FURTHER STUDIES ON THE METABOLISM OF DRUGS BY SUBFRACTIONS OF HEPATIC MICROSOMES

¹ Department of Pharmacology, College of Medicine, University of Iowa, Iowa City, Iowa

Two different methods have been employed to subfractionate hepatic microsomes from rats and rabbits into predominantly smooth-surfaced fractions and predominantly rough-surfaced fractions. Electron micrographs of the pellets have shown them to be essentially homogeneous, but the purest fraction was that of the smooth-surfaced microsomes. Smooth- and rough-surfaced microsomes prepared by both procedures from both species were studied for their capacity to metabolize several drug substrates in the presence of a reduced nicotinamide adenine dinucleotide phosphate (NADPH)-generating system. Employing the Dallner method with rat liver, we found significant concentration of enzyme activity in the smooth-surfaced fraction for the metabolisms of all eight drug substrates studied;smooth/rough ratios ranged between 1.5:1 and 3:1. Using the Rothschild method in this same species, we found significant concentration of enzyme activity in the smoothsurfaced microsomal fraction for the metabolisms of only five of the eight substrates studied; the other three enzyme activities, i.e., nitro-reductase, azo-reductase and benzpyrene hydroxylase, were evenly distributed between the two microsomal subfractions. When rabbit liver microsomes were fractionated by the Dallner method, seven of the eight drug-metabolizing enzyme activities were concentrated in the smooth-surfaced fraction; in general, smooth/rough ratios ranged between 2:1 and 3:1. Benzpyrene hydroxylase was evenly distributed between the two fractions. Fractionation of rabbit liver microsomes by the Rothschild procedure yielded the most consistent and quantitatively significant results; all eight enzymes studied were markedly concentrated in the smooth-surfaced fraction, and smooth/rough ratios of enzyme activity were of the order of 5:1. These results supported the view that in the normal, untreated animal, many hepatic microsomal drug-metabolizing enzymes are concentrated in the smooth-surfaced subfraction. The results also emphasize that species and methodologic differences may influence the results obtained.

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