Development of online microdialysis-microchip system for in vivo monitoring
Issue Date
2009-08-31Author
Nandi, Pradyot
Publisher
University of Kansas
Format
193 pages
Type
Dissertation
Degree Level
Ph.D.
Discipline
Pharmaceutical Chemistry
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This item is protected by copyright and unless otherwise specified the copyright of this thesis/dissertation is held by the author.
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Show full item recordAbstract
Microdialysis is a sampling technique that can be employed to monitor biological events in vivo and chemical reactions in vitro. When it is coupled to an analytical system, microdialysis can provide near real time information on the time dependent concentration changes of analytes in the extracellular space or other aqueous environments. Online systems for the analysis of microdialysis samples enable fast, selective and sensitive analysis while preserving the temporal information. Analytical methods employed for on-line analysis include liquid chromatography (LC), capillary (CE) and microchip electrophoresis and flow-through biosensor devices. This dissertation is focused on the development of microchip systems coupled to microdialysis sampling for online near-real time monitoring with high temporal resolution for the analysis of amino acid neurotransmitters in vivo. Fluorescence detection was utilized for all the studies. Also, naphthalene-2,3-dicarboxaldehyde (NDA) and b-mercaptoethanol (b-ME) or CN- are used as the derivatization reagents for rendering the amino acids fluorescent. Initial studies were performed to evaluate a twin-t design with a 3 cm separation channel that incorporated on-column derivatization with NDA and b-ME. Biogenic amines and peptides were separated on chip demonstrating the feasibility of online microdialysis sampling with online derivatization and analysis in vitro. Subsequently, the chip was further modified by incorporating a 20 cm serpentine separation channel and excitatory amino acid neurotransmitters (glutamate, aspartate) were separated from other analytes in rat brain microdialysis samples. Next, precolumn derivatization was incorporated into the chip and online in vivo experiments were performed for the continuous analysis of glutamate from rat brain microdialysis sample. Also, fluorescein was included in the experiment for the possibility to simultaneously monitor the permeability of the blood brain barrier (BBB) along with changes in levels of excitatory neurotransmitters during online analysis applied to the rat stroke model. The chip was further modified and optimized for experiments that require fast injections and separations thereby attaining high temporal resolution. This work described an online microdialysis-microchip system that can be used for monitoring neurochemical events that involve fast changes in analyte concentration in the brain.
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