Objective: To discuss the effect of butorphanol on lipopolysaccharide induced chondrocyte injury and analyze whether its mechanism is related to the stromal cell-derived factor-1α (SDF-1α)/C-X-C chemokine receptor 4 (CXCR4) pathway. Methods: Human chondrocytes C28/I2 were cultured in vitro and assigned into control group (normal culture), model group (100 μmol/L lipopolysaccharide), model+low-dose butorphanol group (100 μmol/L lipopolysaccharide+1 μmol/L butorphanol), model+medium-dose butorphanol group (100 μmol/L lipopolysaccharide+2 μmol/L butorphanol), model+high-dose butorphanol group (100 μmol/L lipopolysaccharide+4 μmol/L butorphanol), and model+high-dose butorphanol+NUCC-390 group (100 μmol/L lipopolysaccharide+4 μmol/L butorphanol+500 nmol/L CXCR4 agonist NUCC-390). MTT assay, ELISA assay, flow cytometry, and western blot were used to detect cell viability, the IL-6 and TNF-α levels, the cell apoptosis, apoptosis, and the SDF-1α/CXCR4 pathway related proteins, respectively. Results: For the control group, the model group showed decreased chondrocyte survival rate and Bcl-2 protein, and increased TNF-α, IL-6, apoptosis rate, Bax, Cleaved caspase-3, SDF-1α, and CXCR4 proteins (P<0.05). For the model group, the model+low, medium, and high-dose butorphanol groups showed increased chondrocyte survival rate and Bcl-2 protein, and decreased TNF-α, IL-6, apoptosis rate, Bax, Cleaved caspase-3, SDF-1α, and CXCR4 proteins (P<0.05). For the model+high-dose butorphanol group, the model+high-dose butorphanol+NUCC-390 group showed decreased chondrocyte survival rate and Bcl-2 protein, and increased TNF-α, IL-6, apoptosis rate, Bax, Cleaved caspase-3, SDF-1α, and CXCR4 proteins (P<0.05). Conclusions: Butorphanol may improve chondrocyte injury induced by lipopolysaccharide by inhibiting SDF-1α/CXCR4 signaling pathway.