Acute lung injury (ALI) is a common life-threatening lung disease, which is mostly associated with severe inflammatory responses and oxidative stress. Tanreqing injection (TRQ), a Chinese patent medicine, is clinically used for respiratory-related diseases. However, the effects and action mechanism of TRQ on ALI are still unclear. Recently, STING as a cytoplasmic DNA sensor has been found to be related to the progress of ALI. Here, we showed that TRQ significantly inhibited LPS-induced lung histological change, lung edema, and inflammatory cell infiltration. Moreover, TRQ markedly reduced inflammatory mediators release (TNF-α, IL-6, IL-1β, and IFN-β). Furthermore, TRQ also alleviated oxidative stress, manifested by increased SOD and GSH activities and decreased 4-HNE, MDA, LDH, and ROS activities. In addition, we further found that TRQ significantly prevented cGAS, STING, P-TBK, P-P65, P-IRF3, and P-IκBα expression in ALI mice. And we also confirmed that TRQ could inhibit mtDNA release and suppress signaling pathway mediated by STING . Importantly, the addition of STING agonist DMXAA dramatically abolished the protective effects of TRQ. Taken together, this study indicated that TRQ alleviated LPS-induced ALI and inhibited inflammatory responses and oxidative stress through STING signaling pathway.