This study was conducted to compare the effects of fish meal (FM) replacement by two types of soybean meals (SBMs) on activities of digestive enzyme, intestinal brush border enzyme and aminotransferases of juvenile Japanese seabass (initial weight 8.3 ± 0.2 g). Nine isonitrogenous (crude protein 44%) and isoenergetic (20 kJ g−1) experimental diets replacing 0 (the control), 15%, 30%, 45% and 60% FM protein by high-protein SBM (HSB) or commercial SBM (CSB) were formulated. The concentrations of anti-nutritional factors (ANFs) in HSB were relatively lower compared with CSB (P < 0.05). After 10-week feeding experiment, activities of digestive enzymes, intestinal brush border enzymes and aminotransferases in liver decreased with increasing dietary SBM. When the substitution level was above 30% with CSB and 60% with HSB, activities of pepsin, disaccharidase in the intestinal brush border membrane and aspartate aminotransferase (AST) in liver were significantly lower than the control. However, activities of alanine aminotransferase (ALT) and AST in serum were significantly higher than the control when the substitution level was above 30% with CSB and 60% with HSB. Regression analysis showed that effects of FM substitution by HSB on digestion and metabolism of juvenile seabass were relatively smaller compared with CSB.