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Abstract
A simple, rapid and sensitive reversed-phase gradient High-Performance Liquid Chromatographic (HPLC) method is proposed for the determination of anthranilic acid derivatives in anti-inflammatory drugs and biological fluids.
HPLC analyses are performed with a Lichrosorb-RP 18, 10 μm, 250×4 mm I. D., column employing gradient elution for the development of the chromatogram. The gradient system is formed with the following solutions: A = 0.065M ammonium acetate and B = methanol. The flow rate is 0.8 ml/min and the detection is achieved at 282 nm. The retention time is 4.63 min for flufenamic, 4.99 min for mefenamic and 5.46 min for tolfenamic acid. The absolute detection limits are 0.5 ng for flufenamic acid, 0.7 ng for mefenamic acid and 1.0 ng for tolfenamic acid. The linearity is observed up to 70 ng for the flufenamic acid, 70 ng for the mefenamic acid and 80 ng for the tolfenamic acid injected. The method involves the use of solid phase extraction for sample clean-up and subsequent separation of anthranilic acid derivatives and internal standard-caffeine-from endogenous interfering compounds on a reversed-phase column.
The proposed method is applied to the analysis of flufenamic, mefenamic ar. d tolfenamic acid in blood serum (40 μl), urine (200 μl), tablets, capsules, suppositories, suspensions and ointment.