The mode of inactivation of insulin by rat liver extracts.

Studies with insulin-P in vivo have suggested that insulin is rapidly cleaved after intravenous injection into rats (3-5). This apparent degradation was found to be significantly decreased after hepatectomy (3), an observation consistent with the finding that the liver is capable of inactivating or removing insulin in vivo (6). In 1929, Shmidt and Saatchian reported that inactivat,ion of insulin by rat tissue extracts occurred to the greatest extent with extracts of the liver (7). Their finding was confirmed by Mirsky and Broh-Kahn, who carried out extensive studies with the liver extract system at pH 7.5 (8). These workers reported that denatured hemoglobin was not proteolytically degraded by this insulin-inactivating system and that soy bean inhibitor and human serum were not inhibitory to the inactivation of insulin. They therefore suggested the possibility of a non-proteolytic inactivation of insulin. This question of proteolysis was reinvestigated by De Barbieri and Grassi (9), who found that denatured globin and peptic hydrolysates of globin were degraded by rat liver extract at pH 7.5 and 3.5. They suggested that, since proteolytic activity was demonstrable, insulin was probably proteolytically inactivated. In order to try to obtain a definitive answer to this question, a program was initiated to invest,igate the mode of insulin inactivation by rat liver extract. In this study, evidence was obtained which indicates that insulin is probably proteolytically inactivated by rat liver extract. Preparation of Extract-This was essentially the procedure of Mirsky and Broh-Kahn (8). Sprague-Dawley rats weighing 200 to 300 gm. were used without fasting. They mere decapitated, and the liver was excised and placed in ice-cold water. In the cold room, the liver was homogenized for 2 minutes with water or 0.067 M phosphate buffer, pH 7.5. After cen-