Large volume injection: improving the detection capacity of endogenous anabolic steroids by GC-C-IRMS

The 13C/12C ratio, expressed as δ13C (‰) value, is determined through a GC-C-IRMS (gas chromatography combustion isotope ratio mass spectrometry) analysis to discriminate the exogenous or endogenous origin of anabolic androgenic steroids (AAS), included in the list of prohibited substances and methods published yearly by the World Anti-Doping Agency (WADA). The GC-C-IRMS method is a confirmation procedure performed in all accredited doping laboratories when urinary concentration of specific steroid profile markers or metabolites exceeds the limits defined in reference Technical Documents (TD) or after specific request of the Testing Authority. The technical requirements to establish the origin of boldenone and 19-norandrosterone (19-NA, the main metabolite of nandrolone) require high analysis sensitivity: the GC-C-IRMS method must be able to detect very low urinary levels (2 ng/mL). Until now, to meet the WADA criteria, we process a large urine sample volume (21 mL) and we inject 2 or 3 µL to obtain adequate signals within the linear range of the procedure (210 – 6000 mV). The data collected by injecting large sample volume are discussed in order to propose an alternative instrumental method to improve sensitivity and, at the same time, to reduce the needed urine volume.