Abstract

Background Dunhuang Gancao Fuling Xingren decoction (GFXD) is a traditional formulation derived from the Dunhuang Ancient Medical Prescriptions, has been historically utilized for its immunomodulatory and anti-inflammatory properties. However, the protective effect against irinotecan (CPT-11)-induced intestinal mucositis (CIM) remains poorly elucidated. Purpose To investigate the therapeutic efficacy of GFXD in alleviating CIM and elucidate its underlying mechanism and components using  Drosophila melanogaster  and C57BL/6 J mouse models. Methods The therapeutic efficacy of GFXD was assessed in both Drosophila and mouse models by phenotype assay, hematoxylin and eosin (H&E) staining, and Alcian blue-periodic acid schiff (AB-PAS) staining. Transcriptomic profiling combined with 16S rRNA sequencing were employed to identify potential mechanisms of GFXD regulating CPT-11-induced mucositis. Cytokine levels were measured using ELISA, while the expression levels of key signaling pathways, including Toll-Imd and JAK-STAT pathways were analyzed via qRT-PCR, immunofluorescence, fecal microbiota transplantation (FMT) experiment, and antibiotic treatment. Furthermore, functional components of GFXD were characterized via liquid chromatography-mass spectrometry (LC–MS), and their efficacy was validated in CPT-11-treated  Drosophila . Results GFXD significantly mitigated CPT-11-induced systemic and intestinal damage in Drosophila , evidenced by improved survival rate, restored digestive function, elongated intestinal length, reduced acid–base imbalance, and enhanced epithelial and stem cell proliferation. In mice, GFXD alleviated mucositis symptoms, attenuated histopathological damage, and normalized inflammatory cytokine levels. Mechanistically, GFXD suppressed gut microbiota dysbiosis by enriching probiotics ( Lactobacillus, Prevotella ) and reducing pathogens ( Bacteroides ,  Enterobacter, Enterococcus and Helicobacter ). Transcriptomic and molecular analyses revealed