Determination of anabolic hormones multi-residues in animal muscle tissues using liquid chromatography-tandem mass spectrometry

A liquid chromatography-tandem mass spectrometric (LC/MS/MS) method was developed for the simultaneous identification and determination of multi-residues amounts of steroid anabolic hormones epitestosterone (ETS), testosterone 17-propionate (PTS) androlone (17 beta-NT), 17 alpha-methyltestosterone (MTS), progesterone (PG), medroxyprogesterone (MED) 17 beta-estradiot (17 beta-ES), 17 alpha-ethinylestradiol (EES) and estrone (ESN) in the muscle tissues of various animal species. Homogenized tissue. samples were enzymatically digested in acetate buffer (pH 5.0), then methanol was added and the mixtures were extracted under ultrasonication incubation. Clean-up was carried out with at least two times tertiary butyl methyl ether ( MTBE) liquid-liquid partitioning followed by a further reverse phase solid phase extraction (SPE) cartridge purification. After C-18 LC gradient elution separation using acetonitrile-water as mobile phase, the eluents were qualitatively and quantitatively determined under multi-reaction monitoring (MRM) scan type with tandem mass analyzer using positive polarity mode for the androgen and progestational hormones, negative polarity for the estrogen respectively. The limit of quantitation (LOQ) for the above 10 hormones were in the range of 0.5 similar to 1.0 mu g/kg. At the 1.0 mu g/kg L0Q spiked level, the mean recoveries were in the range of 55% and 77%, and accordingly the relative standard deviation range were in the range of 7.1% and 35%. The real sample test showed this method could be used for the sensitive and accurate determination of multi-steroid anabolic hormones residues in biological muscle samples.