ASSAY OF MALONDIALDEHYDE IN BIOLOGICAL-FLUIDS BY GAS-CHROMATOGRAPHY MASS-SPECTROMETRY

Malondialdehyde (MDA) is assayed in femtomole quantities in biological samples by gas chromatography-mass spectrometry (GC-MS). The MDA trap ped in protein as a Schiff base is released by H2SO4, the protein prec ipitated using Na2WO4, and the MDA derivatized with pentafluorophenylh ydrazine to form the stable adduct, N-pentafluorophenylpyrazole. Negat ive chemical ionization (NCI) capability allows the sensitive detectio n of this MDA adduct in biological samples at a level of 5 nM on-colum n. A stable-isotope-labeled MDA, [H-2(2)]MDA, was used as an internal standard for quantitation. MDA recovery from plasma was 76%. This assa y provides two forms of confirmation of the analyte, retention time an d mass ion, thus minimizing error due to interfering compounds. The co mmonly used thiobarbituric acid assay for MDA overestimates the MDA le vels by over 10-fold, possibly resulting from cross-reactivity with ot her aldehydes and artifactual oxidation due to 100 degrees C temperatu re conditions. In our assay, all steps were performed at room temperat ure thereby suppressing artifactual oxidation of the sample. We have s uccessfully applied this assay to biological samples including plasma, tissue homogenates, and sperm. (C) 1994 Academic Press, Inc.