2.6.1.2. Determination of thiobarbituric acid reactive substances (TBARS). The -TBARS value of the lipid extracted from the samples was estimated colorimetrically by the method of Porkony and Dieffenbacher, as described by Kirk and Sawyer (1991). A portion (200 mg) of sample was weighed into a 25-ml volumetric flask. An aliquot (1 ml) of 1-butanol was added to dissolve the sample. The mixture was made to volume and mixed. A portion (5.0 ml) of the mixture was pipetted into a dry stoppered test tube and 5 ml of TBA reagent (prepared by dissolving 200 mg of 2-TBA in 100 ml 1butanol, filtered, stored at 4 C for not more than 7 days) were added. The test tubes were stoppered, vortexed and placed in a water bath at 95 C for 120 min, then cooled. Absorbance (As) was estimated at 530 nm against water blank. A reagent blank was run and absorbance (Ab) measured. TBA value (mg of malonaldehyde equivalents/kg of tissue) was acquired by the formula:
¼ 50*ðAs AbÞ (1)
TBA
200