Synthesis Of An Intrinsically Radiolabeled Enkephalin Analog - [Para-Tritio-Phenylalanyl]4-Norleucyl5-Enkephalin

In a previous study methionine-enkephalin (H-Tyr-Gly-Gly-Phe-Met-OH) and four analogs and homologs were synthesized and compared to morphine with respect to their ability to inhibit electrically evoked contractions of the isolated guinea pig ileum. The compounds were: Tyr-Gly-Gly-Phe-Leu (Leu5-enkephalin), Tyr-Gly-Gly-Phe (des-Met5-enkephalin), Tyr-Gly-Gly-Phe-Nle (Nle5-enkephalin) and Tyr-Gly-Gly-[p-chloro-Phe]-Nle ([p-chloro-Phe]4-Nle5-enkephalin). The IC50 concentrations on the guinea pig ileum assay were 3.0 × 10−7 M, 4.0 × 10−5 M, 1.1 × 10−7 M and 1.6 × 10−7 M respectively. The IC50 for Met5-enkephalin was 5.0 × 10−8 M and for morphine 2.0 × 10−7 M. These data indicate that the methionine residue is important for biological activity and that norleucine is a more acceptable substitution at this position than is leucine. Therefore, the [p-chloro-Phe]4-[Nle]5-enkephalin was synthesized, dehalogenated by catalytic hydrogenation in the presence of 3H2 to yield the intrinsically labeled [p-tritio-Phe]4-Nle5-enkephalin. The labeled peptide (9 Ci/mmole) was completely stable for at least 9 months at −20°C in 50% aqueous acetic acid.