Determination of (+)-catechin in plasma by high-performance liquid chromatography using fluorescence detection.

Abstract

A high-performance liquid chromatographic method, using fluorescence detection, was developed for the determination of (+)-catechin in rabbit plasma. The procedure involved the precipitation of plasma protein using acetonitrile, followed by solid-phase adsorption onto alumina. After washing with water and methanol, the residue was vortex-mixed with perchloric acid solution to release the adsorbed (+)-catechin. Separation was performed on a reversed-phase column using an eluent consisting of phosphoric acid solution with 12% acetonitrile. The excitation and emission wavelengths were set at 280 and 310 nm, respectively. The retention times for (+)-catechin and the internal standard (deoxyhigenamine) were 6.87 and 8.47 min respectively, without any interference. Validations of accuracy and precision were satisfactory in both within- and between-run assays. All coefficients of variance were less than 6% and mean relative errors were within +/- 3.75%. The average recovery was 73.77%. The limit of detection and quantitation were 1 ng and 0.02 micrograms/ml, respectively. Application of this method was successfully assessed by intravenous administration of a 15 mg/kg dose of (+)-catechin in rabbits. This new method provides a simple, specific and sensitive determination for (+)-catechin in rabbit plasma and is suitable for pharmacokinetic studies.