Abstract: To investigate the effects of buffalo milk exosomes on lipid metabolism disorders in the liver of mice induced by high-fat diet. In the experiment, exosomes were obtained through ultra-high speed centrifugation from buffalo milk, and a total 32 male C57BL/J6 mice were selected and randomly divided into normal control group (ND group), high-fat diet group (HFD group), low-dose exosomes group (HFD-L group), and high-dose exosomes group (HFD-H group). The mice in normal control group and the other groups were respectively fed with a 10% kcal control feed and a 60% kcal high-fat diet for 8 weeks. Subsequently, the mice were gavaged with either PBS or buffalo milk exosomes (5 and 20 mg/kg) every 48 h for a duration of 28 d, based on their respective groupings. The following indexes were detected: body weight, total adipose, liver tissue masses, levels of serum, liver triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), as well as serum gluten transaminase (AST) and alanine transaminase (ALT). Furthermore, liver tissue was subjected to HE staining and oil red O staining. The results showed that the intact buffalo milk exosomes could be obtained through ultra-high speed centrifugation, and buffalo milk exosomes could alleviate the weight gain caused by a high-fat diet in mice to a certain degree. Additionally, the exosomes significantly reduced liver and serum levels of TG, TC and LDL-C in a dose-dependent manner in high-fat mice, conversely increased liver and serum levels of HDL-C, and increased the serum levels of AST and ALT (P<0.05), slowing down the liver damage in high-fat mice. In addition, the results from both HE staining and oil red O staining demonstrated that buffalo milk exosomes could improve liver vacuolar degeneration and reduce liver lipid deposition in mice (P<0.05). In summary, the buffalo milk exosomes would possess the potential to alleviate liver lipid disorders induced by a high-fat diet, thereby provide a theoretical basis for the application and advancement of milk-derived exosomes as natural nanomedicines.