title: New clostebol metabolites in human urine by liquid chromatography time-of-flight tandem mass spectrometry and their application for doping control.
abstract: In this study, clostebol metabolic profiles were investigated carefully. Clostebol was administered to one healthy male volunteer. Urinary extracts were analyzed by liquid chromatography quadrupole time-of-flight mass spectrometry (MS) using full scan and targeted MS/MS techniques with accurate mass measurement for the first time. Liquid-liquid extraction and direct injection were applied to processing urine samples. Chromatographic peaks for potential metabolites were found by using the theoretical [M-H](-) as target ion in full scan experiment, and their actual deprotonated ions were analyzed in targeted MS/MS mode. Fourteen metabolites were found for clostebol, and nine unreported metabolites (two free ones and seven sulfate conjugates) were identified by MS, and their potential structures were proposed based on fragmentation and metabolism pathways. Four glucuronide conjugates were also first reported. All the metabolites were evaluated in terms of how long they could be detected and S1 (4ξ-chloro-5ξ-androst-3ξ-ol-17-one-3ξ-sulfate) was considered to be the long-term metabolite for clostebol misuse detected up to 25 days by liquid-liquid extraction and 14 days by direct injection analysis after oral administration. Five conjugated metabolites (M2, M5, S2, S6 and S7) could also be the alternative biomarkers for clostebol misuse.
authors.affiliation: National Anti-Doping Laboratory, China Anti-Doping Agency, 1st Anding Road, ChaoYang District, Beijing, 100029, China.
authors.forename: Jianghai
authors.initials: J
authors.lastname: Lu
authors.forename: María
authors.initials: M
authors.lastname: Fernández-Álvarez
authors.forename: Sheng
authors.initials: S
authors.lastname: Yang
authors.forename: Genye
authors.initials: G
authors.lastname: He
authors.forename: Youxuan
authors.initials: Y
authors.lastname: Xu
authors.forename: R
authors.initials: R
authors.lastname: Aguilera
mesh_terms: D000284:Administration, Oral; D000328:Adult; D002853:Chromatography, Liquid / Q000379:methods*; D004300:Doping in Sports; D020719:Glucuronides / Q000737:chemistry / Q000378:metabolism / Q000652:urine; D006801:Humans; D059625:Liquid-Liquid Extraction; D008297:Male; D015394:Molecular Structure; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013739:Testosterone / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000652:urine; D013997:Time Factors
chemical_list: C000601354:4-chloroandrost-3-ol-17-one-3-sulfate; D020719:Glucuronides; D013739:Testosterone; C076775:clostebol
keywords: clostebol; doping control; human urine; liquid chromatography quadruple time-of-flight mass spectrometry; new metabolites