Increased levels of alcohol consumption are linked to an increased risk of alcoholic liver disease. As a novel source of national food, microalgae are rich in bioactives and have been demonstrated to confer benefits to human health. The effects of microalgae extracellular vesicles (M-EVs) on alcohol-induced liver injury have not been studied previously. In this study, 3 types of M-EVs were isolated using a combination of tangential flow filtration (TFF) and size exclusion chromatography (SEC). The structure and composition of M-EVs were characterized and the safety and liver-accumulation of M-EVs were also confirmed. Results shown that 3 types M-EVs were shown to be colloidal particles ((198.9 ± 7.2), (191.7 ± 5.1), and (259.5 ± 8.1) nm) with negative charge ((–25.0 ± 1.6), (–21.4 ± 0.6), and (–22.4 ± 1.6) mV). M-EVs were able to reduce the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and triglycerides (TGs) in the mice and regulated the levels of malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD), which protected the mice against oxidative damage and inflammation caused by alcohol-induced liver injury. In addition, the M-EVs may also have alleviated alcohol-induced liver injury through activation of the Nrf2/HO-1 signaling pathway and regulation of CYP2E1 expression, as well as by modulating the diversity and composition of the gut microbiota. The potentially beneficial effects of M-EVs on alcoholic liver disease may provide a theoretical basis for further exploration of the health function of M-EVs in the food industry.