Mucus hypersecretion is an important feature of airway inflammatory diseases. In this study, we investigated the effect of Mucirin, a standardized bioactive fraction of Acorus calamus rhizome, on MUC5AC mucin gene expression induced by elastase in lung epithelial cells as well as its mechanism of action. Human lung epithelial NCI-H292 cells were pretreated with elastase to induce MUC5AC expression and treated with Mucirin overnight. We found that 10 μg/mL of Mucirin repressed the expression of MUC5AC at both mRNA and protein levels up to 59 and 60 %, respectively. Mucirin decreased markedly protein expression of molecules involved in MUC5AC transcription such as receptor-ligand EGFR (350%) and TGFα (46%). Moreover, our western blot analysis also demonstrated that Mucirin inhibited TGFα activation, while TNFα and NFκB expressions were significantly decreased as well. These results suggest that Mucirin inhibited elastase-induced MUC5AC expression in human lung epithelial cells through TNFα-EGFR-TGFα-NFκB pathway. The ability of Mucirin on down-regulation proinflammatory cytokine is likely to improve the molecular evidences of A. calamus as a potential drug candidate to treat diseases related to mucus hypersecretion, including cough, bronchitis and COPD.