Abstract
Owing to occasional health damages caused by health food products derived from Pueraria mirifica (PM), the Japanese government has designated PM as an “ingredient calling for special attention”. Miroestrol is a specific isoflavone isolated from PM and possesses very strong estrogenic activity enough to induces side effects in small amount. Therefore, routine analyses for miroestrol quantification is recommended to control the safety and quality of PM products. However, miroestrol content in PM is quite low, and commercial reagent for its detection is rarely available. In this study, we developed a quantitative analysis method for miroestrol in PM without using its analytical standard by using the relative molar sensitivity (RMS) of miroestrol to kwakhurin, another PM-specific isoflavone, as a reference standard. The RMS value was obtained by an offline combination of 1H-quantitative NMR spectroscopy and a LC/PDA and miroestrol content was determined by single-reference LC/PDA using RMS. Furthermore, we investigated miroestrol content in commercially available PM crude drugs and products, and the RMS method was compared with the conventional calibration curve method in terms of performance. The rate of concordance of miroestrol contents determined by two method was 89 – 101%. The results revealed that our developed LC/PDA/MS method with RMS using kwakhurin as a reference standard was accurate for routine monitoring of miroestrol content in PM crude drugs and products to control their quality.
