This study was conducted to investigate the effects of batch sources and the digestive enzyme activity contribution of intestinal fluid dry powder of ducks in the simulated duck intestinal fluid for ducks on the digestive ability of corn, soybean meal and wheat bran, which would provide a reference for the appropriate amount of intestinal fluid dry powder of ducks in the simulated intestinal fluid for ducks. A nested design was adopted with batch of intestinal fluid dry powder of ducks as an independent variable and the percentage of digestive enzyme activities of intestinal fluid dry powder of ducks in simulated intestinal fluid for ducks as a nested variable. There were 3 treatments of batches 1, 2 and 3 in the independent variable, and 4 treatments of 0 (shared), 25%, 50% and 75% in the nested variable. The dry matter digestibility (DMD) and enzymatic hydrolysate gross energy (EHGE) of corn, soybean meal and wheat bran were measured by in vitro digestion using ten simulated intestinal fluid for ducks, respectively. The results showed as follows: 1) there were no significant effects of simulated intestinal fluid for ducks prepared with 3 batches of intestinal fluid dry powder of ducks on the apparent dry matter digestibility (ADMD), true dry matter digestibility (TDMD), apparent enzymatic hydrolysate gross energy (AEHGE), and true enzymatic hydrolysate gross energy (TEHGE) of corn and wheat bran measured using in vitro digestion (P>0.05). No significant difference in the ADMD, AEHGE and TEHGE (P>0.05), but a significant difference in the TDMD (from 71.75% to 72.24%, P<0.05) of soybean meal digested by simulated intestinal fluid for ducks prepared with 3 batches of intestinal fluid dry powder of ducks were observed. 2) In the same batch of intestinal fluid dry powder of ducks, the ADMD and AEHGE of corn, soybean meal and wheat bran digested by 3 simulated intestinal fluid for ducks significantly were decreased on account of the contribution of digestive enzyme activity percentage changed from 25% to 75%, but the TDMD of corn, soybean meal and wheat bran were stable (the max absolute difference was 0.95%, and relative difference was 0.95%). The TEHGE of these 3 feeds digested by simulated intestinal fluid for ducks were stable (max absolute difference was 0.17 MJ/kg, and relative difference was 0.89%) under the contribution of the digestive enzyme activity percentage changed from 25% to 50%. In conclusion, the digestive abilities of simulated intestinal fluid for ducks prepared with 3 batches of intestinal fluid dry powder of ducks are equal. The digestive abilities of simulated intestinal fluid for ducks are stable under the contribution of the digestive enzyme activities changed from 25% to 50%.