This experiment was conducted to investigate the effect of fermented spent mushroom substrate (FSMS) on growth performance, blood profile, intestinal microflora and ammonia gas production of feces in broiler chickens. A total of three hundred sixty, 1-day-old male broiler chicks (Ross) were randomly divided into 3 groups with 6 replicates of 20 birds each. The treatments were control (free FSMS), 15% FSMS (basal diet with 15% FSMS) and 30% FSMS (basal diet with 30% FSMS). The final body weight and body weight gains were slightly improved in 30% FSMS than control (P<0.05). Feed intake and feed conversion were significantly improved as compared to those of the control groups. The Leukocytes of blood serum in FSMS groups were significantly decreased as compared to those of control groups. There are no significant differences among the groups in the contents of albumin (ALB), total cholesterol (TCHO), glucose (GLU), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in blood serum. The content of total glucose (TG) in 30% FSMS containing dietary groups was significantly decreased as compared to that of the control groups. The content of HDLC in 30% FSMS containing dietary groups was significantly increased as compared to that of the control group. The number of lactobacillus in the intestinal microflora were significantly increased in chicks fed FSMS groups. The ammonia gas production in FSMS groups was siginificantly decreased as compard to that the control groups. These results indicated that dietary FSMS exerted growth performance for feeding broiler.