Metabolic stability of synthetic dynorphins [N-terminal fragments of dynorphin A (Dyn A)] were evaluated in vitro and in vivo. These peptides were applied at concentrations 100-1000 times higher than those of the endogenous dynorphins. Degradation kinetics of these peptides were studied in rat brain homogenate by using microbore gradient RP-LC assay, and limited information on their metabolism was obtained by electrospray ionization mass spectrometry (ESI-MS) of the isolated metabolites. In vivo cerebral microdialysis, in which the peptides were introduced via the probe placed in striatum region of the brain of the experimental animals, was used to circumvent contamination arising from autoproteolysis of brain during incubation of the samples in vitro. Metabolites of Dyn A (1-13) and Dyn A (1-11) were identified from electrospray ionization mass spectra of the microdialysates without chromatographic separation; the identification of peptides in the mixtures were supported by medium resolution ESI Fourier-transform ion cyclotron resonance MS. LC-MS was used to fully characterize the complex peptide mixture obtained after the striatal perfusion of Dyn A (1-12).
- Mass spectrometry