Abstract: The peroxyoxalate chemiluminescence(CL) detection method for the evaluation of the CL intensity of malondialdehyde(MDA) condensates with seven 2-thiobarbituric acid derivatives is described. The method consists of a flow injection technique together with a CL detection system using bis(2,4,6-trichlorophenyl) oxalate(TCPO) and hydrogen peroxide as chemiluminogenic reagents. Linear correlations between CL intensity and concentration are obtained for pmol levels of condensates. Among the condensates, 1,3-diethyl-2-thiobarbituric acid(DETBA)-MDA shows the largest CL intensity. High performance liquid chromatography (HPLC)/CL detection of DETBA-MDA and 1,3-diphenyl-2-thiobarbituric acid(DPTBA)-MDA using a mixture of TCPO and hydrogen peroxide in acetonitrile as a post-column reagent solution is also described. The detection limits for DETBA-MDA and DPTBA-MDA are 20 and 200 fmol, respectively, per 20 µL injection at a signal-to-noise ratio of 2. This HPLC/CL detection system was applied to the determination of MDA in rat brains by using DETBA as a fluorescent derivatizing reagent.

Abstract: Plasma was introduced into the PTFE reaction coil (10 mm x 0.5 mm) of the flow injection apparatus (diagram given) and mixed, at 95°C, with thiobarbituric acid (5 g l-1) - H3PO4 (100 mL l-1). This stream was then mixed with the extracting solvent, methylisobutylketone. The organic phase was continuously separated by a successive phase-separation system, and the absorbance was measured at 532 nm. The calibration graph was rectilinear for up to 10 µM of 1,1,3,3-tetraethoxypropane with a detection limit of 0.27 µM. Recovery of malondialdehyde was 84.5 to 90.2%. A simple, precise, and rapid method to measure plasma malondialdehyde (MDA) was developed by use of solvent extraction- flow injection analysis The reagent solution, containing thiobarbituric acid (TBA), 5 g/L in 100 mL/L phosphoric acid, and extraction solvent (methylisobutyl ketone, MIBK) were propelled with a double-plunger micro-pump at a flow rate of 0.3 mL/min, and 20 µL of sample was introduced into the reagent stream. After TBA-MDA reactant was extd. into MIBK, the organic phase was continuously separated by a successive phase-separation system equipped with two phase separators, and the absorbance of the TBA-MDA reactant was measured at 532 nm. This approach resulted in excellent sensitivity, a CV of <1.5%, a good correlation with the conventional manual method, and a sampling frequency of 7 samples/h, suggesting that this semiautomated method is suitable for measuring plasma MDA.