Simultaneous determination of different flavonoids in human plasma by a simple HPLC assay

Abstract

A simple HPLC method for simultaneous determination of five flavonoids in human plasma was developed. Following addition of catechin as internal standard (IS), luteolin-7-glucoside, rutin, apigenin-7-glucoside, quercetin and baicalein were isolated from human plasma by liquid-liquid extraction with acetone. The chromatographic separation was achieved in a DAD-HPLC equipment on a reversed-phase C18 column using a gradient mobile phase consisting of 0.1% acetic acid + methanol: acetonitrile: acetic acid (90:10:1, v/v) and running at a flow rate of 1 mL/min. The effluent was monitored at a wavelength of 280 ηm, as well as IS was well separated from each other and free from interference from blank plasma and other components in plasma. The calibration curve was constructed by plotting peak area ratio of analytes to IS vs. concentration. The method showed good linearity over range of 5 - 30 μg/mL for the five standards used (r>0.996). The developed method may be used not only to identify the analytes assayed but, simultaneously measure plasma concentrations of these herbal compounds.