A SIMPLE, RAPID, HIGHLY SENSITIVE AND REPRODUCIBLE QUANTIFICATION METHOD FOR PLASMA MALONDIALDEHYDE BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

A simple, rapid, highly sensitive and reproducible quantification meth od for plasma malondialdehyde (MDA) was developed using high-performan ce liquid chromatography (HPLC). The present method is based on the th iobarbituric acid (TBA) reaction and reversed-phase HPLC separation wi th fluorescence detection. For sample preparation, an aliquot of 5 mu L plasma was mixed with 0.2% (w/v) TBA in 0.1 M sodium acetate buffer, pH 3.5. After heating at 95 degrees C for 60 min, the reaction soluti on was centrifuged and an aliquot of 10 mu L supernatant was injected into the HPLC system without neutralization or extraction procedures. The TBA-MDA adduct was separated on a reversed-phase column and quanti fied by a fluorescence detection (lambda ex = 515 nm; lambda em = 553 nm). The mobile phase was a mixture of acetonitrile:water (7:3, v/v) u nder isocratic conditions at ambient temperature. These procedures gav e good results with regard to sensitivity (detection limit S/N = 3, 0. 5 fmol per injection), linearity (0.5-500 fmol per injection), precisi on (between-assay C.V = 3.1%, within-assay C.V = 1.2%) and recovery (9 8.6%). The simple sample procedure, the highly sensitive detection lim it and the stability of the TBA-MDA adduct make the present method app licable to numerous clinical samples. As many as 250-300 samples per d ay can be assayed using an autosampler. (C) 1998 John Wiley & Sons, Lt d.