The reduction of Aflatoxin B1 (AF) in juvenile rainbow trout (Oncorhynchus mykiss) diet was analyzed after supplementing Nanostructured Zeolite (NZ) in a 56-day experiment. Two hundred and seventy juveniles with an average weight of 23 ± 3.7 g were placed in 6 different groups of C (control as a basal diet), NZ0.5 (basal diet + 0.5% NZ), NZ1 (basal diet + 1% NZ), AF5 (basal diet + 5 mg AFB1), AF5 NZ0.5 (basal diet + 5 mg AFB1 + 0.5% NZ), AF5 NZ1 (basal diet + 5 mg AFB1 + 1% NZ) with three replications and were fed four times a day based on their satiation. No significant differences were observed in terms of growth performance among the experimental groups (P > 0.05). However, hepatosomatic index in fish fed by AF5 NZ0.5 was reduced compared with NZ0.5 group (P < 0.05). The carcass moisture content showed a higher amount in treatment AF5 NZ0.5 compared to the control group (P < 0.05). There was a decrease in fat content in treatment AF5 compared to that of the control group (P < 0.05). Serum total protein, albumin and globulin levels in fish fed with aflatoxin were lower than in fish fed the diet without AF for all levels of NZ (P < 0.05); however, the interaction between AF and NZ was not significant (P > 0.05). Concentrations of C3, C4 and immunoglobulin M together with serum lysozyme activity showed no significant differences among all treatments (P > 0.05). No considerable histopathological lesions were observed in liver, kidney and spleen for all treatments. Based on the results, NZ showed some effects on physiological functions in juvenile rainbow trout fed by 0.5% dietary NZ which could improve performance in this species.