Normal Rat Liver Microsomes Research Articles

Vitamin K and Biosynthesis of Protein and Prothrombin

The role of vitamin K in the synthesis of prothrombin has been examined with the following results. 1. Vitamin K deficiency has no effect on general protein synthesis as studied by amino acid incorporation into protein in vivo and in vitro and by tryptophan pyrrolase production following tryptophan feeding. 2. Prothrombin activity was found in normal rat liver microsomes with increased release following ultrasonic treatment, but could not be found in microsomes from the livers of vitamin K-deficient rats or dicumarol-treated rats. Administration of suboptimal levels of vitamin K to vitamin K-deficient rats resulted in detectable microsomal prothrombin in 2 hours and essentially normal values within 3 hours. 3. Adequate vitamin K1, given by injection, completely restored the blood prothrombin levels of vitamin K-deficient rats in 1 hour, and of dicumarol- or warfarin-treated rats in 5 to 7 hours depending on the amounts of warfarin and vitamin K1 given. 4. Following administration of actinomycin D or ethionine to vitamin K-deficient rats, treatment with vitamin K1 markedly stimulated prothrombin production, indicating that the site of action of vitamin K is beyond the level of transcription of DNA to prothrombin messenger RNA. 5. Treatment with vitamin K1, following administration to vitamin K-deficient rats of cycloheximide at a level just sufficient to block synthesis of prothrombin for 6 to 8 hours, gave an essentially normal prothrombin response. At higher cycloheximide levels the response to vitamin K1, while much less complete, was clear and definite. 6. The response of vitamin K-deficient rats to vitamin K1, administered following treatment with blocking doses of puromycin, appears significant. 7. These data appear to indicate that the site of function of vitamin K is not at the genetic level, as has been reported, but at a late stage in translation of prothrombin messenger RNA to form a functional prothrombin molecule.

624. Cold affects hydroxylation: Dewhurst, F. (1963). The effect of stress upon the metabolism of 2-naphthylamine in mice. Experientia19, 646

This chapter describes the fluorometric and chromatographic methods for measuring microsomal biphenyl hydroxylation. This method, is the most convenient when biphenyl is used as a routine substrate for the monooxygenase. Biphenyl (I) is a useful model substrate for investigating the cytochrome P-450 (P-448)-mediated, NADPH2-, and oxygen-dependent microsomal aryl hydrocarbon hydroxylase. Microsomal hydroxylation of biphenyl is terminated with acid, which also ensures that the hydroxylated metabolites are not ionized and are extracted into n-heptane. The hydroxybiphenyls are then reextracted into alkali, leaving unreacted biphenyl in the organic phase. The fluorescence of the metabolites is measured at pH 5-6, achieved by adding succinic acid to the alkaline extract. Two compounds, 2-hydroxybiphenyl (II) and 4-hydroxybiphenyl (III) constitute more than 90% of the total biphenyl metabolites with liver microsomes of normal rats or hamsters. Small percentages of dihydroxybiphenyls are formed with rodent liver microsomes, and although 3-hydroxybiphenyl is isolated as a metabolite from normal rabbit urine, it is not formed in significant quantities with normal rodent liver microsomes.