In this study, the effects of arginine on growth, serum antioxidant levels, intestinal digestion physiology, intestinal protein synthesis, and expression of inflammatory-related genes of triploid juvenile rainbow trout (Oncorhynchus mykiss) were investigated. Five diets containing 0.86%, 1.65%, 2.10%, 2.85%, and 3.50% arginine were fed to satiation to triplicate groups of 30 fish per tank for 8 weeks. The results showed that the weight gain rate, specific growth rate, and protein efficiency ratio significantly increased. The optimal dietary arginine requirement was 2.48% (5.97% of dietary protein) based on quadratic regression analysis between specific growth rate and dietary arginine content. There were no significant differences in crude lipid and ash content. Moisture significantly decreased, and crude protein content significantly increased (
P
<
0.05
). Superoxide dismutase, catalase, and lysozyme levels in serum and the liver first significantly increased and then decreased with the level of arginine compared to the control group (
P
<
0.05
), while the malondialdehyde level significantly decreased. Intestinal lipase and amylase levels significantly increased, but there was no significant difference in trypsin level (
P
>
0.05
). The muscular layer thickness and villus width foregut in the 2.85% group were significantly increased. TOR at 2.86% and IGF-1 and S6K1 at 2.11% reached the maximum level, and IL-1β, IL-6, and IL-8 showed a decreasing trend with their lowest levels in the 3.50% and 2.85% groups. The expression of TNF-α first decreased and then increased with the arginine level. Both TGF-β and IL-10 expression first decreased and then increased with the level of arginine and reached the maximum value in the 2.85% group. NF-κB showed an opposite trend and reached the lowest value in the 3.50% group. In conclusion, dietary arginine has a significant effect on growth, serum antioxidant capability, digestion physiology, immunity, digestion, and absorption of nutrients in triploid O. mykiss, and the optimum requirement is 2.48% fed a low-fishmeal diet.