Four total flavonoids extracts were obtained from fresh stem and leaf of Gynura bicolor DC. by method of ethanol extraction-macroporous resin column chromatography-polyamide column  chromatography-recrystallization, and their compositions were analyzed by LC-MS technology. The results show that there are six groups of flavonoid absorption peak in total flavonoids extracts of G. bicolor, which is identified to be eight components including rutin, quercetin-3-O- rubinobioside, hyperoside, isoquercitrin, kaempferol-3-O-rubinobioside, kaempferol-3-O-rutinoside, kaempferol-3-O-galactoside and astragalin. This extraction method can effectively extract and purify total flavonoids from fresh stem and leaf of G. bicolor, so it can be used for factory production.