Monoacylglyceride rich in docosahexaenoic acid (DHA-MAG) as a novel structured lipid can be produced from Schizochytrium oil, which is conductive to accumulate DHA into tissues and perform its biological functions. In this study, the effects of different DHA dose on serum lipid composition, liver fatty acid profile and physiological change, and gut microbiota composition of acute alcoholic liver disease (ALD) mice were deeply investigated. The obtained results showed that the high dose of DHA-MAG promoted the accumulation of DHA in liver tissue and significantly alleviated alcohol-induced liver injury, as evidenced by decreasing serum levels of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase, and increasing the levels of total protein and albumin; meantime, the values of these recorded indexes were close to the ones by the positive control group treated with silymarin. Moreover, DHA-MAG treatment distinctly improved lipid metabolism, as indicated by reducing serum triglycerides, total cholesterol, and low-density lipoprotein levels, and increasing high-density lipoprotein level. Importantly, DHA-MAG normalized the disrupted composition of the gut microbiota caused by alcohol consumption, thereby leading to alterations in the release of metabolites into the systemic circulation. Additionally, DHA-MAG inhibited the TLR₄/NF-κB signaling pathway, thereby modified inflammatory responses in the liver. Overall, this study revealed that DHA-MAG could be potential as a novel therapeutic agent for the prevention and alleviation of ALD.