The objective of this study was to evaluate the immunomodulatory activity of mung bean protein hydrolysate (MBPH) in lipopolysaccharide (LPS)-induced RAW 264.7 cells and discuss the possible immune regulatory mechanism. MBPH was prepared by alcalase, trypsin, neutrase, and flavourzyme. The 3-h alcalase-hydrolyzed hydrolysate with a molecular weight less than 1,450 Da was selected for the immunological tests. Results showed that MBPH possessed strong suppressing activity to proinflammatory mediators in a dose-dependent manner. Compared to the LPS alone group, MBPH (200 mu g/mL) significantly reduced nitric oxide (NO), inducible nitric oxide synthase, interleukin (IL)-6, and IL-1 beta secretion levels by 52.6%, 53.2%, 48.4%, and 49.7%, respectively, in LPS-induced macrophages. It also enhanced IL-10 secretion from 789 to 3,678 pg/mL. MBPH blocked nuclear factor-kappa B (NF-kappa B) translocation in LPS-induced macrophages through the prevention of I kappa B alpha phosphorylation, and this process further prevented p65 translocation into the nucleus. A possible mechanism of MBPH is that it regulated the expression of inflammatory factors via the NF-kappa B pathway, thus inhibiting inflammatory reactions. The results suggested that MBPH is of application potential in the development of immunomodulatory functional food to ameliorate immunosuppression.