Some beneficial effects of grape seed proanthocyanidin extract (GSPE) can be explained by the modulation of enterohormone secretion. As GSPE comprises a combination of different molecules, the pure compounds that cause these effects need to be elucidated. The enterohormones and chemoreceptors present in the gastrointestinal tract differ between species, so if humans are to gain beneficial effects, species closer to humans—and humans themselves—must be used. We demonstrate that 100 mg/L of GSPE stimulates peptide YY (PYY) release, but not glucagon-like peptide 1 (GLP-1) release in the human colon. We used a pig ex vivo system that differentiates between apical and basolateral intestinal sides to analyse how apical stimulation with GSPE and its pure compounds affects the gastrointestinal tract. In pigs, apical GSPE treatment stimulates the basolateral release of PYY in the duodenum and colon and that of GLP-1 in the ascending, but not the descending colon. In the duodenum, luminal stimulation with procyanidin dimer B2 increased PYY secretion, but not CCK secretion, while catechin monomers (catechin/epicatechin) significantly increased CCK release, but not PYY release. The differential effects of GSPE and its pure compounds on enterohormone release at the same intestinal segment suggest that they act through chemosensors located apically and unevenly distributed along the gastrointestinal tract.