Preparation of active, drug-metabolizing, microsomal enzymes under optimal conditions and by iso-electric precipitation

Abstract

The influences of homogenization technique and of relative centrifugal force on the enzyme activity of hepatic microsomes have been investigated to establish optimal conditions for preparation. In general, a glass or Ultra Turrax homogenizer provided a satisfactory homogenate having maximal enzyme activity when nuclear and mitochondrial material were removed by centrifugation at >6500 g for 20 min. Esterase, UDP-transglucuronylase, reductase and oxidase activities of microsomes prepared from liver breis under optimal conditions from guinea-pig, rabbit and rat have been compared. The enzyme activities in microsomes prepared by isoelectric precipitation at pH 5·0 were similar to those prepared by differential centrifugation except for a 50–75% reduction in glucuronyl transferase activity observed in rabbit and guinea-pig.