Metabolism profiles of icariin in rats using ultra-high performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry and in vitro enzymatic study

Icariin (ICA), the major constituent of Epimedium brevicornu Maxim, is recognized as an effective agent for the treatment of osteoporosis. In our previous report, the relatively short half-life (74min) and poor bioavailability (approximately 0.1%) of ICA in rats suggested that not only ICA itself but also its metabolites could be responsible for the observed osteoporosis treatment effect. Therefore, the present study aimed at identifying the metabolites of ICA in rat plasma, bile, urine, and feces after the administration of a single oral dose of ICA (150mg/kg). In this work, ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC/Q-TOF-MS) method was established to identify the metabolic profiles following intragastric administration of pure ICA in rats. The blood, bile, urine, and feces samples of the rats were investigated to explore the complete metabolic pathway of ICA in vivo. A total of 14 metabolites were detected in the bile, revealing that the bile is the main excretion pathway for ICA and its metabolites. The conjugated metabolites as observed in vivo, was further confirmed by the in vitro enzymatic study. Five metabolites of ICA, including demethylicariin, icariside I-3-O-glucuronide, demethylicariside II, demethylicariside II-7-O-glucuronide, and dehydroxyicaritin-glucuronide, were reported for the first time in the literature. In addition, the results revealed that the principal metabolism pathways of ICA in rat were deglycosylation and glucuronidation after deglycosylation.