Abstract: This study was conducted to investigate the difference of intestinal fluid composition along with the intestine of Peking duck fed with wheat-soybean meal diet and corn-soybean meal diet, respectively, which would provide biological basis for simulating intestinal fluid in duck feedstuff evaluation using in vitro digestion. The completely random design was adapted. Thirty adult ducks with similar weight were divided into 2 groups consisting of 5 replicates of 3 birds each and fed ad libitum with wheat-soybean meal and corn-soybean meal diets, respectively. The digesta of duodenum, anterior jejunum, posterior jejunum and ileum were sampled after slaughter at the 19^th day of the test period. Intestinal fluid was made by centrifuging digesta samples at 4 ℃. The activites of digestive enzymes and electrolyte concentration were determined. The results showed that there were higher amylase, lipase activities in duodenal and anterior jejunal fluid than that in posterior jejunal and ileal fluid (P<0.05). The activity of trypsin was higher in anterior and posterior jejunal fluid than that in duodenal and ileal fluid (P<0.05). The anterior jejunal fluid had higher activity of chymotrypsin than that in duodenal, posterior jejunal and ileal fluid (P<0.05). There was significant difference in chymotrypsin activity between in any two of duodenal, posterior jejunal and ileal fluid. Along with intestine of Peking duck, the pH was increased from 6.22 to 7.94. The Na⁺ and Ca²⁺ concentration were increased and then decreased. The K⁺ concentration was slowly decline. The Mg²⁺ concentration in duodenal fluid was lower than that in anterior jejunal, posterior jejunal and ileal fluid. Diet treatments significantly affected amylase, lipase, trypsin, and chymotrypsin activities and K⁺，Mg²⁺ concentration of intestinal fluid. In conclusion, along the duck’s intestine, jejunal fluid had higher 4 digestive enzyme activities than that in duodenal or ileal fluid, and there were significant difference in the ion concentration of intestinal fluid from duodenum, anterior jejunum, posterior jejunum and ileum.