Reduction of 3-keto-5 beta-cholanoic acid to lithocholic and isolithocholic acids by human liver cytosol in vitro.
The formation of lithocholic and isolithocholic acids from 3-keto-5 beta-cholanoic acid by human liver cytosol was examined in vitro. Liver cytosol was incubated at various pH levels with 3-keto-5 beta-cholanoic acid in a phosphate buffer containing NADPH or NADH; the products formed were analyzed by gas chromatography. Results showed that human liver cytosol reduced 3-keto-5 beta-cholanoic acid to lithocholic acid at a pH level of 7.0 or above and to isolithocholic acid at a pH level of 6.0 or below when NADPH was used as a coenzyme, and it was reduced to isolithocholic acid only when NADH was used. Furthermore, two peaks for the reducing enzymes could be clearly found by column chromatography of Affi-Gel Blue. These results indicate that human liver cytosol contains two enzymes acting on reduction of 3-keto-5 beta-cholanoic acid to lithocholic and isolithocholic acids, which are dependent on the pH level and the use of NADPH or NADH in vitro. Since the 3 beta-dehydrogenation was inhibited by the addition of pyrazole, an alcohol dehydrogenase inhibitor or ethanol, and the major peak of 3 beta-hydroxysteroid dehydrogenase coincided with the peak of alcohol dehydrogenase on Affi-Gel Blue chromatography, at least some of the cytosolic 3 beta-hydroxysteroid dehydrogenase seemed to be identical to or to have characteristics similar to alcohol dehydrogenase.
Amuro
Y
Y
Yamade
W
W
Maebo
A
A
Hada
T
T
Higashino
K
K
D000426:Alcohol Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000378:metabolism; D002697:Chloral Hydrate / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D000431:Ethanol / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006913:Hydroxysteroid Dehydrogenases / Q000378:metabolism; D008095:Lithocholic Acid / Q000031:analogs & derivatives* / Q000096:biosynthesis* / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011720:Pyrazoles / Q000494:pharmacology
D011720:Pyrazoles; D000431:Ethanol; C031280:pyrazole; D002697:Chloral Hydrate; D009249:NADP; D008095:Lithocholic Acid; C046230:3-oxocholan-24-oic acid; D000429:Alcohol Oxidoreductases; D006913:Hydroxysteroid Dehydrogenases; D000426:Alcohol Dehydrogenase