This research aimed to investigate the effect of different levels of glycerol butyrate in the diet on growth indices, digestive enzyme activity, antioxidant enzymes, and gene expression related to the immune system of Pacific white shrimp (Litopenaeus vannamei). In the experiment conducted at the aquaculture research station of the Persian Gulf University, four different dietary treatments were considered: 1) commercial feed without glycerol butyrate plus gelatin (control group), 2) commercial feed + 0.25% glycerol butyrate + gelatin, 3) commercial feed + 0.5% glycerol butyrate + gelatin, and 4) commercial feed + 1% glycerol butyrate + gelatin. This experiment examined the immune indices, antioxidant enzymes, and growth and feeding performance of the shrimp. Before the experiment, the culture tanks and equipment were thoroughly disinfected and washed. Then, 150 healthy and disease-free shrimp with an average weight were selected and kept in 1000-liter tanks for 2 weeks to acclimatize and were fed a commercial diet. The culture period was 8 weeks with a photoperiod of 12 hours of light and 12 hours of darkness. Evaluation of growth parameters (WG, WG%, SGR, and FCR) showed that treatments 1, 2, and 3 had a significant difference with the control group (p<0.05), but treatment 1 did not differ significantly from treatments 2 and 3 (p>0.05). The highest values of WG, WG%, and SGR were observed in the control group, and the highest FCR was in treatment 2. Additionally, the comparison of hemolymph immune parameters showed that the control treatment differed significantly from treatments 1, 2, and 3 in the values of Hyaline and Semi granular (p<0.05), but no significant difference was observed in the values of Large granular (p>0.05). The highest values of Hyaline and Semi granular were observed in treatments 2 and 2, respectively, and the lowest values were observed in the control group. Additionally, the comparison of Total Hemocytes values showed that there was a signif