G-Biosciences’ ORAC assay depends on the free radical damage to a fluorescent probe, such as fluorescein, to result in a change of fluorescent intensity and the degree of change is indicative of the amount of radical damage. The presence of antioxidants results in an inhibition in the free radical damage to the fluorescent compound. This inhibition is observed as a preservation of the fluorescent signal.
Antioxidant capacity is an overall ability of organisms or food to catch free radicals and prevent their harmful effect. Antioxidative effect includes protection of cells and cellular structures against harmful effect of free radicals, especially oxygen and nitrogen. Substances with antioxidative properties are called antioxidants. They are contained in food and food supplements, most commonly in fruits, vegetables, rice, wine, meat, eggs, and other foodstuff of plant and animal origin.
Antioxidative systems include antioxidative enzymes, that is, superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase, and nonenzymatic substrates, such as glutathione, uric acid, lipoic acid, bilirubin, coenzyme Q, vitamin C (L-ascorbic acid), vitamin A (retinol), vitamin E (tocopherol), flavonoids, carotenoids, and others. Some biomolecules are also considered biologically active and clinically significant antioxidants, for example, transferrin, ferritin, lactoferrin, ceruloplasmin, hemopexin, haptoglobin, and uric acid.
It is possible quantitate the protection by calculating the area under the curve (AUC) from the experimental sample. After subtracting the AUC for the blank, the resultant difference would be the protection conferred by the antioxidant compound. Trolox®, (6-hydroxy-2,5,7,8-tetrametmethylchroman-2-carboxylic acid) a water-soluble vitamin E analog, is used as the calibration standard and ORAC results are expressed as Trolox® equivalents.
The ORAC assay is unique in that because the assay is driven to completion the AUC calculation combines both the inhibition time as well inhibition percentage of free radical damage by the antioxidant into a single quantity.