The antioxidant and cellular effects of Withania somnifera alone and in combination with selenium

Abstract

The respiratory system is a target for environmental oxidants from airborne pollution and other contaminants. Chronic exposure may result in inflammatory conditions such as allergies and asthma. The ability of the body to counteract oxidation is dependent on the oxidative status of the individual, and can also benefit from the use of medicinal plant preparations such as Withania somnifera (WS), which is accredited with antioxidant and antiinflammatory properties. Selenium is an important cofactor of antioxidant enzymes such as glutathione peroxidase. A deficiency of Se will result in decreased antioxidant enzymes levels and a poor response to oxidative damage. The aim of this study was to determine the antioxidant content and activity of WS, as well as its in vitro effects with seleno-L-methionine (Se-Met), the major component of dietary selenium, in a physiologically relevant cell model system. Little is known about the combinational effects of non-enzymatic and enzymatic antioxidants i.e., WS in combination with Se-Met, and these effects were also quantified. The antioxidant content (total phenolic (TPC) and flavonoid content (TFC)) and activity (2,2- diphenyl-2-picrylhydrazyl (DPPH), trolox equivalent antioxidant capacity (TEAC)) and oxygen radical absorbance capacity (ORAC) of a water extract of WS, as traditionally prepared and used, was determined. The intracellular and total protection of WS alone (0-33.33 mg/ml) and in combination with Se-Met (0-33.33 pg/ml) (range: ½-100x RDA) was evaluated in a standard cell line, the SC-1 fibroblast cell line and in physiologically relevant primary cultures of chick embryo lung cells (CELC) using the 2’,7’-dichlorofluorescein diacetate (DCFH-DA) assay. Water extracts of WS, as traditionally consumed, were found to possess significant antioxidant content and activity. For TPC and TFC, WS was found to contain 4.84 g CE/100 g DW, and 355 mg CE/100 g DW respectively. For DPPH, the IC50 of WS was 56.34 mg/ml, and the antioxidant activity of WS was 274.8 μmol TE/g. For TEAC, the IC50 of WS was 60.3 mg/ml, and the antioxidant activity of WS was 2.2 mmol TE/g. Using ORAC, the antioxidant activity of WS was found to be 15.05 μM TE/g. WS and Se-Met exhibited both pro-oxidant and antioxidant effects in the DCFH-DA assay and this may be a function of concentration, as well as combinations of WS and Se-Met ratios. Data indicates that both WS and Se-Met, alone and in combination, elicit protection via the intracellular (at all concentrations) and extracellular (at higher concentrations) compartment of the SC-1 cell. A synergistic interaction is observed extracellularly between WS and Se-Met, with mostly additive effects occurring intracellularly. For CELC, WS and Se-Met alone both demonstrated total protective effects at almost all concentrations, and intracellular protective effects at higher concentrations. The combination of WS and Se-Met provided total and intracellular protection to the CELC at the highest concentration combination. Antagonism between WS and Se-Met was observed at low concentrations in the total protection assay, with weak synergism and antagonism occurring at higher concentrations. Synergism between WS and Se-Met was observed at low concentrations intracellularly, with antagonistic interactions occurring at higher concentrations. In conclusion, WS and Se-Met, alone and in combination at non-cytotoxic concentrations, displayed significant cellular protection against oxidative damage and this was a function of cell line, concentration and ratios between WS and Se-Met.