DETERMINATION OF TOLFENAMIC ACID IN HUMAN PLASMA BY HPLM

Tolfenamic acid is a potent prostaglandin synthetase inhibitor used cl inically as non-steroidal anti-inflammatory and analgesic-antipyretic agent. A simple, sensitive, accurate, and precise reverse phase high p erformance Liquid chromatographic method has been developed and valida ted for the quantitative determination of tolfenamic acid in small vol umes of human plasma. The chromatographic separation of tolfenamic aci d and the internal standard (phenylbutazone) was performed on a revers ed phase, 5-mu m C18 column (250 x 4 mm) using acetonitrile-10 mM phos phoric acid (60:40, v/v) as mobile phase with a flow rate of L1 ml/min and the chromatographic peaks were detected at 280 nm. Plasma was dep roteinized with acetonitrile, the supernatant fraction was evaporated to dryness and the resulting residue was reconstituted in the mobile p hase and injected into the HPLC system. Calibration curves were linear in the range 0.2-5.0 mu g/ml with a squared correlation coefficient ( r(2)) of 0.999 or better and the detection limit was 50 ng/ml for 100- mu l plasma samples. The method was not interfered with by other endog enous compounds or metabolites and one assay can be completed in 12 mi n. The within-day and between-day assay variation for three different concentrations was found to be less than 6% and the accuracy was nearl y 100%.