High-performance liquid chromatographic determination of quercetin in human plasma and urine utilizing solid-phase extraction and ultraviolet detection

An HPLC method for determining quercetin in human plasma and urine is presented for application to the pharmacokinetic study of rutin. Isocratic reversed-phase HPLC was employed for the quantitative analysis by using kaempferol as an internal standard. Solid-phase extraction was performed on an Oasis™ HLB cartridge (>95% recovery). The HPLC assay was carried out using a Luna ODS-2 column (150×2.1 mm I.D., 5 μm particle size). The mobile phase was acetonitrile–10 mM ammonium acetate solution containing 0.3 mM EDTA–glacial acetic acid, 29:70:1 (v/v, pH 3.9) and 26:73:1 (v/v, pH 3.9) for the determination of plasma and urinary quercetin, respectively. The flow-rate was 0.3 ml/min and the detection wavelength was set at 370 nm. Calibration of the overall analytical procedure gave a linear signal (r>0.999) over a concentration range of 4–700 ng/ml of quercetin in plasma and 20–1000 ng/ml of quercetin in urine. The lower limit of quantification was ∼7 ng/ml of quercetin in plasma and ∼35 ng/ml in urine. The detection limit (defined at a signal-to-noise ratio of about 3) was ∼0.35 ng/ml in plasma and urine. A preliminary experiment to investigate the plasma concentration and urinary excretion of quercetin after oral administration of 200 mg of rutin to a healthy volunteer demonstrated that the present method was suitable for determining quercetin in human plasma and urine.