DPPH assay of vegetable oils and model antioxidants in protic and aprotic solvents.

The rate of reaction of phenolic antioxidants with DPPH depends on solvent composition. The rate constants can differ by more than two orders of magnitude for the same phenolic compound. Reactions are faster in alcohols than in ethyl acetate that is used routinely for the analysis of antioxidant potential (AOP) of nonpolar samples such as vegetable oils. Incorporation of an acid base pair into the assay solvent buffers the system against acid impurities such as free fatty acids and CO2 from the air. This is shown to increase the rate of oxidation and number of electrons of phenolic compounds exchanged with DPPH. Typically, DPPH assays are performed for predetermined time intervals at which phenolic compounds are not fully oxidized and therefore higher reaction rates result in higher values of AOP. More than twofold AOP was obtained for oleuropein, sesamol, sinapic acid, caffeic acid and protocatechuic acid in buffered alcohols than in ethyl acetate. The AOP of sesame, pumpkin seed and extra virgin olive oil is accordingly higher when determined in buffered alcohols. DPPH assays in ethyl acetate result in underestimation of AOP of unrefined vegetable oils.