Doctor of Philosophy / Department of Chemistry / Christopher T. Culbertson / Analytical separations form the bulk of experiments in both research and industry. The choice of separation technique is governed by the characteristics of the analyte and purpose of separation. Miniaturization of chromatographic techniques enables the separation and purification of small volume samples that are often in limited supply. Capillary electrophoresis and immunoaffinity chromatography are examples of techniques that can be easily miniaturized with minimum loss in separation efficiency. These techniques were used in the experiments presented in this dissertation. Chapter 1 discusses the underlying principles of capillary electrophoresis and immunoaffinity chromatography. In the second chapter, the results from immunoaffinity chromatography experiments that utilized antibody-coated magnetic beads to purify serine proteases and serine protease inhibitors (serpins) from A. gambiae hemolymph are presented and discussed. Serine proteases and serpins play a key role in the insect innate immunity system. Serpins regulate the activity of serine proteases by forming irreversible complexes with the proteases. To identify the proteases that couple to these serpins, protein A magnetic beads were coated with SRPN2 antibody and then incubated with A. gambiae hemolymph. The antibody isolated both the free SRPN2 and the SRPN2-protease complex. The purified proteases were identified by ESI-MS from as few as 25 insects. In Chapter 3, an integrated glass/PDMS hybrid microfluidic device was utilized for the transportation and lysis of cells at a high throughput. Jurkat cells were labeled with 6-CFDA (an internal standard) and DAF-FM (a NO specific fluorophore). Laser-induced fluorescence (LIF) detection was utilized to detect nitric oxide (NO) from single Jurkat cells. The resulting electropherograms were used to study the variation in NO production following stimulation with lipopolysaccharide (LPS). 3 h LPS-stimulation resulted in a two fold increase in NO production in both bulk and single cell analysis. A comparison of bulk and single cell NO measurements were performed and the average NO production in single cells compared well to the increase measured at the bulk cell level. Chapter 4 discusses the preliminary experiments with a T-shaped microfluidic device that exploit the property of poly(dimethylsiloxane) (PDMS) as an electroactive polymer (EAP), to enhance fluid mixing. EAPs deform when placed in an electric field. A thin layer of PDMS was sandwiched between chrome electrodes, positioned on the horizontal arms of the T design, and the electrolyte-filled fluidic channel. A potential difference across the PDMS layer caused it to shrink and stretch, thereby increasing the channel volume. The electrodes were actuated at 180[degrees] out of phase and this caused the fluid stream in the vertical channel to fold and stretch resulting in enhanced contact surface area and shorter diffusion distances of the fluid, thereby improving mixing efficiency. All the experiments presented in this dissertation demonstrate the application of miniaturized chromatographic techniques for the efficient analysis of small volume biological samples.

Doctor of Philosophy / Department of Chemical Engineering / Keith L. Hohn / Superparamagnetic iron oxide nanoparticle functionalization is an area of intensely active research, with applications across disciplines such as biomedical science and heterogeneous catalysis. This work demonstrates the functionalization of iron oxide nanoparticles with a quasi-monolayer of 11-sulfoundecanoic acid, 10-phosphono-1-decanesulfonic acid, and 11-aminoundecanoic acid. The carboxylic and phosphonic moieties form bonds to the iron oxide particle core, while the sulfonic acid groups face outward where they are available for catalysis. The particles were characterized by thermogravimetric analysis (TGA), transmission electron microscopy (TEM), potentiometric titration, diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS), inductively coupled plasma optical emission spectrometry (ICP-OES), X-ray photoelectron spectrometry (XPS), and dynamic light scattering (DLS). The sulfonic acid functionalized particles were used to catalyze the hydrolysis of sucrose at 80˚C and starch at 130˚C, showing a higher activity per acid site than the traditional solid acid catalyst Amberlyst-15, and comparing well against results reported in the literature for sulfonic acid functionalized mesoporous silicas. In sucrose catalysis reactions, the phosphonic-sulfonic nanoparticles (PSNPs) were seen to be incompletely recovered by an external magnetic field, while the carboxylic-sulfonic nanoparticles (CSNPs) showed a trend of increasing activity over the first four recycle runs. Between the two sulfonic ligands, the phosphonates produced a more tightly packed monolayer, which corresponded to a higher sulfonic acid loading, lower agglomeration, lower recoverability through application of an external magnetic field, and higher activity per acid site for the hydrolysis of starch. Functionalizations with 11-aminoundecanoic acid resulted in some amine groups binding to the surfaces of iron oxide nanoparticles. This amine binding is commonly ignored in iron oxide nanoparticle syntheses and functionalizations for biomedical and catalytic applications, affecting understandings of surface charge and other material properties.

Doctor of Philosophy / Department of Chemistry / Stefan H. Bossmann / Pancreatic cancer is the fourth leading cause of cancer-related deaths in the USA. Asymptomatic early cancer stages and late diagnosis leads to very low survival rates of pancreatic cancers, compared to other cancers. Treatment options for advanced pancreatic cancer are limited to chemotherapy and/or radiation therapy, as surgical removal of the cancerous tissue becomes impossible at later stages. Therefore, there's a critical need for innovative and improved chemotherapeutic treatment of (late) pancreatic cancers. It is mandatory for successful treatment strategies to overcome the drug resistance associated with pancreatic cancers. Nanotechnology based drug formulations have been providing promising alternatives in cancer treatment due to their selective targeting and accumulation in tumor vasculature, which can be used for efficient delivery of chemotherapeutic agents to tumors and metastases. The research of my thesis is following the principle approach to high therapeutic efficacy that has been first described by Dr. Helmut Ringsdorf in 1975. However, I have extended the use of the Ringsdorf model from polymeric to nanoparticle-based drug carriers by exploring an iron / iron oxide nanoparticle based drug delivery system. A series of drug delivery systems have been synthesized by varying the total numbers and the ratio of the tumor homing peptide sequence CGKRK and the chemotherapeutic drug doxorubicin at the surfaces of Fe/Fe₃O₄-nanoparticles. The cytotoxicity of these nanoformulations was tested against murine pancreatic cancer cell lines (Pan02) to assess their therapeutic capabilities for effective treatments of pancreatic cancers. Healthy mouse fibroblast cells (STO) were also tested for comparison, because an effective chemotherapeutic drug has to be selective towards cancer cells. Optimal Experimental Design methodology was applied to identify the nanoformulation with the highest therapeutic activity. A statistical analysis method known as response surface methodology was carried out to evaluate the in-vitro cytotoxicity data, and to determine whether the chosen experimental parameters truly express the optimized conditions of the nanoparticle based drug delivery system. The overall goal was to optimize the therapeutic efficacy in nanoparticle-based pancreatic cancer treatment. Based on the statistical data, the most effective iron/iron oxide nanoparticle-based drug delivery system has been identified. Its Fe/Fe₃O₄ core has a diameter of 20 nm. The surface of this nanoparticle is loaded with the homing sequence CGKRK (139-142 peptide molecules per nanoparticle surface) and the chemotherapeutic agent doxorubicin (156-159 molecules per surface), This nanoplatform is a promising candidate for the nanoparticle-based chemotherapy of pancreatic cancer.

Master of Science / Department of Chemistry / Jun Li / With the increasing consumption of energy and the depletion of fossil fuels, finding an alternative energy source is critical. Solar energy is one of the most promising energy sources and solar cells are the devices that convert solar radiation into electricity. Currently, the most widely used solar cell is based on p-n junction formed with crystalline silicon materials. While showing high efficiency, the high fabrication cost limits its broad applications. Dye sensitized solar cell (DSSC) is a promising low-cost alternative to the Si solar cell, but its efficiency is much lower. Improvements in materials and interfaces are needed to increase the DSSC efficiency while maintain the low cost. In this thesis, three projects were investigated to optimize the DSSC efficiency and reduce the cost. The first project is to optimize the TiO[subscript]2 barrier layers on Fluorine-doped Tin Dioxide (FTO) surface. Two preparation methods, i.e. TiCl[subscript]4 solution treatment and thermal oxidation of sputtered Ti metal films, were employed and systematically studied in order to minimize electron-hole recombination and electron backflow during photovoltaic processes of DSSCs. TiCl[subscript]4 solution treatment method was found to create a porous TiO[subscript]2 barrier layer. Ti sputtering method created a very compact TiO[subscript]2 blocking layer. Two methods showed different characteristics and may be used for different DSSC studies. The second project is to reduce the DSSC cost while maintaining the efficiency by replacing the expensive Pt counter electrode with a novel vertically aligned carbon nanofiber (VACNF) electrode. A large specific electrode surface area (~125 cm[superscript]2 over 1 cm[superscript]2 geometric area) was obtained by using VACNFs. The relatively high surface area, good electric conductivity and the large numbers of active graphitic edges existed in cone-like microstructure of VACNFs were employed to improve redox reaction rate of I[subscript]3[superscript]-/I[superscript]- mediators in the electrolyte. Faster electron transfer and good catalytic activities were obtained with such counter electrodes. The third project is to develop a metal organic chemical vapor deposition (MOCVD) method to coat TiO[subscript]2 shells on VACNF arrays as potential photoanodes in the DSSC system in order to improve the electron transfer. Fabrication processes were demonstrated and preliminary materials were characterized with scanning electron microscopy and transmission electron microscopy. MOCVD at 300 mTorr vapor pressure at 550° C for 120 min was found to be the optimal condition.

Doctor of Philosophy / Department of Chemistry / Paul E. Smith / Molecular Dynamics (MD) simulations have played an important role in providing detailed atomic information for the study of biological systems. The quality of an MD simulation depends on both the degree of sampling and the accuracy of force field. Kirkwood-Buff (KB) theory provides a relationship between species distributions from simulation results and thermodynamic properties from experiments. Recently, it has been used to develop new, hopefully improved, force fields and to study preferential interactions. Here we combine KB theory and MD simulations to study a variety of intermolecular interactions in solution. Firstly, we present a force field for neutral amines and carboxylic acids. The parameters were developed to reproduce the composition dependent KB integrals obtained from an analysis of the experimental data, allowing for accurate descriptions of activities involved with uncharged N-terminus and lysine residues, as well as the protonated states for the C-terminus and both aspartic and glutamic acids. Secondly, the KB force fields and KB theory are used to investigate the urea cosolvent effect on peptide aggregation behavior by molecular dynamics simulation. Neo-pentane, benzene, glycine and methanol are selected to represent different characteristics of proteins. The chemical potential derivatives with respect to the cosolvent concentrations are calculated and analyzed, and the four solutes exhibit large differences. Finally, the contributions from the vibrational partition function to the total free energy and enthalpy changes are investigated for several systems and processes including: the enthalpy of evaporation, the free energy of solvation, the activity of a solute in solution, protein folding, and the enthalpy of mixing. The vibrational frequencies of N-methylacetamide, acetone and water are calculated using density functional theory and MD simulations. We argue that the contributions from the vibrational partition function are large and in classical force fields these contributions should be implicitly included by the use of effective intermolecular interactions.

Master of Science / Department of Grain Science and Industry / Xiuzhi Susan Sun / Plant oil-based raw materials have become more attractive alternatives in polymer industry as fossil resources depletion and environmental concerns continue to arise. Camelina (camelina sativa L.) seed contains about 45% of oil and about 90% of the oil is unsaturated fatty acids such as linoleic acid, α-linolenic acid, and erucic acids. It also provides the advantages of low cost and low fertilizer demand. Functionalized oils such as epoxidized camelina oil (ECO) and di-hydroxyl camelina oil (DCO) can be used for resins, adhesives, coatings, etc. The objectives of this work were to synthesize and characterize ECO and DCO from camelina oil. The epoxidation reaction of camelina oil was completed with formic acid and hydrogen peroxide. Catalyst ratio, reaction time, and temperature effects on the epoxidation reaction were studied. The optimum epoxy content of 7.52 wt% with a conversion rate of 76.34% was obtained from camelina oil using excess hydrogen peroxide and a molar ratio of formic acid of less than 1 for 5 hours in 50 °C. Camelina oil yields higher epoxy content (7.52 wt%) than soybean oil (6.53 wt%); however, soybean oil had a higher conversion rate of 80.16% compared to camelina oil because of uniform fatty acids distribution. In this study, we found that epoxidation efficiency is significantly affected by fatty acids composition, structure, and distribution. DCO was synthesized from ECO with different reaction parameters. The ring opening of ECO was performed with water, perchloric acid, and THF as proton donor, catalyst, and solvent respectively. Hydroxyl value of DCO was measured, and the maximal hydroxyl value was 369.24 mg KOH/g. physical properties of DCO were characterized by acid value and moisture content; thermal properties of DCO were obtained using different scanning calorimeter (DSC), thermalgravimetric analysis (TGA). Amount of solvent and acid catalyst addition affected the hydroxyl value and residual acid in DCO. Heat capacity, phase transition temperatures, and thermal stability of DCO were obtained and showed higher values than ECO’s. The DCO showed higher peel adhesion when it was formulated with epoxidized soybean oils through UV curing because camelina oil allows higher epoxy content, which results in higher hydroxyl values.

Doctor of Philosophy / Department of Chemistry / Christine M. Aikens / This body of work describes a theoretical study of the growth mechanism of gold thiolate nanoparticles from Au(III) as synthesized in the Brust-Schiffrin method. The Au(III) salt can be reduced to form Au(I) by two thiols or a hydride. Depending on the polarity of the solvent, the Au(I) species will either yield rings and anionic chains, remain in isolation, or create an ionic complex with the phase transfer agent. No matter what form the Au(I) species takes, a second reduction must occur to yield Au(0). If the solvent is polar, such as methanol or water, and the Au(I) species is a ring or anionic chain, then a hydride can reduce the structure and create a gold-gold bond and dissociate a thiol from the structure. The gold atoms involved in the gold-gold bond would have a formal Au(0) oxidation state. However if the Au(I) species can be kept from forming rings or chains in the polar solvent or if the system is in a nonpolar solvent, then two Au(I) species in close proximity in the presence of hydride can react to yield a non-radical Au(0) species. The oxidation of bare gold nanoclusters by thiol will also be examined, such as in the case of SMAD-produced gold nanoparticles. In this process, the gold nanoclusters are initially in the Au(0) oxidation state. However the SR-Au-SR “staple” motifs that are known to passivate gold nanoparticles contain Au(I) species. The adsorption of thiol on various sizes of gold clusters in several charge states will be calculated and the mechanism for the oxidation of Au3 and three-dimensional Au12 will be modeled. The rate-limiting step is found to be the thiol hydrogen dissociation onto the gold cluster. Once this dissociation occurs, the hydrogen can move freely around the surface. Finally, Au25(SH)18- will be investigated as a catalyst for selective hydrogenation of α,β-unsaturated aldehyde. The dependence of the energetics of hydrogen gas dissociation on Au25(SH)18- on the functional and Grimme dispersion correction employed will also be examined.

Doctor of Philosophy / Department of Chemistry / Jun Li / A lot of materials exhibit novel properties when scaled down to nanoscale. Here we explore nanoelectrode arrays (NEAs) and nanoparticles in the application of high performance biosensors. We have developed an electrochemical (EC) method for measuring the activity of proteases using vertically aligned carbon nanofiber (VACNF) NEAs. VACNFs were grown on conductive substrates and encapsulated in SiO₂ matrix. After polishing and plasma etching, controlled VACNF tips are exposed to form an embedded NEA. Tetrapeptides specific to cancer-mediated proteases are covalently attached to the exposed tip, with a ferrocene (Fc) moiety linked at the distal end. The redox signal of Fc can be measured with AC voltammetry (ACV) at ~1 kHz frequency, showing distinct properties from macro-electrodes due to VACNF's unique interior structure. The enhanced ACV properties enable the kinetic measurements of proteolytic cleavage of the surface-attached tetrapeptides by proteases. The well-defined regular VACNF NEAs by e-beam lithography show a much faster kinetics for cathepsin B proteolysis. This EC method was further applied in whole lysate of human breast tissue and breast cells. The detected protease activity was found increased in cancer cells, with the metastatic cancer cell lysate showing the highest cathepsin B activity. The results indicated the potential of this technique as a portable multiplex electronic device for cancer diagnosis and treatment monitoring through rapid profiling of the activity of specific cancer-relevant proteases. In another exploratory study, we modified nanoparticles with luminol and viral nucleic acid to develop chemiluminescence (CL) biosensors for blood-borne pathogens. Luminol-labeled 10-nm-diameter gold nanoparticles (GNPs) served as a nanocarrier for enhancing CL signal. The CL signal can be observed over 8 orders of magnitude variations in GNP concentration. Using the same number of particles, luminol-labeled 30-nm-diameter latex beads showed ~3 orders of magnitude higher CL compared to 10-nm-diameter GNPs. Hybridization of target H1N1 nucleic acid on the latex beads and probe nucleic acid on the glass or optical fiber surface has been achieved. This assay will be incorporated into a simple hand-held device for routine assays in hospitals and clinics, or for large-scale screening of human populations as diagnostic tools to identify specific viral strains.

Master of Science / Department of Chemistry / Christopher T. Culbertson / Circulating tumor cells (CTCs) have become a key component in the identification and treatment of cancer. Once dislodged from the main tumor, CTCs travel through the bloodstream and cause metastasis. Early detection and identification of these cells can help in the evaluation and prognosis of various types of cancer, as well as assisting in patient treatments by determining the spread of the disease. Here, a high-throughput microfluidic analysis technique is described that can efficiently detect and identify cells, with the specific identification of CTCs as a future application through fluorescent labeling in mind. As proof of principle, the device has been shown to detect and characterize individual human Jurkat (T-lymphocyte) cells at a rate of 100 cells/minute. The device employs micro-scale flow focusing to isolate individual cells. The cells are detected using both light scattering and laser-induced fluorescence to evaluate cell size and surface functionality.

Doctor of Philosophy / Department of Chemistry / Christer B. Aakeroy / A family of bis-pyridine based pharmaceutical active ingredients were synthesized and co-crystallized with four iodoperfluoroalkanes. Thirteen new crystal structures that are driven by I‧‧‧N(py) halogen bonds, are presented and compared with that of their hydrogen-bonded analogues. Halogen bonded co-crystals exhibit two different structural arrangements, as opposed to layered architectures observed in hydrogen bonded co-crystals. In order to explore the effect of aromatic stacking interactions on hydrogen and halogen bond driven co-crystallization process, we utilized a series of aromatic hydrogen and halogen bond donors in combination with bis-pyridine based pharmaceutical active ingredients. Aromatic stacking between the donor and the acceptor were limited, due to the lack of complementarity between the donor and the acceptor in terms of size, shape and geometry. In that case, homomeric interactions between the single components were translated into the structure of the binary co-crystals. According to our charge calculations, similarly activated hydrogen and iodine atoms possess similar electrostatics. Therefore, we wanted to investigate the interchangeability of hydrogen bonds and halogen bonds by utilizing 2-aminopyrimidine as the backbone for C(sp)-H and C(sp)-I functionalities which makes self-complementary ribbons via NH‧‧‧N synthons. Our results show that the ethynyl proton is capable of acting as a synthon mimic of ethynyl iodine by interchangeable C(sp)-H‧‧‧N hydrogen bonds and C(sp)-I‧‧‧N halogen bonds. We exploited the halogen bonding donor capability of iodo, bromo and chloro ethynyl functionalities towards a series of halide ions. Based on the grinding experiments these donors showed 90%, 70% and 50% success rates towards halides. Among the halides, chlorides exhibited the highest red shift compared to bromides and iodides. We synthesized a series of cavitands functionalized with hydrogen bond donor and acceptor groups and studied their binding preferences towards a series of active ingredients. We have shown that suitably functionalized cavitands can act as carriers of active ingredients and especially, selective binding of aspirin is demonstrated using a two-point binding mode.