Abstract: Objective:This study was to evaluate the hepatoprotective effect of polysaccharides from Pleurotus ostreatus (POP) on acute liver injury rats.Methods:An animal model of acute liver injury in rats was induced by carbon tetrachloride.The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) in blood were determined by biochemical analyzer.The effects of Pleurotus ostreatus polysaccharides on urinary metabolic markers and metabolic pathways in rats with liver injury were analyzed by gas chromatography-mass spectrometry.Results:The levels of ALT, AST and ALP were markedly decreased in POP group compared with model group (p < 0.05).Meanwhile, after treatment with POP, the levels of citric acid, malic acid, D-glucuronic acid and 1, 2, 3-propanetricarboxylic acid decreased (p < 0.05) and hexadecanoic acid level increased (p < 0.05). These biomarkers belonged to TCA cycle, glyoxylate and dicarboxylate metabolism, ascorbate and aldarate metabolism. Conclusion:POP could ameliorate the abnormal changes of liver function and regulate the disorder of biomarkers and pathways on acute liver injury rats.Conclusion:POP had a hepatoprotective effect on acute liver injury and had certain application value to be developed as liver protection functional foods.