Development of Bioluminescent Enzyme Immunoassay for S-Equol Using Firefly Luciferase and Its Application to the Assessment of Equol-Producer Status

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In this study, we developed a specific bioluminescent enzyme immunoassay (BLEIA) for S-equol, employing firefly luciferase as a labeling enzyme, as an alternative to HPLC methods. Satisfactory correlation (r=0.992) was shown when this S-equol BLEIA was compared with HPLC. The cross-reactivity with R-equol as its diastereoisomer is <5%, and that with daidzein, which is the substrate of equol, is 0.02%. Frequencies of Japanese equol producers determined using two distinct approaches were compared: a threshold value for urinary S-equol concentration of 232 ng/ml gave frequencies of 32% of men and 19% of women. These values correspond to the results for log10-transformed urinary S-equol to daidzein ratio threshold of −1.75, namely, 34% of men and 19% of women. When the changes in concentration of urinary equol and daidzein were measured after ingestion of isoflavone, the maximum concentration (Cmax) of urinary equol appeared after 9.6 h of isoflavone consumption; this Cmax was 2 h later than that for daidzein. The S-equol BLEIA documented in this study is expected to be an important tool for the assessment of equol producer status and demonstration of the bioavailability of isoflavone.