BACKGROUND Acute chemical liver injury needs to be further explored. The present study aimed to compare the effects of intraperitoneal injection with carbon tetrachloride on acute liver toxicity after 24 h in male and female Kunming mice. MATERIAL AND METHODS In this study, female and male mice were simultaneously divided into 3 different groups. Each group was treated differently, and after 24 h, blood samples were collected to check for changes in the activity of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), which were used to assess liver toxicity. Liver samples were used for hematoxylin-eosin staining, and periodic acid Schiff reagent staining was performed to detect the pathological changes of each group. The expression level of biomarker molecules in liver cells was also systematically analyzed. RESULTS Our results showed that, compared with male mice, female mice showed more serious damage: reduced glycogen and higher degree of necrosis, and the levels of heatshock protein 27 (HSP27), heat-shock protein 70 (HSP70), proliferating cell nuclear antigen (PCNA) and B cell lymphoma/lewkmia-2 (Bcl-2) were significantly lower than in the male group (P<0.05 or P<0.01), while the results of Bcl-2-associated X protein (Bax), cysteinyl aspartate specific proteinase 3 (Caspase3), and cytochrome P450 2E1 (CYP2E1) were the opposite (P<0.05 or P<0.01). CONCLUSIONS The findings from this study showed that, compared with male mice, at 24 h after CCl₄ toxicity, female mice showed more severe changes of hepatocyte necrosis and PAS-positivity, with significantly reduced expression of HSP27, HSP70, PCNA, and Bcl-2, and significantly increased expression of Bax, caspase-3, and CYP2E1.