SEQUENTIAL MASS-SPECTROMETRY APPLIED TO THE STUDY OF THE FORMATION OF INTERNAL FRAGMENT IONS OF PROTONATED PEPTIDES

Internal fragment ions of protonated peptides arise by charge retention on a portion of the structure excised from the peptide chain. Their formation appears to be favored by the extended time scale and multiple collision conditions associated with decomposition experiments which use the r.f.-only quadrupole of a hybrid sector/quadrupole mass spectrometer. Sequential product ion scanning and reaction intermediate scanning techniques of MS-MS-MS were used to probe these necessarily multistep fragmentations. With an instrument of BEqQ geometry, mass-analyzed ion kinetic energy spectrometry analyses were used to determine the population of candidate intermediates available for subsequent decomposition to internal fragments in the r.f.-only quadrupole. Reaction intermediate scanning then indicates the relative quantitative significance of competing pathways. In the examples studied, certain Y"-type ions show a high inherent tendency to fragment further to give internal fragments, consistent with the formal equivalence of these intermediates to protonated truncated peptides. Internal fragments can also arise, however, through a number of multistep pathways which can predominate in situations where the requisite intermediates are formed in high abundance.