Instrumentation and development of a mass spectrometry system for the study of gas-phase biomolecular ion reactions (2015)

Gas-phase reactions of biomolecular ions are highly relevant to the understanding of structures and functions of the biomolecules. Mass spectrometry is a powerful tool in investigating gas-phase ion chemistry. Various mass spectrometers have been developed to explore ion/molecule reactions, ion/ion reactions, ion/photon reactions, ion/radical reactions etc., both at atmospheric pressure and in vacuum. In-vacuum reactions have an advantage of involving pre-selecting the ions for the reactions using a mass analyzer. Over the decades, a variety of mass analyzers have been employed in the research of ion chemistry. Hybrid configurations, such as quadrupole ion trap with a time-of-flight and or a quadrupole ion trap tandem with an Orbitrap, have been utilized to improve the performances for both the reaction (in trapping mode) and the mass analysis (accurate mass measurements). Complicated instrument structures, including ion optics, multiple mass analyzers and differential pumping for high vacuum, are typically required for the mass spectrometers for gas phase ion chemistry study. An alternative approach is to simplify the instrumentation by using pulsed discontinuous atmospheric pressure interfaces for introducing ionic or neutral reactants and a single ion trap as both the reactor and the mass analyzer. Such a simple mass spectrometry system was set up and demonstrated using two discontinuous atmospheric pressure interfaces in the study for this thesis. It was capable of carrying out ion/molecule and ion/ion reactions at an elevated pressure without the needs of ion optics or differential pumping system. Together with a pyrolysis radical source, in-vacuum ion/radical reactions were performed and their associated chemistry was studied. Radicals are important intermediates related to biochemical processes and biological functions. There are very limited techniques to monitor the reactive intermediates in-situ during a multi-step reaction in aqueous phase. On the other hand, these intermediates can be "cooled down" and preserved into a single-step procedure in gas-phase reactions since they only occur via collisions. Therefore, the fundamental study of gas-phase radical ion chemistry will provide evidences of the reactivity, energetics, and structural information of biological radicals, which has the potential to solve puzzles of aging, disease biomarker identification, and enzymatic activities. Using the system described above, a new reaction between protonated alkyl amines and pyrolysis formed cyclopropenylidene carbene was discovered, as the first experimental evidence of the reactivity of cyclopropenylidene. Given the important role of cyclopropenylidene in the combustion chemistry, organic synthesis, and interstellar chemistry, it is highly desirable to establish a fundamental understanding of their physical and chemical properties. The amine/cyclopropenylidene reactions were systematically studied using both theoretical calculation and experimental evidences. A proton-bound dimer reaction mechanism was proposed, with the amine and the carbene sharing a proton to form a complex as the first step, which was closely related to the high gas-phase basicity of cyclopropenylidene. Subsequent unimolecular dissociation of the complex yielded three possible reaction pathways, including proton-transfer to the carbene, covalent product formation, and direct separation. These reactions were studied with a variety of alkyl amines of different gas-phase basicities. For the covalent complex formation, partial protonation on cyclopropenylidene within the dimer facilitates subsequent nucleophilic attack to the carbene carbon by the amine nitrogen and leads to a C-N bond formation. The highest yield of covalent complex was achieved with the gas-phase basicity of the amine slightly lower but comparable to cyclopropenylidene. The results demonstrated a new reaction pathway of cyclopropenylidene besides the formation of cyclopropenium, which has long been considered as a "dead end" in interstellar carbon chemistry. Further reactivity study of cyclopropenylidene towards biomolecular ions was also carried out for nucleobases, nucleosides, amino acids, peptides, proteins, and lipids. The reaction to form proton-bound dimer for protonated biomolecular ions remained as the dominant reaction pathway. Interestingly, other possible reaction pathways, such as modifications of thiyl group or disulfide bonds, double bond addition, and single bond insertion, were inhibited in gas-phase ion/carbene reactions. Such results inferred that the reactivity of neutral species was not directly applicable to ion reactions, with the proton involved in the gas-phase biomolecular ion reactions.