Abstract: In this study, extract of male Eucommia flowers was obtained by ethyl acetate.Six different polar extracts were further isolated from ethyl acetate extract of Eucommia by silica gel column chromatography.This study was conducted to investigate the antibacterial activities of these seven extract by using punch method or dilution method.Minimal inhibitory concentration ( MIC) of extracts was also determined. Results showed that ethyl acetate extract had good antibacterial activities against Staphylococcus aureus, Bacillus anthracis, Bacillus subtilis, Escherichia coli and Aspergillusniger. The MIC was 0.0156, 0.0156, 0.0313, 0.0313, 0.1250 g / m L, respectively. No antibacterial effect was observed on Rhizopus or Pseudomonas sp. The six compounds were named sample obtained by 10% methanol, sample obtained by 20% methanol, sample obtained by 30% methanol, sample obtained by pure methanol, sample obtained by pure methanol with a few ammonia, and sample obtained by water according to their polarities isolated by silica gel column chromatography.The activities of sample obtained by 10% methanol and sample obtained by 20% methanol were higher than that of ethyl acetate extract.While, the sample obtained by 30% methanol and sample obtained by pure methanol did not show antibacterial activities on most of strains.Strikingly, the sample obtained by water exhibited antibacterial activities on Pseudomonas sp only.Thus, the ethyl acetate extract and six compounds isolated by silica gel column chromatography had good antimicrobial activities.Silica gel column chromatography provided good separation of active extract.This research was profitable for the study and use of the male Eucommia flowers.