Biochemical and morphological comparison of microsomal preparations from rat, quail, trout, mussel, and water flea

Abstract

Differential centrifugation methods already in use were applied to purify rat, quail, and trout liver microsomes and modified as necessary to purify microsomes from mussel digestive gland and whole water flea. All these microsomal preparations were comparatively characterized with respect to protein and RNA content, levels of markers of subcellular contaminants, ultrastructural morphology, differential spectra of cytochromes P-450 and b 5, monoxygenase activity, and in vitro metabolism of p-dichlorobenzene. Yields of microsomal proteins of the tested organisms differed widely, with mussel showing the lowest yield. Very low levels of nuclear and mitochondrial contaminants were detected in all microsomal preparations, but cell membrane contaminants were clearly present in most preparations. Daphnia microsomes were significant amounts of partially disrupted secretory granules and plasma membrane. From a qualitative standpoint differential spectra of cytochrome b 5 were very similar for all the preparations, whereas cytochrome P-450 spectra were largely dependent on the microsomal preparation as well as on the assay method used. The content of cytochrome P-450 was highest for rat liver microsomes and very low or absent in Daphnia and mussel preparations; the range of cytochrome b 5 contents was much narrower. Significant differences were observed among monoxygenase activities of the different preparations. In Daphnia and mussel microsomes, aniline hydroxylase was absent and benzo[ a]pyrene hydroxylase activity was much lower than in rat and quail microsomes. Benzo[ a]pyrene hydroxylase activity of trout liver microsomes was similar to that of rat and quail microsomes, whereas hydroxylation of substrates which in rat liver are preferentially metabolized by phenobarbital-inducible forms of cytochrome P-450 was much lower in trout microsomes.