Oral mucositis is a common side-effect caused by chemotherapy or radiotherapy occurring in the majority of cancer patients and is characterized by inflammation and ulcers in the oral mucosa. In the present study, we examined the protective effects of Salvia miltiorrhiza Bunge (SM) on oral mucositis induced by 5-fluorouracil (5-FU) in human pharyngeal cells and golden Syrian hamsters. We investigated the proliferation and antioxidant abilities of SM using MTT, 2-diphenyl-1-picrylhydrazyl (DPPH) and reactive oxygen species (ROS) assays in vitro. Additionally, TUNEL assay was performed, and the expression levels of nuclear factor-κB (NF-κB), caspase-3 and proinflammatory cytokines were assessed by immunoblotting. The results showed that SM increased the cell proliferation rate in human pharyngeal cells up to 128.97±9.7% compared with this rate in the untreated cells and exerted protective effects on mucosal injury caused by 5-FU treatment. In addition, all concentrations of SM increased DPPH scavenging ability and blocked ROS generation in the treated cells. Taken together, following SM treatment, expression of NF-κB and cleaved caspase-3 were significantly decreased followed by inhibition of cell death. These data suggest that SM could be used for the prevention and treatment of oral mucositis caused by cancer therapies.