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Background and objective Phelligridin D is a hispidin analogue from the mushroom Phellinus baumii that is widely used as a food source in East Asia. This study tested phelligridin D for the anti-inflammatory effect and mechanism in lipopolysaccharide (LPS)-induced human periodontal ligament cells (HPDLCs). The objective of this study was to clarify whether the anti-inflammatory function of phelligridin D affects periodontal regeneration for supporting the HPDLCs of teeth. Material and methods Primary HPDLCs were isolated from healthy teeth and then cultured. The anti-inflammatory function, mechanism and differentiation molecules were verified with reactive oxygen species generation and western blot analysis in LPS-induced HPDLCs. Results HPDLCs showed increased inflammatory molecules (intracellular adhesion molecule-1 and vascular cell adhesion molecule-1) and decreased osteogenic proteins (bone morphogenetic protein-7, Osterix and runt-related transcription factor 2) by LPS treatment. Phelligridin D decreased inflammatory molecules and increased osteogenic molecules via downregulation of the extracellular signal-regulated kinase and c-jun N-terminal kinases pathway among the mitogen-activated protein kinase, followed by blocking of nuclear factor kappa-B translocation from cytosol to nucleus. In addition, phelligridin D showed antioxidant properties by reducing reactive oxygen species activity. Finally, the anti-inflammatory and antioxidant function of phelligridin D promoted the periodontal differentiation of HPDLCs. Conclusion These results suggest that phelligridin D supports teeth on the alveolar bone against outside stress, and may be used as an anti-inflammatory compound for the prevention of periodontitis or periodontal regenerative related disease.