The leaves of Hippophae rhamnoides were extracted with methanol and then further fractioned with hexane, ethyl acetate, butanol and water in order to investigate biological activity. Two flavonol glycosides were isolated and identified from ethyl acetate soluble fraction, which showed the strongest antioxidant activity (RC50 = 4.33 μg/ml). Isolated two compounds have shown strong free radical scavenging activity. Especially, quercetin 3-O-glucoside (RC50 = 2.60 μg/ml) was more active than α-tocopherol (RC50 = 4.67 μg/ml). Total phenol and flavonoid contents were the highest to 4.17 and 1.14 mg/ml in the ethyl acetate soluble fraction. In addition, ethyl acetate soluble fraction was shown to the strongest α-glucosidase inhibitory activity (IC50 = 137.88 μg/ml) among the fractions. The results suggest that leaves of H. rhamnoides could be a potential source of natural antioxidant.