About the Book
The latest edition of the authoritative reference to HPLC High-performance liquid chromatography (HPLC) is today the leading technique for chemical analysis and related applications, with an ability to separate, analyze, and/or purify virtually any sample. Snyder and Kirkland's Introduction to Modern Liquid Chromatography has long represented the premier reference to HPLC. This Third Edition, with John Dolan as added coauthor, addresses important improvements in columns and equipment, as well as major advances in our understanding of HPLC separation, our ability to solve problems that were troublesome in the past, and the application of HPLC for new kinds of samples.
This carefully considered Third Edition maintains the strengths of the previous edition while significantly modifying its organization in light of recent research and experience. The text begins by introducing the reader to HPLC, its use in relation to other modern separation techniques, and its history, then leads into such specific topics as:
The basis of HPLC separation and the general effects of different experimental conditions
Equipment and detection
The column—the "heart" of the HPLC system
Reversed-phase separation, normal-phase chromatography, gradient elution, two-dimensional separation, and other techniques
Computer simulation, qualitative and quantitative analysis, and method validation and quality control
The separation of large molecules, including both biological and synthetic polymers
Chiral separations, preparative separations, and sample preparation
Systematic development of HPLC separations—new to this edition
Troubleshooting tricks, techniques, and case studies for both equipment and chromatograms
Designed to fulfill the needs of the full range of HPLC users, from novices to experts, Introduction to Modern Liquid Chromatography, Third Edition offers the most up-to-date, comprehensive, and accessible survey of HPLC methods and applications available.
Table of Contents:
Preface xxxi
Glossary of Symbols and Abbreviations xxxv
1 Introduction 1
1.1 Background Information 2
1.2 A Short History of HPLC 6
1.3 Some Alternatives to HPLC 8
1.4 Other Sources of HPLC Information 12
References 15
2 Basic Concepts and the Control of Separation 19
2.1 Introduction 20
2.2 The Chromatographic Process 20
2.3 Retention 24
2.4 Peak Width and the Column Plate Number N 35
2.5 Resolution and Method Development 54
2.6 Sample Size Effects 69
2.7 Related Topics 74
References 83
3 Equipment 87
3.1 Introduction 88
3.2 Reservoirs and Solvent Filtration 89
3.3 Mobile-Phase Degassing 92
3.4 Tubing and Fittings 96
3.5 Pumping Systems 104
3.6 Autosamplers 113
3.7 Column Ovens 125
3.8 Data Systems 127
3.9 Extra-Column Effects 131
3.10 Maintenance 131
References 144
4 Detection 147
4.1 Introduction 148
4.2 Detector Characteristics 149
4.3 Introduction to Individual Detectors 160
4.4 UV-Visible Detectors 160
4.5 Fluorescence Detectors 167
4.6 Electrochemical (Amperometric) Detectors 170
4.7 Radioactivity Detectors 172
4.8 Conductivity Detectors 174
4.9 Chemiluminescent Nitrogen Detector 174
4.10 Chiral Detectors 175
4.11 Refractive Index Detectors 177
4.12 Light-Scattering Detectors 180
4.13 Corona-Discharge Detector (CAD) 184
4.14 Mass Spectral Detectors (MS) 185
4.15 Other Hyphenated Detectors 191
4.16 Sample Derivatization and Reaction Detectors 194
References 196
5 The Column 199
5.1 Introduction 200
5.2 Column Supports 200
5.3 Stationary Phases 217
5.4 Column Selectivity 227
5.5 Column Hardware 238
5.6 Column-Packing Methods 240
5.7 Column Specifications 244
5.8 Column Handling 246
References 250
6 Reversed-phase Chromatography for Neutral Sam- Ples 253
6.1 Introduction 254
6.2 Retention 256
6.3 Selectivity 263
6.4 Method Development and Strategies for Optimizing Selectivity 284
6.5 Nonaqueous Reversed-Phase Chromatography (narp) 295
6.6 Special Problems 297
References 298
7 Ionic Samples: Reversed-phase Ion-pair and Ion- Exchange Chromatography 303
7.1 Introduction 304
7.2 Acid–Base Equilibria and Reversed-Phase Retention 304
7.3 Separation of Ionic Samples by Reversed-Phase Chromatography (RPC) 319
7.4 Ion-Pair Chromatography (IPC) 331
7.5 Ion-Exchange Chromatography (IEC) 349
References 357
8 Normal-phase Chromatography 361
8.1 Introduction 362
8.2 Retention 363
8.3 Selectivity 376
8.4 Method-Development Summary 385
8.5 Problems in the Use of NPC 392
8.6 Hydrophilic Interaction Chromatography (HILIC) 395
References 401
9 Gradient Elution 403
9.1 Introduction 404
9.2 Experimental Conditions and Their Effects on Separation 412
9.3 Method Development 434
9.4 Large-Molecule Separations 464
9.5 Other Separation Modes 465
9.6 Problems 470
References 471
10 Computer-assisted Method Development 475
10.1 Introduction 475
10.2 Computer-Simulation Software 481
10.3 Other Method-Development Software 491
10.4 Computer Simulation and Method Development 492
References 497
11 Qualitative and Quantitative Analysis 499
11.1 Introduction 499
11.2 Signal Measurement 500
11.3 Qualitative Analysis 516
11.4 Quantitative Analysis 520
11.5 Summary 529
References 529
12 Method Validation 531
with Michael Swartz
12.1 Introduction 532
12.2 Terms and Definitions 534
12.3 System Suitability 542
12.4 Documentation 543
12.5 Validation for Different Pharmaceutical-Method Types 546
12.6 Bioanalytical Methods 548
12.7 Analytical Method Transfer (AMT) 554
12.8 Method Adjustment or Method Modification 561
12.9 Quality Control and Quality Assurance 564
12.10 Summary 565
References 566
13 Biochemical and Synthetic Polymer Separations 569
with Timothy Wehr, Carl Scandella, and Peter Schoenmakers
13.1 Biomacromolecules 570
13.2 Molecular Structure and Conformation 571
13.3 Special Considerations for Biomolecule HPLC 579
13.4 Separation of Peptides and Proteins 584
13.5 Separation of Nucleic Acids 618
13.6 Separation of Carbohydrates 625
13.7 Separation of Viruses 630
13.8 Size-Exclusion Chromatography (SEC) 631
13.9 Large-Scale Purification of Large Biomolecules 641
13.10 Synthetic Polymers 648
References 658
14 Enantiomer Separations 665
with Michael Lämmerhofer, Norbert M. Maier and Wolfgang Lindner
14.1 Introduction 666
14.2 Background and Definitions 666
14.3 Indirect Method 670
14.4 Direct Method 675
14.5 Peak Dispersion and Tailing 681
14.6 Chiral Stationary Phases and Their Characteristics 681
14.7 Thermodynamic Considerations 715
References 718
15 Preparative Separations 725
with Geoff Cox
15.1 Introduction 726
15.2 Equipment for Prep-LC Separation 730
15.3 Isocratic Elution 736
15.4 Severely Overloaded Separation 748
15.5 Gradient Elution 751
15.6 Production-Scale Separation 754
References 755
16 Sample Preparation 757
with Ronald Majors 757
16.1 Introduction 758
16.2 Types of Samples 759
16.3 Preliminary Processing of Solid and Semi-Solid Samples 760
16.4 Sample Preparation for Liquid Samples 764
16.5 Liquid–Liquid Extraction 764
16.6 Solid-Phase Extraction (SPE) 771
16.7 Membrane Techniques in Sample Preparation 790
16.8 Sample Preparation Methods for Solid Samples 791
16.9 Column-Switching 796
16.10 Sample Preparation for Biochromatography 797
16.11 Sample Preparation for LC-MS 800
16.12 Derivatization in HPLC 802
References 805
17 Troubleshooting 809
Quick Fix 809
17.1 Introduction 810
17.2 Prevention of Problems 811
17.3 Problem-Isolation Strategies 819
17.4 Common Symptoms of HPLC Problems 821
17.5 Troubleshooting Tables 865
References 876
Appendix I. Properties of HPLC Solvents 879
I.1 Solvent-Detector Compatibility 879
I. 2 Solvent Polarity and Selectivity 882
I. 3 Solvent Safety 885
References 886
Appendix II. Preparing Buffered Mobile Phases 887
II.1 Sequence of Operations 887
II.2 Recipes for Some Commonly Used Buffers 888
Reference 890
Index 891
About the Author :
Lloyd R. Snyder, PhD, is a Principal at LC Resources in Walnut Creek, California. He is the author or coauthor of several books including An Introduction to Separation Science, Introduction to Modern Liquid Chromatography, Second Edition, the bestselling Practical HPLC Method Development, Second Edition, and the comprehensive High-Performance Gradient Elution, all published by Wiley. Joseph J. Kirkland, PhD, is Vice President of Research and Development for Advanced Materials Technology, Inc., and coauthor of Introduction to Modern Liquid Chromatography, Second Edition, Practical HPLC Method Development, Second Edition, and Modern Size-Exclusion Liquid Chromatography, Second Edition, all published by Wiley.
John W. Dolan, PhD, is a Principal at LC Resources. He is author of the popular "LC Troubleshooting" column in LCGC magazine and coauthor with Lloyd Snyder of Troubleshooting LC Systems and High-Performance Gradient Elution.
Review :
"In summary, I would strongly recommend this book for anyone working with LC and LC/MS. It is a finely crafted introduction that adequately covers nearly every aspect of the science. The emphasis on basic principles and practical aspects ensures that it will be a useful reference for many years to come." (J Am Soc Mass Spectrom, 2011) "This third edition is highly cross-referenced, so as to allow the reader to follow up on topics of special interest, or to clarify questions that may arise during reading. The third edition of Introduction to Modern Liquid Chromatography will continue to be the HPLC reference book for thousands of readers, either experienced workers who may wish to explore topics of his/her choice, or find an answer to specific problems, or beginners who would like to understand and know the possibilities offered by the
technique." (Chemistry Journals, 11 April 2011)
"This classic text on liquid chromatography has been thoroughly updated through the addition of information on modern instrumentation, columns, and troubleshooting. It is a valuable resource for practicing chromatographers at all levels.." (Anal Bioanal Chem, 2011)
"In summary, I would strongly recommend this book for anyone working with LC and LC/MS. It is a finely crafted introduction that adequately covers nearly every aspect of the science. The emphasis on basic principles and practical aspects ensures that it will be a useful reference for any years to come." (American Society for Mass Spectrometry, 21 Januay 2011)
"The text is illustrated with many figures and tables originating from authors' own work or taken from the literature . . . both groups of readers will find in this book plenty of information and inspiration." (Journal of Separation Science, 1 June 2010)
"Following their highly successful second edition (1979), which appeared more than 30 years ago, Snyder (LC Resources) and Kirkland (Advanced Materials Technology) have teamed with Dolan (LC Resources) and additional collaborators to provide an excellent update of their earlier work." (CHOICE, July 2010)
"It is difficult in these times to ever call a scientific book a "bargain", but this truly is. The wealth of information contained in these almost 1000 pages is invaluable. The book is comprehensive, eminently readable and approachable, and highly useful for both the skilled chromatographer and those new to the technique. Everyone using a liquid chromatographic instrument would benefit from owning a copy." (JACS, 2010)
"It's current, clearly-indexed, well-referenced, and comprehensive. The inclusion of John Dolan as author has significantly expanded the sections on troubleshooting, and additional expert contributors have enhanced the coverage of the specialist chapters." - Bruce Hamilton posted on the Chromatography Forum
