Abstract: In this study, the healthy human fecal microbiota was used as the inoculum to establish an in vitro fecal fermentation system with inulin (INU) as the sole carbon source. Metagenomic sequencing and untargeted metabolomics were integrated to systematically evaluate the regulatory effects of inulin INU on the gut microbial ecosystem of healthy individuals from multiple perspectives, including microbial composition, microbial interaction networks, and metabolic responses. The results showed that the pH of the fermentation system with INU significantly decreased after 24 h, accompanied by pronounced alterations in microbial community structure across multiple taxonomic levels. β-Diversity analysis revealed a clear separation between the INU and control groups in the Bray-Curtis distance space, whereas α-diversity analysis indicated a reduction in richness index with no significant change in Pielou’s evenness index. Analysis of key responsive species showed that the abundances of Bifidobacterium pseudocatenulatum, Ligilactobacillus salivarius, Limosilactobacillus reuteri, and Bacteroides uniformis increased following INU intervention, whereas those of Prevotella copri and Faecalibacterium prausnitzii decreased. Microbe-microbe co-occurrence network analysis further indicated that the microbial interaction network formed three major functional clusters after INU intervention, with distinct association patterns among clusters. Untargeted metabolomics revealed that INU markedly reshaped the metabolic profile of the microbial community, with 169 metabolites significantly upregulated and 120 metabolites significantly downregulated. Among these, tryptophan metabolism was the most significantly enriched pathway. The results of correlation analyses between functional pathways and metabolites, as well as between functional genes and metabolites, demonstrated that carbohydrate metabolism-related pathways were significantly correlated with multiple organic acids and amino acid metabolites, while tryptophan metabolism-related genes were significantly correlated with various indole metabolites. Collectively, inulin may influence the metabolic profile of the gut microbiota in healthy individuals by altering microbial composition, restructuring microbial interaction networks, and modulating key metabolic pathways. This study reveals the regulatory characteristics of inulin on the healthy gut microbial ecosystem from a multi-omics perspective, providing experimental evidence for elucidating its microbiota-mediated mechanisms and for evaluating its potential applications in precision nutrition.

Key words: inulin; in vitro fecal fermentation; species level; gut microbiota; microbial interaction networks