The invention provides an antibacterial composition and a preparation method thereof, and relates to the technical field of antibiosis. The antibacterial composition is prepared from the following raw materials in parts by weight: 30 to 102 parts of antibacterial substances, 600 to 720 parts of olive oil, 250 to 330 parts of soybean oil, 5 to 15 parts of vitamin E and 40 to 60 parts of ozone. The antibacterial composition has the advantages of being antibacterial, mild in property and harmless to human bodies.