New carboxyalkyl compounds derived from Phe-Leu and Phe-Ala were synthesized and checked as inhibitors of "enkephalinase", a metalloendopeptidase cleaving the Gly3-Phe4 bond of enkephalins from mouse striatal membranes. Differential recognition of both brain enkephalinase and angiotensin-converting enzyme (ACE) catalytic sites by these carboxylalkyl compounds lead to potent (KI approximately 0.5 microM), competitive and selective inhibitors of the enkephalin-degrading enzyme. The most interesting compound, N-[(RS)-2-carboxy-3-phenylpropanoyl]-L-leucine (3, KI = 0.34 microM), is 10000 times more potent on enkephalinase than on ACE activities. Intracerebroventricular (icv) injection of 3 in mice leads to a high potentiation of the analgesic effect of the exogenously administered D-Ala2-Met-enkephalin, evidencing the in vivo inhibition of enkephalinase. Moreover, icv administration of 3 alone induces a dose-dependent analgesia in mice measured on both hot-plate and writhing tests. In the former assay, the ED50 was approximately 10 micrograms per animal, slightly higher than that of thiorphan. All the antinociceptive effects were antagonized by naloxone, demonstrating the involvement of enkephalins in analgesia and their in vivo protection from enkephalinase by 3. The described compounds can be considered as first examples of a new series of analgesics and potentially psychoactive agents.