OBJECTIVE： To investigate the effect of aqueous extraction of Lonicerae Flos （SYHW） on anti-influenza A virus H1N1 （H1N1 virus） in vitro. METHODS： Using Madin-Darby canine kid ney （MDCK） cells cultured in vitro by H1N1 virus， half of the tissue culture infection dose （TCID50） was calculated. Culturing MDCK cells for 24 h with different mass concentrations of SYHW， the maximum non-toxic concentration was investigated. And then test was divided into normal cell group， virus control group， SYHW preventive administration group， therapeutic administration group and direct killing group （given SYHW of maximum non-toxic concentration， infecting cells by 100 TCID50 H1N1 virus）， and antiviral effective rate （ER） of SYHW was determined. Test was divided into normal cell group， virus control group， SYHW therapeutic group and direct killing group （the same administration and infection as above）， changes of cell proliferation index （PI） and cell apoptosis rate were respectively determined. RESULTS： 100 TCID50 of H1N1 virus was 1.26×10－7， and the maximum non-toxic concentration of SYHW on MDCK cells was 50 μg/mL （cell survival rate was 91.3％）. ERs of preventive administration group， therapeutic administration group and direct killing group were 0， 80.3％ and 52.7％， respectively. Compared with normal cell group， PI value in virus control group was significantly reduced （P＜0.05）， early and late apoptotic rates were significantly increased （P＜0.05）. Compared with virus control group， PI value in directly killing group was significantly increased （P＜0.05）， and early apoptotic rate was significantly reduced （P＜0.05）； early apoptotic rates in therapeutic administration group were significantly reduced （P＜0.05）. CONCLUSIONS： SYHW shows anti-H1N1 virus effect in vitro， therapeutic administration and directly killing are preferred in antiviral effect.