Xiuping Wan, Juyan Sun, Qifan Zhang, Meiwen Sun, Lisha Wang, Guangjing Chen
(NSP), 1 M KOH (HF1), 4 M KOH (HF4), and a cellulose-rich residue (CF), and were comprehensively characterized for their composition, architecture, and prebiotic potential. Pectin fractions (WSP, CSP, NSP) were found to be enriched in galacturonic acid and to exhibit domain-specific features: WSP and CSP were identified as high-methoxy, homogalacturonan (HG)-dominant pectins, whereas NSP was rich in rhamnogalacturonan-I (RG-I) and showed greater aggregation. Hemicellulose fractions (HF1, HF4) were observed to possess the highest molecular weights and thermal stability, while partial crystallinity was retained in CF. In vitro fecal fermentations demonstrated rapid, structure-dependent carbohydrate utilization and acidification, accompanied by sustained increases in total short-chain fatty acids (SCFAs), including acetate, propionate, and butyrate. All CWP fractions significantly modulated the gut microbiota compared to blank control, generally enriching Firmicutes while reducing Proteobacteria, increasing beneficial genera such as Faecalibacterium and Bifidobacterium, and suppressing Escherichia-Shigella. Pectin fractions-particularly HG-rich WSP and CSP-promoted early acidogenesis and robust SCFA production, while RG-I-rich NSP selectively enriched known pectin-degraders. Hemicelluloses were highly fermentable but exhibited weaker pathogen modulation. Spearman correlation analyses indicated that higher levels of rhamnose, arabinose, and galacturonic acid, together with lower branching, were linked to elevated acetate and propionate production. Collectively, a unified workflow was established by integrating sequential extraction, structural profiling, and human fecal fermentation assays, thereby enabling specific polysaccharide domains to be directly linked to gut microbial decomposition and metabolic outcomes, with greater translational relevance than conventional single-assay in vitro approaches.