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Intestinal epithelial differentiation plays an important role in maintaining gut health and is impaired due to many diseases. Purple potatoes are rich in polyphenols that promote the differentiation of intestinal epithelial cells. In this study, we found that purple potato (PP) extract up-regulated the level of peroxisome proliferator-activated receptor γ coactivator (PGC)-1α, the expression of mitochondrial-related enzymes, the content of mitochondrial DNA, and the levels of tricarboxylic acid cycle metabolites in human intestinal epithelial Caco-2 cells. Inhibition of PGC-1α by either PGC-1α inhibitor SR-18292 or siRNA-mediated knockdown eliminated the enhancing effects of PP extract on the expression of differentiation markers, mitochondrial metabolite contents and mitochondrial DNA copy number, as well as the expression of mitochondrial DNA-encoded enzymes. In summary, PP extract promoted mitochondrial biogenesis and the differentiation of intestinal epithelial cells, and these promotion effects are dependent on the activation of PGC-1α.