Abstract:The method of gas chromatography/combustion/isotope ratio mass spectrometry was developed for the determination of progesterone in beef samples in this study. The beef samples were extracted by acetonitrile oscillation and ultrasonic-assisted extraction, and were dehydrated by NaCl to induce separation of water and organic layers. After centrifugation and evaporation, the extracts were further treated with ZnCl2 to remove lipids and purified by solid phase extraction using LC-C18, LC-Si and LC-NH2 cartridges. After Pre-HPLC purification using a C18 column, the samples were analyzed for progesterone by GC/C/IRMS system. The δ13C value of endogenous progesterone in beef was about -25.70±0.13‰ (n=6), and the δ13C value of exogenous progesterone spiked in the sample was about -30.64±0.24‰ (n=6). One-way analysis of variance(ANOVA, p=2.23×10-14<0.05) indicated that there was a highly significant difference between the endogenous and exogenous δ13C value of progesterone, while there was no significant difference between the exogenous and standard solution δ13C value of progesterone. Meanwhile, the simulation experiment showed that the δ13C value of mixture (endogenous and exogenous) of progesterone in actual sample was homologous with the exogenous δ13C value of progesterone. The results showed that the method was specificity and reliable. GC/C/IRMS can be a powerful tool to investigate the origin of natural hormones. The method will fill the domestic technical gap for identifying the origin of hormones.
