The intestinal epithelium is a single‐cell layer on the mucosal surface that absorbs food‐derived nutrients and functions as a barrier that protects mucosal integrity. Hesperidin (hesperetin‐7‐rhamnoglucoside) is a flavanone glycoside composed of the flavanone hesperetin and the disaccharide rutinose, which has various physiological benefits, including antioxidative, anti‐inflammatory, and antiallergic effects. Here, we used human intestinal Caco‐2 cell monolayers to examine the effect of hesperidin on intestinal barrier function. Hesperidin‐treated Caco‐2 cell monolayers displayed enhanced intestinal barrier integrity, as indicated by an increase in transepithelial electrical resistance (TEER) and a decreased apparent permeability (Papp) for fluorescein. Hesperidin elevated the mRNA and protein levels of occludin, MarvelD3, JAM‐1, claudin‐1, and claudin‐4, which are encoded by tight junction (TJ)‐related genes. Moreover, hesperidin significantly increased the phosphorylation of AMP‐activated protein kinase (AMPK), indicating improved intestinal barrier function. Thus, our results suggest that hesperidin enhances intestinal barrier function by increasing the expression of TJ‐related occludin, MarvelD3, JAM‐1, and claudin‐1 via AMPK activation in human intestinal Caco‐2 cells.

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