Simultaneous doping analysis of main urinary metabolites of anabolic steroids in horse by ion-trap gas chromatography-tandem mass spectrometry

The use of anabolic steroids in racehorses is strictly regulated. We have developed a method for the simultaneous analysis of 11 anabolic steroids: fluoxymesterone, 17 alpha-methyltestosterone, mestanolone, methandienone, methandriol, oxymetholone, boldenone, furazabol, methenolone, nandrolone, and stanozolol, for possible application to a doping test in racehorses. We selected 15 kinds of target substances for a doping test from the main metabolites of these anabolic steroids, and established a method for simultaneous analysis. Urine was hydrolyzed and subjected to solid-phase extraction. Then, the residue from the extracts was derivatized by trimethylsilylation. The derivatized samples were subjected to ion-trap gas chromatography-tandem mass spectrometry, and their mass chromatograms and product ion spectra were obtained. The limit of detection of the target substances was 5 - 50 ng/mL, and the mean recovery and coefficient of variation were 71.3 - 104.8% and 1.1 - 9.5%, respectively.