Previously, dietary humic substances (HSs) or protease complexes (PCs) in fingerling channel catfish Ictalurus punctatus (Rafinesque) have demonstrated potential for health enhancement. Two trials were conducted to elucidate an optimal HS or PC administration plan for growth and health promotion. The first feeding trial was conducted within a recirculating aquaculture system. Catfish fingerlings were fed a commercial control (28% protein) or supplemented diet (control + PC or HS). Diets were administered through four feeding regimens of either HS, or PC‐supplemented diet. After 60 days, continuous feeding of diets with HS or PC contributed to improved feed efficiency and weight gain of the fish. Similarly, catfish growth was enhanced following the application of HS or PC for the final 4 weeks. Serum lysozyme activity was relatively improved in catfish when a PC or HS diets were offered for 60 days. Feeding either of the supplements for the final 4 weeks improved skin lysozyme activity compared to the control group. In the second trial, catfish were fed either the commercial control diet, PC‐supplemented diet, or HS‐supplemented diet in an outdoor production environment for 60 days. Both HS and PC catfish groups had higher survival rates than the control group. Furthermore, growth performance was superior in additive‐fed groups. After HS or PC application, gut microbiota analysis from catfish fecal at two time points revealed significant variation in alpha diversity, community abundance, and beta diversity. However, the association of dietary HS or PC on improved survival, immunity, and gut microbiota modulations presents their importance as functional immunostimulants in catfish aquaculture.