BACKGROUND: Ulcerative colitis (UC) is an inflammatory bowel disease marked by epithelial barrier dysfunction and hyperactivation of immune signaling, particularly in the NF-kappaB pathway. Conventional therapies are limited by side effects and relapse risk, highlighting the need for safer, plant-based interventions. Oldenlandia umbellata L., a traditional herb, has demonstrated anti-inflammatory potential but remains underexplored in gastrointestinal disorders. OBJECTIVE: This study investigates the therapeutic efficacy of a methanolic extract of O.umbellata L. (MEOU) in a dextran sulfate sodium (DSS)-induced mouse model of colitis, focusing on its effects on clinical symptoms, histopathology, intestinal permeability, cytokine expression, and NF-kappaB signaling. METHODS: Acute colitis was induced in BALB/c mice with 5% DSS for 7 days, followed by oral treatment with MEOU (100, 200 or 400 mg/kg) and prednisolone (5 mg/kg) for another 7 days. Clinical parameters (body weight, Disease Activity Index (DAI), colon and spleen morphology, and histopathology were evaluated. Intestinal permeability was measured by FITC-dextran assay, while TNF-alpha, IL-6 mRNA levels, and NF-kappaB p65 protein levels were analyzed by RT-qPCR and Western blot, respectively. RESULTS: MEOU significantly attenuated weight loss, reduced DAI scores, and reversed colon shortening and splenomegaly in a dose-dependent manner. Histological analysis revealed preserved mucosal structure and reduced inflammatory infiltration. FITC-dextran assays confirmed improved barrier integrity. Molecular analyses showed that MEOU downregulated pro-inflammatory cytokines and suppressed NF-kappaB p65 activation. CONCLUSION: MEOU exhibits potent anti-colitic activity through anti-inflammatory and barrier-protective mechanisms by NF-kappaB pathway inhibition, supporting its potential as a plant-based therapeutic for UC.