► TFPL possesses considerable amounts of flavonoids. ► TFPL possess potent antioxidant and free radical scavenging activities. ► Protect effect of TFPL on cell viability and antioxidative status in MC3T3-E1 cells. Persimmon leaves are commonly consumed as beverages, but are also used as a popular folk medicine in China. The purpose of this work is to assess the antioxidant activity of an extract of total flavonoids from persimmon leaves (TFPL). The effect of TFPL on total antioxidant activity, reducing power, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, superoxide anion (O2-) radical scavenging, hydroxyl (OH) radical scavenging and metal chelating activities was examined. We found that TFPL possesses considerable amounts of flavonoids (192μg catechin equivalent/g of extract). The effect of this extract in total antioxidant activity, scavenging activity of superoxide anion and hydroxyl radical, reducing power and iron chelating activity was significantly better than that of rutin. However, the effect of TFPL in free radical scavenging of DPPH was significantly not as good as than rutin. In addition, TFPL significantly decreased the level of reactive oxygen species (ROS) and malondialdehyde (MDA), while increasing the activity of catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in MC3T3-E1 cells in a dose-dependent manner. In conclusion, TFPL possess potent antioxidant and free radical scavenging activities. These antioxidant activities could contribute, at least in part, to the traditionally claimed therapeutic benefits of persimmon leaves.