Magister Scientiae (Biodiversity and Conservation Biology) / Combating antimicrobial resistant infections caused by nosocomial pathogens poses a major public health problem globally. The widespread use of broad-spectrum antibiotics for the treatment of wound infections has led to the appearance of multidrug-resistant (MDR) microbes which further exacerbates the growth of microbes amongst patients. It may result in prolonged debility of the patient and an increase in healthcare costs due to prolonged hospital stays and expensive treatment regimens to avoid patient-patient transmission. Therefore, it is imperative that alternative sources of treatment to antimicrobial use in wound infections needs to be developed in order to inhibit or kill resistant microbes and to provide point of care medical treatment to the less fortunate at an affordable cost.

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In this work, we use optical approaches to study and provide mechanistic insight into electrochemical reactions occurring at the surface of single nanoparticles. Correlated optical-electrochemical studies offer several advantages over single nanoparticle electrochemical studies including, higher spatial resolution, the ability to interrogate many nanoparticles at the same time and identify populations of inactive nanoparticles. Throughout this dissertation, two optical techniques are discussed in detail, dark-field scattering and super-localization imaging. In the first set of experiments, we describe calcite-assisted localization and kinetics (CLocK) microscopy, a multiparameter super-localization imaging technique. By placing a rotating birefringent calcite crystal in the infinity space of an optical microscope, CLocK provides immediate polarization and orientation information while still maintaining the ability to localize a single nanoparticle with < 10 nm resolution. Additionally, we demonstrate that the CLocK point spread function encodes kinetic information that we quantified to be an order of magnitude shorter than the integration time of the camera. In this work, CLocK provides new mechanistic insight into dynamic processes such as the dissolution of single gold nanorods as well as single-molecule surface-enhanced Raman scattering. In the second work, dark-field scattering was employed to monitor a proposed post-synthesis silver nanoparticle surface cleaning strategy to improve homogeneity across a population. Here, a sacrificial silver-sulfide sulfide shell is chemically grown on single silver nanoparticles to outcompete surface impurities. We demonstrate that upon electrochemical removal of the shell, a more reactive and reproducible silver surface can be achieved as revealed by enhanced electrodissoluion of the freshly cleaned silver nanoparticles. In these experiments, we additionally found a sulfide-dependent formation of multiple sulfide-species as well as mixed character sulfide shells on single nanoparticles themselves, thus demonstrating the sensitivity provided by optical microscopy at identifying multiple surface chemistries. Overall, the work in this dissertation highlights the ability of optical tools at revealing heterogeneity in single particle studies providing insight into structure-function relationships. / Physiology

<p>    </p> <p>Surface functionalization of metallic nanoparticles is an attractive route to tailor the ensemble geometry and redox properties of active sites in heterogeneous catalysts. However, it is challenging to generate well-defined interfaces through conventional impregnation and one-pot colloidal synthesis methods. In this work, we utilize ligand exchange reactions for post synthetic surface modification of colloidal nanoparticles to generate unique core-shell and surface alloy structures. We use halometallate and metal chalcogenide complexes to create surface sites that are active for electrocatalytic hydrogen evolution reaction (HER). </p> <p>We synthesize a self-limiting monolayer of metal chalcogenides on colloidal Au nanoparticles through biphasic ligand exchange reaction between ammonium tetrathiomolybdate (NH₄)₂MoS₄ complex and Au nanoparticles. Through a combination of spectroscopy techniques and computational methods, we show that strong Au-S interactions introduce electronic and geometric distortion to the geometry and bond metrics of MoS₄^2-complex. Moreover, proximal MoS₄ units adsorbed on the Au surface interlink to form small MoSx oligomers with highly active bridging disulfide sites. Consequently, these core-shell AuMoS₄ nanoparticles exhibit significantly higher HER activity than MoS₄^2- supported on non-interacting carbon supports under highly acidic electrolyte conditions. Although post catalysis characterization reveals partial hydrolysis of surface adsorbed MoSx species, stable HER activity under bulk electrolysis condition indicates that active sites remain persistent. </p> <p>In an effort to extend these ligand exchange reactions to create metal/metal interfaces on other coinage metal nanoparticles such as Ag, we design metal-ligand coordination complexes to mitigate undesired galvanic replacement reactions. By varying the strength and number of coordinating ligands, we fine-tune the redox potential of oxidized noble metal precursors and confine the deposition of noble metals to a few surface layers of the Ag nanoparticles. We utilize organic amine and phosphine ligands to generate Ag@AgM core-shell nanoparticles, where M = Pd, Pt, and Au. Surface alloy or pure metal shells of Pd and Pt on Ag nanoparticles generated through this ligand-based strategy exhibited high precious metal atom utilization in electrocatalytic hydrogen evolution reaction. </p>

As demand for food increases, rapid testing methods are becoming increasingly important. In the past few years, yogurt has become popular. Yeast species are the most common spoilage organism, costing consumers and food companies money. A novel lateral flow assay has been developed to detect yeast oligonucleotide sequences. Gold nanoparticles were used as the standard reporter and fluorescent nanoparticles were developed as the novel reporter. The fluorescent nanoparticles were ruthenium-doped silica nanoparticles synthesized using the modified Stöber method. Visual analysis of assays using standard reporters showed the limit of detection to be 10 femtomoles of target sequence. Analysis of the fluorescent nanoparticles using a plate reader showed the limit of detection to be 0.027 femtomoles. The fluorescent reporter’s limit of detection is 1000 fold lower due to a sophisticated, more sensitive analysis method. Gold nanoparticles are appropriate for presence or absence testing, but fluorescent nanoparticles are best for obtaining quantitative data with low detection limits. Pathogens have been used as biological warfare for centuries. A brief review of common biowarfare agents is included. Yersinia pestis, the causative agent of the Plague, and Bacillus anthracis, the causative agent of Anthrax, are the focus. Additional work using gold nanoparticles as reporter in a sandwich assay is also included. The novel dye covered reporter was compared to the control, which was a single dye molecule linked to the reporter sequence. Repeated testing showed the novel reporter had a lower limit of detection and higher sensitivity due to increased ability to bind target.

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In this report we studied the properties of the dye IR806 and possible mechanisms of the dye sensitization effect on ytterbium-erbium co-doped upconversion nanoparticles. We found that the dye IR806 has two primary emission peaks in the NIR spectral range at around 850 nm and at around 950 nm. The intensity of these peaks were observed to be affected by the concentration of the dye and the addition of Gadolinium(III) chloride and Yttrium(III) chloride. Specifically increases in the intensity of the 950 nm peak relative to the 850 nm were of interest since ytterbium readily absorbs 950 nm and transfers this energy in the upconversion process. Our hypothesis is that the change in the intensity of the 850 nm and the 950 nm peak is associated with aggregation of the dye IR806 and the amount of monomers and dimers. Results from adding ytterbium-erbium co-doped upconversion nanoparticles in IR806-ethanol solution points to the picture of dimers being formed on the surface on the nanoparticles. This analysis is however based on the assumption that the 850 nm emission peak of IR806 is associated with monomers and that the 950 peak is associated with dimers, which is yet to be confirmed and further studies are therefore needed.

The ability of engineered cerium oxide nanoparticles to confer radioprotection was examined. Rat astrocytes were treated with cerium oxide nanoparticles to a final concentration of 10 nanomolar, irradiated with a single 10 Gy dose of ionizing radiation and cell death was evaluated by propidium iodine uptake at 24 and 48 hours after radiation insult. Treatment of rat astrocytes with nanoceria resulted in an approximate 3-fold decrease in radiation induced death. These results suggest that the nanoceria are conferring protection from radiation induced cell death. Further experiments with human cells were conducted. Human normal and tumor cells (MCF-7 and CRL8798) were treated with the same dosage of cerium oxide nanoparticles, irradiated and evaluated for cell survival. Treatment of normal cells (MCF-7) conferred nearly 99% protection from radiation-induced cell death while the same concentration of nanoceria showed almost no protection in tumor cells (CRL8798). TUNEL analysis results of similarly treated cells demonstrated that nanoceria reduced radiation-induced cell death by 3-fold in normal breast cells but not in MCF-7 tumor cell lines when cultured under the same conditions. We concluded that cerium oxide nanoparticles confer radioprotection in a normal human breast line (CRL 8798) but not in a human breast tumor line (MCF-7). It is hoped that the outcome of this study will guide future endeavors toward a better elucidation of the molecular pathways involved in the protection of cells with nanoceria against radiation-induced cell death, as well as the minimization of the bystander effect in radiation therapy.