Analysis of Phytoestrogens by High Performance Liquid Chromatography

Phytoestrogens are biochemicals synthesized in plants which mimic steroidal estrogen activity in mammals. Analysis of these compounds in the legumes which produce them and in body fluids is important to the study of their physiological effects. High pressure liquid chromatography (HPLC) has been found to be an efficient and sensitive method of identification and quantitation of isoflavonoids, one class of phytoestrogen. Here we report the separation of three isoflavonoids, biochanin A, genistein and daidzein using an HPLC system with a Cg reverse phase column and a linear gradient mobile phase containing acetonitrile and acetic acid/water (10/90, v/v) over 60 minutes. Minimum detection limits for the three isoflavonoids were 0.556 mug/mL, 0.314 mug/mL, and 0.377 mug/mL, respectively. This method was used to measure the concentrations of isoflavonoids in two types of soy meal and in several animal feeds. Projected use of this assay includes studies of reproductive ability following ingestion of these isoflavonoids in domestic ruminants and in wild rodents