We synthesized fentanyl analogues that possess key groups common to the opioid peptides to investigate whether or not these two classes of compounds interact with common subsites on opioid receptors. The design of the analogues was based on the possibility of structural analogy between the two aromatic rings of fentanyl and the Tyr1 and Phe4 residues of the opioid peptides. The synthesized compounds showed very weak or no opioid activity as tested in the electrically stimulated longitudinal muscle of the guinea pig ileum or mouse vas deferens preparations. These results, together with those of reported studies, suggest that fentanyl and the opioid peptides interact with different subsites on either mu or sigma receptors. Studies using the irreversible mu opioid receptor antagonist, beta-funaltrexamine, indicate that fentanyl interacts preferentially with mu opioid receptors.