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Determination of 13 C/ 12 C ratios of endogenous urinary steroids: method validation, reference population and application to doping control purposes
Author(s) -
Piper Thomas,
Mareck Ute,
Geyer Hans,
Flenker Ulrich,
Thevis Mario,
Platen Petra,
Schänzer Wilhelm
Publication year - 2008
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.3601
Subject(s) - chemistry , epitestosterone , chromatography , detection limit , analyte , etiocholanolone , gas chromatography , population , mass spectrometry , isotope ratio mass spectrometry , pregnanediol , androsterone , urine , analytical chemistry (journal) , steroid , biochemistry , demography , sociology , hormone
The application of a comprehensive gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS)‐based method for stable carbon isotopes of endogenous urinary steroids is presented. The key element in sample preparation is the consecutive cleanup with high‐performance liquid chromatography (HPLC) of underivatized and acetylated steroids, which allows the isolation of ten analytes (11 β ‐hydroxyandrosterone, 5 α ‐androst‐16‐en‐3 β ‐ol, pregnanediol, androsterone, etiocholanolone, testosterone, epitestosterone, 5 α ‐androstane‐3 α ,17 β ‐diol, 5 β ‐androstane‐3 α ,17 β ‐diol and dehydroepiandrosterone) from a single urine specimen. These steroids are of particular importance to doping controls as they enable the sensitive and retrospective detection of steroid abuse by athletes. Depending on the biological background, the determination limit for all steroids ranges from 5 to 10 ng/mL for a 10 mL specimen. The method is validated by means of linear mixing models for each steroid, which covers repeatability and reproducibility. Specificity was further demonstrated by gas chromatography/mass spectrometry (GC/MS) for each analyte, and no influence of the sample preparation or the quantity of analyte on carbon isotope ratios was observed. In order to determine naturally occurring 13 C/ 12 C ratios of all implemented steroids, a reference population of n = 61 subjects was measured to enable the calculation of reference limits for all relevant steroidal Δ values. Copyright © 2008 John Wiley & Sons, Ltd.