The extensive use of antibiotics has caused the global spread of multidrug-resistant bacteria and genes, seriously reducing antibiotic efficacy and threatening animal and human health. As an alternative, traditional Chinese veterinary medicine (TCVM) was used in this study for its lack of drug resistance and low toxicity. Huangqin-honghua-pugongying-jinyinhua extract (HHPJE), a novel TCVM, consists of the extracts of Huangqin (), Honghua (), Pugongying () and Jinyinhua (), and was developed to treat bovine mastitis. In this study, we evaluated the toxicity, bacteriostatic, analgesic, anti-inflammatory, and antipyretic activities of HHPJE. Our results show that HHPJE did not show any acute oral toxicity and can be considered safe for oral administration. Additionally, HHPJE possessed a dose-dependent antibacterial effect on , , and . HHPJE (60, 30 and 15 g/kg) can reduce the abdominal pain by 44.83 ± 7.69%, 43.15 ± 9.50% and 26.14 ± 4.17%, respectively. The percentages of anti-inflammation inhibition (60, 30 and 15 g/kg) were 35.34 ± 2.17%, 22.29 ± 2.74% and 12.06 ± 3.61%, respectively. The inhibition rates (60, 30 and 15 g/kg) of antipyretic activity were 82.05%, 65.71% and 52.80%, respectively. The evaluation of pharmacodynamics and toxicity indicate that HHPJE possesses significant bacteriostatic, analgesic, anti-inflammatory and antipyretic potential, and also that it is safe for acute oral toxicity, which means it has potential value for treating bovine mastitis in future and alleviating clinical symptoms with no drug resistance or side effects.