The effects of biguanides on thrombokinase, thrombin and trypsin.
A purified preparation of bovine thrombokinase (activated Factor X) loses the ability to hydrolyze TAME (p-toluenesulfonyl-L-arginine methyl ester) when it is incubated at 37 degrees in 0.25 M Tris. HCl buffer, pH 7.4 with lauroxypropyl biguanide, N1, N5-dimethyl, N1-lauroxypropyl biguanide, N1-p-chlorophenethyl, N5-phenethyl biguanide, or N1-methyl, N1-p-chlorobenzyl, N5-o,p-dichlorobenzyl biguanide. Activity is lost much more slowly when 0.15 M NaCl is also present. Lauroxypropyl biguanide is the most potent of the compounds tested, 0.22 mM causing thrombokinase to lose almost all of its activity in about 30 minutes at 37 degrees in pH 7.4 buffered saline. Topical bovine thrombin also loses activity when incubated with either of the lauroxypropyl biguanides but not with the diphenethyl or the dibenzyl compound. Instead, the latter biguanides accelerate thrombin's hydrolysis of TAME. The percent acceleration is not affected or only slightly decreased by the presence of 0.15 M NaCl or KCl, and it is also unaffected by incubating the enzyme with the compounds in buffered saline for 4 to 120 minutes. Purified bovine trypsin is stabilized by both lauroxypropyl and the diphenethyl biguanide when incubated at 37 degrees in pH 7.4 buffered saline for the 60 minute test period but neither compounds has any effect on its rate of hydrolysis of TAME. It is postulated that the enzymes first react rapidly and reversibly with all of the test biguanides and, depending upon the enzyme and the substrate, the rate of hydrolysis of the substrate is unaffected, accelerated or inhibited. The lauroxypropyl biguanides also undergo a second, slower reaction with both thrombokinase and thrombin that produces loss of enzymatic activity. The dibenzyl and diphenethyl biguanides also undergo this second slow reaction with thrombokinase but not with thrombin, and none of the biguanides undergo this second reaction with trypsin.
D000818:Animals; D001645:Biguanides / Q000494:pharmacology*; D001769:Blood; D002417:Cattle; D055598:Chemical Phenomena; D002621:Chemistry; D004355:Drug Stability; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D012965:Sodium Chloride; D013329:Structure-Activity Relationship; D013696:Temperature; D013917:Thrombin / Q000037:antagonists & inhibitors*; D013925:Thromboplastin / Q000037:antagonists & inhibitors*; D014106:Tosylarginine Methyl Ester; D014357:Trypsin
D001645:Biguanides; D012965:Sodium Chloride; D014106:Tosylarginine Methyl Ester; D013925:Thromboplastin; D014357:Trypsin; D013917:Thrombin