Abstract: Objective: To explore the effect of ursolic acid (UA) on the gut microbiota of rats with alcoholic liver injury for the purpose of providing new insights into controlling alcoholic liver injury. Methods: A total of 30 healthy male Wistar rats at the age of 2 months were randomly divided into 3 groups with 10 animals in each group including normal control group (saline), alcohol model group and UA-treated group (alcohol + UA) throughout the 8-week experiment. The rats were weighed and anesthetized with 10% chloral hydrate at twelve hours after the last treatment. Then blood samples were collected by abdominal aorta puncture to determine biochemical parameters. Meanwhile, liver tissue and feces were also taken. Pathological changes of liver tissue were evaluated by hematoxylin-eosin (HE) staining. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum were determined by the method of Reitman and Frankel. The endotoxin level in serum was tested by tachypleus amebocyte lysate kit. The content of D-lactic acid (D-LA) in plasma was determined by a modified enzymatic method at an ultraviolet wavelength. The genomic DNA of the intestinal flora was used as the template through enterobacterial repetitive intergenic consensus (ERIC)-polymerase chain reaction (PCR) amplification and PCR-denaturing gradient gel electrophoresis (DGGE). At the same time, the diversity of gut microbiota was analyzed in rats. Quantitative real-time PCR was used to quantify the changes of gut microbiota following ethanol and UA treatments. Results: The pathological results obtained by HE staining showed that the hepatic lobular structure in the alcohol model group was vague, and fatty degeneration and inflammatory infiltration were increased. Compared with the alcohol model group, liver tissue damage was significantly improved and the activity of serum transaminase was significantly reduced in the UA-treated group (P < 0.05). In addition, D-LA and endotoxin levels in serum induced by alcohol were significantly suppressed after supplementing UA (P < 0.05). The results of ERIC-PCR and PCR-DGGE indicated that the alcohol-induced disorder of the gut microbiota structure was brought back to almost normal after the supplementation of UA. The results of quantitative real-time PCR indicated that the numbers of Enterococcus faecalis and Escherichia coli in the UA-treated group were significantly decreased to almost normal levels when compared with the alcohol model group (P < 0.05). Moreover, the numbers of Lactobacillus and Bifidobacterium were significantly higher in UA-treated group than in the alcohol model group (P < 0.05). Conclusion: Supplementation of UA can effectively improve alcohol liver injury in rats, likely by modulating gut microbiota distribution and improving intestinal microecology.

Key words: ursolic acid, alcohol-induced liver injury, gut microbiota, D-lactic acid, endotoxin