Physico-Chemical Studies On Oligosaccharides From Procine Mucosal Heparin

Our recent studies have revealed that low molecular weight antithrombotic (in vivo) and anti Xa fractions and fragments can be obtained from porcine mucosal heparin (PMH) using three different processes: 1) Direct alcoholic fractionations, 2) depolymerization with nitrous acid and 3) by bacterial heparinase digestion. Ihe crude products are further fractionated by anti thrombin-III affinity chromatography and gel filtration. Physiocochemical studies were carried on each fragment employing nitrous acid degradation, colorimetric and NMR analysis. A potent antithrombotic octasaccharide (h-ULMF) is isolated from heparinase depolymerized fractions and in the NMR analysis gave some unexpected signals one of which was consistant to a 3-0-sulfated glucosamine unit (unsaturated sulfated uronic acid/N-Sulfo-Glucosamine/Iduronic acid/N- Acetyl-G1ucosamine/Glucouronic acid/N-Sulfoglucosami ne/Sulfated Idouronic acid/N-Sulfo-Glucosamine). ULMFs obtained by extraction and clemical degradation were also found to exhibit similar signals. Further degradation of h-ULMF-8 resulted in formation of smaller fragments with biological activity. Our results suggest that a smaller oligosaccharide fragment contained in the biologically active octasaccharide molecule is primarily responsible for the antithrombotic actions. Furthermore these studies provide additional data on the structure activity relationship for the antithrombotic actions of heparin, its fraction and fragments.