The aim of this study was to investigate the acidic polysaccharide GiP-B1 from Glycyrrhiza inflata on activation of RAW 264.7 through TLR4/MyD88/NF-κB signaling pathway.RAW 264.7 were transfected with TLR4 interfering RNA (TLR4 siRNA) for 24 h in vitro,and then treated with GiP-B1 (100 μg/mL) for 12 h.The effects of proliferation and secretion of inflammatory factors in RAW 264.7 were detected by CCK-8 and ELISA kits,respectively.RT-PCR and Western blot method was used to detect the effects of GiP-B1 on the mRNA and the protein expression levels of TLR4,MyD88,NF-κB p65 and IκBα.The results showed that compared with the blank control group,GiP-B1 could significantly increase the proliferation of RAW 264.7 and the secretion of interleukin 1 beta (IL-1β),interleukin 6 (IL-6),and tumor necrosis factor alpha (TNF-α),and significantly up-regulate the mRNA and protein expressions of TLR4,MyD88,NF-κB p65 and IκBα (P＜0.05).After TLR4 gene silencing was performed for each group,compared with the normal cultured GiP-B1 intervention group,the transfection group could inhibited the proliferation and cytokine secretion of RAW 264.7 induced by GiP-B1,which also inhibited the mRNA and protein expression levels of TLR4,MyD88,NF-κB p65 and IIκBα (P＜0.05).In summary,GiP-B1 activates TLR4/MyD88/NF-κB signaling pathway to promote the expression of related genes,and finally promotes NF-κB into the nuclear to regulate the transcription level of cytokines,and then regulates the macrophage immune function.