Human infection by enterovirus type 71 (EV71) can cause life-threatening meningo-encephalitis. Currently, there is no effective anti-EV71 therapy available. Since EV71 infection commonly involves skin lesions, we tested our hypothesis that water extract of Glycyrrhiza uralensis (G. uralensis) could inhibit the cytopathic effects of EV71 in a human foreskin cell line by using an XTT-based method. Our results showed that the water extract of G. uralensis at 3,000 microg/ml has only 30% cytotoxicity on host cells, and furthermore, that the water extract of G. uralensis at 0.1 microg/ml could effectively protect host cells against EV71 infection (p < 0.0001). The half maximal inhibitory concentration (IC(50)) was 0.056 microg/ml with a selective index greater than 50,000. The water extract of G. uralensis exerted its effects not only by preventing viral attachment (p < 0.0001), but also by inhibiting the penetration of the virus (p < 0.0001). EV71 infection caused cells to produce significant amounts of IFN-beta (p = 0.0003). However, the anti-EV71 activity of the water extract of G. uralensis was not mediated by IFN. In conclusion, the water extract of G. uralensis possesses potent anti-EV71 effects with less cytotoxicity. Its low IC(50) and high 50% cytotoxic concentration (CC(50)) values suggest that it is a promising anti-EV71 agent.