[Determination of tiopronin in rat plasma by HPLC following fluorescent derivatization]

A sensitive, rapid method for determining reduced tiopronin concentration in rat plasma has been developed by using a high-performance liquid chromatography (HPLC) technique in conjunction with the derivatizing agent N-(1-pyrenyl) maleimide (NPM). The analytes were separated on a Kromasil C18 column (250 mm x 4.6 mm, 5 microm) using 0.2% glacial acetic acid aqueous solution including 0.015 mol x L(-1) KH2PO4 and acetonitrile (56:44) as a mobile phase at a flow-rate of 0.8 mL x min(-1), and fluorescence detection wavelength were set at lamda(e x) = 340 nm and lamda(e m) = 375 nm, the column temperature was 30 degrees C. The calibration curve was found to be linear over a range of 0.1 - 10.0 microg x mL(-1), the limit of quantitation was 0. 1 mg x L(-1). The coefficients of the variation for the within-run and between-run precisions ranged from 5.3% to 10.8% and 7.0% to 10.8%, respectively. The percentage of absolute recovery ranged from 73.7% to 79.7%. The method was used to determine the concentration of tiopronin in rat plasma after a single intragastric administration of 25 mg x kg(-1) tiopronin to 6 healthy male Wistar rats. The pharmacokinetic process was fitted to a two-compartment model. The method has been successfully applied to the determination of tiopronin in rat plasma.