Development of fragmentation techniques in mass spectrometry for biological applications Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/cv43p0257

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  • In this dissertation, I present the work of developing more efficient and economical techniques for fragmenting large bio-molecules, and ultimately making mass spectrometry a more powerful tool in proteomics, DNA sequencing, and other areas of biomolecular analysis. Aiming at lowering the installation and operating cost, and delivering electron capture dissociation (ECD) technique to general users for the study of proteomics, I present the results of developing ECD on a triple-quadrupole mass spectrometer in the second chapter of this thesis. With a cross-flow configuration design, I demonstrate that zero-energy electrons (i.e. electron with energies on order of 1/2kT) can be transmitted into the reaction volume of the ECD chamber in numbers sufficient to produce detectable resonance electron capture (REC) reactions while maintaining ion transmission at a high enough level to perform cross-beam experiments. With a parallel-flow configuration design, on the other hand, I obtained the ECD spectrum of a peptide called substance P and demonstrate that the cross section for ECD can be increased by increasing both the interaction time and effective volume for interaction. I also demonstrate that with further improvements, in particular increasing the magnetic field strength it should be possible to turn ECD into a high-throughput tool for applications in proteomics. In chapter three I present the results of the first observation ever of the charge remote fragmentation (CRF) of a series of long-chain saturated and monounsaturated fatty acid anions during fast atom bombardment (FAB) of analyte-matrix mixture without collisional activation. I demonstrate that the process is efficient enough to allow recording of collision induced dissociation (CID) and metastable ion decomposition MS/MS spectra of any charge-remote [M-H2-(CH2)n]¯ fragments as well as of spectra of neutral losses. These new results indicate that a multi-step radical mechanism is involved in CRF ion formation. The first step of the process appears to be accompanied by hydrogen elimination that occurs randomly throughout the molecule. The primary fragment radical ions formed can decompose further with the formation of the next generation of CRF ions. In the last chapter I present our new discovery that FAB mass spectrometry in the negative ion mode can be used to unambiguously distinguish between cis- and trans- isomers of monounsaturated fatty acids by the relative strengths of two signals in the mass spectrum. I show the results of six fatty acids and demonstrate that under normal FAB ionization/desorption conditions, the deprotonated molecules [M-H]– of fatty acids undergo CRF. A characteristic fragmentation pattern of two intense clusters of mass peaks with three weak intervening peak-clusters can be used in each case to identify the position of the double bond. I also discuss the possibility of REC occurring during the FAB process.
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  • description.provenance : Submitted by Hong Ji (jih@onid.orst.edu) on 2007-10-05T06:27:19Z No. of bitstreams: 1 thesis.pdf: 3071233 bytes, checksum: a40bea50d5f0fa5e93d87fc035fd1e56 (MD5)
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