SIMULTANEOUS DETERMINATION, IN CALF URINE, OF 12 ANABOLIC AGENTS AS HEPTAFLUOROBUTYRYL DERIVATIVES BY CAPILLARY GAS-CHROMATOGRAPHY MASS-SPECTROMETRY

A method to determine twelve anabolic hormones (diethylstilbestrol, he xestrol, dienestrol, 17 beta-estradiol, 19-nortestosterone, testostero ne, 1-dehydrotestosterone, 17 alpha-methyltestosterone, progesterone, estrone, 17 alpha-ethynilestradiol, and trenbolone) is presented. Urin e samples were extracted with octadecylsilica columns and clean-up was performed in two steps with basic alumina and silica solid-phase extr action cartridges. The extracts obtained were derivatized with heptafl uorobutyric anhydride and analyzed by GC-MS. Stability of derivatives was good and compounds having keto groups produced enol derivatives th at were stable also. SIM mode was applied to increase the sensitivity and, when possible, the higher mit ions were selected to improve ident ification. Repeatability of the chromatographic analysis was evaluated on the basis of area repeatability, and the coefficient of variation obtained was lower than 13%. Absolute recoveries were in the range 35- 60% (dehydrotestosterone and estrone <20%) with coefficients of variat ion between 14 and 37% for the whole procedure. [H-2(3)]Testosterone a nd [H-2(8)]diethylstilbestrol were evaluated to improve quantitative d ata. The recovery of [H-2(3)]testosterone was found to be equal to or slightly higher than that of the other hormones, but the recovery of [ H-2(8)]diethylstilbestrol was lower than any other. [H-2(3)]Testostero ne was the most suitable for use as an internal standard, as its addit ion at the beginning of analytical procedure, corrected recovery resul ts and greatly improved precision. Corrected recoveries from urine ran ged from 72-110%, and coefficients of variation ranged from 6-15%, exc ept for testosterone which yielded slightly higher values. The limit o f detection was 0.5 ng/ml for all the compounds studied.