title: Isolation and quantification by high-performance liquid chromatography-ion-trap mass spectrometry of androgen sulfoconjugates in human urine.
abstract: Together with steroid glucuronides, sulfoconjugates may be used as markers of steroid administration as well as endogenous steroid production. A fast and sensitive analytical procedure has been developed for the simultaneous separation, determination and quantification of sulfate and glucuronide derivatives of testosterone (T), epitestosterone (E), androsterone (A), etiocholanolone (Etio) and dehydroepiandrosterone (DHEA) in human urine. First, a weak anion-exchange solid-phase extraction support (SPE Oasis WAX) was used for complete and rapid separation of sulfates and glucuronides in two extracts after loading of urine sample (2 mL). Then sulfates were analyzed directly by high-performance liquid chromatography-ion-trap mass spectrometry (LC-MS/MS) with electrospray ionization in negative mode. Chromatographic separation of the targeted sulfoconjugates was achieved using a Waters XBridge C18 column (150 mm x 4.6 mm I.D., 5 microm) with gradient elution. Assay validation demonstrated good performance for instance for T sulfate (TS) and E sulfate (ES) in terms of trueness (89-107%), repeatability (3.4-22%) and intermediate precision (5.8-22%) over the range of 2-200 ng/mL (corresponding to 1.5-147 ng/mL as free steroids). Results obtained on biological samples demonstrated the suitability of this analytical strategy for direct measurement of androgen sulfoconjugates and glucuroconjugates in human urine.
authors.affiliation: Laboratoire Suisse d'Analyse du Dopage, Institut Universitaire de Médecine Légale, Chemin des Croisettes 22, 1066 Epalinges, Vaud, Switzerland.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Strahm
authors.forename: Isabelle
authors.initials: I
authors.lastname: Kohler
authors.forename: Serge
authors.initials: S
authors.lastname: Rudaz
authors.forename: Sophie
authors.initials: S
authors.lastname: Martel
authors.forename: Pierre-Alain
authors.initials: PA
authors.lastname: Carrupt
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Veuthey
authors.forename: Martial
authors.initials: M
authors.lastname: Saugy
authors.forename: Christophe
authors.initials: C
authors.lastname: Saudan
mesh_terms: D000728:Androgens / Q000032:analysis / Q000302:isolation & purification / Q000652:urine*; D000738:Androsterone / Q000032:analysis / Q000302:isolation & purification / Q000652:urine; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003687:Dehydroepiandrosterone / Q000032:analysis / Q000302:isolation & purification / Q000652:urine; D004845:Epitestosterone / Q000032:analysis / Q000302:isolation & purification / Q000652:urine; D005043:Etiocholanolone / Q000032:analysis / Q000302:isolation & purification / Q000652:urine; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D015203:Reproducibility of Results; D052616:Solid Phase Extraction; D013739:Testosterone / Q000032:analysis / Q000302:isolation & purification / Q000652:urine
chemical_list: D000728:Androgens; D013739:Testosterone; D003687:Dehydroepiandrosterone; D004845:Epitestosterone; D005043:Etiocholanolone; D000738:Androsterone