title: Untargeted metabolomics in doping control: detection of new markers of testosterone misuse by ultrahigh performance liquid chromatography coupled to high-resolution mass spectrometry.
abstract: The use of untargeted metabolomics for the discovery of markers is a promising and virtually unexplored tool in the doping control field. Hybrid quadrupole time-of-flight (QTOF) and hybrid quadrupole Orbitrap (Q Exactive) mass spectrometers, coupled to ultrahigh pressure liquid chromatography, are excellent tools for this purpose. In the present work, QTOF and Q Exactive have been used to look for markers for testosterone cypionate misuse by means of untargeted metabolomics. Two different groups of urine samples were analyzed, collected before and after the intramuscular administration of testosterone cypionate. In order to avoid analyte losses in the sample treatment, samples were just 2-fold diluted with water and directly injected into the chromatographic system. Samples were analyzed in both positive and negative ionization modes. Data from both systems were treated under untargeted metabolomic strategies using XCMS application and multivariate analysis. Results from the two mass spectrometers differed in the number of detected features, but both led to the same potential marker for the particular testosterone ester misuse. The in-depth study of the MS and MS/MS behavior of this marker allowed for the establishment of 1-cyclopentenoylglycine as a feasible structure. The putative structure was confirmed by comparison with synthesized material. This potential marker seems to come from the metabolism of the cypionic acid release after hydrolysis of the administered ester. Its suitability for doping control has been evaluated.
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: Montse
authors.initials: M
authors.lastname: Raro
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Ibáñez
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: Rubén
authors.initials: R
authors.lastname: Gil
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Andreu
authors.initials: A
authors.lastname: Fabregat
authors.affiliation: §DoCoLab, Ghent University, Technologiepark 30, 9052, Zwijnaarde, Belgium.
authors.forename: Eva
authors.initials: E
authors.lastname: Tudela
authors.affiliation: §DoCoLab, Ghent University, Technologiepark 30, 9052, Zwijnaarde, Belgium.
authors.forename: Koen
authors.initials: K
authors.lastname: Deventer
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.|∥Department of Experimental and Health Sciences, Universitat Pompeu Fabra, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Rosa
authors.initials: R
authors.lastname: Ventura
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.|∥Department of Experimental and Health Sciences, Universitat Pompeu Fabra, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Jordi
authors.initials: J
authors.lastname: Segura
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.|∥Department of Experimental and Health Sciences, Universitat Pompeu Fabra, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Marcos
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.|⊥Department of Biological Chemistry and Molecular Modelling, Institute of Advanced Chemistry of Catalonia, Spanish Council for Scientific Research (IQAC-CSIC), Jordi Girona 18-26, 08034 Barcelona, Spain.
authors.forename: Aristotelis
authors.initials: A
authors.lastname: Kotronoulas
authors.affiliation: ⊥Department of Biological Chemistry and Molecular Modelling, Institute of Advanced Chemistry of Catalonia, Spanish Council for Scientific Research (IQAC-CSIC), Jordi Girona 18-26, 08034 Barcelona, Spain.
authors.forename: Jesús
authors.initials: J
authors.lastname: Joglar
authors.affiliation: #Human Pharmacology and Neurosciences Research Group, IMIM, Clinical Pharmacology Unit, Hospital Universitari Germnans Trias I Pujol-IGTP and Universitat Autònoma de Barcelona, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Magi
authors.initials: M
authors.lastname: Farré
authors.affiliation: ○National Anti-Doping Laboratory, China Anti-Doping Agency, 1, An Ding Road, Beijing, 100029, China.
authors.forename: Sheng
authors.initials: S
authors.lastname: Yang
authors.affiliation: ○National Anti-Doping Laboratory, China Anti-Doping Agency, 1, An Ding Road, Beijing, 100029, China.
authors.forename: Yanyi
authors.initials: Y
authors.lastname: Xing
authors.affiliation: #Human Pharmacology and Neurosciences Research Group, IMIM, Clinical Pharmacology Unit, Hospital Universitari Germnans Trias I Pujol-IGTP and Universitat Autònoma de Barcelona, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Peter
authors.initials: P
authors.lastname: Van Eenoo
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: Elena
authors.initials: E
authors.lastname: Pitarch
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: Félix
authors.initials: F
authors.lastname: Hernández
authors.affiliation: †Research Institute for Pesticides and Water, University Jaume I, Av. Sos Baynat S/N, 12071 Castellón, Spain.
authors.forename: Juan Vicente
authors.initials: JV
authors.lastname: Sancho
authors.affiliation: ‡Bioanalysis Research Group, IMIM, Hospital del Mar, Doctor Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Óscar J
authors.initials: ÓJ
authors.lastname: Pozo
mesh_terms: D002851:Chromatography, High Pressure Liquid; D006801:Humans; D008297:Male; D055432:Metabolomics; D015394:Molecular Structure; D012015:Reference Standards; D053719:Tandem Mass Spectrometry; D013739:Testosterone / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D016482:Urinalysis / Q000379:methods*
chemical_list: D013739:Testosterone