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Recurrent aphthous stomatitis leads to ulcers that cause severe pain, which is a substantial burden on patients. Equisetum arvense extract (EA) is a crude drug that promotes wound healing of mucous membranes caused by perineal incision during childbirth and alleviates pain. Here, we elucidated the effects of EA on wound healing and pain in a stomatitis hamster model. After stomatitis induction, two different EA doses were continuously applied to the wound area through the intramucosal injection of acetic acid into the cheek pouch (stomatitis/100*EA group and stomatitis/EA group). The body weight and wound area were measured over time, and histological evaluation was performed after stomatitis induction. The wound area was harvested 10 h after stomatitis induction, and gene expression associated with pain and inflammation was analyzed using qPCR. The dorsal root ganglia of the rat spinal cord were isolated, dispersed, and cultured to examine the inhibitory effect of EA on the K+-evoked release of neurotransmitter substance P. In the stomatitis/100*EA group, a significant reduction in wound size was observed compared with the stomatitis/physiological saline (PS) group, and the weight gain rate was considerably higher. The stomatitis/EA group revealed similar histological changes in the wound and wound size as the stomatitis/PS group; however, the weight gain rate was considerably higher on day 7. The stomatitis/EA group suppressed the expression of inflammatory cytokine mRNA, such as Tnf-α and Il-6, and Cox-2 mRNA in the wound area compared with the stomatitis/PS group. EA treatment reduced the upregulated substance P release from the dorsal root ganglia following high-concentration K+ stimulation. EA alleviates pain in a stomatitis model by suppressing inflammatory cytokine expression in the wound area and substance P release from primary sensory neurons. Therefore, using oral care products containing EA is expected to suppress stomatitis pain., Competing Interests: F.S. and M.M. belonged to a collaborative research laboratory of Earth Corporation (https://corp.earth.jp/en/index.html) and Hiroshima University, which used joint research funds to conduct this study. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2024 Shiba et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)