Severe inflammatory damage and permeability changes in colon are the main features of inflammatory bowel disease (IBD) which is a serious digestive system disease. SiNi San (SNS) originated from Treatise on Febrile Diseases, is usually used to treat gastrointestinal diseases. In this study, we explored the protective effect of SNS on IBD. IBD rats were induced by trinitrobenzene sulfonic acid. 125, 250 and 500 mg/kg of oral SNS significantly alleviated weight loss and diarrhea, reduced colitis inflammatory cell infiltration and inhibited pro-inflammatory cytokine production. SNS also revealed anti-inflammatory effects on lipopolysaccharide (LPS)-induced RAW cells. The mechanism may be related to the inhibition of tumor necrosis factor- α (TNF-α)-induced nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway to suppress cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase(iNOS) expressions. Also colons were collected for immunofluorescence assay and LPS-induced cell-co-culture system of Caco-2 and RAW264.7 cells were used to determine protective effect on intestinal epithelial monolayers damage. Our results showed SNS enhanced expression of claudin-2 and zona occludens-1 (ZO-1), elevated trans epithelial electric resistance, decreased the permeability in LPS-damaged epithelial monolayers system. SNS showed regulatory effect on IBD by anti-inflammatory and protecting intestinal mucosal barrier. It is suggested SNS may be new supplements against IBD.