Bacteria living as biofilm are frequently reported to exhibit the resistances to the common antimicrobial agents. Hence, the in vitro antimicrobial activities of the compound 1,8-cineole were evaluated to explore the novel agent. The bioactivities were estimated by the MIC and MBC determinations on 8 species of bacteria and fungi, as well as the confocal laser scanning on 3 types of bacterial biofilms. It was found that the essential oil has good inhibiting activities against fungi rather than bacteria, especially Candida albicans, with the lowest MIC value (0.156 %). Both of gram-positive and gram-negative bacterial biofilms are sensitive to 1,8-cineole. In addition, there is linear relationship between inhibition activities and concentration of 1,8-cineole. Further analysis revealed that 1,8-cineole can prompt the biofilm-surface bacteria to die off or live as planktonic, in order to help the immune system to remove the bacteria. Thus, 1,8-cineole could be judged as a kind of potential drug with rather antimicrobial activities.