Effects of dietary supplemental stachyose on cecal skatole concentration, hepatic cytochrome P450 (CYP450, CYP) mRNA expressions and enzymatic activities in broilers were evaluated. Arbor Acre commercial mixed male and female chicks were assigned randomly into six treatments. The positive control (PC) diet was based on corn-soybean meal, the negative control (NC) diet was based on corn-nonsoybean meal. The NC diet was then supplemented with 4, 5, 6, and 7 g/kg stachyose to create experimental diets, named S-4, S-5, S-6, and S-7, respectively. Each diet was fed to 6 replicates of 10 birds from days 1 to 49. On day 49, the cecal skatole concentrations in the PC, S-4, S-5, S-6, and S-7 groups were lower than those in the NC group by 42.28%, 23.68%, 46.09%, 15.31%, and 45.14% (P < 0.01). The lowest pH value was observed in the S-5 group (P < 0.05). The stachyose-fed groups of broilers had higher cecal acetate and propionate levels compared to control groups, and propionate levels in the S-6, S-7 groups were higher than those in the S-4 and S-5 groups (P < 0.001). The highest CYP3A4 expression was found in the S-7 group (P < 0.05), but this was not different from PC, S-4, S-5 and S-6 treatment. There was no significant difference in CYP450 (1A2, 2D6, 3A4) enzymatic activities among the groups (P > 0.05). In conclusion, cecal skatole levels can be influenced by dietary stachyose levels, and 5 g/kg of stachyose in diet was suggested.