Determination of "in vitro" degradation of protamine in plasma by three different methods

Three techniques for the quantitative or semi-quantitative determination of the degradation of protamine in plasma are described. One is based on the measurement of liberated arginine, since arginine is the single most important constituent of protamine (80% in weight). The second utilizes successive estimations of protamine by addition to a secondary heparinized medium in which excess heparin is measured by thrombin time and polybrene titration. The third method employs electrophoresis on cellulose acetate, and offers direct visualization of the soluble complexes formed between protamine and albumin, and of their degradation. When applied to an incubation mixture containing diluted plasma (1 : 8) and protamine 0.8 mg/ml, the first two methods were well correlated and showed that protamine degradation proceeded linearly with time. The third method had good semiquantitative agreement with the two former. The rate of protamine degradation was different when estimated by each of the three methods, due probably to the different physico-chemical reactions involved.