Erythrocytes and cell line-based assays to evaluate the cytoprotective activity of antioxidant components obtained from natural sources

Abstract

Oxidative stress can damage cellular components including DNA, proteins or lipids, and may cause several skin diseases. To protect from this damage and addressing consumer's appeal to natural products, antioxidants obtained from algal and vegetal extracts are being proposed as antioxidants to be incorporated into formulations. Thus, the development of reliable, quick and economic in vitro methods to study the cytoactivity of these products is a meaningful requirement. A combination of erythrocyte and cell line-based assays was performed on two extracts from Sargassum muticum, one from Ulva lactuca, and one from Castanea sativa. Antioxidant properties were assessed in erythrocytes by the TBARS and AAPH assays, and cytotoxicity and antioxidant cytoprotection were assessed in HaCaT and 3T3 cells by the MTT assay. The extracts showed no antioxidant activity on the TBARS assay, whereas their antioxidant capacity in the AAPH assay was demonstrated. On the cytotoxicity assays, extracts showed low toxicity, with IC50 values higher than 200μg/mL. C. sativa extract showed the most favourable antioxidant properties on the antioxidant cytoprotection assays; while S. muticum and U. lactuca extracts showed a slight antioxidant activity. This battery of methods was useful to characterise the biological antioxidant properties of these natural extracts.

Keywords: 2,2-azobis(2-amidinopropane) dihydrochloride; 3-[4,5-dimethylthiazol-2-yl]-2-5-diphenyltetrazolium bromide; AAPH; Antioxidant extract; CmaxA-OX; Cytotoxicity; DMEM; DPPP; Dubecco’s modified Eagle’s medium; FBS; Haemolysis; MTT; ROS; TBARS; concentration with a maximal antioxidant effect; diphenyl-1-pyrenylphosphine; foetal bovine serum; reactive oxygen species; thiobarbituric acid reactive species.