An integrated microbiome-drug interaction and bioinformatics approach identifies active deglycosylated and glycosidic chain metabolites of Platycodin D targeting PTPN2/HIF1A in acute lung injury

Platycodin D (PD) is a typical triterpenoid saponin with low oral bioavailability. Despite its potent anti-ALI effects, its active metabolites and underlying mechanisms remain unclear, thereby significantly impeding its further development. This study aimed to establish a new integrated method combining microbiome-drug interaction analysis, bioinformatics approaches, and experimental validation to elucidate the active metabolites and mechanisms by which PD exerts anti-ALI effects. Microbiome-drug interaction analysis identified PD-derived metabolites, including deglycosylated metabolites (DGMs), glycosidic chain metabolites (GCMs), and short-chain fatty acids (SCFAs) derived from the glycosidic chains, while simultaneously assessing PD’s modulatory effects on the intestinal microbiota. Bioinformatics approaches predicted the potential anti-ALI targets of the metabolites in the blood. The bioactivities and mechanisms of these metabolites were subsequently validated using LPS-induced and pseudo-sterile ALI mouse models and molecular docking. PD and its 9 DGMs were identified in vitro, while only Deapio-platycodin D (DPD), 3-O-β-D-glucopyranosyl platycodigenin (GPN) and platycodigenin (PN) were detected in serum; 9 GCMs including 1 trisaccharide, 2 disaccharides and 5 monosaccharides derived from the glycosidic chain of PD were identified in vitro. Only 5 monosaccharides of Glucose (Glu), Arabinose (Ara), Rhamnose (Rha), Xylose (Xyl) and Apiose (Api) were detected in serum. Notably, saccharide-to-SCFA transformation was markedly inhibited both in vitro and in serum. PD also modulated intestinal microbiota by increasing probiotics and reducing pathogens. Bioinformatics analysis showed DGMs targeted PTPN2, whereas GCMs co-targeted both PTPN2 and HIF1A. In vivo models confirmed the activities and mechanisms of PD, while molecular docking further verified the active metabolites were 2 DGMs of GPN, PN and 5 GCMs of Glu, Ara, Rha, Xyl and Api. Novel active metabolites and mechanisms of PD against ALI were elucidated and validated by the proposed strategy. Bioactivity of PD extends beyond its metabolites with parent nucleus, as GCMs of PD also showed significant activities. SCFAs are not necessarily active metabolites of PD despite its saccharide-rich structures. In addition, this study establishes a new paradigm for elucidating active metabolites and mechanisms of glycosides, especially those with low oral bioavailability, advancing natural product-based ALI treatment strategies.