Differential anticoagulant activity of heparin fragments prepared using microbial heparinase

Heparin of an average molecular weight of 13,000 with known polydispersity was degraded using microbial heparinase. The kinetics of this degradation were followed by four assays which measured the anticoagulant activity of the heparin digestion products. Both clotting and amidolytic chromogenic assays were used to measure heparin-potentiated inhibition of both thrombin and Factor Xa. These assays showed different profiles throughout the digestion and were related to the average molecular weight of the digestion products. The final products of this enzymatic digestion were fractionated on the basis of size and their anticoagulant activities were measured. Fragments causing Factor Xa inhibition but not thrombin inhibition were isolated. Anticoagulant activity was found in a fragment as small as a tetrasaccharide.