Hydroxylation of Deoxycorticosterone at the 6β, 11α, 15α, 15β, and 16α Positions by Surviving Liver Slices of the Bullfrog^*

Abstract

6p-, 1 la-, 15a-, 15p-, and 16a-Hydroxyby infrared spectroscopy and further characterized deoxycorticosterone were isolated following the aerobic by paper chromatographic methods. The results were incubation of deoxycorticosterone with surviving liver compared with those hydroxylations observed where slices of the bullfrog. These metabolites were identified mammalian tissues were employed.I n an earlier comparative study of the 7a-hydroxylation of deoxycorticosterone (compound I) by surviving liver slices of various species (Schneider, 1962), it was noted that, while this reaction occurred only in rodent tissue, certain nonrodent tissue introduced hydroxyl groups into several positions other than C-7. The paper chromatographic pattern obtained in the case of the bullfrog (Ranu catesbieuna) was particularly complex and, since this species is readily available, several largescale incubations were carried out with the object of recovering the metabolites in pure form. These were identified as 6p-, lla-, 15a-, 15p-, and 16a-hydroxydeoxycorticosterone. It is the purpose of this paper to detail their isolation and identification. MethodsThe incubation, extraction, and paper chromatographic procedures followed those described earlier (Schneider, 1962). The reference steroids were supplied by Dr. Robert Neher. Extinction coefficients were obtained in methanol solution using a Zeiss PMQ I1 spectrophotometer. The infrared spectra of both the reference and isolated steroids were determined as KBr pellets by the use of a Beckman IR-9 instrument for the former and a Perkin-Elmer Model 21 instrument for the latter. The use of two different instruments did not lead to difficulties in comparing the spectra. The bullfrogs used in these incubations were obtained from commercial sources in Wisconsin and New Jersey. ResultsIn the representative experiment here described, 500 mg of free compound I was incubated (3 hours at 38",

Studies on the Metabolism of C21 Steroids in Rat Liver

“…Finally, Lisboa and Gustafsson have shown the presence of 6a,68-and 1601-hydroxylases active on progesterone in human foetal liver microsomes [13, 19,271. 15-Hydroxylation of 3-0x0-A4-C,, steroids has been reported to occur in the liver of reptilia. Thus, Schneider has shown that deoxycorticosterone is hydroxylated in positions 68,11a,15a and 16a when incubated with liver slices from the bull frog (Rana catesbieana), red racer (Masticophis flagellum piceus) and bull snake (Pituophis catenifer affinis) [26,28]. The bull frog was also able to hydroxylate deoxycorticosterone in position 15b.…”

In vivo metabolism of 15α-hydroxyprogesterone in man

“…It is of interest to note that 15a hydroxylation of neutral steroids is not confined to the human fetoplacental unit. Schneider (1965) found that liver slices of the American bullfrog (Rana catesbiana) was capable of converting deoxycorticosterone into a number of hydroxy la ted products, among which were 15aand 15/3-hydroxydeoxycorticosterone. Recently, it has been found that the feces of conventional rats contain 3a,15a-dihydroxy-15a-pregnan-20-one as the predominant C-21 steroid (Gustafsson, 1968a,b) and that this metabolite is also present in the feces of the germ-free rat.…”

Isolation of 15α-Hydroxyprogesterone from Human Pregnancy Urine*

“…More recently Schneider (1965) found that liver slices of the American bullfrog (Rana catesbiana) was capable of converting deoxycorticosterone to a number of hydroxylated products, among which were 15aand 15/3-hydroxydeoxycorticosterone. Schneider (1965) regarded 15a-and 15/3-hydroxylations as primitive reactions in an evolutionary sense because he could not demonstrate such hydroxylations in mammalian tissues.…”

“…More recently Schneider (1965) found that liver slices of the American bullfrog (Rana catesbiana) was capable of converting deoxycorticosterone to a number of hydroxylated products, among which were 15aand 15/3-hydroxydeoxycorticosterone. Schneider (1965) regarded 15a-and 15/3-hydroxylations as primitive reactions in an evolutionary sense because he could not demonstrate such hydroxylations in mammalian tissues. With the demonstration that the adult human adrenal (Knuppen et al, 1965b) and the human fetal liver (Schwers et al, 1965) can 15a-hydroxylate estrogens and with the isolation of 15a-hydroxyprogesterone from late-pregnancy urine reported in this paper it is clear that such hydroxylations are not confined to the lower species.…”