Melanoidins are considered to have several biological activities. In this study, black garlic melanoidins (MLDs) were collected using ethanol solution extraction; 0%, 20%, and 40% ethanol solutions were used for chromatography. Three kinds of melanoidins were produced by macroporous resin, named MLD-0, MLD-20, and MLD-40. The molecular weight was determined, and the infrared and microscopic structures were studied. In addition, Balb/c mice were induced with cyclophosphamide (CTX) to establish an immune deficiency model to evaluate the immune efficacy of black garlic melanoidins (MLDs). The results showed that MLDs restored the proliferation and phagocytosis ability of macrophages, and the proliferation activity of B lymphocytes in the MD group was 63.32% (♀) and 58.11% (♂) higher than that in the CTX group, respectively. In addition, MLDs alleviated the abnormal expression of serum factors such as IFN-γ, IL-10, and TNF-α. 16SrDNA sequencing of intestinal fecal samples of mice showed that MLDs changed the structure and quantity of intestinal flora, and especially that the relative abundance of Bacteroidaceae was significantly increased. The relative abundance of Staphylococcaceae was significantly reduced. These results showed that MLDs improved the diversity of intestinal flora in mice, and improved the adverse state of immune organs and immune cells. The experiments confirm that black garlic melanoidins have potential value in immune activity, which provides an important basis for the development and utilization of melioidosis.