Tandem mass spectrometry of peptides: Mechanistic aspects and structural information based on neutral losses. II—Tri‐ and larger peptides

The neutral products arising during the collisionally activated dissociation of protonated oligopeptides (MH + ) are post‐ionized by collision and detected in neutral fragment‐reionization ( + N f R + ) mass spectra. For the isomeric tripeptides Ala‐Gly‐Gly, Gly‐Ala‐Gly and Gly‐Gly‐Ala, the amino acid and dipeptide losses from the C ‐terminus and the diketopiperazine losses from the N ‐terminus allow for differentiation. These neutral fragments are identified in the corresponding + N f R + spectra by comparison to reference collision‐induced dissociative ionization (CIDI) mass spectra of individual amino acids, dipeptides and diketopiperazines. Peptides with distinct C ‐termini but otherwise identical sequences are found to yield + N f R + products that are characteristic of the respective C ‐terminal amino acid. This is demonstrated for several peptide pairs, including leucine‐ and methionine‐enkephalin. In general, + N f R + spectra are dominated by the heavier neutral losses; further, + N f R + and CIDI cause extensive dissociation, indicating that the collisional ionization process imparts high average internal energies.