Collision induced dissociation of peptide ions.: Part 3. Comparison of results obtained using sector—quadrupole hybrids with those from tandem double-focusing instruments

Abstract

Two series of peptides, viz., dynorphin fragments (up to 9 residues) and oligomers of phenylalanine, have been studied by collision induced dissociation (CID) of their protonated forms produced by fast atom bombardment (FAB) ionization. Spectra were obtained using both a four-sector tandem instrument (CID in the kiloelectronvolt range) and a tandem hybrid instrument (CID at a few tens of electronvolts). The four-sector instrument provided fragment spectra of excellent quality under conditions simulating a real-world analytical situation, i.e. 1–2 nmol of material available, although the FAB efficiency for Phe 8, was so low (precursor ion beam current ∼ 10 −14 A) that no useful fragment spectra were obtained. The hybrid instrument provided abundant fragment ions for smaller precursors (molecular weights < 500–600 Da) but for larger precursors (≥ 800 Da or so) the CID efficiency was much reduced and highly susceptible to composition effects, particularly the presence of highly basic amino acid residues. The information content of these spectra was evaluated objectively via two complementary computer algorithms. The limitations of CID in r.f.-only quadrupole collision cells are not due primarily to ion transmission effects, since examples of good quality spectra for large precursors are presented. The low efficiency of collisional activation of organic ions of m/z ≥ 800 Da or so, under conditions typical of quadrupole collision cells, was confirmed by the observation that in those cases where abundant fragment ions were obtained the corresponding spectra obtained in absence of collision gas were of comparable quality. The differences between collisional activation of such ions in the kiloelectronvolt range and in the electronvolt range, as well as the dramatic differences between low energy CID of large and small precursor ions, are discussed in terms of fundamental considerations of inelastic collisions. Finally, the qualitatively different kinds of fragmentation reactions, observed in the two regimes of collision energy, are described. In both cases the well-known peptide sequence fragments were observed. The high-energy CID also produced intense fragments arising from cleavage of all or part of sidechains (d, s, v and w series) which were entirely absent from the low-energy spectra. The latter contained abundant internal cleavage ions (both N- and C-terminus residues lost) which were relatively weak in the high-energy spectra. Loss of the C-terminus residue was observed in both regimes as a low-energy process in the absence of collision gas, although the compositional conditions giving rise to this process have not been determined in the present work.