Analysis of 3-O-sulfo group containing heparin tetrasaccharides in heparin by liquid chromatography-mass spectrometry

摘要

Complete heparin digestion with heparin lyase 2 affords a mixture of disaccharides and resistant tetra-saccharides with 3-O-sulfo group containing glucosamine residues at their reducing ends. Quantitative on-line liquid chromatography-mass spectrometric analysis of these resistant tetrasaccharides is described. The disaccharide and tetrasaccharide composition of seven porcine intestinal heparins and five low molecular weight heparins were analyzed by this method. These resistant tetrasaccharides account for from 5.3 to 7.3 wt% of heparin and from 6.2 to 8.3 wt% of low molecular weight heparin. Since these tetrasaccharides are derived from heparin’s antithrombin III-binding sites, we examined whether this method could be applied to estimate the anticoagulant activity of heparin. The content of 3-O-sulfo group containing tetrasaccharides in a heparin positively correlated (r = 0.8294) to heparin’s anticoagulant activity.