The inhibitory effects of garlic oil self-microemulsion（GO-SMEDDS） on Escherichia coli， Staphylococcus aureus， Bacillus subtilis， Pseudomonas aeruginosa， Candida albicans and Saccharomyces cerevisiae were investigated in this paper. The minimum inhibitory concentration （MIC） and the minimum bactericidal concentration （MBC） of GO-SMEDDS were determined by measuring the diameter of the inhibitory circle. Escherichia coli and Staphylococcus aureus were applied to use as test strains， and then the inhibitory effect of GO-SMEDDS on bacteria was evaluated by the combination of morphology， electrical conductivity and the activity of Alkaline phosphatase（AKP）， Alanine Aminotransferase （ALT） and Aspartic Transaminase（AST）. The results indicated that the potential ability of the bacteriostatic effects of GO-SMEDDS were in high to low order as follows： fungi， gram-positive bacteria and gram-negative bacteria. The MIC of GO-SMEDDS against Escherichia coli and Staphylococcus aureus were 1.875 mg/mL and 0.469 mg/mL， respectively， and the MBC were 3.750 mg/mL and 0.938 mg/mL， respectively. The antibacterial mechanism of GO-SMEDDS on Escherichia coli and Staphylococcus aureus is that the cell walls and membranes are damaged， which causes the leakage of AKP， ALT， AST in the bacteria cells， thus leading to the death of the bacteria.