The ion-neutral complex-mediated fragmentation reaction in electrospray ionization tandem mass spectrometric analysis of N-phenyl-3-(phenylthio)propanamides

Rationale Amides are the fundamental units of both peptides and proteins, and also important functional groups of medical chemicals. Investigation of the fragmentation mechanism of amides in the gas phase is scientifically important for structural analysis. However, understanding of this problem is still elusive. Methods Protonated N-phenyl-3-(phenylthio)propanamide and its derivatives were investigated using positive ion tandem mass spectrometry (ESI-MS/MS) with an LCQ mass spectrometer. Accurate mass analysis was conducted with a micrOTOF-QII mass spectrometer. Density functional theory (DFT) calculations using the Gaussian 03 program and deuterium-labelling (D-labelling) experiments were performed to verify the proposed fragmentation mechanism. Results Interpretation of the fragment ions in the collision-induced dissociation mass spectra showed that the ionizing proton in the protonated ion transferred from the most thermodynamically favorable carbonyl oxygen to the dissociative protonation site at amide nitrogen or sulfur atom upon collisional activation. The dissociation of the amide or the C-S bond was induced by such proton transfer. An ion-neutral complex (INC) was generated via the dissociation of the amide bond. In the INC, it was observed that the carbocation of the ionic part attacked the ortho phenyl carbon atom adjacent to the sulfur atom, and proton transfer from the carbon atom to the nitrogen atom led to the formation of protonated aniline. Conclusions The fragmentation mechanism of protonated N-phenyl-3-(phenylthio)propanamide and its derivatives was proposed and elucidated. All the compounds studied showed similar fragmentation pathways, and the competitive formation of two ions, RC9H9OS+ and C6H8N+, was observed. The generation of protonated aniline is mediated by INC in ESI-MS/MS.