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

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Abstract
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 µg flavone and 15 µg rutin (internal standards) and the phytoestrogens were extracted by using solid‐phase (C18) extraction cartridges. Blank or spiked serum samples were extracted by using either C18 cartridges or trichloroacetic acid–methanol extraction. The extracts were analyzed by the HPLC equipped with a reverse‐phase (250 × 4 mm, C18) 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 C18 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 © 2004 John Wiley & Sons, Ltd.