title: Detection and structural investigation of metabolites of stanozolol in human urine by liquid chromatography tandem mass spectrometry.
abstract: The applicability of LC-MS/MS in precursor ion scan mode for the detection of urinary stanozolol metabolites has been studied. The product ion at m/z 81 has been selected as specific for stanozolol metabolites without a modification in A- or N-rings and the product ions at m/z 97 and 145 for the metabolites hydroxylated in the N-ring and 4-hydroxy-stanozolol metabolites, respectively. Under these conditions, the parent drug and up to 15 metabolites were found in a positive doping test sample. The study of a sample from a chimeric uPA-SCID mouse collected after the administration of stanozolol revealed the presence of 4 additional metabolites. The information obtained from the product ion spectra was used to develop a SRM method for the detection of 19 compounds. This SRM method was applied to several doping positive samples. All the metabolites were detected in both the uPA-SCID mouse sample and positive human samples and were not detected in none of the blank samples tested; confirming the metabolic nature of all the detected compounds. In addition, the application of the SRM method to a single human excretion study revealed that one of the metabolites (4xi,16xi-dihydroxy-stanozolol) could be detected in negative ionization mode for a longer period than those commonly used in the screening for stanozolol misuse (3'-hydroxy-stanozolol, 16beta-hydroxy-stanozolol and 4beta-hydroxy-stanozolol) in doping analysis. The application of the developed approach to several positive doping samples confirmed the usefulness of this metabolite for the screening of stanozolol misuse. Finally, a tentative structure for each detected metabolite has been proposed based on the product ion spectra measured with accurate masses using UPLC-QTOF MS.
authors.affiliation: DoCoLab, UGent, Department of Clinical Chemistry, Microbiology and Immunology, Technologiepark 30, B-9052 Zwijnaarde, Belgium. oscar.pozomendoza@ugent.be
authors.forename: Oscar J
authors.initials: OJ
authors.lastname: Pozo
authors.forename: Peter
authors.initials: P
authors.lastname: Van Eenoo
authors.forename: Koen
authors.initials: K
authors.lastname: Deventer
authors.forename: Leen
authors.initials: L
authors.lastname: Lootens
authors.forename: Susana
authors.initials: S
authors.lastname: Grimalt
authors.forename: Juan V
authors.initials: JV
authors.lastname: Sancho
authors.forename: Felix
authors.initials: F
authors.lastname: Hernández
authors.forename: Philip
authors.initials: P
authors.lastname: Meuleman
authors.forename: Geert
authors.initials: G
authors.lastname: Leroux-Roels
authors.forename: Frans T
authors.initials: FT
authors.lastname: Delbeke
mesh_terms: D000328:Adult; D045930:Anabolic Agents / Q000737:chemistry* / Q000378:metabolism / Q000652:urine*; D000818:Animals; D002853:Chromatography, Liquid; D004300:Doping in Sports; D006801:Humans; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D013197:Stanozolol / Q000737:chemistry* / Q000378:metabolism / Q000652:urine*; D053719:Tandem Mass Spectrometry
chemical_list: D045930:Anabolic Agents; D013197:Stanozolol