Identification of equol‐7‐glucuronide‐4′‐sulfate, monoglucuronides and monosulfates in human plasma of 2 equol producers after administration of kinako by LC‐ESI‐MS

Equol is a product formed during the biotransformation of the naturally occurring isoflavone daidzein by intestinal bacteria. The role of equol in the prevention of several hormone‐dependent diseases such as prostate cancer and osteoporosis as well as vasomotor symptoms has been extensively investigated. Equol primarily occurs in the form of major metabolites such as glucuronides and sulfates, while intact equol has been detected at only ca. 1% in human plasma. However, to date, conjugated metabolites have been evaluated by measuring the free equol obtained after selective enzymatic hydrolysis. Thus, the precise types of conjugates circulating in vivo and the position(s) of the conjugation sites on the equol skeleton have yet to be clarified. Our study describes the identification of polar equol metabolites in the plasma of 2 equol‐producers obtained at 8 hours after consuming 50 g of kinako (approximately 37 mg of daidzein). The structural identification of these conjugated metabolites in plasma was performed by comparison to the LC‐ESI‐MS (n) and (1)H‐NMR spectral data of the corresponding chemically synthesized compounds. The results of the LC‐ESI‐MS/MS analysis indicated that the main conjugated metabolite in plasma was (S)‐equol‐7‐glucuronide‐4′‐sulfate along with lower amounts of 7‐ and 4′‐monoglucuronides as well as 7‐ and 4′‐monosulfates.