Certification of steroid carbon isotope ratios in a freeze‐dried human urine reference material

An accurate method for the measurement of carbon isotope ratios of steroids in human urine has been developed at the National Measurement Institute, Australia (NMIA) for the certification of a freeze-dried human urine reference material (CRM NMIA MX005). The method measures δ13C values by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) analysis following hydrolysis, solvent extraction and high performance liquid chromatography (HPLC) purification. Reference δ13C values for testosterone metabolites etiocholanolone, androsterone, and endogenous reference compounds (ERCs) 11β-hydroxyandrosterone and pregnanediol were determined, as well as information δ13C values for testosterone, epitestosterone, 11-oxoetiocholanolone, and a range of differences (Δ13C) between testosterone metabolites and ERCs. The measurement uncertainty was rigorously evaluated with expanded uncertainties for the reference δ13C values between 1.1 and 1.6 ‰ at the 95% coverage level. Copyright © 2012 John Wiley & Sons, Ltd.