Establishment of an HPLC Method for Determination of Coumarin-3-Carboxylic Acid Analogues in Rat Plasma and a Preliminary Study on Their Pharmacokinetics.

ABSTRACT

A simple high performance liquid chromatography (HPLC) method was developed and validated for the determination of coumarin-3-carboxylic acid analogues (C3AA) in rat plasma and a preliminary study on pharmacokinetics. Ferulic acid (FA) was used as the internal standard substance, and coumarin-3-carboxylic acid (C3A) was used as a substitute for quantitative C3AA. After protein precipitation with methanol, the satisfactory separation was achieved on an ODS2 column when the temperature was maintained at 30 ± 2°C. The correlation coefficient r in the C3A linear equation is equal to 0.9990. Pharmacokinetic parameters for t1/2, Tmax, Cmax, area under the curve (AUC)0-t, average residence time (MRT), apparent volume of distribution (V z/F) and clearance (Cl/F) were 1.89 ± 0.03 h, 0.39 ± 0.14 h, 1.81 ± 0.10 g· mL-1 ·h, 7.88 ± 0.24 g·mL-1·h, 3.23 ± 0.14 h, 0.43 ± 0.03 (mg·kg-1)·(g·mL-1)-1·h-1, respectively. The high performance liquid chromatography-photo diode array detector (HPLC-PDA) method established in this study can be used to separate and determine the content of C3AA in plasma of rats after 60% ethanol extraction by gavage. The plasma concentration-time curve and pharmacokinetic parameters reflect the absorption of C3AA in rat blood after oral administration to some extent.