High-Performance Gradient Elution (eBook)

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, and the bestselling Practical HPLC Method Development, 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.

PREFACE.

GLOSSARY OF SYMBOLS AND TERMS.

1 INTRODUCTION.

1.1 The "General Elution Problem" and the Need forGradient Elution.

1.2 Other Reasons for the Use of Gradient Elution.

1.3 Gradient Shape.

1.4 Similarity of Isocratic and Gradient Elution.

1.5 Computer Simulation.

1.6 Sample Classification.

2 GRADIENT ELUTION FUNDAMENTALS.

2.1 Isocratic Separation.

2.2 Gradient Separation.

2.3 Effect of Gradient Conditions on Separation.

2.4 Related Topics.

3 METHOD DEVELOPMENT.

3.1 A Systematic Approach to Method Development.

3.2 Initial Experiments.

3.3 Developing a Gradient Separation: Resolution versusConditions.

3.4 Computer Simulation.

3.5 Method Reproducibility and Related Topics.

3.6 Additional Means for an Increase in SeparationSelectivity.

3.7 Orthogonal Separations.

4 GRADIENT EQUIPMENT.

4.1 Gradient System Design.

4.2 General Considerations in System Selection.

4.3 Measuring Gradient System Performance.

4.4 Dwell Volume Considerations.

5 SEPARATION ARTIFACTS AND TROUBLESHOOTING.

5.1 Avoiding Problems.

5.2 Method Transfer.

5.3 Column Equilibration.

5.4 Separation Artifacts.

5.5 Troubleshooting.

6 SEPARATION OF LARGE MOLECULES.

6.1 General Considerations.

6.2 Biomolecules.

6.3 Synthetic Polymers.

7 PREPARATIVE SEPARATIONS.

7.1 Introduction.

7.2 Isocratic Separation.

7.3 Gradient Separation.

7.4 Severely Overloaded Separation.

8 OTHER APPLICATIONS OF GRADIENT ELUTION.

8.1 Gradient Elution for LC-MS.

8.2 Ion-Exchange Chromatography.

8.3 Normal-Phase Chromatography.

8.4 Ternary- or Quaternary-Solvent Gradients.

9 THEORY AND DERIVATIONS.

9.1 The Linear Solvent Strength Model.

9.2 Second-Order Effects.

9.3. Accuracy of Gradient Elution Predictions.

9.4 Values of S.

9.5 Values of N in Gradient Elution.

Appendix I: THE CONSTANT-S APPROXIMATION IN GRADIENTELUTION.

Appendix II: ESTIMATION OF CONDITIONS FOR ISOCRATIC ELUTION,BASED ON AN INITIAL GRADIENT RUN.

Appendix III: CHARACTERIZATION OF REVERSED-PHASE COLUMNS FORSELECTIVITY AND PEAK TAILING.

Appendix IV: SOLVENT PROPERTIES RELEVANT TO THE USE OF GRADIENTELUTION.

Appendix V: THEORY OF PREPARATIVE SEPARATION.

Appendix VI FURTHER INFORMATION ON VIRUS CHROMATOGRAPHY.

Index.

"This book is clear, well written, and easy to understand despite
the complexity of the subject." (Journal of the American
Chemical Society, July 2007)