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Chromogenic peptide substrates S-2222 designed for F. Xa assay is also amidolyzed relatively slowly by thrombin; in the usual F. Xa assay system however a considerable amount of thrombin is inevitably formed. In the present study, therefore, No. 805, highly selective synthetic thrombin inhibitor, (4 methyl- (methyl-tetrahydro-quino- linesulfonyl-arginyl ][ 2-piperidinecarboxylic acid), having extremely potent stereostructure (2R, 4R) at 4-methyl-2- piperidinecarboxylic acid portion, which was reported by S. Okamoto et al. in 1980, was used to improve selectivity of S-2222 for F. Xa assay. F. Xa in human plasma activated by either Russel’s viper venom or thromboplastin was estimated according to the methods by Aurell et al. . Ki of No. 805 for thrombin was found 0.019 juM and presence of No. 805 of 5 mM inhibited thrombin at least by 99.5%. However, No. 805 of this concentration showed no inhibition for purified F. Xa. Results obtained from chromogenic assay for F. Xa by S-2222 with or without No. 805 of 5 mM indicated that 10̴20% of amidolysis was derived from the action of thrombin formed in the assay system. Concerning high Km value of S-2222 to thrombin, molecular concentration of thrombin formed in the assay system was calculated as 3̴6 times of the F. Xa molecular concentration. Use of highly selective and potent synthetic thrombin inhibitor seems to be promissing to improve the selectivity of some other chromogenic assay also.

thrombosis and haemostasis

Selectivity Increase Of Chromogenic Assay Of Factor Xa By Use Of Highly Selective Synthetic Thrombin Inhibitor Having Extremely Potent Stereostructure (No. 805)