Detection of Stanozolol and Its Major Metabolites in Human Urine by Liquid Chromatography-Tandem Mass Spectrometry

The determination of stanozolol and its metabolites in human urine has been of particular interest in sports drug testing due to its frequently revealed misuse. A simple and rapid sample preparation procedure based on consecutive solid-phase and liquid–liquid extraction with subsequent re-extraction followed by liquid chromatography and electrospray ionization tandem mass spectrometry was established. It allowed the determination of stanozolol and its metabolic products 16β-OH-stanozolol and 4β-OH-stanozolol in human urine at detection limits of 0.1, 0.2 and 0.2 ng mL−1, respectively, with recoveries ranging from 5 to 38%. The robust nature of the assay and the efficient removal of interfering biological matrix provides excellent signal-to-noise ratios, and, thus, a rapid alternative to established procedures utilizing multiple solid-phase extraction or immunoaffinity chromatography strategies. More than 15 doping control urine specimens tested positive for stanozolol during the last 12 months have been confirmed using the described approach.