The intestinal H⁺/peptide cotransporter 1 (PEPT1) plays important roles as a nutrient and drug transporter. Previously, we reported that rat intestinal PEPT1 showed a diurnal rhythm, and that this rhythm is closely related to the feeding schedule. Furthermore, we also demonstrated that transcription factors, Sp1, Cdx2, and PPAR, contribute to the basal, intestine-specific, and fasting-induced expression of PEPT1, respectively. In this study, to clarify the molecular mechanism governing the diurnal rhythm of PEPT1 expression, we compared expression profiles of these transcription factors under two kinds of feeding schedules. The intestinal Sp1 and Cdx2 did not show a circadian accumulation of mRNA or response to the daytime feeding regimen. Plasma free fatty acids, endogenous PPAR ligands, exhibited a robust circadian fluctuation in phase with that of PEPT1. However, subsequent experiments using PPAR null mice revealed the absence of any association between PEPT1's circadian rhythm and PPAR. We then focused on the clock genes (Clock, Bmal1, Per1-2, and Cry1) and clock-controlled gene, DBP. A robust and coordinated circadian expression of the clock genes was observed and daytime feeding entirely inverted the phase except for Clock. The expression of DBP was in phase with that of PEPT1 in both groups. Electrophoretic mobility shift assays and reporter assays revealed that DBP has the ability to bind the DBP-binding-site located in the distal promoter region of the rat PEPT1 gene and induce the transcriptional activity. These findings indicate that DBP plays pivotal roles in the circadian oscillation of PEPT1.