The aim of this study was to evaluate the effect of supplementing feed with a functional mixture of immunomodulators, including β-glucans, nucleotides, ascorbic acid, and alpha-tocopherol, associated with a diet with lower levels of animal protein (11.5%) and higher levels of soybean meal (43.5%), on the zootechnical performance, health, hematological and immunological parameters, intestinal morphology, centesimal composition, and intestinal microbiome of juvenile Nile tilapia (initial weight 1.88 g ± 0.25 g, mean ± standard deviation). Two isocaloric and isoproteic diets (35% crude protein) were formulated, one with the inclusion of the immunostimulant functional mixture (40 kg·t−1), composed of 150 mg·kg−1 of nucleotides, 1000 mg·kg−1 of β-glucans, 1000 mg·kg−1 of ascorbic acid (vitamin C), and 20 mg·kg−1 of alpha-tocopherol (vitamin E), and another without. The combined supplementation of nucleotides, β-glucans, ascorbic acid, and alpha-tocopherol resulted in a 59.95% increase in final weight, 64% weight gain, 66% daily gain, a 21.31% decrease in feed conversion rate, and double the retention of body protein. Supplementation also improved intestinal morphology and modulated the intestinal microbiome, increasing Chao-1 diversity. Transmission electron microscopy confirmed that fish fed with both diets exhibited intact intestinal mucosal membranes. Supplementation did not alter the hematological and immunological parameters, suggesting that there was no overstimulation of the fish’s immune system. This work allows us to evaluate the effect of reducing the use of animal protein in the diets of fish, along with the effects of nucleotides, β-glucans, ascorbic acid, and alpha-tocopherol. Together, these compounds can provide fish with the necessary tools to achieve optimal health and growth.