Quantitative analysis of phytoestrogens in kudzu-root, soy and spiked serum samples by high-pressure liquid chromatography

A sensitive and reliable HPLC method that allows simultaneous quantification of phytoestrogens extracted from kudzu-root and soy preparations, and serum samples has been developed. Kudzu-root and soy preparations were mixed with 5 mug flavone and 15 mug rutin (internal standards) and the phytoestrogens were extracted by using solid-phase (C-18) extraction cartridges. Blank or spiked serum samples were extracted by using either C, cartridges or trichloroacetic acid-methanol extraction. The extracts were analyzed by the HPLC equipped with a reverse-phase (250 x 4 mm, C-18) column and UV, diode-array or MS detector. A linear gradient of acetic acid and acetonitrile provided excellent separation of glycoside and aglycone-phytoestrogens from kudzu root and soy preparations. The C-18 cartridge extraction of serum yielded excellent recovery of both glycoside- and aglycone-phytoestrogens, while the trichloroacetic acid-methanol extraction yielded excellent recovery of glycoside: but poor recovery of aglycone compounds. UV and MS detectors were suitable for phytoestrogen analysis in plant and serum samples, while the diode-array detector was suitable for generating the UV absorbance curve for phytoestrogens. Copyright (C) 2004 John Wiley Sons, Ltd.