On-tissue chemical derivatization: Improving metabolic detection for MALDI-MS imaging

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Forsman, Trevor Thomas
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Lee, Young Jin
Anderson, Jared
Potoyan, Davit
Smith, Emily
Gundlach-Graham, Alexander
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Mass spectrometry imaging (MSI) provides chemical and spatial information of molecules in complex samples such as animal and plant tissue. Some molecules of low abundance are lost in MSI analysis, especially those that ionize poorly. On-tissue chemical derivatization (OTCD) is a method that improves analyte signal by chemically binding a poorly ionizing analyte with a molecule that either ionizes well or has a permanent positive charge. Herein this dissertation, further developments in derivatization chemistry are made. In the first chapter, background is provided for MSI and OTCD, and the final chapter provides conclusions and future directions for the works discussed. The second chapter outlines a method to improve the signal of vicinal diol compounds in MSI. Vicinal diols suffer from poor ionization efficiency, although they have biological significance with compounds such as catecholamines and sugars. An on-tissue chemical derivatization method utilizing 4-dimethylaminophenyl boronic acid (DBA) achieved signal improvements for vicinal diol compounds. In addition, a side reactivity screening of DBA was made, revealing reactivity with phenols in vicinal position to carboxylic acids, and carboxylic acids. This method was applied to maize tissue samples where dozens of chemically modified metabolites were tentatively identified using CornCyc and Metlin databases. The third chapter examines the side reactions for three amine-targeting OTCD reagents: coniferyl aldehyde, 5-hydroxy-2-nitrobenzaldehyde, and tetramethylpyryllium tetrafluoroborate. Side reactions can lead to false annotations in untargeted and semi-targeted MSI analysis. Through monitoring reactions between reagents and standards, side reactions that were likely to annotate were determined. These side reaction mechanisms were then elucidated through MS/MS. The three OTCD reactions were performed on maize root tissue sections to examine the effects of side reactions on chemical annotations. The fourth chapter proposes a method to observe volatile metabolites in MALDI-MSI. Volatile metabolites vaporize in MALDI-MSI analysis due to sample vacuum induction during sample preparation and due to the vacuum of the ionization source. To alleviate the loss of volatiles, a precoating derivatization strategy is adapted to display the stabilization of volatiles through chemical modification. Short-chain fatty acids were analyzed using the precoating derivatization method in chicken cecum, ileum, and jejunum tissue sections.
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