Abstract
Although stereospecificity is characteristic of the reactions of single enzymes, both optical antipodes are usually metabolized in a qualitatively similar manner in the intact animal (l-4) where numerous possibilities of enzyme attack exist. When dl-aldosterone and other racemic steroids are added to enzyme or microbial systems (5, 6), only the naturally occurring d antipodes undergo reaction, whereas the 1 isomers are recovered unchanged. The fate of the 1 steroids in vivo, however, is not known. Since aldosterone has been available chiefly in racemic form,’ the metabolism of its I antipode was of interest. In the present study, a method for comparing the metabolism of a pair of antipodes has been applied to measure the conversion of I-aldosterone to each of the two known urinary products of the d isomer in man, the conjugate which is hydrolyzed at pH 1 (10) and tetrahydroaldosterone2 (11). The method consisted of administering a known mixture of d- and dl-aldosterone and comparing the ratio of d to dl forms in the administered mixture with that found in the urinary metabolites. Two methods were used to measure the ratio of d-aldosterone to dl-aldosterone. 1. When the endogenous production of aldosterone was negligible, the d isomer was administered in labeled form and racemic aldosterone in unlabeled form. The ratio of d- to racemic aldosterone was determined from the specific activity. 2. Alternatively, d- and racemic aldosterone were each labeled with different isotopes, H3 and CY4. The ratio of d- to racemic aldosterone was determined from the ratio of H3 to Cl*. Both methods demonstrated that over-all metabolism of aldosterone in man is stereospecific. Z-Aldosterone was not converted either to the conjugate hydrolyzed at pH 1 or to tetrahydroaldosterone.
Highlights
MethodsMeasurement of Radioactivity-Tritium and Cl* were measured in a Tri-Carb liquid scintillation spectrometer in 5 ml of phosphor solution
The administration of racemic aldosterone to human subjects would be expected to lead to a resolution of the mixture and to the excretion of optically active urinary products of the d configuration, the excretion of only one antipode need not have been due to the absence of enzymes capable of acting on the 2 form
The failure to account for I-aldosterone among the unconjugated steroids of urine, as well as the failure of I-aldosterone to be excreted either as the conjugate hydrolyzed at pH 1 or the tetrahydrometabolite, suggests another explanation
Summary
Measurement of Radioactivity-Tritium and Cl* were measured in a Tri-Carb liquid scintillation spectrometer in 5 ml of phosphor solution. The phosphor consisted of 0.3 y0 2,5-diphenyloxazole and 0.01 y0 1,4 - bis - 2(5 - phenyloxazolyl) -benzene in toluene. Both tritium and Cl4 were measured at a constant photomultiplier voltage of 1230 (tap 8). Tritium was measured in channel 1 at 10 to 100 volts, and Cl4 was measured in channel 2 at 100 volts to infinity. The approximate counting efficiencies at these discriminator settings were 20% for tritium in channel 1 and 70%.
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