UDP glucuronosyltransferase and mixed function oxidase activity in microsomes prepared by differential centrifugation...

Microsomes were prepared from the liver, kidney and lung of phenobarbital or 20-methylcholanthrene treated and control rats with the conventional ultracentrifugation and calcium aggregation methods. The two methods were compared as to the yield of microsomal protein, amount of cytochrome P-450/448 and activity of UDP-GLUCURONOSYLTRANSFERASE, BENZOPYRENE HYDROXYLASE AND P-NITROANISOLE O-demethylase. The absolute amount of cytochrome P-450/448 (nmol/g wet weight), as well as the enzymatic activities dependent on it (nmol produced/g wet weight) did not differ significantly in any tissue of either treated or control animals nor did that of UDPglucuronosyltransferase. However, the ultracentrifugation method resulted in a slightly smaller yield of the hepatic microsomal protein and a correspondingly higher yield of cytochrome P-450/448 per mg protein as well as higher specific enzymatic activities of both the consecutive drug biotransformation reactions studied. The specific activity of UDPglucuronosyltransferase in digitonin treated microsomes was twice as high in the conventional microsomes as in the calcium aggregated microsomes; no differences was found in the trypsin treated microsomes. The specific activity of the hepatic benzpyrene hydroxylase of the benzpyrene treated animals in the calcium harvested microsomes was 55 per cent of that in the ultracentrifugated microsomes.