Dietary supplementation of herbal essential oils is recommended for aquatic animals due to their growth-promoting, immune, and antioxidative potential. Hence, the present study evaluated the inclusion of menthol essential oil (MNT) at 0.1%, 0.2%, 0.3%, and 0.4% on the growth performance, digestive enzyme activity, immunity, and antioxidative responses of Nile tilapia. After eight weeks, samples were collected, and fish was exposed to acute ammonia challenge at 5 mg TA-N/L for 6 h. The results displayed that dietary MNT significantly enhanced the final body weight (FBW), weight gain, and feed conversion ratio (FCR) in a linear and quadratic manner (P < 0.05). The regression analysis revealed that the optimum rate of dietary MNT required for the best FBW is 0.26% and 0.30% for FCR. Dietary MNT enhanced the protease activity at the rate of 0.30% (P < 0.05) while lipase and amylase activities did not influence by MNT supplementation. Before and after the ammonia challenge, lysozyme (LZM), phagocytic activities (PA), phagocytic index (PI), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) were higher in fish fed MNT than fish fed MNT free diet. On the other hand, cortisol, glucose, and malondialdehyde (MDA) were decreased by dietary MNT in a linear and quadratic manner (P < 0.05). Ammonia stress induced reduced LZM, PA, and PI but increased the SOD, CAT, GPx, MDA, cortisol, and glucose levels. The related inflammatory genes (IL-8, TNF-α, and IL-1β) were reduced by dietary MNT either before or after the ammonia exposure, and the mRNA levels of these genes were increased after ammonia stress. The ammonia exposure upregulated the expression of HSP70, and dietary MNT displayed reduced HSP70. The study assumed that MNT inclusion at the rate of 0.22% to 0.30% is recommended to increase the growth performance, protease activity, immunity, antioxidative response, and the resistance against ammonia exposure in Nile tilapia.