Gradient HPLC for Practitioners - RP, LC-MS, Ion Analytics, Biochromatography, SFC, HILIC
This practical guide for analytical scientists explains the use of gradients in liquid chromatography. The fundamentals of gradient separations, as well as the most common application scenarios are addressed, from LC-MS coupling to biochromatography to the separation of ionic substances. Throughout,...
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| Médium: | E-kniha |
| Jazyk: | English |
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Newark
John Wiley & Sons
2019
John Wiley & Sons, Incorporated Wiley-Blackwell Wiley-VCH |
| Vydanie: | 1 |
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| ISBN: | 9783527344086, 352734408X |
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- Title Page The Structure of the Book Preface Table of Contents 1. Aspects of Gradient Optimization 2. Instrumental Influences on the Quality and Performance of Gradient Methods and Their Transfer between Different HPLC Devices 3. Optimization of a Reversed-Phase Gradient Separation Using Excel 4. Gradient Elution of Ionic Compounds 5. The Gradient in Biochromatography 6. Specifications of Gradients in Hydrophilic Interaction Liquid Chromatography (HILIC) 7. Specifications of Gradients in Supercritical Fluid Chromatography 8. Aspects of Gradient Elution in LC-MS Analysis 9. Additional Tools for Method Development: Flow and Temperature Gradients Index
- 5.4.5 Affinity Chromatography (AC) -- 5.5 Summary -- 6 Specifications of Gradients in Hydrophilic Interaction Liquid Chromatography (HILIC) -- 7 Specifications of Gradients in Supercritical Fluid Chromatography -- 7.1 Types of Gradients in SFC -- 7.1.1 Mobile Phase Gradients -- 7.1.2 Pressure Gradients -- 7.1.3 Temperature Gradients -- 7.2 Effects of gradients -- 8 Aspects of Gradient Elution in LC-MS Analysis -- 8.1 Role and Importance of Gradient Elution for LC-MS -- 8.2 Technical Aspects of Gradient Elution in LC-MS Analysis -- 8.2.1 Technical Impact of the LC System: System Dispersion, Gradient Proportioning Precision, and How they Affect MS Results -- 8.2.2 Technical Impacts of a Mass Spectrometer: LC Gradients and Signal Generation in the MS -- 8.2.3 Quantitation in Mass Spectrometry Within a Gradient Separation: Matrix Effects and How to Address Them -- 8.2.4 MS Workload Balancing in Gradient Elution - Column Equilibration as a Throughput Bottleneck -- 8.2.5 Gradient Delay, Flow Rate, and Column Dimension - How Far Can we Get With Downsizing of Gradient Separations in LC-MS? -- 8.3 Summary -- 8.4 Abbreviations -- 9 Additional Tools for Method Development: Flow and Temperature Gradients -- 9.1 Introduction -- 9.2 Temperature Gradients -- 9.3 Flow Gradients -- 9.4 Combination of Flow and Temperature Gradients -- 9.5 Case Example -- 9.6 Conclusions -- Index -- EULA
- Cover -- Main title -- Copyright page -- Contents -- Preface -- The Structure of the Book -- Notes on Contributors -- List of Contributors -- Part 1 Principles of Gradient Elution -- 1 Aspects of Gradient Optimization -- 1.1 Introduction -- 1.2 Special Features of the Gradient -- 1.3 Some Chromatographic Definitions and Formulas -- 1.4 Detection Limit, Peak Capacity, Resolution - Possibilities for Gradient Optimization -- 1.4.1 Detection Limit -- 1.4.2 Peak Capacity and Resolution -- 1.5 Gradient "Myths" -- 1.6 Examples for the Optimization of Gradient Runs: Sufficient Resolution in an Adequate Time -- 1.7 Gradient Aphorisms -- 2 Instrumental Influences on the Quality and Performance of Gradient Methods and Their Transfer Between Different HPLC Devices -- 2.1 Technical Implementation of the Gradient Elution and the Respective Characteristics -- 2.1.1 Low-Pressure and High-Pressure Gradient Proportioning -Two Fundamentally Different Technical Principles -- 2.1.2 The Role of the Mixing Device in HPG and LPG Systems -- 2.1.3 The Operational Principle of Mixing Devices and the Systematic Characterization of Their Effectiveness -- 2.1.4 Effects of Volume Contraction when Mixing Water and Organic Solvents in Gradient Instruments -- 2.1.5 Effects of Minimum Leak Rates of Pump Heads in Sensitive Applications and HPG Synchronization Techniques to Correct Them -- 2.2 The Determination and Significance of the Gradient Delay Volume of the System -- 2.2.1 The Determination of the GDV and its Dependence on the Specific Operation Conditions of the System -- 2.2.2 The Influence of GDV on the Chromatographic Results -- 2.2.3 Possibilities of the User to Influence System GDV and its Impact on Chromatography -- 2.3 The Transfer of Gradient Methods Between Different HPLC Systems -- 2.3.1 Practical Tips for Dealing with Deviating GDVs and Possible Countermeasures
- 2.3.2 The Relevance of the Pressure Dependence of the GDV in Method Transfer -- 2.3.3 Effect of a too High Elution Strength of the Sample Solvent in the Presence of Weakly Eluting Solvent at the Gradient Start -- 2.4 Influence of Fluctuations of the Eluent Composition on the Quality of the Detection -- 2.4.1 Influence of a Reference Channel on the Baseline in Diode Array Detectors -- 2.4.2 The Special Challenge in Methods with UV-Absorbing Retained Additives in the Mobile Phase such as TFA -- 2.5 Other Kinds of Practical Application of Gradient Systems in HPLC -- 2.5.1 Alternative and Combined Gradient Modes in HPLC -- 2.5.2 Advantages in the Implementation of Isocratic Methods with Gradient Instruments -- 2.5.3 Use of Gradient Systems in Method Development and Method Optimization -- 3 Optimization of a Reversed-Phase Gradient Separation Using EXCEL -- Part 2 Specifics of the Gradient in Different Elution Modes -- 4 Gradient Elution of Ionic Compounds -- 4.1 Introduction -- 4.2 Theoretical Aspects -- 4.3 Gradient Types in Ion Chromatography -- 4.4 Choice of Eluent -- 4.4.1 Possibilities for Optimizing Concentration Gradients -- 4.5 Gradient Elution of Anions on Anion Exchangers -- 4.6 Gradient Elution of Cations on Cation Exchangers -- 4.6.1 pH Gradients for the Separation of Monoclonal Antibodies -- 4.7 Gradient Elution of Anions and Cations on Mixed-Mode Stationary Phases -- 5 The Gradient in Biochromatography -- 5.1 Biomolecules -- 5.2 Biochromatography -- 5.3 The Gradient in Biochromatography -- 5.3.1 A Gradient you Should Definitely Avoid … -- 5.4 Gradients for Different Biochromatographic Techniques -- 5.4.1 Gel-Filtration, Size-Exclusion Chromatography (SEC) -- 5.4.2 Ion Exchange Chromatography (IEX) -- 5.4.3 Hydrophobic Interaction Chromatography (HIC) -- 5.4.4 Reversed-Phase Chromatography of Biomolecules