Characterization of the metabolites of irisflorentin by using ultra-high performance liquid chromatography combined with quadrupole/orbitrap tandem mass spectrometry.

Irisflorentin is one of the bioactive constituents from the root of Belamcanda chinensis (L.) DC, which displayed anti-inflammatory and anti-tumor activities. In this work, the in vitro metabolism of irisflorentin was investigated using liver microsomes and hepatocytes. The metabolites were identified by ultra-high performance liquid chromatography combined with quadrupole/orbitrap tandem mass spectrometry. Under the current conditions, a total of 11 metabolites were detected and structurally identified according to accurate masses, fragment ions and retention times. Metabolite M10, identified as 6,7-dihydroxy-5,3',4',5'-tetramethoxy isoflavone, was biosynthesized and unambiguously characterized by nuclear magnetic resonance spectroscopy. The metabolic pathways of irisflorentin included oxidation, demethylation and glucuronidation. M10 was the most abundant metabolite in all tested species. Further phenotyping studies revealed that α-naphthoflavone and ketoconazole displayed significant inhibitory effect on the formation of M10. Cytochrome P450 (CYP) 1A2 and 3A4 were the major enzymes responsible for the formation of M10 by using individual recombinant human CYP450 enzymes. For the first time the current study provides an overview of the in vitro metabolic fates of irisflorentin, which is helpful for us to predict the human metabolism and the potential drug-drug interactions caused by irisflorentin.