Glucuronidation of the red clover isoflavone irilone by liver microsomes from different species and human UDP-glucuronosyltransferases.

Red clover (Trifolium pratense L.) is used as a source for isoflavone (IF) dietary supplements. In this study, we focused on the red clover IF irilone (IRI), because of its reported comparatively high bioavailability. Because the conjugative metabolism plays a key role in the elimination of IF, we investigated the species-specific differences and glucuronidation kinetics of IRI using different liver microsomes as well as the recombinant UDP-glucuronosyltransferases (UGTs) 1A1, 1A7, 1A8, 1A9, 1A10, and 2B15. Both possible monoglucuronides, the IRI-O-4'-monoglucuronide (IRI-G4') and the IRI-O-5-monoglucuronide (IRI-G5), were detected. Human liver microsomes (HLM) as well as rat liver microsomes predominantly formed IRI-G5, whereas for porcine liver microsomes, IRI-G4' prevailed. HLM showed an apparent V(max) value of 0.43 nmol/min · mg and an apparent K(m) value of 9.8 μM for the formation of IRI-G5 and a V(max) of 0.35 nmol/min · mg and a K(m) of 64.7 μM in the case of IRI-G4'. Formation of both glucuronides was best fit using the substrate inhibition equation. The glucuronidation of IRI by UGTs led to values for the intrinsic clearance varying between 4 and 100 ml/min · mg, with UGT1A7 showing the lowest and UGT1A10 the highest IRI conversion rate. The results indicate that IRI undergoes an efficient glucuronidation, presumably in the intestine and liver, following atypical kinetic profiles.