Antioxidative properties of pure compounds and phytogenic feed additives in a chemical (ORAC) and a cell-based assay (yeast)

Antioxidants in animal feed prevent oxidation of essential components like vitamins or fatty acids, confer benefits to animal health by scavenging excess free radicals, and improve storage stability of animal products. Phytogenic feed additives (PFAs) are derived from plant material and may contain considerable amounts of antioxidants. Chemical assays to assess antioxidative properties usually involve reduction of oxidants or radical scavenging. Innate antioxidative defence mechanisms are observed in living cells and yeast serves as a model organism for testing antioxidants in a cell-based assay comprising a wide range of potential modes of action.

In this study, the oxygen radical absorbance capacity assay (ORAC) and an assay based on growth arrest of Saccharomyces cerevisiae exposed to oxidative stress caused by hydrogen peroxide were applied. Both assays were employed to test natural and synthetic antioxidants and PFAs.

Antioxidative effects of gallic acid, quercetin and butylhydroxytoluene determined by the ORAC assay corresponded well to expected values, while ascorbic acid surprisingly did not show activity up to 25 µM. Three PFAs (see Table 1), which are complex mixtures of herbs, spices, extracts and essential oils, displayed values of 226, 320 and 603 µmol trolox equivalents/g. In the yeast assay, 8 mM ascorbic acid successfully countered growth arrest induced by 4 mM hydrogen peroxide, as expected. Other commonly used antioxidants like butylhydroxyanisole and ethoxyquin however were unable to reverse growth arrest, even at high concentrations. The same phytogenic feed additives which showed clear antioxidative activity in the ORAC assay were unable to prevent growth arrest in the yeast assay.

Well-established antioxidants and PFAs yielded diverging results in a chemical and a cell-based assay for antioxidant activity. Depending on the chemical environment, the same substance may act as pro- or antioxidant, which might explain the observed results.

Tab. 1: Composition of tested phytogenic feed additives (PFA)

PFA	Major ingredients
1	Preparation from Thymus vulgaris, Cinnamomum verum, and Allium sativum
2	Preparation from Carum carvi, Mentha x piperita and Citrus sinensis
3	Preparation from Mentha x piperita, Syzyghium aromaticum and Pimpinella anisum