2.10. Determination of malondialdehyde (MDA)

The concentration of malondialdehyde was quantified for the assessment of lipid peroxidation level [38]. Spleen and liver samples (0.4 mL) were mixed with 1.6 mL of Tris-KCl buffer (0.15 M, pH-7.4) to which 0.5 mL of 30% trichloroacetic Acid (TCA) was added. Then 0.5 mL of 0.75% Thiobarbituric Acid (TBA) was added and incubated in a water bath for 45 min at 80 °C. After incubation a pink colored reaction mixture was cooled in ice and centrifuged at 10,000 rpm for 15 min. The absorbance of the clear pink supernatant was read at 532 nm.

Calculation:

Where, E₅₃₂ is extinction coefficient of TBA at 532 nm = 1.56 × 10⁵.