METHOD FOR DETERMINATION OF URINARY 5β-PREGNANE-3α,17α,20α-TRIOL AND Δ5-PREGNENE-3β,17α,20α-TRIOL*

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Abstract
During studies of adrenal function in human hypertension, Bongiovanni and Eberlein's procedure for determination of urinary pregnanetriol was found inaccurate. Pregnanetriol eluted from an alumina column with benzene-6% ethanol, after enzymatic and acid hydrolysis of the steroids in urine, was contaminated in most instances by 2 substances interfering with the determination in concentrated sulfuric acid. One of them was a compound described in a previous communication and identified as Compound III. The second was characterized as Δ5-pregnene-3β,17α,20α-triol. Modifications of the original procedure are proposed for the specific determination of 5β-pregnane-3α,17α,20α-triol and Δ5-pregnene-3β,17α,20α-triol. The modified method involves successive chromatographic separation of the crude neutral-extract residue of urine, first on a silica-gel column, second on an ethylene-glycol/toluene paper system, and finally on an alumina column. Spectrophotometric determination of the highly purified compound, free from interfering substances, is achieved by use of the sulfuric-acid color reaction.

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