The invention provides a breast perfusion preparation for treating mammitis of milk cow in lactation period, which belongs to the field of medical preparation for veterinary use; every 3g of the perfusion preparation contains 150-250 mg of amoxicillin, 50-125 mg of sulbactam, 5-15 mg of prednisolone and the rest of substrate; the substrate contains 0.1-12% of suspending aid, 0.1-2% of wetting agent, 0.01-1% of antioxidant, 0.05-1% of preservative and the rest of dispersion medium; the invention can be obtained via the following steps: firstly, dissolving or dispersing suspending aid, antioxidant, wetting agent and preservative into proper amount of dispersion medium, then slowly adding the rest of the dispersion medium into a colloid mill to obtain the substrate, adding medicine into the substrate, grinding via the alternating method of circularly grinding and non-circularly grinding. The preparation of the invention is simple in preparation technique, easy to realize industrialization, low in production cost, low in expenditure for treatment, low in toxicity, wide to use and high in efficiency; the invention can be clinically used for treating mammitis in lactation period caused by Gram positive bacterium and Gram negative bacterium.