Pomegranate seed oil attenuates palmitic acid‐induced hepatic injury through modulation of oxidative stress: insights from GC ‐Q‐ TOF ‐ MS ‐based metabolomics

BACKGROUND Non‐alcoholic fatty liver disease (NAFLD) is a leading cause of chronic liver disease worldwide, with oxidative stress and lipid dysregulation as central pathogenic drivers. Pomegranate seed oil (PSO), rich in the conjugated fatty acid punicic acid, has demonstrated potential antioxidant and lipid‐modulating properties, yet its hepatoprotective mechanisms remain incompletely characterized at the metabolic level. RESULTS Gas chromatographic–mass spectrometric analysis of the fatty acid composition of supercritical CO 2 ‐extracted PSO revealed that punicic acid was the predominant fatty acid, comprising 74.47% of the total identified fatty acids, which collectively accounted for approximately 95% of the fatty acid profile. In palmitic acid (PA)‐challenged C57BL/6 mice, PA administration significantly increased serum and hepatic lipid levels, liver function markers, oxidative stress indicators, and inflammatory cytokines. PSO intervention dose‐dependently ameliorated several of these abnormalities and markedly reduced hepatic lipid accumulation. Gas chromatography–quadrupole time‐of‐flight mass spectrometry‐based serum metabolomics revealed that PSO markedly reversed PA‐induced metabolic disturbances, identifying 27 key differential metabolites predominantly associated with amino acid and lipid metabolism. Pathway enrichment analysis highlighted oxidative stress‐related pathways, including glutamate, taurine, and fatty acid metabolism. PSO significantly upregulated antioxidant metabolites (cysteine, glutamate, hypotaurine, α‐tocopherol) while downregulating oxidative markers (uric acid, xanthine). Complementary in vitro experiments in PA‐treated L02 human hepatocytes further demonstrated that PSO alleviated lipotoxicity by attenuating reactive oxygen species generation and preserving membrane integrity. CONCLUSION These findings demonstrate that PSO exerts hepatoprotective effects through multi‐target, multi‐pathway synergistic mechanisms, particularly via enhancing endogenous antioxidant defense systems, providing scientific evidence for PSO as a promising nutritional intervention strategy for NAFLD prevention and treatment.