Analysis of exogenous dehydroepiandrosterone excretion in urine by gas chromatography/combustion/isotope ratio mass spectrometry

Abstract

A detailed procedure for the analysis of exogenous dehydroepiandrosterone (DHEA) in urine by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) has been established for detecting doping with DHEA. The average delta-value (parts per thousand difference of (13)C/(12)C ratio from the isotope ratio standard) of 26 synthetic steroids commercially available was -30.1 +/- 2.6, and was significantly lower than that of human endogenous DHEA in urine of the world class athletes who had participated in the XVIIth Olympic Winter Games (-20.3 +/- 2.1, n = 446). Although large inter-individual variations of urinary DHEA excretion were observed following a single oral administration of 50 mg of DHEA, no significant inter-individual difference was found when the excretion of exogenous DHEA was monitored in terms of delta-values using GC/C/IRMS; the minimum delta-values were observed around 6-8 h after the administration, and the values returned to the base level at over 72 h after the dosing. Thus, the deviations in delta-values of DHEA and its diol metabolites are considered to be conclusive evidence for detecting doping with DHEA. Some successful cases of detection of doping with DHEA from athletes are also reported.

Recent developments in the use of isotope ratio mass spectrometry in sports drug testing

“…Derivatisation with O-ethylhydroxylamine was investigated for determination of etiocholanolone (E) and androsterone (A) [51], and Ueki and Okano [17] investigated PD, 5a and 5b excreted glucuronidated and added dehydroepiandrosterone sulfate (DHEA_S) to the scope of IRMS methods using a TEAP-LH20 anion exchange column.…”

“…2, the difference between endogenous and synthetic steroids is not always large enough to allow them to be distinguished. Whereas endogenous steroids reflect the CIR of a mixture of C 3 and C 4 plants, synthetic steroids should exhibit a clear single C 3 signature, resulting in values lower than −26.5‰ [17,18]. Some rare cases have been reported with steroids exhibiting δ 13 C values in the endogenous range [61,82] and recently a comprehensive investigation on seized T samples showed that nearly 10% of all T preparations were enriched more than −25‰ [81].…”

“…C values of endogenous steroids fall between −26‰ and −16‰, whereas pharmaceutical preparations have δ 13 C values below −27‰ [17][18][19]. As the absolute difference between endogenous and exogenous steroids can be quite small, WADA established difference (Δ) values between endogenous reference compounds (ERC) and target compounds (TC) to distinguish the source of urinary steroids.…”

Combination of carbon isotope ratio with hydrogen isotope ratio determinations in sports drug testing

“…Within the last years, the number of investigations on carbon isotope ratios (CIR) of endogenous urinary steroids in the context of sports drug testing has considerably increased [1][2][3][4][5][6][7][8][9][10][11][12][13][14]. Regarding absolute values expressed as δ 13 C VPDB against the international standard Vienna Pee Dee Belemnite (VPDB) based on Eq.…”

Toxicology of Specific Substances

“…Differentiation of natural (endogenous) from synthetic (exogenous) hormones -AASs steroidal products yielded δ 13 C values of -30.1‰ ± 2.6‰ and -25.9‰ to -32.8‰ (Ueki and Okano 1999;de la Torre et al 2001); however, δ 13 C values reflecting typical 'human' CIRs were also reported (Cawley et al 2010;Forsdahl et al 2011) representing an additional challenge for doping control authorities. To account for the fact that diets vary considerably, and thus also δ 13 C values of urinary steroids, endogenous reference compounds (ERCs), i.e.…”