Volpi Capillary Electrophoresis of Carbohydrates

From Monosaccharides to Complex Polysaccharides
1. Auflage 2010
ISBN: 978-1-60761-875-1
Verlag: Humana Press
Format: PDF
Kopierschutz: 1 - PDF Watermark

E-Book, Englisch, 277 Seiten

Simple carbohydrates, complex oligosaccharides and polysaccharides all belong to a class of ubiquitous (macro)molecules that exhibit a wide range of biological functions, and the recent advent of enhanced enzymatic, chemical and analytical tools used to study these sugars has inaugurated a genuine explosion in the field of glycomics. Specifically, it has led to a deeper understanding of how specific sugar structures modulate cellular phenotypes, and that breakthrough has led to the discovery of new pharmaceuticals for the treatment of many serious diseases, such as cancer. The subsequent rapid expansion of this research holds high promise for future therapeutic regimens, and capillary electrophoresis (CE) refers to the range of related separation techniques that are integral to this vital research. CE uses narrow-bore fused-silica capillaries to separate a complex array of large and small molecules, and Capillary Electrophoresis of Carbohydrates offers a comprehensive look at the latest breakthroughs and improvements in CE and CE techniques applied to monosaccharides up to complex oligosaccharides and polysaccharides. It begins with an overview of the application of CE and CE- mass spectrometric in the analysis of simple carbohydrates without any previous derivatization step before discussing various detection techniques such as spectrophotometric detection, electrochemical detection and other less common techniques. It then covers in detail an array of related topics and numerous applications. It is an essential text for anyone exploring the myriad possibilities of this rapidly expanding field.

Autoren/Hrsg.

Volpi, Nicola

Inhaltsverzeichnis

1;Preface;6
2;Contents;10
3;Contributors;12
4;Chapter 1: Analysis of Simple Carbohydrates by Capillary Electrophoresis and Capillary Electrophoresis–Mass Spectrometry;16
4.1;1.1 Introduction;17
4.2;1.2 Determination of Simple Carbohydrates by CE;17
4.2.1;1.2.1 Detection Systems in CE;24
4.2.1.1;1.2.1.1 Spectrophotometric Detection for the Determination of Underivatized Carbohydrates by CE;24
4.2.1.2;1.2.1.2 Electrochemical Detection for the Determination of Carbohydrates by CE;27
4.2.1.3;1.2.1.3 Other Detection Methods for the Determination of Carbohydrates by CE;29
4.2.2;1.2.2 Electrolyte Systems for the CE-Separation of Simple Carbohydrates;30
4.2.3;1.2.3 Analysis of Simple Carbohydrates in Real Samples by CE;32
4.3;References;32
5;Chapter 2: Fluorophores and Chromophores for the Separation of Carbohydrates by Capillary Electrophoresis;37
5.1;2.1 Introduction;38
5.2;2.2 Reductive Amination;40
5.3;2.3 Amination Via Formation of Glycosylamine;53
5.4;2.4 Derivatization with N-Methyl-glycamine Derivatives;53
5.5;2.5 Formation of Hydrazones;55
5.6;2.6 Derivatization with Pyrazolone Compounds;56
5.7;2.7 Derivatization at Carboxylic Acid Functionalities;58
5.8;2.8 On-Column Derivatization;59
5.9;2.9 Conclusion and Future Directions;61
5.10;References;61
6;Chapter 3: Capillary Electrophoresis of Bacterial (Lipo)Polysaccharides;66
6.1;3.1 Introduction;68
6.2;3.2 Capillary Electrophoresis of Lipopolysaccharides;69
6.2.1;3.2.1 On-Gel Hydrolysis and CE-LIF Monosaccharide Composition of Excised Lipopolysaccharides;71
6.2.2;3.2.2 CE Chips for Screening of Endotoxin Chemotypes from Whole-Cell Lysates;72
6.2.3;3.2.3 Capillary Zone Electrophoresis (CZE) Separation of O-Deacylated LPS and Polysaccharide Derivatives;74
6.2.4;3.2.4 CE for Quantitative Analysis of Extracted Lipopolysaccharides;75
6.2.5;3.2.5 CZE for the Determination of Meningococcal Polysaccharides;75
6.2.6;3.2.6 CE to Monitor Lipopolysaccharides (Endotoxins) by Protein Complexation;78
6.2.7;3.2.7 CE of Alginic Acid;80
6.2.8;3.2.8 Characterization of Lipopolysaccharides by CE-MS;82
6.2.8.1;3.2.8.1 Characterization of Intact LPS from the H. influenzae by Electrophoresis-Assisted Open-Tubular Liquid Chromatography-MS;82
6.2.8.2;3.2.8.2 Carbohydrate Analysis of Typical and Atypical Isolates of A. salmonicida for the LPS-Based Classification;83
6.2.8.3;3.2.8.3 CE and High-Field Asymmetric Waveform Ion Mobility Spectrometry MS for the Analysis of LPS;84
6.2.8.4;3.2.8.4 Glycotyping by CE-MS of Pseudomonas aeruginosa Isolates from Patients with Cystic Fibrosis;85
6.2.8.5;3.2.8.5 Characterization of Short-Chain LPS Glycoforms by CE-MS;86
6.2.8.6;3.2.8.6 Isomer Separation and Characterization by CE-MS;86
6.2.8.7;3.2.8.7 In-Source Fragmentation Strategy for Polysaccharide Analysis;87
6.3;3.3 CE Analysis of Bacterial Glycosaminoglycan-Like Polysaccharides;87
6.3.1;3.3.1 General Considerations;87
6.3.2;3.3.2 CE Analysis of E. coli K4 Polysaccharide;88
6.3.3;3.3.3 CE Analysis of E. coli K5 Polysaccharide (Heparosan);90
6.4;3.4 Conclusion;91
6.5;References;92
7;Chapter 4: Capillary Electrophoresis and Its Microchip Format for the Analysis of Glycosaminoglycans;96
7.1;4.1 Introduction;97
7.2;4.2 CE Analysis of GAGs;99
7.2.1;4.2.1 Analysis of Hyaluronic Acid and Its Oligosaccharides;99
7.2.2;4.2.2 Analysis of Sulfated GAGs;101
7.2.2.1;4.2.2.1 Intact Sulfated GAGs;101
7.2.2.2;4.2.2.2 Unsaturated Disaccharides;103
7.3;4.3 Microchip Electrophoresis of GAGs;104
7.3.1;4.3.1 Analysis of Di- and Oligosaccharides;106
7.3.2;4.3.2 Analysis of GAG Polysaccharides;109
7.4;4.4 Conclusion;112
7.5;References;113
8;Chapter 5: Extracellular Polysaccharides in Microbial Biofilm and Their Influence on the Electrophoretic Properties of Microbial Cells;117
8.1;5.1 Biofilm: The Mode of Bacterial Life;118
8.1.1;5.1.1 Extracellular Polymeric Matrix;118
8.1.2;5.1.2 Extracellular Polysaccharide Matrix in Staphylococci;119
8.1.3;5.1.3 Extracellular Matrix in Pseudomonas Biofilm and Alginate;120
8.1.4;5.1.4 Extracellular Polysaccharides in Candida Biofilms;120
8.1.5;5.1.5 Production of Extracellular Polysaccharides in Streptococci, Lactobacilli, and Multispecies Oral Biofilms;121
8.2;5.2 Capillary Electrophoresis of Microbial Cells;122
8.2.1;5.2.1 The Surface Charge of Microorganisms;122
8.2.2;5.2.2 The Isoelectric Point of Microorganisms;124
8.2.3;5.2.3 Capillary Electromigration Techniques;124
8.2.3.1;5.2.3.1 Capillary Surface Modification;125
8.2.3.2;5.2.3.2 Detection of the Microorganisms in Capillary Electrophoresis;126
8.2.3.3;5.2.3.3 CZE of Microorganisms;126
8.2.3.4;5.2.3.4 CIEF of Microorganisms;128
8.3;5.3 Capillary Electromigration Techniques: A Useful Tool for the Detection of Biofilm Formation, an Important Virulence Factor in Microorganisms;128
8.4;References;132
9;Chapter 6: Capillary Electrophoresis Applied to Polysaccharide Characterization;139
9.1;6.1 Introduction;140
9.2;6.2 Structural Studies Via Degradation Mechanisms;141
9.3;6.3 Studies of Macromolecular Features;148
9.4;6.4 Capillary Electrophoresis for Practical Uses;153
9.5;6.5 Capillary Electrophoresis in Biomedical and Biological Application Fields;155
9.6;6.6 Conclusion;157
9.7;References;158
10;Chapter 7: Capillary Electrophoresis for Monitoring Natural and Synthetic Processes of Saccharide-Bearing Molecules;162
10.1;7.1 Introduction;163
10.2;7.2 Synthesis Involving Mono- and Oligo-saccharide Species;165
10.3;7.3 Glycoconjugate Preparation;170
10.3.1;7.3.1 Saccharidic Backbone;170
10.3.2;7.3.2 Non-saccharidic Backbone;172
10.4;7.4 Saccharidic Species as Enzyme Substrate: Studies on Enzyme Activity and Mechanism of Action;172
10.5;7.5 Studies of Natural Processes and of Reactions in Single Cells;177
10.6;7.6 Conclusion;179
10.7;References;179
11;Chapter 8: Analysis of Intact Glycoprotein Biopharmaceuticals by Capillary Electrophoresis;184
11.1;8.1 Introduction;185
11.2;8.2 General Considerations;187
11.3;8.3 Capillary Electrophoresis and Glycoprotein Pharmaceuticals;189
11.3.1;8.3.1 Impact of the Process;189
11.3.1.1;8.3.1.1 Host Cell or Expression System;189
11.3.1.2;8.3.1.2 Cell Culture Conditions;191
11.3.1.3;8.3.1.3 Purification Process;193
11.3.2;8.3.2 Quality Control;194
11.3.2.1;8.3.2.1 In-Process Monitoring;194
11.3.2.2;8.3.2.2 Purified Products;195
11.3.2.3;8.3.2.3 Finished Product;198
11.3.3;8.3.3 Stability Testing;204
11.4;8.4 CE in Biological Fluids for Clinical Application;206
11.5;8.5 Conclusion;209
11.6;References;210
12;Chapter 9: Capillary Electrophoresis and Capillary Electrophoresis–Mass Spectrometry for Structural Analysis of N-Glycans Derived from Glycoproteins;216
12.1;9.1 Introduction;218
12.2;9.2 Isolation and Direct Analysis of Intact Glycoproteins;220
12.3;9.3 Release of N-Glycans from Glycoproteins;223
12.3.1;9.3.1 Chemical Release;223
12.3.2;9.3.2 Enzymatic Release;224
12.4;9.4 Analysis of N-Glycans by Capillary Electrophoresis;224
12.4.1;9.4.1 Derivatization of Released N-Glycans and the Detection;224
12.4.2;9.4.2 Separation Modes;228
12.4.2.1;9.4.2.1 Capillary Zone Electrophoresis;229
12.4.2.2;9.4.2.2 Capillary Gel Electrophoresis;231
12.4.2.3;9.4.2.3 Capillary Isoelectric Focusing;231
12.4.2.4;9.4.2.4 Micellar Electrokinetic Chromatography;231
12.4.2.5;9.4.2.5 Capillary Electrochromatography;232
12.4.2.6;9.4.2.6 Capillary Affinity Electrophoresis;232
12.5;9.5 Analysis of N-Glycans by Capillary Electrophoresis–Mass Spectrometry;233
12.5.1;9.5.1 Interfacing with Mass Spectrometry;235
12.5.2;9.5.2 Derivatization of N-Glycans for Capillary Electrophoresis–Mass Spectrometry Analysis;236
12.5.3;9.5.3 Capillary Zone Electrophoresis–Mass Spectrometry;237
12.5.4;9.5.4 Capillary Electrochromatography–Mass Spectrometry;239
12.6;9.6 Conclusion;239
12.7;References;240
13;Chapter 10: Monosaccharide Compositional Analysisof Glycoproteins and Glycolipids: Utilityin the Diagnosis/Prognosis of Diseases;247
13.1;10.1 Introduction;248
13.2;10.2 Chemical and Enzymatic Release for Monosaccharide Compositional Analysis;249
13.3;10.3 Chromatographic and Electrophoretic Methods Developed for Monosaccharide Compositional Analysis of Glycoproteins and Glycolipids;251
13.3.1;10.3.1 High-Performance Anion Exchange Chromatography with Pulsed Amperometric Detection;251
13.3.2;10.3.2 High-Performance Liquid Chromatography with Fluorescence Detection;256
13.3.3;10.3.3 High-Performance Liquid Chromatography with Mass Spectrometry;262
13.3.4;10.3.4 Capillary Electrophoresis with Laser-Induced Fluorescence;265
13.4;10.4 Monosaccharide Compositional Analysis of Glycoproteins and Glycolipids in the Diagnosis/Prognosis of Diseases;268
13.5;10.5 Conclusion;270
13.6;References;271
14;Index;278