Determination of piroximone in human plasma by high-performance liquid chromatography

A rapid and selective high-performance liquid chromatographic (HPLC) method using solid-phase extraction (SPE) for measuring piroximone in plasma samples has been developed. Human plasma and internal standard were pipetted onto a Bond Elut C18 extraction cartridge conditioned with methanol and water. Impurities and proteins were washed out with water. Piroximone and internal standard were eluted with methanol. After evaporation, the residue was dissolved in the mobile phase and the aliquot injected into the HPLC system. Piroximone and its related compounds were separated on a reversed phase C18 HPLC column maintained at 50 degrees C using a mobile phase consisting of phosphate buffer and methanol. Piroximone-N-oxide, piroximone, and internal standard were detected spectrophotometrically at 340 nm. The extraction recovery for piroximone was 94%. The within- and between-run coefficients of variation were < 3% in the concentration range of 320-5,000 ng/ml and < 17.5% at lower concentrations, e.g., 20 ng/ml. The limit of detection was 10 ng/ml.