Purpose: Bidens pilosa L.var.radiata Sch Bip (BP) is widely distributed in tropical,subtropical and temperate regions.The present study aimed to investigate the chemical composition of BP, as well as the antioxidant and anti-proliferative activities.Methods: The whole herb of Bidens pilosa L.var.radiata.Sch Bip (BP) was extracted with 95% ethanol, which was then partitioned sequentially with petroleum ether, EtOAc and n-BuOH to afford a petroleum ether fraction, ethyl acetate fraction (EE-BP), n-BuOH fraction and the remainder of the water fraction.EE-BP was further isolated and purified by repeated column chromatography, the obtained compounds were identified by comparing 1H and 13C NMR spectrum with published chemical.The flavonoids were evaluated for the DPPH and ABTS free radical scavenging activity by colorimetric method, as well as their anti-proliferative effect against RKO cells by MTT method.Results: Our phytochemical investigation resulted in a total of 6 compounds first isolated from BP.They were identified as flavonoids or flavonoid glycosides,including isoquercitrin (1), vitexin (2), astragalin (3), 5,6,7,4-tetramethoxylflavone (4), 5,3′,4′-trihydroxy-3,7-dimethoxylflavone (5) and quercetin (6).The result indicated that compound 6 was the most effective in both DPPH (IC50 value of 10.7μmol/L) and ABTS (IC50 value of 17.5 μmol/L) free radical-scavenging assays.Moreover, both compound 4 and 5 exhibited significant growth inhibitory effect against RKO cells with ICs0 value of 39.08 μmol/L and 17.68 μmol/L, respectively (p<0.01).Conclusion: BP is a noteworthy material containing a series of bioactive flavonoid compounds, thus suggesting that it can be utilized as a potential health supplement and an easily available resource of natural antioxidants, as well as an effective material in pharmaceutical applications.