Abstract: Objective: The aim of this study is to investigate the antioxidant activity in vitro of anthocyanins extracted from the fruits of Habenaria ciliolaris Kranzl and the protective effect of the extract on H2O2-induced oxidative stress in V79-4 cells. Methods: The anthocyanin composition was detected by high performance liquid chromatography-mass spectrometry, and the antioxidant activity was evaluated by using 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assays. Moreover, the protective effect of the extract on H2O2-induced oxidative damage in V79-4 cells was measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl- 2-H-tetrazolium bromide, thiobarbituric acid reactive substances and 2’,7’-dichlorofluorescein diacetate assays. Results: The anthocyanins in the extract were identified as cyanidin 3-galactoside, cyanidin 3-(6”-coumaroyl) glucoside, peonidin 3-galactoside and peonidin 3-(6”-coumaroyl) glucoside by retention time and mass spectral data. The anthocyanins extract exhibited strong ABTS and DPPH free radical scavenging capacity with half maximal inhibitory concentrations of (1.72 ± 0.26) and (0.74 ± 0.24) μg/mL, respectively, which was superior to that of soluble VE. The pretreatment of V79-4 cells with the extract in the ranges of 50-200 μg/mL significantly reduced H2O2-induced cytotoxictiy, potentially inhibited lipid peroxidation and reduced the level of intracellular reactive oxygen species. Conclusion: These findings confirm the in vitro free radical scavenging activity of anthocyanins extracted from H. ciliolaris Kranzl and their cytoprotective effect on H2O2-induced oxidative stress in V79-4 cells.

Key words: Habenaria ciliolaris Kranzl, anthocyanins extract, antioxidant, free radical, oxidative stress damage