Development and validation of a sensitive UFLC−MS/MS method for quantification of quercitrin in plasma : application to a tissue distribution study

Abstract

Quercitrin, a glycosylated form of the flavonoid quercetin, is one of the major constituents of Copaifera langsdorffii leaves and potentially contributes to the medicinal properties reported for this plant species, including the treatment and prevention of kidney stones. To better understand the pharmacokinetics of quercitrin, a simple, rapid, and sensitive ultra fast liquid chromatography−tandem mass spectrometry method was developed and validated for the quantification of quercitrin in rat plasma and applied to a tissue distribution study. Sample preparation involved simple liquid−liquid extraction by ethyl acetate with high efficiency, using afzelin as internal standard. The chromatographic separation was performed on a Phenomenex Synergi Polar-RP (100 × 3.0 mm2, 2.5 μm), with a gradient elution of acetonitrile and 0.5% formic acid in water. The mass spectrometry analysis was conducted in negative ionization mode with multiple reaction monitoring transitions at m/z 447 → 300 for quercitrin and m/z 431 → 281 for afzelin. The method showed linearity in the concentration range of 5−100 ng/mL (r2 > 0.9959) and the lower limit of quantification was 5 ng/mL. The intraday and interday precision (relative standard deviation) were less than 10.73%, whereas the accuracy ranged from 81.4 to 111.0%. The extraction recovery, stability, matrix effect, and integrity dilution involved in the method were also validated. In addition, tissue distribution was assessed after an intravenous administration of 1 mg/kg quercitrin. This is the first report quantifying quercitrin in kidneys, demonstrating that the free tissue/plasma ratio was 23.7.