Phenolic-enriched ethanol-soluble fraction of Codonopsis pilosula attenuates LPS-induced acute lung injury in mice via the gut-lung axis

Background Codonopsis pilosula (Dangshen), a traditional Chinese medicine targeting the Lung Meridian, is commonly steeped in 50% (alcohol by volume) distilled spirits (Baijiu) in China as a medicinal liquor for protecting against lung diseases, including acute lung injury (ALI). This alcohol-steeping preparation yields an ethanol-soluble fraction with a distinct phytochemical profile from conventional aqueous extracts. However, its bioactive compounds and the underlying protective mechanisms against ALI remain unclear. Objective This study aimed to characterize the bioactive compounds of the ethanol-soluble fraction of C. pilosula (designated as CPPP) and investigate its protective mechanisms against lipopolysaccharide (LPS)-induced ALI in mice. Methods CPPP was characterized by non-targeted metabolomic profiling and targeted quantitative analysis using UPLC-ESI-MS/MS, performed sequentially to achieve comprehensive metabolite coverage and precise quantification of identified constituents. Male C57BL/6J mice were pretreated with CPPP for 30 days prior to intraperitoneal injection of LPS to induce ALI. Comprehensive assessments included 16S rRNA sequencing of the gut microbiota, untargeted metabolomic profiling of feces and serum, enzyme-linked immunosorbent assay (ELISA) quantification of serum inflammatory factors and LPS, histological evaluation and immunohistochemistry (IHC) analysis of gut and lung tissues. Results CPPP was enriched in phenolic acids and derivatives (25.3%, w/w). Compared to the normal control, CPPP significantly (P < 0.05) enriched beneficial gut bacteria Clostridium, Bifidobacterium and Akkermansia . CPPP increased fecal levels of butyric acid, serotonin and spermidine, elevated serum level of serotonin, and reduced serum levels of trimethylamine N-oxide, phenylacetylglycine, and phenylacetylglutamine. CPPP significantly (P < 0.001) decreased the levels of LPS-induced pro-inflammatory factors in serum (CRP, TNF-α, IL-6) and lung tissue (MPO, TNF-α, IL-1β, IL-6, IL-18), while increasing the level of anti-inflammatory IL-10 in serum (P < 0.001) and lung tissue (P < 0.01). These anti-inflammatory effects of CPPP, together with the microbiota-derived metabolic alterations, correlated strongly with the CPPP-mediated preservation of intestinal barrier parameters (villus height, goblet cell density, and tight junction protein expression) and attenuation of lung injury parameters (pulmonary inflammation, edema, and histopathological damage). Conclusion CPPP attenuated LPS-induced ALI in mice by remodeling gut microbiota, reprogramming key metabolites and suppressing pro-inflammatory factors along the gut-lung axis. These findings validate the lung-protective potential of phenolic compounds from traditional alcohol-steeping preparation of C. pilosula and provide mechanistic insights for developing microbiota-targeted interventions in inflammatory lung diseases.