High performance capillary electrophoresis method to characterize heparin and heparan sulfate disaccharides

A rapid, sensitive and accurate high-performance capillary electrophoresis method is described for the determination of the sulfation pattern of heparin and heparan sulfate disaccharides. The analysis, performed after enzymic degradation of the polysaccharides with heparinase and heparinases II and III in combination, yields highly UV-absorbing delta-disaccharides. The separation is performed with reversed polarity using 15 mM phosphate buffer, pH 3.50. This method is superior to others since all known 12 disaccharides carrying N-acetylated, N-sulfated or unsubstituent glucosamine can be separated in a single run of 15 min. At the highest sensitivity the analysis consumes only a few femtograms of glycosaminoglycan and allows a determination of delta-disaccharides at the attomole level.