Green-synthesized zinc nanoparticles (ZnNP) have attracted considerable attention for use as aqua-feed supplements. The effect of ZnNPs on cellular energy, amino acid metabolism, and gene expression of major histocompatibility-II (MHC-II) is poorly understood. Our study explored the effect of dietary ZnNPs supplementation on hepatic function and overall health status. Farmed Nile tilapia cultured in a hapa-culturing system were fed ZnNPs with two ascending doses of 30 and 60 mg/kg dry feed for 8 weeks. Liver function enzymes, hepatic energy, amino acid profiles, gene expression analysis, and hepatic morphometry were evaluated. Non-significant changes were observed in the estimated biochemical indices, and a significant decrease (P < 0.05) in lactate dehydrogenase (LDH) activity was recorded in ZnNPs60 mg/kg dry feed compared to the control non-supplemented fish. Hepatic energy adenosine triphosphate (ATP) and CoQ10 were increased (P < 0.05) in the supplemented groups, whereas adenosine monophosphate (AMP) showed the opposite trend. The hepatic amino acid profile indicates the selectivity of dietary ZnNPs in modulating specific protein-metabolic pathways. A higher relative expression of IL-1β, along with no statistical changes in IL-10, HSP70, and MHC-II, was recorded. Normal liver histology and a significant dose-dependent increase in mean hepatocytic area were observed. Our results suggest that incorporating ZnNPs as a functional Nile tilapia supplement is beneficial. However, further comparative investigations of different fish species are required to determine the optimal dose.