The protein extracted from fermented Zanthoxylum bungeanum seeds mitigates DSS-induced ulcerative colitis by orchestrating inflammatory cytokine modulation, counteracting oxidative stress, and safeguarding gut microbiota homeostasis.

Zanthoxylum bungeanum seeds (ZBS) are protein-rich by-products with limited high-value utilization, and fermentation may provide an effective strategy to improve the functional properties and bioactivity of their proteins. In this study, fermented ZBS protein (FZP) was prepared, and its physicochemical properties, antioxidant activity, gastrointestinal digestibility, and protective effects against colitis were systematically investigated. FZP exhibited a high purity of 94.37% and was predominantly composed of proteins with molecular weights ranging from 15 to 25 kDa. Ultrafiltration revealed that the FZP > 10 kDa fraction possessed superior antioxidant activity. In vitro gastrointestinal digestion tests showed progressive protein degradation, with a final degree of hydrolysis of 62%, indicating the high digestibility of FZP. Arginine, aspartic acid, and glutamic acid were identified as the most abundant amino acids in FZP. In a dextran sulfate sodium (DSS)-induced colitis mouse model, FZP markedly alleviated colitis symptoms by mitigating body weight loss, decreasing disease activity index scores, and preventing colon shortening. Moreover, FZP effectively ameliorated colonic histopathological injury and attenuated goblet cell depletion. Mechanistically, FZP reduced the levels of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α), oxidative stress-related markers (MDA and MPO), and serum LPS, which is associated with endotoxin exposure and intestinal barrier damage, while elevating anti-inflammatory IL-10 and antioxidant enzymes (SOD and CAT). Furthermore, FZP restored the expression of intestinal tight junction proteins (ZO-1, occludin, claudin-1) and ameliorated gut microbiota dysbiosis by increasing microbial diversity and enriching beneficial intestinal bacteria. Collectively, FZP alleviates colitis via the coordinated regulation of inflammatory responses, oxidative stress, intestinal barrier function, and gut microbiota homeostasis. These findings suggest that FZP is a promising bioactive fermented protein ingredient and provides a feasible approach for the high-value utilization of ZBS by-products.