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Clinical grade heparin is a very heterogeneous mucopolysaccharide, containing molecules with Mr ranging from 6,000 to 30,000 that have either a high affinity or a low affinity for antithrombin III (AT). In this study, the antithrombotic properties of intact high-affinity heparin (Mr = 15,000) and of two heparin fragments (h16, a 16-monosaccharide fragment, with Mr = 4,300, and h12, a 12-monosaccharide fragment, with Mr = 3,200) and of their functional covalent stoichiometric complexes with human AT were compared in a venous thrombosis stasis model in rabbits. Thrombosis was induced by injection of glass-activated human plasma and measured in a segment of the jugular vein that was isolated between two vascular clamps for 10 min. Injections of 55 micrograms/kg resulted in a clear antithrombotic effect for intact heparin, but not for the two fragments. Equivalent amounts (carbohydrate moiety) of covalent complexes of heparin or of both heparin fragments with human AT resulted in an antithrombotic effect lasting for 45-60 min. Injection of 110 micrograms/kg of heparin and of the heparin fragments yielded an antithrombotic effect, lasting 45-60 min; the corresponding amounts of covalent complexes caused an anti-thrombotic effect for 60-120 min. The free and conjugated fragments produced equal antithrombotic effects at equal plasma levels of anti-Factor Xa activity, but the specific antithrombotic activities of free and complexed intact heparin, on a molar basis, were 10-20-fold greater than those of the free and complexed heparin fragments. The plasma half-life of the covalent complexes of the heparin fragments with AT is, however, 10 times longer than that of the complex between intact heparin and AT and 30 times longer than that of free intact heparin. Covalent complexes between AT and heparin fragments could, therefore, be useful to maintain more stable levels of antithrombotic activity in plasma.

Antithrombotic properties in rabbits of heparin and heparin fragments covalently coupled to human antithrombin III.