Previously, we confirmed that dietary fructooligosaccharides (FOS) increase calcium absorption in rats. In this study, we examined the influence of FOS feeding on the concentration of calbindin-D9k of several intestinal segments in rats. Rats in the control group were fed a diet without FOS. Rats in the other two groups were fed the diet containing FOS at either 50 or 100 g/kg for 10 d and subjected to a calcium absorption study. On the final day of feeding, the rats were killed and the entire intestine was removed. The intestinal mucosa was collected from four segments, i.e., the proximal and distal segments of the small intestine, the cecum and the colorectum, respectively. The apparent absorption of calcium increased dose dependently (r = 0.9256, P < 0.0001). Significant positive correlations between apparent calcium absorption and the relative amounts of calbindin in both large intestinal segments were observed (cecum, r = 0.8956, P = 0.0011; colorectum, r = 0.8828, P = 0.0016). Also, significant negative correlations between apparent calcium absorption and the relative amounts of calbindin-D9k in both small intestinal segments were observed (proximal, r = -0.7149, P = 0. 0304; distal, r = -0.8740, P = 0.0021). In conclusion, FOS feeding increases levels of calbindin-D9k in the large intestine, but decreases those in the small intestine. Moreover, these results suggest that part of the stimulatory effect of fructooligosaccharides relates to the transcellular route of calcium absorption in the large intestine of rats.