Coagulation protein function VI - Augmentation of anticoagulant function by acetaldehyde-treated heparin

Acetaldehyde (AcH) at preincubation concentrations of 447, 89.4, and 17.9 m M potentiates the effects of heparin on the clotting time of plasma. While control plasma clotted in the range of 12.6 +/- 0.1 to 13.8 +/- 0.1 sec, an d heparin-treated plasma clotted in a range from 131.5 +/- 2.5 to 168.2 +/- 1.2 sec, heparin that was preincubated at room temperature for 30 min with 89.4 or 447 mM AcH did not clot plasma in 300 sec. Heparin exposed to 17.9 mM AcH clotted plasma in 193 +/- 1.1 sec, Ethanol at a 404 mM concentratio n also prolonged the clotting time of heparin-treated plasma >300 sec, whil e 202 mM ethanol prolonged the clotting time of heparin-treated plasma from 149.0 +/- 2.0 sec to 219.5 +/- 1.7 sec, It is suggested that AcH alters th e tertiary structure of heparin by adduct formation, possibly by formation of cyclic acetals with iduronic and glucuronic acids, thereby more readily affecting binding of the glycosaminoglycan to antithrombin III and/or throm bin, prolonging clotting time, Ethanol, which does not react covalently wit h heparin, might affect its conformation as a consequence of an organic sol vent effect. Protamine sulfate prolonged the clotting time of plasma from 1 3.6 +/- 0.1 sec to 17.9 +/- 0.2 sec. Protamine sulfate-treated heparin clot ted plasma in 21.0 +/- 0.4 sec relative to heparin-treated plasma (160.4 +/ - 1.7 sec). In subsequent experiments, AcH-treated protamine sulfate extend ed the clotting time of protamine sulfate from 17.9 +/- 0 sec to 33.7 +/- 0 .6 sec. Prior addition of protamine sulfate to AcH-heparin mixtures or hepa rin to protamine sulfate-AcH mixtures before addition to plasma resulted in clotting times of 22.0 +/- 0.4 sec and 24.1 +/- 0.5 sec, respectively, rel ative to control clotting times of 162.3 +/- 2.6 sec for plasma-heparin mix tures. These results confirm both the reduction in coagulation time of hepa rin-treated plasma by protamine sulfate and the prolongation of clotting ti me of plasma by protamine sulfate. Furthermore, and importantly, they indic ate that acetaldehyde-treated protamine sulfate is a more effective anticoa gulant than protamine sulfate. It is suggested that reversible adduct forma tion between acetaldehyde, heparin, and protamine sulfate may occur as a me ans explaining the essentially identical coagulation time of these mixtures when added to plasma regardless of the order of premixing. Ethanol (404 mM ) did not influence protamine sulfate effects. Lastly, the potentiation of the anticoagulant function of heparin by acetaldehyde suggests that a struc tural modification of the glycosaminoglycan may occur in alcoholics.