ISSN:
0173-0835
Keywords:
Capillary electrophoresis
;
Cyclodextrins
;
Glycoproteins
;
Oligosaccharides
;
Chemistry
;
Biochemistry and Biotechnology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Chemistry and Pharmacology
Notes:
Oligomers of glucose and oligosaccharides released from glycoproteins were derivatized with 2-aminobenzamide. As this fluorophore imparts no charge to the oligosaccharides, several strategies were investigated to achieve capillary electrophoresis (CE) separation of both neutral and charged derivatized glycans. Micellar electrokinetic capillary chromatography (MEKC) with the addition of anionic surfactants was evaluated as a first approach: sodium dodecyl sulfate (SDS) produced the best separation of the oligoglucose fragments, where the migration was inversely related to their degree of polymerization. To demonstrate the applicability of this method for complex carbohydrate analysis, oligosaccharide mixtures derived from ribonuclease B (RNase B) and α-acid glycoprotein (α-AGP) were analyzed. A satisfactory separation for the high-mannose structures found in RNase B could be obtained, whereas charged oligosaccharides from α-AGP were poorly resolved. Cyclodextrin-modified CE was chosen as the second approach: the effect of the addition of sulfobutylether-β-cyclodextrin (SBE-β-CD) or sulfobutylether-γ-cyclodextrin (SBE-γ-CD) on the electrophoretic mobilities and resolution of neutral and charged oligosaccharides was then studied. Selectivity of sialylated structures could be further improved by using anionic cyclodextrins (CDs) instead of micelles. However, this latter approach failed to baseline-resolve the different high-mannose structures of RNase B. A successful separation of the complex mixture of oligosaccharides from α-AGP was obtained with the addition of 4% of SBE-γ-CD and triethylamine (TEA) in a phosphate buffer, pH 6.7.
Additional Material:
8 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/elps.1150191511
Permalink
