Isolation and identification of the liver microsomal cytochrome P 450 isoen zymes responsible for the formation of diazepam main metabolites nordiazepam and temazepam in rats were studied. The effects of P 450 ind...Isolation and identification of the liver microsomal cytochrome P 450 isoen zymes responsible for the formation of diazepam main metabolites nordiazepam and temazepam in rats were studied. The effects of P 450 inducers and inhibitors on the protein contents in SDS poly acrylamide gel electrophoresis and thin layer chromatography to the corresponding diazepam me tabolizing activities of rat liver microsomes were observed. The P 450 contents were dramatically re duced by ip diazepam, cimetidine or propranolol. Diazepam and propranolol inhibited temazepam formation, high dose of propranolol also inhibited nordiazepam formation. Phenobarbital increased the P 450 contents and induced the production of both nordiazepam and temazepam. It also induced proteins with molecular weight (m) of 51 and 59 kDa in SDS PAGE and those with m ranging from 45 to 55 kDa and from 55 to 65 kDa in TLC. Propranolol inhibited both fractions, especially that of m 55～65 kDa, whereas diazepam tended to inhibit the fraction of 45～55 kDa. The protein of m 51 kDa could be mainly involved in diazepam C3 hydroxylation, whereas those of m 59 kDa could be responsible for the N demethylation of diazepam in rats.展开更多
A dual wavelength differential first derivative spectrophotometric method has been developed to standardize the concentration of a saturated aqueous solution of carbon monoxide (CO) as the standard and to identify and...A dual wavelength differential first derivative spectrophotometric method has been developed to standardize the concentration of a saturated aqueous solution of carbon monoxide (CO) as the standard and to identify and to determine CO formed during the microsomal metabolism of xenobiotics in vitro. The method can significantly eliminate the background interference in the assay media and increase the quantitative accuracy and the sensitivity. There is a good linear relationship between CO concentration in the range of 2～10 μmol·L 1 CO and the distance D between the first derivative peak at 415 nm amd valley at 426 nm with r=0.9999(n=5),the regression equation being C (mmol·L 1 )=17.6D 0.4, the detection limit lower than 0.1 μmol·L 1 CO. The average recoveries of CO from the assay system and the sample were 102.1%, RSD=2.9% (n=7) and 79.7%, RSD=6.8% (n=12),respectively. The RSD of within day was 4.4%(n=18),and the RSD of day to day was 6.1%(n=16). By this method, four trihaloanilines and one trihalobenzene were tested, the results showed that only 2,4,5 trifluoroaniline could be converted to CO by the incubation with rat hepatic microsomes, NADPH and oxygen, the ability of phenobarbital or dexamethasone to induce rat hepatic microsomes to catalyze CO formation was 3 or 8 times higher than that of the control.展开更多
文摘Isolation and identification of the liver microsomal cytochrome P 450 isoen zymes responsible for the formation of diazepam main metabolites nordiazepam and temazepam in rats were studied. The effects of P 450 inducers and inhibitors on the protein contents in SDS poly acrylamide gel electrophoresis and thin layer chromatography to the corresponding diazepam me tabolizing activities of rat liver microsomes were observed. The P 450 contents were dramatically re duced by ip diazepam, cimetidine or propranolol. Diazepam and propranolol inhibited temazepam formation, high dose of propranolol also inhibited nordiazepam formation. Phenobarbital increased the P 450 contents and induced the production of both nordiazepam and temazepam. It also induced proteins with molecular weight (m) of 51 and 59 kDa in SDS PAGE and those with m ranging from 45 to 55 kDa and from 55 to 65 kDa in TLC. Propranolol inhibited both fractions, especially that of m 55～65 kDa, whereas diazepam tended to inhibit the fraction of 45～55 kDa. The protein of m 51 kDa could be mainly involved in diazepam C3 hydroxylation, whereas those of m 59 kDa could be responsible for the N demethylation of diazepam in rats.
文摘A dual wavelength differential first derivative spectrophotometric method has been developed to standardize the concentration of a saturated aqueous solution of carbon monoxide (CO) as the standard and to identify and to determine CO formed during the microsomal metabolism of xenobiotics in vitro. The method can significantly eliminate the background interference in the assay media and increase the quantitative accuracy and the sensitivity. There is a good linear relationship between CO concentration in the range of 2～10 μmol·L 1 CO and the distance D between the first derivative peak at 415 nm amd valley at 426 nm with r=0.9999(n=5),the regression equation being C (mmol·L 1 )=17.6D 0.4, the detection limit lower than 0.1 μmol·L 1 CO. The average recoveries of CO from the assay system and the sample were 102.1%, RSD=2.9% (n=7) and 79.7%, RSD=6.8% (n=12),respectively. The RSD of within day was 4.4%(n=18),and the RSD of day to day was 6.1%(n=16). By this method, four trihaloanilines and one trihalobenzene were tested, the results showed that only 2,4,5 trifluoroaniline could be converted to CO by the incubation with rat hepatic microsomes, NADPH and oxygen, the ability of phenobarbital or dexamethasone to induce rat hepatic microsomes to catalyze CO formation was 3 or 8 times higher than that of the control.
