Identification of in vitro metabolites of boldenone using Camel liver microsomes – A HR-LCMS approach

Abstract

Anabolic steroids are widely abused in animal sports to improve their performance. The present study identifies the possible metabolites of boldenone in camel liver. A high-resolution accurate QE mass spectrometer was used to identify the parent boldenone and its metabolites. To investigate the phase 1 biotransformation of boldenone in camel, sodium phosphate buffer and NADPH were used. Chromatographic separation was carried out on a Thermo Hypersil C18 column using acetonitrile and formic acid as mobile phases. The current study, helped in unequivocal detection of six metabolites (Phase 1) for boldenone. The 17b-boldenone is an 3-oxo-Delta (1), Delta(4)-steroid substituted by an oxo group at position 3 and a beta-hydroxy group at position 17. It is prone to oxidation, which results in three hydroxylated metabolites with protonated parent ion of m/z 303.1954 (C19H27O3)+. Androsta-1,4-diene-3,17-dione [M+H]+ of m/z 285.1848 (C19H25O2+), hydroxyandrosta-1,4-diene-3,17-dione [M+H]+ of m/z 301.1797 (C19H25O3+) in addition to 17-hydroxy-androsta-1-en-3-one [M+H]+ of m/z 289.2162 (C19H29O2+) were also identified. The structures of the detected metabolites were identified based on their accurate mass, fragmentation pattern, and chromatographic retention time. In this research, camel liver was successfully used for in vitro experiments as an alternative to liver microsomes.