[Leu-enkephalin homogeneously labeled with tritium in studying the Selank inhibiting effect on the enkephalin-degrading enzymes of human plasma].

A method of analysis of enkephalinase activity in blood plasma based on the application of Leu-enkephalin generally labeled with tritium at all its amino acid residues was developed. The method allows the simultaneous estimation of activity of several peptidases in microquantities of tissues. [G-3H]Leu-enkephalin was prepared by the method of solid phase catalytic isotope exchange (120 Ci/mmol) and subjected to proteolysis by the treatment with blood plasma. The resulting radioactive metabolites were separated by HPLC in the presence of the mixture of unlabeled fragments of Leu-enkephalin as internal standards. It was shown that aminopeptidases, dipeptidylaminopeptidases, and dipeptidylcarboxypeptidases respond for approximately 80%, 2%, and 10% of the total enzymatic activity, respectively. The new pathway of degradation of Leu-enkephalin by carboxypeptidase that provides for approximately 6% of the total enkephalin-degrading activity was discovered. Bestatin was shown to predominantly inhibit aminopeptidases and carboxypeptidases, whereas Selank is more specific for carboxypeptidases and dicarboxypeptidases. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2004, vol. 30, no. 3; see also http://www.maik.ru.