A new steroidal saponin, named as (25R)-6α-[(β-D-glucopyranosyl) oxy]-5α-spirostan-3β-yl β-D-glucopyranosyl (1''→3')-O-β-D-glucopyranosid (2) was isolated from the flowers of Cestrum elegans. In addition, a known steroidal saponin (1), a flavonoid compound (3) along with a triterpene saponin (4) were isolated for the first time in genus Cestrum and identified based on their chromatographic properties, chemical and spectral data (HR-ESI-MS, 1H, 13C NMR, 1H-1H COSY, HSQC, and HMBC). The four compounds are characterized by different effects against Gram positive, Gram negative bacteria, antifungal and antiviral activities. The maximum non-toxic concentration (MNTC) on vero cell line was 1.56 µg/mL for all compounds. While compound 1 and 3 showed the highest biological activities against hepatitis A virus possess 34.3% and 25% antiviral activity respectively..