METABOLITES OF 4-CHLOROTESTOSTERONE ACETATE IN CATTLE URINE AS DIAGNOSTIC MARKERS FOR ITS ILLEGAL USE

Seven metabolites of 4-chlorotestosterone acetate were identified in u rine of cattle that received a single injection of the drug. The stero ids were isolated by means of a series of clean-up steps carried out b efore and after enzymatic hydrolysis. The obtained extract was fractio nated by high-performance liquid chromatography and each fraction was examined both by high-performance thin-layer chromatography and by cap illary gas chromatography-mass spectrometry of the m-ethoxime-trimethy lsilyl derivatives. The metabolites were tentatively identified by stu dying the mass spectra of selected peaks not found in blank samples. T he structures of two metabolites, viz. 4-chloroandrost-4-ene-3,17-dion e and 4-chloroandrost-4-ene-3alpha,17beta-diol were confirmed by chemi cal synthesis. The synthesized metabolites and 4-chloro-17alpha-testos terone, a third metabolite which was identified tentatively, were loca ted on the thin-layer chromatograms obtained. This study led to the co nclusion that the illegal use of 4-chlorotestosterone acetate can be d etected by identifying one or more of its metabolites in urine.