Capillary Electrophoresis For Total Glycosaminoglycan Analysis

A capillary zone electrophoresis-laser-induced fluorescence detection (CZE-LIF) method was developed for the simultaneous analysis of disaccharides derived from heparan sulfate, chondroitin sulfate/dermatan sulfate, hyaluronan, and keratan sulfate. Glycosaminoglycans (GAGs) were first depolymerized with the mixture of GAG lyases (heparinase I, II, III and chondroitinase ABC and chondroitinase AC II) and GAG endohydrolase (keratinase II) and the resulting disaccharides were derivatized by reductive amination with 2-aminoacridone. Nineteen fluorescently labeled disaccharides were separated using 50 mM phosphate buffer (pH 3.3) under reversed polarity at 25 kV. Using these conditions, all the disaccharides examined were baseline separated in less then 25 min. This CZE-LIF method gave good reproducibility for both migration time (a parts per thousand currency sign1.03 % for intraday and a parts per thousand currency sign4.4 % for interday) and the peak area values (a parts per thousand currency sign5.6 % for intra- and a parts per thousand currency sign8.69 % for interday). This CZE-LIF method was used for profiling and quantification of GAG derivative disaccharides in bovine cornea. The results show that the current CZE-LIF method offers fast, simple, sensitive, reproducible determination of disaccharides derived from total GAGs in a single run.