The characteristics of the microsomal hydroxylation of tolbutamide

1991 ◽  
Vol 69 (3) ◽  
pp. 400-405 ◽  
Author(s):  
Pierre M. Bélanger ◽  
Serge St-Hilaire
Keyword(s):  
Carbon Monoxide ◽  
Liver Microsomes ◽  
Hepatic Metabolism ◽  
Species Variation ◽  
Rat Liver Microsomes ◽  
Hydroxylase Activity ◽  
Microsomal Metabolism ◽  
Microsomal Hydroxylation ◽  
Cytochrome P 450

The in vitro metabolism of tolbutamide to the hydroxymethyl derivative was studied using hepatic microsomal homogenates. The hydroxymethyl metabolite was quantitated by HPLC. The hepatic microsomal hydroxylase was completely inhibited by carbon monoxide and was NADPH dependent. Metyrapone, α-naphthoflavone, phenelzine, mercuric chloride, and nitrogen significantly inhibited the reaction indicating the involvement of the cytochrome P-450 monooxygenase. Species variation showed that the order of hepatic microsomal activity was rat > rabbit >> guinea pig >> mouse and hamster. The reaction increased with time up to 40 min and followed Michaelis–Menten kinetics in rat liver microsomes with apparent Km and Vmax values of 224.4 μM and 359.9 pmol∙mg−1∙min−1, respectively. The reaction was induced by phenobarbital but was depressed after pretreatment with 3-methylcholanthrene and isosafrole. However, expression of the hydroxylase activity per nanomoles of cytochrome P-450 showed that the activity was much higher in liver microsomes of isosafrole pretreated rats. These results indicate the involvement of different isozymes of cytochrome P-450 in the microsomal hydroxylation of tolbutamide.Key words: tolbutamide metabolism, tolbutamide hydroxylation, microsomal hydroxylation, microsomal metabolism of tolbutamide, hepatic metabolism of tolbutamide.

IN-VITRO METABOLISM OF 4-ANDROSTENE-3,17-DIONE BY HEPATIC MICROSOMES FROM THE RAINBOW TROUT (SALMO GAIRDNERII): EFFECTS OF HYPOPHYSECTOMY AND OESTRADIOL-17β

1981 ◽  
Vol 90 (1) ◽  
pp. 103-112 ◽  
Author(s):  
TIIU HANSSON ◽  
JAN-ÅKE GUSTAFSSON
Keyword(s):  
Rainbow Trout ◽  
Liver Microsomes ◽  
The Other ◽  
Sham Operation ◽  
Hydroxylase Activity ◽  
Oxidoreductase Activity ◽  
Hepatic Microsomes ◽  
Cytochrome P 450 ◽  
Reductive Metabolism

The metabolism of 4-androstene-3,17-dione by liver microsomes from juvenile rainbow trout, Salmo gairdnerii, was studied in vitro. Hypophysectomy of the fish significantly increased mean hepatic 17-hydroxysteroid oxidoreductase activity when compared with that from sham-operated fish but none of the other enzyme activities investigated were affected. Administration of oestradiol-17β resulted in a significant decrease in mean hepatic 6β-hydroxylase activity and total cytochrome P-450 content but had no effect on the 16-hydroxylation or on the reductive metabolism of androstenedione. The effect of oestradiol-17β on hepatic 6β-hydroxylase activity was as pronounced after hypophysectomy as after sham-operation indicating that these effects of oestradiol-17β are mainly direct and independent of the pituitary gland. The results indicate that hypophysial hormone(s) as well as oestradiol-17β play a role in the regulation of hepatic steroid metabolism in trout.

scholarly journals Androgen hydroxylation catalysed by a cell line (SD1) that stably expresses rat hepatic cytochrome P-450 PB-4 (IIB1)

1989 ◽  
Vol 260 (1) ◽  
pp. 81-85 ◽  
Author(s):  
D J Waxman ◽  
D P Lapenson ◽  
J J Morrissey ◽  
S S Park ◽  
H V Gelboin ◽  
...  
Keyword(s):  
Cell Line ◽  
Gene Product ◽  
Rat Liver ◽  
Membrane Fraction ◽  
Liver Microsomes ◽  
Parental Cell Line ◽  
Rat Liver Microsomes ◽  
Hydroxylase Activity ◽  
Specific Content ◽  
Cytochrome P 450

Androgen hydroxylation catalysed by Chinese hamster fibroblast SD1 cells, which stably express cytochrome P-450 form PB-4, the rat P450IIB1 gene product, was assessed and compared to that catalysed by purified cytochrome P-450 PB-4 isolated from rat liver. SD1 cell homogenates catalysed the NADPH-dependent hydroxylation of androstenedione and testosterone with a regioselectivity very similar to that purified by P-450 PB-4 (16 beta-hydroxylation/16 alpha-hydroxylation = 6.0-6.8 for androstenedione; 16 beta/16 alpha = 0.9 for testosterone). Homogenates prepared from the parental cell line V79, which does not express detectable levels of P-450 PB-4 or any other cytochrome P-450, exhibited no androgen 16 beta- or 16 alpha-hydroxylase activity. The hydroxylase activities catalysed by the SD1 cell homogenate were selectively and quantitatively inhibited (greater than 90%) by a monoclonal antibody to P-450 PB-4 at a level of antibody (40 pmol of antibody binding sites/mg of SD1 homogenate) that closely corresponds to the P-450 PB-4 content of the cells (48 pmol of PB-4/mg of SD1 homogenate). Fractionation of cell homogenates into cytosol and microsomes revealed that the P-450 PB-4-mediated activities are associated with the membrane fraction. Although the P-450 PB-4-specific content of the SD1 microsomes was 15% of that present in phenobarbital-induced rat liver microsomes, the P-450 PB-4-dependent androstenedione 16 beta-hydroxylase activity of the SD1 membrane fraction was only 2-3% of that present in the liver microsomes. This activity could be stimulated several-fold, however, by supplementation of SD1 microsomes with purified rat NADPH P-450 reductase. These studies establish that a single P-450 gene product (IIB1) can account for the hydroxylation of androgen substrates at multiple sites, and suggest that SD1 cells can be used to assess the catalytic specificity of P-450 PB-4 with other substrates as well.

ANDROGENIC REGULATION OF HEPATIC METABOLISM OF 4-ANDROSTENE-3,17-DIONE IN THE RAINBOW TROUT, SALMO GAIRDNERII

1982 ◽  
Vol 92 (3) ◽  
pp. 409-417 ◽  
Author(s):  
TIIU HANSSON
Keyword(s):  
Rainbow Trout ◽  
Reductase Activity ◽  
Juvenile Fish ◽  
Liver Microsomes ◽  
Hepatic Metabolism ◽  
Suppressive Effect ◽  
Hydroxylase Activity ◽  
Androgen Treatment ◽  
Androgenic Regulation

The metabolism of 4-androstene-3,17-dione by liver microsomes from the juvenile rainbow trout, Salmo gairdnerii, was studied in vitro. Administration of testosterone, 11-oxotestosterone, dihydrotestosterone or 17α-methyltestosterone to juvenile fish significantly increased mean hepatic 17-hydroxysteroid oxidoreductase (17-HSOR) activity. Androgen treatment tended to increase the total cytochrome P-450 content significantly in liver microsomes from 11-oxotestosterone-treated fish. On the other hand, androgen treatment decreased mean hepatic 6β-hydroxylase activity but did not affect 16-hydroxylase or 5α-reductase activity. The suppressive effect of simultaneous administration of testosterone and oestradiol-17β on 6β-hydroxylase activity was more pronounced than when these steroids were administered separately. Furthermore, oestradiol-17β diminished the effect of testosterone on 17-HSOR activity. Testosterone treatment of hypophysectomized fish caused a significant increase in 17-HSOR activity when compared with activity in untreated hypophysectomized fish, indicating that this effect of testosterone is mainly direct and independent of the pituitary gland. The results indicate that androgens as well as oestradiol-17β play a role in the control of sexual differences in hepatic steroid metabolism in trout.

scholarly journals The effects of unsaturated fatty acids on hepatic microsomal drug metabolism and cytochrome P-450

1969 ◽  
Vol 115 (3) ◽  
pp. 547-554 ◽  
Author(s):  
Richard P. Di Augustine ◽  
James R. Fouts
Keyword(s):  
Fatty Acids ◽  
Linoleic Acid ◽  
Rat Liver ◽  
Unsaturated Fatty Acids ◽  
Liver Microsomes ◽  
Type I ◽  
Rat Liver Microsomes ◽  
Microsomal Metabolism ◽  
Nitrobenzoic Acid

1. The effects of unsaturated fatty acids on drug-metabolizing enzymes in vitro were measured by using rat and rabbit hepatic 9000g supernatant fractions. 2. Unsaturated fatty acids inhibited the hepatic microsomal metabolism of ‘type I’ drugs with inhibition increasing with unsaturation: arachidonic acid>linolenic acid>linoleic acid>oleic acid. Inhibition was independent of lipid peroxidation. Linoleic acid competitively inhibited the microsomal O-demethylation of p-nitroanisole and the N-demethylation of (+)-benzphetamine. 3. The hepatic microsomal metabolism of ‘type II’ substrates, aniline and (−)-amphetamine, was not affected by unsaturated fatty acids. 4. The rate of reduction of p-nitrobenzoic acid and Neoprontosil was accelerated by unsaturated fatty acids. 5. Linoleic acid up to 3·5mm did not decelerate the generation of NADPH by rat liver soluble fraction, nor the activity of NADPH–cytochrome c reductase of rat liver microsomes. Hepatic microsomal NADPH oxidase activity was slightly enhanced by added linoleic acid. 6. No measurable disappearance of exogenously added linoleic acid occurred when this fatty acid was incubated with rat liver microsomes and an NADPH source. 7. The unsaturated fatty acids used in this study produced type I spectra when added to rat liver microsomes, and affected several microsomal enzyme activities in a manner characteristic of type I ligands.

scholarly journals Metabolism in Vitro of 3,4,3',4'- and 2,5,2',5'-Tetrachlorobiphenyl by Rat Liver Microsomes and Highly Purified Cytochrome P-450.

1991 ◽  
Vol 14 (5) ◽  
pp. 276-284 ◽  
Author(s):  
Chuzo ISHIDA ◽  
Nobuyuki KOGA ◽  
Nobumitsu HANIOKA ◽  
Helena Kazue SAEKI ◽  
Hidetoshi YOSHIMURA
Keyword(s):  
Rat Liver ◽  
Liver Microsomes ◽  
Rat Liver Microsomes ◽  
Cytochrome P 450

Oxygenation of 18-hydroxycorticosterone as the final reaction for aldosterone biosynthesis

1984 ◽  
Vol 107 (3) ◽  
pp. 395-400 ◽  
Author(s):  
Itaru Kojima ◽  
Etsuro Ogata ◽  
Hiroshi Inano ◽  
Bun-ichi Tamaoki
Keyword(s):  
Carbon Monoxide ◽  
Hydroxysteroid Dehydrogenase ◽  
Dependent Manner ◽  
In Vitro Production ◽  
Mitochondrial Preparation ◽  
Final Reaction ◽  
Vitro Production ◽  
Dose Dependent ◽  
Cytochrome P 450

Abstract. Incubation of 18-hydroxycorticosterone with the sonicated mitochondrial preparation of bovine adrenal glomerulosa tissue leads to the production of aldosterone, as measured by radioimmunoassay. The in vitro production of aldosterone from 18-hydroxycorticosterone requires both molecular oxygen and NADPH, and is inhibited by carbon monoxide. Cytochrome P-450 inhibitors such as metyrapone, SU 8000. SU 10603, SKF 525A, amphenone B and spironolactone decrease the biosynthesis of aldosterone from 18-hydroxycorticosterone. These results support the conclusion that the final reaction in aldosterone synthesis from 18-hydroxycorticosterone is catalyzed by an oxygenase, but not by 18-hydroxysteroid dehydrogenase. By the same preparation, the production of [3H]aldosterone but not [3H]18-hydroxycorticosterone from [1,2-3H ]corticosterone is decreased in a dose-dependent manner by addition of non-radioactive 18-hydroxycorticosterone.

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