QUANTITATION OF CORTISOL AND RELATED 3-OXO-4-ENE STEROIDS IN URINE USING GAS-CHROMATOGRAPHY MASS-SPECTROMETRY WITH STABLE ISOTOPE-LABELED INTERNAL STANDARDS

A method for the profiling of several important 3-oxo-4-ene urinary st eroids is reported. The methodology is combined gas chromatography/mas s spectrometry (GC/MS) utilizing stable isotope-labeled internal stand ards. The following standards were obtained or easily synthesized: [9, 11,12,12(2)-H-4]cortisol, [1,2-H-2(2)] and [9,12,12- H-2(2)]cortisone, [1,2-H-2(2)]6 beta-hydroxycortisol, and [1,2-H-2(2)]18-hydroxycortiso l. We found the following excretions of free steroids for normal adult males and females: cortisol (males mean +/- SD, 35 +/- 13; females me an +/- SD, 23 +/-: 13), cortisone (males mean +/- SD, 58 +/- 23;female s mean +/- SD, 50 +/- 22), 6 beta-hydroxycortisol (males mean +/- SD, 164 +/- 59; females mean +/- SD, 108 +/- 55), and 18-hydroxycortisol ( males mean +/- SD, 148 +/- 55; females mean +/- SD, 71 +/- 30). For 18 -hydroxycortisol in particular, the excretions were much higher for ma les than for females. We Sound that the larger part of urinary cortiso l and cortisone is not free but is released from conjugation by enzyme s present in snail digestive juice. Using a pooled urine sample from a n equal number of male and female subjects, we found that for cortisol 29% was excreted free, 28% as glucuronide and 43% as other conjugates (probably sulfates). For cortisone 41% was free, 45% beta-glucuronide and 14% as other conjugates. Relatively little (3-8%) of the hydroxyl ated cortisols were excreted conjugated. (C) 1996 by Elsevier Science Inc.