Linoleic acid emulsion system-thiocyanate method
Antioxidant activity of PM and PMK extracts was also
evaluated using the thiocyanate method (10). The reaction
mixture consisted of 0.28 g linoleic acid, 0.28 g of Tween
20 and 50 mL of phosphate buffer (0.2 M, pH 7.0). To
2.5 mL of the above linoleic acid emulsion, 0.5 mL of test
sample (28 mg/mL) and 2.5mL of phosphate buffer (0.2 M,
pH 7.0) were added and incubated at 50oC in the dark for
72 h. The mixture prepared, as above, without test sample
was the control. The readings were taken every 24 h. The
mixture (0.1 mL) was taken and mixed with 4.7 mL of
75% ethanol, 0.1 mL of 30% ammonium thiocyanate and
0.1 mL of 20 mM ferrous chloride in 3.5% HCl and allowed
to stand at room temperature. Precisely 3 min after the
addition of ferrous chloride to the reaction mixture, the
absorbance was measured at 500 nm. The relative inhibition
of linoleic acid peroxidation was calculated as (%)=[1−
(absorbance of the sample at 500 nm)/(absorbance of the
control at 500 nm)]×100. A greater percentage figure
indicates a greater level of antioxidant activity.