Bioassay-guided fractionation of an ethanolic extract from aerial parts of Athrixia phylicoides using silica and sephadex column chromatography led to the isolation of three flavonoids. The compounds were identified as: 5-hydroxy-6,7,8,3’,4’,5’-hexamethoxyflavon-3-ol (1), 3-0-demethyldigicitrin (2), and Quecertin (3). Isolated compounds together with ethanol crude extract were tested for antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH)-spectrophotometric assay, while cytotoxicity effect was determined using XTT colorimetric assay. The crude extract showed a concentration-dependent radical scavenging activity with EC50 value of 10.64 ± 0.08 μg/ml. Compound 3 was the most potent radical scavenger, exhibiting EC50 value of 1.27 ± 0.25 μg/ml, followed by compound 1 and 2 showing 2.74 ± 0.10 and 3.41 ± 0.09 μg/ml respectively. The crude extract showed no or little toxicity on Vero cells at lower concentrations tested exhibiting the IC50 value of 107.8 ± 0.13 μg/ml. Compound 3 showed minimal toxicity effect by exhibiting IC50 value of 81.38 ± 0.33 μg/ml as compared to compound 2 (IC50,
28.92 ± 0.12 μg/ml) and compound 1 (IC50, 27.91 ± 0.18 μg/ml). The results obtained from this study
provide a clear rationale for the medicinal uses of A. phylicoides.