Development, Validation, and Application of HPLC Method for Quantification of Antihyperuricemic Compounds from Lippia nodiflora in Rat Plasma

Abstract

An HPLC method for simultaneous determination of arenarioside (1 ), verbascoside (2), 6-hydroxyluteolin (3), 6-hydroxyluteolin-7-O-glycoside (4), and nodifloretin (5) from Lippia nodiflora in rat plasma was developed and validated. The optimal chromatographic separation was achieved with a gradient mobile phase comprising 0.1% aqueous acetic acid and acetonitrile. The limit of detection was 78.1 ng/mL for 3 and 39.1 ng/mL for the other compounds (signal-to-noise ratio=3), whereas the limit of quantification was 312.5 ng/mL for 3 and 156.3 ng/mL for the other compounds (signal-to-noise ratio=12). The recovery values of compounds 1–5 ranged from 89.37–100.92%. Their accuracy values were between 96.48 and 105.81%, while their corresponding precision values were in the range of 0.75–9.06% for both intraday and inter-day analysis. The method was then applied in the first pharmacokinetic study of 1–5. Following intravenous administration, 1–5 were eliminated slowly from the body with a mean clearance value of 0.11, 0.13, 0.30, 0.09, and 0.23 L/kg h, respectively. Meanwhile, their peak plasma concentration upon oral administration was 8.97, 1.07, 1.06, 0.65, and 0.38 µg/mL, respectively. Compound 3 (5.97%) exhibited the highest absolute oral bioavailability value, followed by 1 (5.22%), 4 (3.13%), 2 (2.10%), and 5 (0.93%).