Non-direct sequence ions in the tandem mass spectrometry of protonated peptide amides--an energy-resolved study.

The fragmentation reactions of the MH(+) ions of Leu-enkephalin amide and a variety of heptapeptide amides have been studied in detail as a function of collision energy using a QqToF beam type mass spectrometer. The initial fragmentation of the protonated amides involves primarily formation of bn ions, including significant loss of NH3 from the MH(+) ions. Further fragmentation of these bn ions occurs following macrocyclization/ring opening leading in many cases to bn ions with permuted sequences and, thus, to formation of non-direct sequence ions. The importance of these non-direct sequence ions increases markedly with increasing collision energy, making peptide sequence determination difficult, if not impossible, at higher collision energies.