Theses and Dissertations from UMD
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New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM
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Item Towards the synthesis of PNAG crosslinkers to identify protein binding partners(2019) Mrugalski, Kevin R; Poulin, Myles; Chemistry; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Bacterial biofilms are an area of major concern in the medical field due to natural drug resistance. Many pathogenetic species of bacteria that infect humans including Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Vibrio cholera form biofilms and their associated infections are becoming harder to treat. Poly β-(1→6)-N-acetyl-D-glucosamine (PNAG) is a major component of biofilms across multiple species and has been found to play a key role in the early stages of the biofilm life-cycle. However, little information is known about what proteins interact with this important polysaccharide. Our goal is to create small PNAG analogues to covalently capture and identify PNAG binding partners in E. coli, an important model organism. PNAG analogues will contain photoaffinity groups, that when activated, covalently link associated proteins to the probe. Then, using a proteomics-mass spectrometry-based approach, we will identify PNAG binding partners. Here, we describe the efforts and challenges encountered synthesizing the final PNAG probes. New synthetic routes are proposed based on literature precedent that will enable synthesis of the desired compounds.Item Bio-Inspired Polymer Microparticles for Targeted Recognition and Response(2014) Arya, Chandamany; Raghavan, Srinivasa R.; Chemical Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Microbeads and microcapsules are container structures that are frequently used in biomedical applications. In this dissertation, we have sought to impart new functionalities to these particles, inspired by phenomena observed with biological cells. We have engineered polymer microparticles that recognize and respond to specific species from the surroundings (e.g. cells, polymer chains, metal ions). Three classes of new microparticles are reported, which are each reminiscent of a different type of biological cell in terms of recognition capabilities and response. In the first part of this dissertation, we create functionalized microbeads from the biopolymer, chitosan, and use these to selectively recognize and capture Circulating Tumor Cells (CTCs) from blood. The microbeads are functionalized with a protein (streptavidin) and packed into an array within a microfluidic device. Blood samples with biotin-labeled CTCs are flowed over the packed bed of chitosan beads. Similar to how macrophages adhere to foreign bacteria (i.e. antibody-antigen interactions), the streptavidin-labeled chitosan beads can selectively recognize and adhere to the biotin-labeled CTCs. We show that such a packed bed of chitosan beads could serve as an inexpensive platform for customized capture of different rare cells (cancer cells, stem cells etc) from blood. In the next study, we develop a class of microbeads that undergo clustering (aggregation) in the presence of specific polymers. The inspiration for this comes from the cells (e.g., platelets) and polymers involved during the formation of blood clots. Our system consists of chitosan microbeads coated with cyclodextrins (sugar molecules with a hydrophobic binding pocket), which are then exposed to a polymer that is decorated with hydrophobic units. The particles bind to the polymer chains via hydrophobic interactions and in turn, the particles are induced to form clusters. Subsequently, the polymer precipitates and forms a matrix around the particle clusters, leading to a structure that is reminiscent of a blood clot (platelets enveloped by a mesh of fibrin chains). Lastly, we develop a class of microparticles that have the ability to selectively destroy other microparticles. The inspiration here is from the body's immune system, where cells like the killer T cells selectively destroy cancer and virus infected cells without harming healthy cells. Towards this end, we synthesize two types of microparticles: chitosan capsules that contain the enzyme glucose oxidase (GOx), and beads of a different biopolymer, alginate that are crosslinked with copper (Cu2+) ions. The chitosan capsules enzymatically convert glucose from the surroundings into gluconate ions. When these capsules approach the alginate/copper beads, the gluconate ions chelate the copper ions, leading to the disintegration of the alginate beads. Other beads that do not contain Cu2+ are not affected in this process.Item Optimization of xanthan chitosan polyelectrolytic hydrogels for microencapsulation of probiotic bacteria(2011) Soma, Pavan Kumar; Lo, Martin; Food Science; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The effectiveness of microencapsulation system for targeted delivery of probiotics depends on its ability to protect cells from harsh gastrointestinal conditions of stomach followed by effectively releasing the cells in intestinal conditions. Oppositely charged xanthan and chitosan form stable polyelectrolytic hydrogels capable of encapsulating enzymes and cells. The present study aims at developing an effective microencapsulation system for probiotics by screening and optimizing the factors critical to xanthan-chitosan hydrogel (XCH) capsule formation. The changes in the core pH of the hydrogel capsule in response to simulated gastric juice (SGJ) were characterized. Increase in xanthan concentration and chitosan molecular weight improved the barrier properties, however, increasing complexation time beyond 40 min had the opposite effect. Increase in molecular weight of chitosan resulted in improved viability of probiotic bacteria, Lactobacillus acidophilus, after SGJ treatment, which could be attributed to the differences in hydrogel membrane thickness at the surface of capsule, as evidenced by scanning electron micrographs (SEM). Introducing XCH capsules made with high molecular weight (HMW) chitosan into xanthan solution resulted in the formation of xanthan-chitosan-xanthan hydrogel (XCXH) capsules. Unlike HMW and medium molecular weight (MMW) chitosan, low molecular weight (LMW) chitosan did not form the outer layer beyond XCH, suggesting the significance of chitosan molecular weight in the formation of XCXH. The increased hydrogel thickness of XCXH capsules formed with HMW chitosan compared to XCH capsules rendered better retention of cells in SGJ treatment for a longer period of time, further suggesting the importance of membrane thickness on the hydrogel stability and its barrier properties. Furthermore, complete release of cells from XCXH in simulated intestinal fluid (SIF) was extended by approximately an hour compared to XCH capsules. Smaller, nozzle-sprayed XCXH capsules using HMW chitosan protected probiotic bacteria in SGJ albeit one-log reduction in its protective efficacy compared to syringe extruded capsules. When incorporated into stirred yogurt, XCXH microcapsules improved the viability of L. acidophilus by ~1 log CFU/ml between 15 and 30 days of storage. The stability of bacteria against bile salts was significantly improved, enabling the delivery of prescribed number of cells to attain the claimed health benefits.Item EFFECT OF CHITOSAN ON THE INDUCTION OF DNA DAMAGE RESPONSE BY SELENIUM COMPOUNDS.(2009) Zhang, Shu; Cheng, Wen-Hsing; Nutrition; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Selenium (Se), a nutrient trace mineral, plays important roles in optimizing human health. Chitosan is an effective, natural-oriented material for synthesizing nanopolymers, with preferable properties such as biocompatibility, biodegradation and resistance to certain enzymes. In this study, encapsulated Na2SeO3 and methylseleninic acid (MSeA) with low and medium molecular weight chitosan were used to determine the efficacy of Se in mitigating tumorigenesis. We applied Se compounds, which is from sub-lethal to lethal dose, to colon cancer cell line HCT-116 and normal fibroblasts cell line MRC-5. Analysis of cellular selenium content demonstrated that: 1) Na2SeO3, but not MSeA, treatment resulted in a greater Se retention in HCT-116 than in MRC-5 cells, 2) chitosan encapsulation enhanced Se contents in cells treated with the various Se preparations. Cell survival analysis showed that chitosan encapsulation protected HCT-116 and MRC-5 cells from Na2SeO3 or MSeA induced toxicity. Moreover, this beneficial effect was greater in MRC-5 cells. MSeA encapsulated with chitosan induced phosphorylated ATM Ser-1981 formation in MRC-5 and HCT-116 cells to a less extent as compared to MSeA alone treatment. Taken together, the results suggest that, when encapsulated with chitosan, cells are less susceptible to Se treatment, possibly through a mechanism by which the presence of chitosan attenuates Se-induced activation of ATM and corresponding DNA damage response pathway.Item Characterization of Electrodeposited Chitosan: an Interfacial Layer for Bio-assembly and Sensing(2009) Buckhout-White, Susan Lynn; Rubloff, Gary W; Material Science and Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Microfluidics and Lab-on-a-Chip devices have revolutionized the field of analytical biology. To fully optimize the potential of the microfluidic environment it is critical to be able to isolate reactions in specific locations within a channel. One solution is found using chitosan, an amine-rich biopolymer with pH responsive solubility. Induction of hydrolysis at patterned electrodes within the fluidic channel provides a means to spatially control the pH, thus enabling biochemical functionalization that is both spatially and temporally programmable. While chitosan electrodeposition has proven to be reliable at producing films, its growth characteristics are not well understood. In situ optical characterization methods of laser reflectivity, fluorescence microscopy and Raman spectroscopy have been employed to understand the growth rate inter diffusion and lateral resolution of the deposition process. These techniques have also been implemented in determining where a molecule bound to an amine site of the polymer is located within the film. Currently, electrodeposited chitosan films are primarily used for tethering of biomolecules in the recreation of metabolic pathways. Beyond just a biomolecular anchor, chitosan provides a way to incorporate inorganic nanoparticles. These composite structures enable site-specific sensors for the identification of small molecules, an important aspect to many Lab-on-a-Chip applications. New methods for creating spatially localized sites for surface enhanced Raman spectroscopy (SERS) has been developed. These methods have been optimized for particle density and SERS enhancement using TEM and Raman spectroscopy. Through optimization, a viable substrate with retained chitosan amine activity capable of integration into microfluidics has been developed.Item An Optical MEMS Sensor for On-chip Catechol Detection(2008-12-08) Dykstra, Peter Hume; Ghodssi, Reza; Electrical Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)This thesis reports the successful design, fabrication and testing of an optical MEMS sensor for the detection of the toxic phenol, catechol. Catechol's presence in food and drinking water posses a health concern due to its harmful effects on cell respiration. By-products of catechol oxidation have demonstrated increased absorbance changes in a chitosan film in the UV and near UV range. Our reported sensor utilizes patterned SU-8 waveguides and a microfluidic channel to deliver catechol samples to an electrodeposited chitosan film for absorbance measurements at 472 nm. Concentrations as low as 1 mM catechol are detected while control experiments including ascorbic acid display no measurable response. By using optical detection methods, our device does not suffer from many of the problems which plague conventional electrochemical based sensors.