Abstract: The present study assessed the acute toxicity of intramammary infusion of rifaximin on mice by using standard method.The results revealed that the mice received the highest dosage of 5 000 mg/（kg·BW）of rifaximin had no adverse reactions and their dietary behavior was normal. No deaths occurred during the observation period in the highest dosage group. Therefore, it is concluded that the LD50 of intramammary infusion of rifaximin by intragastric administration was higher than 5 000 mg/（kg·BW）,indicating that it is an actually non-toxic level substance.