An 8-week feeding trial was conducted to investigate the effects of partial replacement of fish meal by soy protein concentrate (SPC) on the growth performance, immune responses, intestine morphology and relation gene expression of intestinal inflammation for juvenile hybrid grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) (initial weight 12.5 ± 0.00 g). Eight isonitrogenous and isolipidic diets (48.61% protein and 11.17% lipid) were formulated by replacing 0% (the control), 11%, 22%, 33%, 44%, 55%, 66%, and 77% of fish meal (FM) with SPC, respectively (the eight dietary be named FM, S11, S22, S33, S44, S55, S66, and S77, respectively). With the replacement level increased, the final body weight, weight gain ratio (WGR), specific growth rate (SGR), and survival rate of fish were significantly decreased (P < 0.05) compared with the group FM. By contrast, the feed conversion ratio (FCR) of fish was significantly increased (P < 0.05) when the replacement level up to 44%. Partial FM replacement by SPC (ranging from 11% to 77%) substantially reduced (P < 0.05) the serum total protein, albumin, and total cholesterol contents compared with the group FM. Liver total superoxide dismutase, glutathione peroxidase, catalase activities, and total antioxidant capacity showed the same trend of gradual increase first and then decrease. Their highest values were found in the replacement levels of 55%, 33%, 22%, and 55% and were significantly higher (P < 0.05) than the control group. The lowest malondialdehyde content was observed in group S77 and was significantly lower (P < 0.05) than that of the control group. The complements C3 and C4 contents of fish fed with experimental diets (replacement level ranged from 11% to 66%) were significantly higher (P < 0.05) than the group FM. The liver lysozyme activity of the control group was the lowest and was significantly lower than that of other dietary treatments (P < 0.05). Villus length and muscle thickness in the intestine of fish were significantly lower (P < 0.05) than other groups when the replacement level exceeded 44%. With dietary replacement levels increased, the TLR22, MyD88, p65, pro-inflammatory cytokines (IL-1β, TNF-α, IL-12P40 and INF-γ) and anti-inflammatory cytokines (TGF-β, IL-10, epinecidin, MHCIIβ and hepcidin) mRNA levels in the proximal intestine were significantly up-regulated (P < 0.05). The TLR22, MyD88, p65, pro-inflammatory cytokines (IL-1β, TNF-α, IL-12P40 and INF-γ) and anti-inflammatory cytokines (TGF-β, IL-10, MHCIIβ and hepcidin) mRNA levels in the mid intestine were significantly up-regulated (P < 0.05). The mRNA levels of TLR22, anti-inflammatory cytokines (IL-1β, TNF-α, IL-12P40, INF-γ) and anti-inflammatory cytokines (TGF-β, IL-10, epinecidin, MHCIIβ and hepcidin) in the distal intestine were significantly down-regulated (P < 0.05). The mRNA expression of MyD88 and p65 mRNA were showed a tend increased first and then decreased, and the highest values were observed in group S33 and S55 (P < 0.05), respectively. Based on the present work, the correlation between WGR and FM replacement level with SPC was described using the broken-line model, which estimated the optimum FM replacement to 37.23% for juvenile hybrid grouper dietary.