The experiment was conducted to study whether the garlic volatile oil (GO) as a safe and high quality feed additives could be widely used in livestock and poultry production. The antibacterial effects of two kinds of refined GO preparations（GO-Ⅰ and GO-Ⅱ）, marketing natural GO and synthetic GO were tested in vitro with double dilution method on four pathogenic bacteria, i.e. Avian Pathogenic Escherichia coli, Avian Salmonella, Swine E.coli and Staphylococcus aureus. The results showed that the antibacterial effects of GO-Ⅰ and GO-Ⅱ were both better than marketing natural GO and synthetic GO. The minimum inhibitory concentration (MIC) of GO-Ⅱ, which contained more than 50% DADS in proportion, were 62.5 μg/mL on Swine Escherichia coli and Avian Escherichia coli and 31.3 μg/mL on Avian Salmonella and Staphylococcus aureus; and MIC of GO-Ⅰ was the same as GO-Ⅱ on Avian Salmonella and Staphylococcus aureus. This study demonstrated that the different proportion of DADS and DATS in GO preparations had different antibacterial effects.