Activation and fragmentation of peptides in time-of-flight mass spectrometry using optical and non-optical ionization sources

• Main Author: Fountain, Scott Timothy
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-1994
• Subjects: Analytical chemistry
• ISBN: 9798209085652

Four ionization sources, including electron impact (EI), vacuum ultraviolet (VUV), resonance-enhanced multiphoton ionization (REMPI), and matrix-assisted laser desorption/ionization (MALDI), are interfaced to a reflectron time-of-flight mass spectrometer (TOFMS). A comparison of these techniques for the analysis of small peptides (200-2200 amu) is described. Low energy ($<$70 eV) EI ionization provides isomeric discrimination of peptides containing leucine and isoleucine, via unique side-chain cleavage, only in limited cases. Soft ionization is not easily induced. Similar results are obtained with pulsed VUV lamp sources, where secondary ionization processes caused by photoelectron emission limits selectivity. Variable-wavelength REMPI (193 nm and 266 nm) provides discrimination, via metastable ion analysis, of isomeric dipeptide pairs (e.g., Leu-Tyr/Ile-Tyr, Tyr-Leu/Tyr-Ile, Ala-Trp/Trp-Ala) when coupled with supersonic cooling in CO$\sb2$ or He. The reflectron is used as an energy analyzer in order to determine the metastable fragmentation mechanism associated with isomer discrimination. A series of biologically active oligopeptides (e.g., bradykinin, substance P, and neurotensin) are studied by MALDI using a quadrupole ion trap/time-of-flight mass spectrometer (IT/TOFMS). This apparatus allows metastable decay of desorbed peptides to occur in the ion source, rather than the TOFMS drift tube, thereby providing structural information without calibration or reflectron energy analysis. The ion trap buffer gas pressure and RF field potential applied during desorption are found to influence ion activation, as is the desorption laser power. The source of unimolecular peptide fragmentation is determined to be from the pulsed desorption and pulsed extraction events. The ion trap source is coupled to a series of analog waveform generators to induce selective ion analysis via resonance ejection. These circuits allow peptide molecular ions to be isolated from matrix background and sample mixtures. Applications for tandem mass spectrometry are discussed.