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New Ionization Methods for the Analysis and Imaging of Microbial Populations and Biological Tissues by Mass Spectrometry Open Access

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To evaluate the complex and diverse biochemical processes within organisms, continued advances in chemical analysis are needed for the rapid detection, identification and quantitation of biomolecules. Due to its exquisite sensitivity and specificity, mass spectrometry (MS) based methods have become an indispensable tool for the analysis and spatial mapping of metabolites, lipids, peptides, and xenobiotics. This dissertation describes my efforts in advancing the analysis of biological samples, cell populations, and tissue sections using MS-based methods based on direct laser sampling and photonic nanostructures. Chapter 1 provides background information on well established and emerging ambient and laser desorption ionization mass spectrometric methods for the analysis of small molecules. The emphasis is on high-throughput analysis directly from microorganisms, matrix-free molecular imaging from biological tissue sections, and enhancement of ionization by matrix-free nanostructure arrays. Chapter 2 describes using laser ablation electrospray ionization with ion mobility separation (LAESI-IMS)-MS as a high-throughput method for the metabolic and lipidomic profiling of genetically modified algae, as well as for exploring environmental perturbations for increased lipid production. Benefits of IMS include aiding in lipid classification and identification by assigning unique drift times to lipid classes and to the degree of saturation, as well as helping structure elucidation by IMS linked to tandem-MS. Chapter 3 presents pulse-chase analysis in combination with LAESI-IMS-MS to determine metabolite, lipid, and peptide turnover rates directly from algal cell populations. Here IMS was used to help identify the nitrogen isotopologues in which the relaxation of isotope distribution patterns to natural abundances was monitored as a function of time. Chapter 4 explores the symbiotically induced metabolites involved in the interaction between legumes and soil rhizobia by LAESI-IMS-MS. By laser ablation sampling, the spatial mapping of biomolecules was demonstrated on whole soybean root nodules by depth profiling through the different anatomical regions of the nodule. Chapter 5 demonstrates the feasibility of vacuum-based matrix-free MS imaging by laser desorption ionization (LDI) from silicon nanopost arrays (NAPA)through determining the spatial distributions of biomolecules in biological tissues and small cell populations. The mechanism of ion formation and material removal from thin tissue sections was explored using electron microscopy. Chapter 6 introduces a new nanophotonic LDI MS platform, elevated bowtie (EBT) arrays, that exhibits enhanced near-field effects, higher ion intensities, as well as greater control over fragmentation for small molecules and peptides. The effect of the bowtie antenna apex angle on ion production and near-field enhancement was also evaluated. Chapter 7 surveys the current state of the MS field for emerging ambient- and vacuum- based ionization technologies with a focus on overcoming their major challenges and providing future improvements. Future directions were explored with emphasis on potential applications of these MS-based technologies.

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