions for Separation 131

6.1.1 Columns 132

6.1.2 Separation Conditions 135

6.2 Suppressed Anion Chromatography 138

6.2.1 Electrolytic Suppressors 140

6.2.2 Solid-Phase Reagents, 1990[7] 141

6.2.3 Typical Separations 142

6.2.3.1 Isocratic and Gradient Elution 144

6.2.3.2 Influence of Organic Solvents 146

6.3 Nonsuppressed Ion Chromatography 147

6.3.1 Principles 147

6.3.2 Explanation of Chromatographic Peaks 150

6.3.3 Eluent 150

6.3.3.1 General Considerations 150

6.3.3.2 Salts of Carboxylic Acids 151

6.3.3.3 Basic Eluents 152

6.3.3.4 Carboxylic Acid Eluents 153

6.3.4 System Peaks 154

6.3.5 Scope of Anion Separations 155

6.3.6 Sensitivity 155

6.3.6.1 Conductance of a Sample Peak 158

6.4 Coated Columns 160

6.5 Optical Absorbance Detection 163

6.5.1 Introduction 163

6.5.2 Direct UV Absorption 163

6.5.3 Indirect Absorbance 164

6.6 Detection 166

6.7 Pulsed Amperometric Detector (PAD) 168

6.8 Evaporative Light Scattering Detector (ELSD) 170

6.9 Inductively Coupled Plasma Methods (ICP) 172

6.9.1 Atomic Emission Spectroscopy (AES) 172

6.9.2 Inductively Coupled Plasma Mass Spectrometry (ICP-MS) 173

7 Cation Chromatography 175

7.1 Introduction 175

7.2 Columns 176

7.2.1 Historical Development 178

7.2.2 Phosphonate Columns 179

7.2.3 Macrocycle Columns 181

7.2.4 Surfactant Columns 182

7.3 Separations 184

7.3.1 Suppressed-Conductivity Detection 184

7.3.2 Non-Suppressed-Conductivity Detection 187

7.3.3 Spectrophotometric Detection 188

7.4 Effect of Organic Solvents 191

7.4.1 Separation of Alkali Metal Ions 193

7.5 Separation of Metal Ions with a Complexing Eluent 195

7.5.1 Principles 195

7.5.2 Separations 196

7.5.3 Use of Sample Masking Reagents 197

7.5.4 Weak-Acid Ion Exchangers 198

7.6 Chelating Ion-Exchange Resins and Chelation Ion Chromatography 201

7.6.1 Fundamentals 201

7.6.2 Examples of Metal-Ion Separation 204

8 Ion-Exclusion Chromatography 207

8.1 Principles 207

8.1.1 Equipment 209

8.1.2 Eluents 209

8.1.3 Detectors 210

8.2 Separation of Organic Acids 211

8.2.1 Effect of Alcohol Modifiers 214

8.2.2 Separation of Carboxylic Acids on Unfunctionalized Columns 216

8.3 Simultaneous Determination of Anions and Cations 217

8.4 Conclusions 220

8.5 Determination of Carbon Dioxide and Bicarbonate 222

8.5.1 Enhancement Column Reactions 222

8.6 Separation of Bases 223

8.7 Determination of Water 226

8.7.1 Determination of Very Low Concentrations of Water by HPLC 229

8.8 Separation of Saccharides and Alcohols 230

8.8.1 Introduction 230

8.8.2 Separation Mechanism and Control of Selectivity 230

8.8.3 Detection 235

8.8.4 Contamination 235

9 Ion Pair Chromatography 239

9.1 Principles 239

9.2 Typical Separations 242

9.3 Mechanism 246

10 Zwitterion Stationary Phases 251

10.1 Introduction 251

10.2 Simultaneous Separation of Anions and Cations 253

10.3 Separation of Anions 255

10.4 Separation of Cations 256

10.5 Mechanism 259

11 Capillary Electrophoresis 263

11.1 Introduction 263

11.1.1 Steps in Analysis 264

11.1.1.1 Capillary Pretreatment 264

11.1.1.2 Sample Introduction 264

11.1.1.3 Sample Run 265

11.1.1.4 Detection 265

11.2 Principles 265

11.2.1 Terms and Relationships 265

11.2.2 Zone Broadening 267

11.2.3 Sample Injection 267

11.3 Electrosmotic Flow (EOF) 268

11.3.1 Effect of EOF on Separations 270

11.3.2 Control of EOF 271

11.4 Separation of Ions 274

11.4.1 Separation of Anions 274

11.4.1.1 Separation of Isotopes 276

11.4.2 Separation of Cations 278

11.4.3 Separations at Low pH 279

11.4.4 Capillary Electrophoresis at High Salt Concentration 280

11.5 Capillary Electrophoretic Ion Chromatography 283

11.5.1 Micellar Electrokinetic Chromatography (MEKC) 284

11.5.2 Partial Complexation 285

11.5.3 Effect of Ionic Polymers 287

11.5.4 Effect of Alkylammonium Salts 291

11.5.4.1 Separation Mechanism 294

11.6 Summary 294

12 DNA and RNA Chromatography 299

12.1 Introduction 299

12.1.1 Importance of DNA and RNA Chromatography 299

12.1.2 Organization of this Chapter 300

12.2 DNA and RNA Chemical Structure and Properties 301

12.3 DNA and RNA Chromatography 303

12.3.1 Development of DNA and RNA Chromatography 303

12.3.2 Column Properties 305

12.3.3 Ion-pairing Reagent and Eluent 306

12.4 Temperature Modes of DNA and RNA Chromatography 307

12.4.1 Nondenaturing Mode 307

12.4.2 Fully Denaturing Mode 308

12.4.3 Partially Denaturing Mode 309

12.5 Instrumentation 310

12.5.1 Effect of Metal Contamination 310

12.5.2 The Column Oven 313

12.5.3 UV and Fluorescence Detection 313

12.5.4 Fragment Collection 314

12.6 Applications of DNA Chromatography 314

12.6.1 Dhplc 314

12.6.2 Nucleic Acid Enzymology 315

12.6.2.1 Telomerase Assays 315

12.6.2.2 Polynucleotide Kinase Assays 316

12.6.2.3 Uracil DNA Glycosylase Assays 317

12.7 Applications of RNA Chromatography 317

12.7.1 Separation of Messenger RNA from Ribosomal RNA 318

12.7.2 Analysis of Transfer RNA 319

13 Sample Pretreatment 323

13.1 Dilute and Shoot or Pre-treat the Sample? 323

13.2 Particulate and Column-contaminating Matter 324

13.3 Preconcentration 325

13.3.1 Collection of Ions from Air 325

13.3.2 Preconcentration of Ions in Water 326

13.4 Sample Pretreatment 328

13.4.1 Anions in Acids 328

13.4.2 Neutralization of Strongly Acidic or Basic Samples 328

13.4.3 Dialysis Sample Preparation 329

13.4.3.1 Passive Dialysis 330

13.4.3.2 Donnan (Active) Dialysis 330

13.4.4 Isolation of Organic Ions 333

14 Method Development and Validation 335

14.1 Choosing a Method 335

14.1.1 Define the Problem Carefully 335

14.1.2 Experimental Considerations 336

14.1.3 Example of Method Development 338

14.2 Some Applications of Ion Chromatography 340

14.3 Statistical Evaluation of Data 341

14.3.1 Common Statistical Terms 341

14.3.2 Distribution of Means 344

14.3.3 Confidence Intervals 345

14.4 Validation of Analytical Procedures 347

14.4.1 Analytical Control 349

15 Chemical Speciation 353

15.1 Introduction 353

15.2 Detection 355

15.3 Chromatography 356

15.4 Valveless Injection IC 357

15.5 Speciation of some Elements 359

15.5.1 Chromium 359

15.5.2 Iron 360

15.5.3 Arsenic 361

15.5.4 Tellurium 362

15.5.5 Selenium 363

15.5.6 Vanadium 364

15.5.7 Tin 364

15.5.8 Mercury 365

15.5.9 Other Metals 365

Index 369