An alternative metabolic pathway of 11-deoxycorticosterone in bovine adrenal in vitro: evidence for the presence of a pathway of 11-deoxycorticosterone oxidation at 19-position.
Comparative studies of 11 beta-, 18-, and 19-hydroxylation activities of 11-deoxycorticosterone (DOC) by bovine adrenal mitochondria revealed that an appreciable level of hydroxylation rate was observed in 19-hydroxylation (0.32 nmol/min/mg mitochondrial protein), as well as in 11 beta- and 18-hydroxylations (4.7 and 0.27 nmol/min/mg mitochondrial protein, respectively), at saturated substrate concentration in vitro. Also, the rates of the oxidation reactions of 19-hydroxy-11-deoxycorticosterone (19-OH-DOC) and 19-oxo-11-deoxycorticosterone (19-oxo-DOC) at the 19-position were about 5 times higher than the 19-hydroxylation rate of DOC. Although the affinities of 19-OH-DOC and 19-oxo-DOC for the enzyme(s) involved in the C-19 oxidation were about one-fifth those of DOC, these results strongly suggest the presence of the following pathway in bovine adrenal in vitro: DOC----19-OH-DOC----19-oxo-DOC----19-oic-DOC. This was further confirmed by a dynamic study of the formation and subsequent decay of the C-19 oxidized metabolites produced from DOC. At maximum concentrations of 19-OH-DOC and 19-oxo-DOC, the rates of production of, respectively, 19-oxo-DOC and 19-oic-DOC reached maximum. Furthermore, at the beginning of the incubation (1-4 min), an induction period in the formation of 19-oxo-DOC and 19-oic-DOC was observed and the formation of 19-oxo-DOC always preceded the appearance of 19-oic-DOC. These observations strongly support the existence of the pathway of the C-19 oxidation of DOC as mentioned above. It was also established that reduced pyridine nucleotide (NADPH) and molecular oxygen were required for these oxidation reactions. In addition, these three oxidation reactions were uniformly inhibited by the presence of carbon monoxide or metyrapone (0.01-1.0 microM), which is known to bind specifically with cytochrome P-450, while potassium cyanide (0.01-0.1 mM) did not affect them. These results suggest the possibility of the involvement of cytochrome P-450 in the C-19 oxidation reactions of DOC, 19-OH-DOC, and 19-oxo-DOC. We also showed that 19-oic-DOC is not further metabolized to other steroids such as 19-nor-11-deoxycorticosterone in bovine adrenal cortex.
Kobayashi
Y
Y
Clinical Chemistry Laboratory, Kobe Women's College of Pharmacy, Japan.
Nishiguchi
Y
Y
Kiguchi
T
T
Ninomiya
I
I
Watanabe
F
F
D000311:Adrenal Glands / Q000378:metabolism*; D000818:Animals; D002248:Carbon Monoxide / Q000494:pharmacology; D002417:Cattle; D003577:Cytochrome P-450 Enzyme System / Q000502:physiology; D003900:Desoxycorticosterone / Q000031:analogs & derivatives / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008797:Metyrapone / Q000494:pharmacology; D010084:Oxidation-Reduction; D011190:Potassium Cyanide / Q000494:pharmacology
C027855:19-hydroxydeoxycorticosterone; D003900:Desoxycorticosterone; C033576:19-oxo-deoxycorticosterone; D002248:Carbon Monoxide; D003577:Cytochrome P-450 Enzyme System; D011190:Potassium Cyanide; D008797:Metyrapone