This study investigated the protective effect of aplysin on the liver and its influence on inflammation and the gut microbiota in rats with ethanol-induced liver injury.Male Sprague-Dawley rats were randomly assigned to an alcohol-containing liquid diet, control liquid diet or treatment with aplysin for 8 weeks. Hepatic and intestinal histopathological analysis was performed, and cytokine levels and the intestinal mucosal barrier were assessed. Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) and 16S rDNA high-throughput sequencing were performed to provide an overview of the gut microbiota composition.Chronic alcohol exposure caused liver damage in rats. Serum aspartate aminotransferase (AST), aminotransferase (ALT), alkaline phosphatase (ALP) and triglyceride (TG) activities in liver tissue were higher than in the control group. Alcohol administration elevated the levels of serum transforming growth factor-β (TGF-β) and tumor necrosis factor-α (TNF-α) and reduced interleukin-10 (IL-10) levels compared with those of control rats. In addition, the levels of plasma endotoxin, diamine oxidase (DAO), and fatty acid-binding protein 2 (FABP2) in the alcohol group were higher than in the control group. The results of ERIC-PCR indicated that aplysin treatment shifted the overall structure of the ethanol-disrupted gut microbiota toward that of the control group. One hundred twenty to 190 genera of bacteria were detected by high throughput sequencing. Alcohol-induced changes in the gut microbial composition were detected at the genus level. These alcohol-induced effects could be reversed with aplysin treatment.These results suggest that aplysin exerts a protective effect on ethanol-induced hepatic injury in rats by normalizing fecal microbiota composition and repairing intestinal barrier function.