Method development and validation for isoflavones quantification in coffee

This paper reports the development and validation of an analytical method to quantify three isoflavones (daidzein, genistein and formononetin) in coffee beans and coffee brews, by HPLC with diode-array detection. The aglycones were released by methanolic acid hydrolysis in the presence of the antioxidant BHT and the internal standard 2′-methoxyflavone. The method showed high correlation coefficients (r > 0.999) for standards subjected to the entire procedure. For samples, good intra- and inter-day precisions (<8%), and accuracies (recoveries of 95 ± 1% for ground coffee and 92 ± 2% for coffee brew) were achieved. Detection and quantification limits ranged from 5 to 8 ng/mL and from 14 and 25 ng/mL, respectively. The proposed method proved to be sensitive, precise and accurate.