Synthesis and biological activity of delta Phe4-enkephalins

The delta Phe4-enkephalins have been synthesized and examined in an in vitro receptor binding assay and an in vivo tail flick analgesia test. The delta Phe4 residue was derived from Boc-Gly-Phe(beta-OH)-OH by spontaneous dehydration and azlactonization. The dipeptide azlactone was coupled directly with H-Leu-OBzl to yield a tripeptide which was converted into the pentapeptides after stepwise coupling with two amino acids using the water soluble EDC-HOBt method. Dehydroenkephalins were liberated with hydrogen fluoride in the presence of anisole. In the radioligand binding assay which did not contain an enzyme inhibitor [D-Ala2, delta Phe4, Leu5] enkephalin was almost twice as active as saturated [D-Ala2, D-Leu5]-enkephalin. The delta Phe4-enkephalins exhibited a considerably diminished activity as compared with the saturated peptide in the in vivo analgesic assay. These results are discussed with regard to the enzyme stability and receptor preference of dehydroenkephalins.