Abstract: Objective: The antibacterial activity and its mechanism were investigated on acetone extracts from the Rubia Cordifolia to provide a scientific basis for development and utilization of the rubia cordifolia resources.Method: The antibacterial activities were evaluated by using the filter method and resazurin indicator method to measure the inhibition zone,MIC and MBC.The antibacterial mechanisms were studied by drawing growth curve,the change of conductivity and leakage of cellular contents.Result: The results showed that the sensitivities to the extract were different for different tested bacteria.The antibacterial activity of the extract was particularly strongest against B subtilis,and was weakest against B.Cereus.The following test results including growth curve of B subtilis and its variation trend of liquid conductivity and the cell contents showed that antibacterial mechanism may due to inhibit split speed of bacterial exponential phase and result in the increase in permeability of cell membranes and the leakage of intracellular constituents on the basis of the cell constituents' release assay.The data of this study suggested that the extract from Rubia Cordifolia had great potential for application as a natural antimicrobial agent.