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Background Ulcerative colitis is a refractory inflammatory bowel disease characterized by complex pathology, with limited therapeutic options available. Ferroptosis, an iron-dependent form of regulated cell death, has been implicated in damage to intestinal epithelial cells. Targeting ferroptosis may represent a promising strategy for the treatment of colitis. Arachidonate 15-lipoxygenase (ALOX15) serves as a key regulator of ferroptosis; however, natural compounds that directly inhibit ALOX15 remain underexplored.Methods We initially identified ALOX15 as a central regulator of ferroptosis in colonic epithelial cells through bioinformatic analysis. Subsequently, machine learning was employed to screen natural product libraries, leading to the identification of rosmarinic acid (RA) as a potential ALOX15 inhibitor. This was further validated by molecular docking, which demonstrated stable binding of RA to the ALOX15 active site. Cellular and biochemical assays, including Cellular Thermal Shift Assay (CETSA), Drug Affinity Responsive Target Stability (DARTS), and co-immunoprecipitation (Co-IP), were utilized to confirm the direct interaction between RA and ALOX15. The anti-ferroptotic activity of RA was assessed in vitro by measuring markers such as GPX4, SLC7A11, ACSL4, lipid peroxidation, and intracellular iron levels. The therapeutic efficacy of RA was evaluated in vivo using mouse models of colitis.Results RA directly bound to ALOX15 and effectively inhibited ferroptosis in intestinal epithelial cells, as evidenced by restored expression of GPX4 and SLC7A11, reduced levels of ACSL4, decreased lipid peroxidation, and diminished iron accumulation. Genetic knockout of ALOX15 abolished the protective effects of RA, confirming its target specificity. In mouse colitis models, administration of RA significantly alleviated disease severity, including improved weight recovery, reduced colon shortening, and enhanced intestinal barrier integrity through the upregulation of tight junction proteins. Mechanistically, RA was found to inhibit the ALOX15-VDAC1 axis, leading to restoration of mitochondrial membrane potential and a reduction in mitochondrial reactive oxygen species (ROS) production.Conclusions This multidisciplinary study identifies the natural compound rosmarinic acid as a novel inhibitor of ferroptosis that ameliorates ulcerative colitis by directly targeting the ALOX15-VDAC1 axis ( IDDF2026-ABS-0073 Figure 1. Graphical Abstract). Our findings underscore the therapeutic potential of RA and validate the ALOX15-VDAC1 pathway as a compelling target for the treatment of inflammatory bowel diseases. (Acknowledgements: This study was funded by National Natural Science Foundation of China (82370564))Abstract IDDF2026-ABS-0073 Figure 1