Measuring Malondialdehyde Levels in Rat Tissues

The concentration of MDA in the various rat homogenates was measured as previously described [21 (link)]. This assay relies on the reaction of the 2-thiobarbituric acid with malondialdehyde at 70°C. A single molecule of malondialdehyde complexes two molecules of 2-thiobarbituric acid Knoevenagel-type condensation to yield a chromophore with absorbance maximum at 532 nm. A volume of 2 mL of MDA working solution (Trichloroacetic acid (10 · 10⁻³ M) (Sigma-Aldrich, Germany) and 1 ml of 2-thiobarbituric acid (67 · 10⁻³ M) (Sigma-Aldrich, Germany) were added to a test tube containing 100 of the sample. The mixture was vortexed and incubated at 100°C for 15 min. Then the tubes were allowed to cool at room temperature and centrifugated at 3000 rpm for 5 min. The supernatant of each tube was collected and the OD were read at 532 nm [8 (link)].