Profiling of 19-norsteroid sulfoconjugates in human urine by liquid chromatography mass spectrometry.
19-Nortestosterone (nandrolone) major metabolites in human urine are excreted as sulfoconjugated and glucuroconjugated forms. A sensitive and selective liquid chromatography/tandem mass spectrometry (LC/MS/MS) method in negative ESI mode was developed for direct quantification of 19-norandrosterone sulfate (19-NAS) and 19-noretiocholanolone sulfate (19-NES). For both sulfoconjugates, the [M-H](-) ion at m/z 355 and the fragment ion at m/z 97 were used as the precursor and product ions, respectively. The purification method involved a complete and rapid separation of sulfates and glucuronides in two extracts after loading the sample on a weak anion exchange solid phase extraction support (SPE Oasis WAX). Then, sulfates were separated by LC (Uptisphere ODB, 150 mm x 3.0 mm, 5 microm) and analyzed on a linear trap and a triple quadrupole mass spectrometer. The lower limit of detection (LLOD) and lowest limit of quantification (LLOQ) were of 100 pg mL(-1) and 1 ng mL(-1), respectively. Assay validation demonstrated good performances in terms of trueness (92.0-104.9%), repeatability (0.6-7.2%) and intermediate precision (1.3-10.8%) over the range of 1-2500 ng mL(-1). Finally, 19-NAS and 19-NES in urine samples collected after intake of 19-norandrostenedione (nandrolone precursor) were quantified. This assay may be easily implemented to separate glucuronide and sulfate steroids from urine specimens prior to quantification by LC/MS/MS.
Strahm
Emmanuel
E
Laboratoire Suisse d'Analyse du Dopage, Institut Universitaire de Médecine Légale, Chemin des Croisettes 22, 1066 Epalinges, Switzerland.
Rudaz
Serge
S
Veuthey
Jean-Luc
JL
Saugy
Martial
M
Saudan
Christophe
C
D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013256:Steroids / Q000652:urine*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D013256:Steroids