Based on classical drug design theory, a novel series of gentiopicroside derivatives was designed and synthesized. All synthesized compounds were then biologically evaluated for their inhibition of influenza virus and anti-HCV activity in vitro. Some of the gentiopicroside derivatives, such as 11a, 13d and 16 showed interesting anti-influenza virus activity with IC50 at 39.5 μM, 45.2 μM and 44.0 μM, respectively. However, no significant anti-HCV activity was found for all of gentiopicroside derivatives. The preliminary results indicate that modification of the sugar moiety on gentiopicroside was helpful for enhancing the anti-influenza activities. Our works demonstrate the importance of secoiridoid natural products as new leads in the development of potential antiviral inhibitors.