<p> This dissertation details original work focused on the DNA adduct N2-(1-carboxyethyl)-2'-deoxyguanosine (CEdG). This DNA adduct results from the spontaneous reaction of DNA with the endogenous and exogenous formed, carbohydrate-derived, reactive carbonyl species, methylglyoxal. Using <i>in vitro</i> steady state kinetics, we have shown that CEdG in template DNA leads to DNA miscoding effects when the model replicative polymerase, exonuclease-free Klenow fragment (KF-) is used. The development, validation and application of a novel stable isotope dilution, triple quadrupole mass spectrometric method for the quantitation of CEdG is also detailed. This method was used to quantitate CEdG in urine from diabetic rats, urine from human patients, human tumor and adjacent biopsy tissue, diabetic animal tissue and DNA treated with methylglyoxal. Finally, we detail the adaptation, validation and application of a novel, commercially-available microfluidic HPLC-chip for increased sensitivity in the quantitation of CEdG and also apply it to the quantitation of the RNA analogue, CEG. Combined, these studies establish CEdG as a potential biomarker for glycation and point to a viable avenue for connecting chronic glycolytic flux with genetic instability. </p>

This thesis develops and applies size exclusion chromatography (SEC) for the accurate characterization and size separation of semiconductor and metallic nanoparticles. Size exclusion chromatography is employed for the first time to determine the size and size distribution of cadmium selenide nanocrystals. The method accurately determines nanocrystal size as well as the length of any surface capping ligands. The separation resolution is enhanced using recycling SEC and a shape separation of quantum dots and multipods is performed. SEC is further applied to measure the hydrodynamic diameter of gold and cadmium selenide nanocrystals passivated with polystyrene. Polystyrene occupies a brush conformation on the surface of these nanoparticles and the polymer length is found to scale linearly with molecular weight and as the cube root of polymer coverage. Using these measurements, a general formula is proposed to predict the hydrodynamic diameter of polymer coated nanoparticles. Gold and cadmium selenide nanocrystals discretely functionalized with different numbers of polystyrene molecules are also separated. Fluorescence and 1H-NMR measurements of collected fractions confirm the separation and the population of the species is studied as a function of polystyrene concentration. These discrete nanocrystal-polymer conjugates form ordered microphases upon drying that are dependent upon the tether number. Such data illustrates the potential of these species as building blocks for self-assembly.

Plasmon resonances in metal nanoparticles control the far field and near field optical properties of these metallic structures. The enhanced electromagnetic near field is strongest at the surface of the nanoparticles and rapidly decays away from the surface. This enhanced near field is exploited in surface enhanced spectroscopies including Surface Enhanced Raman Spectroscopy (SERS) and Metal Enhanced Fluorescence Spectroscopy (MEFS). A measurement of the decay profile of the fringing field is important both for further development of surface enhanced spectroscopy for sensor device application, and for understanding from a fundamental physics point of view. Gold nanoshells are spherical colloidal nanoparticles with a silica core covered by a thin gold shell. The plasmon resonance of nanoshells can be controllably tuned in the visible and infrared parts of the spectrum. The near field profile of nanoshells can be theoretically calculated on the basis of Mie scattering theory. The thesis describes a series of experiments designed to experimentally verify the near field profile of nanoshells. A scaffold of ss-DNA is used to place a fluorescein dye molecule at varying distances from the nanoshell surface. The SERS intensity from both the scaffold molecules and the fluorescein placed at the end of the tether is measured simultaneously and self consistently. The fluorescein-ss-DNA nanoshell conjugate structures are also used to study the distance dependence of the fluorescence emission from fluorescein. The thesis discusses the results of the SERS intensity profile agreement with the intensity profile calculated using Mie scattering theory. The quenching and enhancement of the fluorescence emission at varying distances from the nanoshell surface are also discussed.

With the ever growing applications of nanomaterials, fast, accurate and cost effective characterization methodologies are needed to ensure high quality materials and practical manufacturing. This problem is a challenging one as nanoparticle shape, size and size distribution are all important features which govern properties as diverse as sample melting point and chemical reactivity. New methods that provide key analytical information, along with a means of improving size and shape distributions would have substantial impact on this growing area. Chromatography is a technique which traditionally has been used to both provide analysis as well as separation of high valued added chemicals. Its applications in nanoscience are to date quite limited, and the objective of this thesis is to demonstrate that chromatographic techniques can be used to measure the dimensions of gold nanocrystals in solution as well as provide methods for physically separating complex distributions of particles. Recycling size exclusion chromatography (RSEC) has been employed to effect physical, base-line separation of subpopulations of spherical gold nanocrystals. This method can be easily applied on other nanocrystalline systems and is scalable to large production. Anisotropic gold nanocrystals, such as gold nanorods, exhibit spectacular optical properties that can be utilized in medical and biological fields. An analytical method, based on aqueous SEC that determines the dimensions of gold nanorods has been developed. The method couples the hydrodynamic volume obtained from SEC and aspect ratio from absorption spectra to provide a complete assessment of nanorod ensembles directly in liquid phase media. Additionally, a close analysis of the hydrodynamic values reveals that the nanorods are aligned with the flow, which makes the separation responsive to variations in aspect ratio. Finally, conventional chromatography provides a resolution which is not large enough to sharpen the distributions of the most monodisperse nanoparticles now produced directly in solution. The application of super high-resolution methods, such as critical point chromatography, can substantially improve the separation power and its application to polymer stabilized gold nanocrystals has been demonstrated.

The high-resolution structures of two separate RNA molecules were determined using heteronuclear nuclear magnetic resonance (NMR) spectroscopy. Both RNA molecules, in their respective systems, perform critical functions of regulating gene expression. To explore the dynamic properties of one molecule, an extensive investigation into its fast (&lt;5ns) intra-molecular motions was carried out. The dynamics investigation was performed using a novel application of heteronuclear relaxation measurements from several base and ribose sites within the molecule. The results of this work were then interpreted using various forms of the Lipari and Szabo motional model. The significance of this research is two fold: (1) it provides detailed structures of two RNA molecules and a basis for their roles in protein recognition and binding and (2) augments the structural characterization with direct measurements of fast internal motions using a novel application of heteronuclear NMR relaxation analysis.

No description available.

The development of a framework for the automated analysis of inductively couple plasma atomic emission spectroscopy is present. Some of the research that lead to current state of this framework is presented. A small expert system that uses information about the current sample to generate a line search strategy which minimizes the number of emission lines which need to be measured, and avoids spectral overlaps when possible. A program is presented that evaluates the minimum number of spectral windows required to perform elemental analysis by ICP-AES, given a certain spectral window width. A method with the potential for rapidly ascertaining the physical properties of the sample matrix is presented. This system has the potential to help reduce sample introduction related system failures. Finally, three optimization algorithms are compared in their ability to optimize ICP-AES performance, from this an optimization module was developed for inclusion in the automated analysis framework.

The solubility of Zr-bearing minerals in peralkaline, H2O-saturated alkali aluminosilicate melts with or without added F reaches a maximum of about 4 and 3.5 wt.% ZrO2 respectively at approximately 57 wt.% SiO2. The saturating phase for melts with SiO2 content above this threshold is zircon. In the halogen-free experiments, the saturating phase for melts with lower silica content is wadeite (K2ZrSi 3O9), while ZrO2 crystallizes from melts with 1 wt.% added F. Experiments with Cl-bearing melts indicate no maximum solubility of Zr minerals; the solubility remains at 2 to 2.2 wt.% ZrO2 across a wide compositional range. The saturating phases in the Cl-bearing melts are the same as those of the F-bearing melts and the transition from ZrO 2 to zircon occurs at the same composition. The positive slope of the wadeite saturation curve on XZrO2-XSiO2 plots for the halogen-free melts suggests that increasing SiO2 activity increases the solubility of wadeite. / Peralkaline Ti,Zr-bearing sodium aluminosilicate glasses have been analyzed by Raman spectroscopy to determine the effect of Cl on glass structure. The spectra of the Ti-bearing glasses show a significant difference between the Cl-free and the Cl-bearing composition. The Cl-free glass spectrum contains a strong, asymmetric peak at 900 cm--1 which is associated with Ti in five-fold coordination. This peak is shifted to higher frequency and becomes more symmetric with the addition of 0.3 wt.% Cl. Deconvolution of the high-frequency waveband suggests that differences between spectra result from the contribution of a peak at 945 cm--1. This peak is believed to be the result of Ti-O vibrations in fully-polymerized titanate tetrahedra. It is proposed that the addition of Cl destabilizes [5] Ti in favour of tetrahedral coordination as a result of competition between Cl and titanate groups for alkalis. / 23Na NMR MAS analyses of a suite of Na-aluminosilicate glasses with Na/Al = 2 and varying SiO2 content has revealed a trend toward more negative chemical shift (greater shielding of the nucleus) as the glass structure becomes more polymerized, i.e. the average number of non-bridging oxygen atoms per tetrahedron (NBO/T) decreases. This trend is observed only for glasses with NBO/T &le; 0.3. For more polymerized glasses no change in chemical shift is measured. (Abstract shortened by UMI.)

Surface Enhanced Raman Scattering (SERS) is a process by which molecules near certain special metal surfaces exhibit Raman scattering that is measurably more intense than is the normal Raman scattering exhibited by molecules in the absence of the surface. The majority of SERS-active metal substrates, such as roughened electrodes and colloidal sols, are fabricated from coinage metals, Au, Ag and Cu. The principal subject of this thesis is the surface enhanced Raman spectroscopic study of the adsorbates; pyrazinamide, 2- and 4-mercaptopyridine. A range of SERS-active Ag substrates was used, viz., Ag colloids, metal liquid-like films (MELLFs), and roughened Ag electrodes. In addition, an original technique that encompasses both SERS and waveguide Raman spectroscopy (WRS), known as integrated optics, evanescent wave, surface enhanced Raman spectroscopy (IOEW-SERS) was developed. For this technique, a waveguide heterostructure was fabricated on nano-scale dimensions by self-assembly of silver colloidal particles on thin glass slides. The unique optical behaviors of both SERS and WRS systems are combined such that optically guided light propagates in the thin film, with an evanescent wave and couples with surface plasmon modes of metal particles. A comparative SERS study of 2- and 4-mercaptopyridine (MPy) was undertaken using all of the above methods. The purpose of this comparison was to determine the similarities between IOEW-SERS and other Ag SERS-active systems. These studies were extended by using a 4-MPy modified SERS-active optical waveguide as a thin film chemical sensor. Intermolecular interactions between probe ions, Cu$ sp{2+}$ and H$ sp+,$ with 4-MPy were observed by vibrational perturbations to the IOEW-SERS spectrum. X-ray photoelectron spectroscopy (XPS) allowed a layer-by-layer examination of the 4-MPy derivatized waveguide from the substrate to the Cu$ sp{2+}$ ion. A potential dependent SERS study of 4-MPy modified Ag electrodes, including the interact

In this study the effectiveness of quantitative 31P nuclear magnetic resonance (NMR) spectroscopy in conjunction with spin trapping to detect and quantify various free radical species was investigated. Initially the research was focussed on assigning the 31P NMR signals for hydroxyl and superoxide radicals and on identifying the quantitative reliability of the technique. The radical adduct reaction products (diamagnetic species), of hydroxyl (HO·) and superoxide (O2 -·) radicals with phosphorus containing nitroxides, were determined to be relatively stable and as a result the 31P NMR chemical shifts for these species were determined. Their structures were verified and quantified using 31P NMR in the presence of a phosphorus containing internal standard. The 31P NMR chemical shifts for 5-diisopropylphosphoryl-5-methyl-1-pyrroline-N-oxide (DIPPMPO) and 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) spin traps with a variety of carbon centered radicals were also determined. / A novel equilibrium reaction involving molecular oxygen and alkaline hydrogen peroxide was investigated and it was found that the concentration of superoxide radicals present in a system of alkaline hydrogen peroxide is affected by the nature of gas with which it is in equilibrium. The concentration of superoxide radicals was determined to be significantly larger than that of hydroxyl radicals at elevated pH values. / Experiments have shown that NMR in conjunction with spin trapping can be a valuable tool for the detection and quantification of radical species.