ETHNOPHARMACOLOGICAL RELEVANCE: Acute lung injury causes high mortality in clinical settings. Biyuan Tongqiao Granules (BYTQ), serves as a modified prescription which is extensively applied in clinical practice for the remedy of related respiratory diseases, has been proved for curing a series of inflammatory-induced diseases in clinical setting. However, the underlying mechanism has not been reported and understood. AIM OF STUDY: This study is in an effort to elucidate the therapeutic effects of BYTQ against ALI and its related molecular mechanisms in lipopolysaccharides (LPS) - induced ALI mouse model. MATERIALS AND METHODS: The mice were treated BYTQ by means of intragastric administration for 7 continuous days, then the LPS was used to induce ALI model. The lung and intestinal injury were detected by H&E staining, lung index, inflammation, levels of tight junction proteins and other means to evaluate the changes of gut and lung tissue, respectively. Microbiota flora was analyzed by 16S rRNA and the alteration of metabolites were analyzed by metabolomics in both serum and lung tissue. The mechanisms of BYTQ were demonstrated by the Astral Mass Analyzer for quantitative proteomics and western blotting. RESULTS: BYTQ alleviated ALI symptoms in both lung and intestinal tissues, as validated by pathological scores, lung index, inflammatory factors containing TNF-α, IFN-γ, IL-1β and IL-6, combined with oxidative factors containing SOD, CAT, MDA, GSH and MPO, as well as the goblet cell, levels of LPS, D-LA, DAO and SIgA, etc. Also, BYTQ reversed the dysregulated microbiota flora to normal trends by 16S rRNA. Unsaturated fatty acid pathway was distinguished be means of untargeted metabolomic analysis. Proteomics and Western blot were integrated and the results confirmed that the molecular targets of BYTQ for treating ALI were centered on PI3K/AKT/mTOR pathway. CONCLUSION: BYTQ ameliorated lung and intestinal injury induced by LPS in ALI mice. The mechanisms were likely to be associated with mitigating inflammatory response, protecting intestinal barrier function and imbalanced intestinal flora, regulating unsaturated fatty acid pathway through PI3K/AKT/mTOR pathway, which are likely to be correlated with the modulation via gut-lung axis. This investigation highlights the anti-ALI effects of BYTQ and provides a solid foundation for its further application in ALI treatment.