Non-viral gene delivery agents, such as cationic polyelectrolytes, are attractive replacements to viruses due to the absence of potential immunogenic risk and the ability to tune their macromolecular structure. Although non-viral vectors possess numerous design advantages, several investigators have shown that transfer efficiencies are considerably lower when compared to viral vectors. The work reported in this dissertation aims to fundamentally understand the underlying structure-transfection relationships involved in polycation-mediated gene delivery. Efforts focused on the influence of molecular weight, macromolecular topology, carbohydrate modifications, and charge density on the overall transfection activity in vitro. Several families of polycations were synthesized in order to correlate chemo-physical characterization with transfection results. Results revealed that seemingly small changes in the structure of cationic polyelectrolytes can have profound consequences on their transfection activity. / Ph. D.

In this project, we have designed and synthesized a new series of long-chain, water-soluble, dendritic, anionic amphiphiles [3CAmn, RCONHC(CH₂CH₂COOH)₃, R= CnH2n+1] to alleviate the low aqueous solubility of fatty acids. The dendritictricarboxlyato headgroup improves aqueous solubility and allows us to measure the intrinsic biological activity of our amphiphiles without the potential hindrance of low aqueous solubility. The aqueous solubilities of the anionic amphiphiles have been measured and were vastly higher than that of fatty acids. For example, 3CAm17 (1700 μM at pH 7.2) has much better aqueous solubility than the C₁₈ fatty acid analog (<<1 micromol at pH 7.4). Following the determination of aqueous solubility, both anionic and nonionic amphiphiles were tested against a wide variety of microorganisms. The anionic amphiphiles were mostly active against Candida albicans (4.4 microgram/mL), Saccharomyces cerevisiae (4.4 μg/mL), and Mycobacterium smegmatis (18 microgram/mL) and exhibited modest activity against both Gram-negative (71–280 microgram/mL) and Gram-positive bacteria (36– >6300 microgram/mL). With the exception of Neisseria gonorrhoeae (9.8 microgram/mL), the nonionic amphiphiles were mostly minimally active or inactive against Gram-negative bacteria (630–5000 microgram/mL). The nonionic amphiphiles were similarly inactive against fungi (625–5000 microgram/mL). However, the nonionic amphiphiles exhibited good activity against M. smegmatis (20 microgram/mL) and exhibited the best activity against Grampositive bacteria, such as MRSA (22 microgram/mL), Staphylococcus aureus (20 microgram/mL), and Micrococcus luteus (20 microgram/mL). The anionic and nonionic amphiphiles were also tested for possible spermicidal and anti-human immunodefiency virus (HIV) activity. The anionic amphiphiles exhibited anti-HIV activity (EC₅₀, 73–340 microgram/mL), but lacked spermicidal activity. The series had comparable anti-HIV activity to the commercial product N-9 (80 microgram/mL). Except 3CAm13, all anionic amphiphiles (1.4–4) had better selectivity indices than that of N-9 (0.9). The nonionic amphiphiles exhibited both anti-HIV (44–67 microgram/mL) and spermicidal activity (226–2000 microgram/mL). The nonionic amphiphile were more spermicidal and antiviral than Nonoxynol-9. In addition to biological activity, we determined whether the anionic amphiphiles could be utilized as corrosion inhibitors or ore flotation enhancers. The anionic amphiphiles formed stable thin films on silver oxide that were resistant to ethanol washings. We also measured the water contact angles of the anionic amphiphiles on mineral surfaces [apatite (95°), calcite (92°)]. / Ph. D.

One of the most significant challenges faced when modeling mass transfer from contaminant source zones is uncertainty regarding parameter estimates. These rate parameters are of particular importance because they control the connectivity between a simulated contaminant source zone and the aqueous phase. Where direct observation has fallen short, this study attempts to interpret field data using an inverse modeling technique for the purpose of constraining mass transfer processes which are poorly understood at the field scale. Inverse modeling was applied to evaluate parameters in rate-limited models for mass transfer. Two processes were analyzed: (i) desorption of hydrophobic contaminants and (ii) multicomponent Non-Aqueous Phase Liquid (NAPL) dissolution. Desorption was investigated using data obtained from elution experiments conducted with weathered sediment contaminated with 2,4,6 trinitrotoluene (TNT) (Sellm and Iskandar, 1994). Transport modeling was performed with four alternative source models, but predictive error was minimized by two first-order models which represented sorption/desorption using a Freundlich isotherm. The results suggest that first-order/Freundlich models can reproduce dynamic desorption attributed to high-and-low relative energy sorption sites. However, additional experimentation with the inversion method suggests that mass constraints are required in order to appropriately determine mass transfer coefficients and sorption parameters. The final portion of this research focused on rate-limited mass transfer from multicomponent NAPLs to the aqueous phase. Previous work has been limited to bench and intermediate scale findings which have been shown to inadequately translate to field conditions. Two studies were conducted in which numerical modeling was used to reproduce dissolution from multicomponent NAPL sources. In the first study, a model was generated to reproduce dissolution of chloroform (TCM), trichloroethylene (TCE) and tetrachloroethylene (PCE) observed during an emplaced-source field experiment conducted within a flow cell (Broholm et al., 1999). In the second study, a methodology was developed for analyzing benzene, toluene, ethylbenzene and xylene (BTEX) data during a field-scale mass transfer test conducted within a vertically-smeared source zone (Kavanaugh, 2010). The findings suggest that the inversion technique, when provided appropriate characterization of site and source parameters and when given appropriate dataset resolution, represents a viable method for parameter determination. Furthermore, the findings of this research suggest that inversion-based modeling provides an innovative predictive method for determining mass transfer parameters for multicomponent mixtures at the field scale. / Ph. D.

Corneal ulceration with prolonged healing following Nd:YAG laser cyclophotocoagulation in dogs is a frequent complication. It is hypothesized that these corneal ulcerations may be a form of neurotrophic keratitis due to laser-induced damage to corneal innervation. Fifteen clinically normal dogs had the neodymium:yttrium aluminum garnet(Nd:YAG) laser cyclophotocoagulation performed on the left eye. Each treated eye received 100 Joules of laser energy. Corneal touch threshold (CTT) and Schirmer I tear tests (STT) were performed before the surgery and on days 1,3,5,7,9,11, and 13 post-laser treatment. Applanation tonometry was performed before surgery and twice daily for 14 days post-laser treatment. Eyes were enucleated after 14 days and corneal nerves were stained using a gold chloride technique. Major nerve bundles entering the cornea were quantitated by quadrant, using camera lucida reproductions. Nerve bundle diameters were measured using NIH image computer software on computer-scanned images. Statistical methods included repeated values for analysis of variance for CTT, STT and IOP, and a paired t-test for nerve diameters and bundles. All laser treated eyes had significantly higher CTTs (P<0.05) compared to control eyes for all measurements. Six out of fifteen dogs had evidence of ulcerative keratitis. Intraocular pressure was significantly lower in laser treated eyes compared to control eyes in the a.m. on days 2-9,and 14, and in the p.m. on days 2-11 using a Bonferroni-corrected alpha level (P<0.0039). A significant decrease of one nerve bundle per corneal quadrant was found between the laser treated and control eyes. There was no significant difference in STT or nerve bundle diameters between laser treated and control eyes. Nd:YAG laser cyclophotocoagulation effectively reduces IOP while increasing CTT. The procedure also causes a significant decrease in the number of major nerve bundles entering the cornea, but has no effect on the diameter of those bundles. These findings support the hypothesis that nerve damage and corneal hypoesthesia are etiologic factors in ulcerative keratitis following Nd:YAG laser cyclophotocoagulation. / Master of Science

Biopharmaceutical manufacturing is a rigorous and expensive process. Due to the medicinal nature of the product, a high purity level is required and several expensive purification steps must be utilized. Cost-effective production and purification is essential for any biopharmaceutical product to be successful and development of the fastest, most economical, and highest-yielding purification scheme is a constant engineering challenge. Commercial-scale purification schemes currently revolve around the use of multiple chromatography steps for the purification of biopharmaceutical products. Chromatography has many shortcomings including high cost, limited throughput, and complex scale up. The goal of this research was to develop an alternative, non-chromatography purification step for the separation of an acidic model protein, recombinant β-glucuronidase (rGUS), from transgenic tobacco with high yield and purity. Aqueous two-phase extraction (ATPE) is a powerful technique for separation and purification of proteins, and has the potential to replace an expensive chromatography step for the initial purification of recombinant proteins. ATPE enables high levels of target protein recovery and concentration while removing large amounts of impurities from the initial extract. Fractional factorial designs and response surface methodology were used to determine an optimized aqueous two-phase system for the purification of rGUS from transgenic tobacco. In a 13.4 % (w/w) PEG/18% (w/w) potassium phosphate system, 74% of the rGUS was recovered in the top PEG-rich phase while 90% of the native tobacco proteins were removed in the interphase and the bottom phase. A purification factor of about 20 was achieved in this process. / Master of Science

Background: Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used for their analgesic, anti-pyretic and anti-inflammatory properties in both human and veterinary patients. Topical ophthalmic NSAIDs are commonly employed in the management of intraocular inflammation (uveitis), corneoconjunctival inflammatory disease and pre-operatively to prevent intraoperative miosis during cataract surgery. Despite their routine application in these clinical scenarios, little is known regarding the corneal penetration and relative anti-inflammatory efficacy of the available topical ophthalmic NSAIDs in the dog. Decisions regarding which of these agents to employ are therefore based upon factors such as cost and ease of acquisition as opposed to established efficacy. Objectives: To investigate the relative intraocular penetration and anti-inflammatory efficacy of two commonly utilized topical ophthalmic NSAIDs in dogs, diclofenac 0.1% and ketorolac 0.5%. Animals: Twenty-two client owned dogs (22 operated eyes) presenting to the VTH ophthalmology service for routine cataract surgery for mature or hypermature cataract. Methods: Subjects were randomized to be treated with either topical ketorolac 0.5% or topical diclofenac 0.1% ophthalmic solutions at specified times in the 24-hour period pre-operatively. Aqueous humor samples were obtained intra-operatively and stored for subsequent evaluation of drug concentrations and prostaglandin E2 (PGE2) concentrations via ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS) and enzyme-linked immunoassay (ELISA) analysis, respectively. Results: Median aqueous humor drug concentrations were significantly higher in dogs treated with ketorolac 0.5% (1311.6 ng/mL) compared to those treated with diclofenac 0.1% (284.9 ng/mL). There was no significant difference in aqueous humor PGE2 concentrations between the two treatment groups. No significant association was determined between aqueous humor drug concentration and PGE2 concentration. There was no significant association between diabetic status and aqueous humor drug concentration or PGE2 concentration in either group. Conclusions and clinical importance: This study suggests that topical ketorolac 0.5% and diclofenac 0.1% are efficacious in decreasing aqueous humor PGE2 concentrations and are equally suitable for use based on their comparable anti-inflammatory profiles. The results of these assays provide clinically relevant information regarding intraocular penetration and anti-inflammatory efficacy of these medications in dogs with cataract. / Master of Science / Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used for their analgesic, anti-pyretic and anti-inflammatory properties in both human and veterinary patients. Topical ophthalmic NSAIDs are commonly employed in the management of intraocular inflammation (uveitis), corneoconjunctival inflammatory disease and pre-operatively to prevent intraoperative miosis during cataract surgery. Despite their routine application in these clinical scenarios, little is known regarding the intraocular penetration and relative anti-inflammatory efficacy of the available topical ophthalmic NSAIDs in the dog. Decisions regarding which of these agents to employ are therefore based upon factors such as cost and ease of acquisition as opposed to established efficacy. Efficacy of topical anti-inflammatory medications in controlling intraocular inflammation is primarily related to the ability of the medication to penetrate the cornea and its efficacy at suppressing inflammatory mediators. The purpose of this study, therefore, is to investigate the relative intraocular penetration and anti-inflammatory efficacy of two commonly utilized topical ophthalmic NSAIDs in dogs, diclofenac 0.1% and ketorolac 0.5%. Twenty-two dogs presenting to the VTH ophthalmology service for routine cataract surgery with the presence of a mature or hypermature cataract were enrolled in a prospective, randomized clinical trial. Subjects were treated with either topical ketorolac 0.5% or topical diclofenac 0.1% ophthalmic solutions at specified times in the 24-hour period pre-operatively. Aqueous humor samples were obtained intra-operatively and stored for subsequent evaluation of drug concentrations (n=22) and prostaglandin E2 (PGE2) concentrations (n=19) via ultra performance liquid chromatography (UPLC) and enzyme-linked immunoassay (ELISA) analysis, respectively. Treatment with topical ketorolac 0.5% resulted in higher median aqueous humor drug concentrations when compared to treatment with diclofenac 0.1% (1311.6 ng/mL vs. 284.9 ng/mL). However, there was no significant difference in anti-inflammatory efficacy when comparing PGE2 concentrations between the two groups. Furthermore, no significant association was determined when drug concentration was directly compared with PGE2 concentration. The results of these assays suggest that topical ketorolac 0.5% and diclofenac 0.1% are equally suitable for use based on their comparable anti-inflammatory profiles, and provides clinically relevant information regarding intraocular penetration and anti-inflammatory efficacy of these medications in dogs with cataract.

Aqueous film-forming foam (AFFF) used for fuel firefighting contains polyfluoroalkyl substances (PFAS) that have been identified as environmentally persistent and bioaccumulative resulting in phase out of AFFF. Currently, there are no environmentally friendly foams available that can perform at the same level as AFFF. Fuel transport has been recognized as a potential mechanism behind poor fire extinguishment, but the key features are yet unidentified. To fill these knowledge gaps, identifying the properties and features of surfactants used in firefighting foam that will prevent the transport of liquid fuel through the surfactant solution was imperative. To achieve that, this research was performed exclusively on single surfactants that have applications in firefighting foam. Impact of single surfactants on fuel transport was evaluated. Thermodynamics of the interaction between single surfactants and fuel; and kinetics of fuel transport through single surfactant solutions was observed. It was hypothesized that the liquid fuel transport would influence microstructure in the bulk of the surfactant solution. Experiments were conducted for different single surfactant structures. Various methods were applied to identify the microstructure and interfacial properties of surfactants with and without exposure to liquid fuel. The factor affecting microstructure, identified through this study was further used to evaluate the firefighting performance of single surfactants through ignition test. The thermodynamics of the interaction between fuel and single surfactants helped us to understand the fuel transport mechanism and role of micelle on fuel transport. Surfactant and fuel interaction has been studied below, at, and above the critical micelle concentration of surfactants. The effect of surfactant concentration, convection, and surfactant types were observed on the fuel transport. Moreover, an ignition test was conducted to evaluate the firefighting performance of single surfactants for various fuel types. Overall, the findings from this study will help design a new type of superefficient, environmentally acceptable surfactant for firefighting foam application. / Doctor of Philosophy / Aqueous film-forming foam (AFFF) used for fuel firefighting contains fluorinated compounds which are environmentally persistent and bioaccumulative. Therefore, AFFF has been phased out. There are no environmentally friendly foams available as efficient as current AFFF. Researchers have found that fuel transport through surfactant foam solution is the reason for foam collapse and poor fire extinguishment performance. However, the key parameters affecting fuel transport through foam solution have not been identified. Therefore, new formulations have become challenging, and it is important to identify the parameters affecting fuel transport through the firefighting foams. Surfactants are the key components of firefighting foam. The liquid fuel transport affects the microstructure of the surfactants in the bulk solution. Through this research microstructural and interfacial properties of single surfactants have been studied with and without exposure to liquid fuel. The factors affecting microstructure and firefighting performance of surfactants have been identified. Moreover, the interaction between fuel and single surfactants has been evaluated. The effect of surfactant concentration and fuel type on fuel transport has been observed. Moreover, the effect of convection (at the foam-fuel interface) on fuel transport has been observed. Overall, an understanding of factors affecting fuel transport of firefighting foam is achieved through this research, which can guide new types of efficient, environmentally friendly surfactant design.

The low temperature phase behavior of the system H2O-NaCl-FeCl2 was examined using synthetic fluid inclusions. Experiments were conducted along the 5 wt% NaCl (relative to the total solution) pseudobinary, with FeCl2 concentrations varying from 2 to 33 wt%, and along the pseudobinary defined by mixing known amounts of FeCl2-4H2O with a 5 wt% NaCl solution, with final FeCl2 concentrations varying from 0 to 29 wt%. Synthetic fluid inclusions in quartz were prepared in cold-seal pressure vessels at 500 degrees C - 800 degrees C and 2 or 3 kilobars. The fO2 conditions were controlled by the Ni-NiO equilibrium curve. The liquid released from the capsule upon opening was initially colorless, but turned yellow-orange after contact with atmospheric O2. The clear color is characteristic of ferrous iron solutions, whereas the yellow-orange color is consistent with the presence of Fe3+ in solution. This color change suggested that the unopened capsules initially contained ferrous iron in solution, which oxidized to ferric iron when exposed to the atmosphere. Borisenko (1977) reported a eutectic temperature of -37 degrees C for the system H2O-NaCl-FeCl2. In this study, it was not possible to verify this temperature due to the persistence of a metastable liquid down to liquid N2 temperatures (~-196 degrees C). Final ice melting temperatures were obtained for concentrations less than 24 wt% FeCl2 and show a decrease in temperature with increase in FeCl2 concentration. For more concentrated solutions, final melting temperatures could not be obtained because the samples could not be frozen. / Master of Science

Polyimides epitomize one of the most versatile high-performance engineering polymers. Polyimides are inherently mechanically robust, chemically inert, and thermooxidatively stable to 400+ °C depending on their chemical structure, enabling their function in numerous aerospace, electronic, medical, and flame-retardant applications. Polyimides can be highly modular even within synthetic limitations, which promotes and sustains innovative research. One recent interest concerns the innovation of fire suppression foams. Aqueous film-forming foams (AFFFs) are regularly sought when engaging liquid fuel (gasoline, jet fuel) fires. AFFFs utilize perfluorinated compounds (PFCs) like perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA), which exhibit toxicity, bioaccumulation, and persistence in the environment resulting in the presence of fluorosurfactant chemicals in environments either through direct or secondary exposure via chemical migration. Recently, the USEPA has even detected PFAS in drinking water at hundreds of military training facilities and civilian airports. While fluorinated compounds provide desirable thermooxidative stability and excellent fire retardancy, the environmental impact imposed by these chemicals strongly encourages research that targets the complete removal of PFCs in conventional formulations. This thesis focuses on the fundamental development of water-soluble sulfonated polyimide (sPI) and poly(amic acid) (sPAA) systems for next-generation polymer-based fire suppression foams. The use of sulfonated monomers and poly(amic acid) salt formation enables tunable structures and water solubilities. The polymers maintain competitive thermal stabilities to conventional polyimides and, when combined with readily available, non-toxic surfactants (SDS), produce stable foams. The MIL-F-24385F performance requirement evaluates foam quality/stability, drainage time, and burnback resistance to access viability and provides comparison to other systems; preliminary testing shows that sPI/sPAA formulations perform well. Solution rheology offers insights into fundamental scaling relationships of specific viscosity vs. concentration in both salt and salt-free solution that are important to future foam development. Additionally, the structural nature of the sPIs/ sPAAs allows for their modification with phosphonium moieties or siloxanes, which are slated to have positive effects on performance. Overall, these sPIs and sPAAs provide a promising platform for the future direction of fire suppression foams. / Master of Science / High-performance polymers are used in the most demanding of engineering applications. Polyimides represent one of the most versatile high-performance polymers. Polyimides are mechanically strong, chemically inert, and resistant to extreme temperatures depending on their chemical structure, allowing their use in numerous aerospace, electronic, medical, and flame-retardant applications. Polyimides are synthetically versatile, which enables the discovery of new uses after decades of research. One new targeted application is fire suppression foams. Aqueous film-forming foams (AFFFs) are the standard when battling liquid fuel (gasoline, jet fuel) fires. AFFFs contain perfluorinated compounds (PFCs), which are toxic and persist in the environment; they migrate easily to affect indirectly exposed ecosystems. Recently, the USEPA has even detected PFAS in drinking water at hundreds of military training facilities and civilian airports. While AFFFs with PFCs are highly effective, replacement materials are needed. This thesis focuses on the fundamental development of water-soluble sulfonated polyimide (sPI) and poly(amic acid) (sPAA) systems for fire suppression foams. The polymers remain thermally stable, and when combined with readily available surfactants (SDS), produce stable foams. Preliminary fire testing shows that sPI/sPAA formulations perform well against military specifications. Solution rheology (study of flow) explores the solution behavior of sPI, which offers insights into fundamental concentration-viscosity relationships that are important to future foam development. Additionally, the structural nature of the sPIs/ sPAAs allows for their modification with phosphonium groups or siloxanes, which changes their characteristics. Overall, these sPIs and sPAAs are initially promising for the future direction of fire suppression foams.

No / Ten different mathematical models representing the electrophoretic mobility of analytes in capillary electrophoresis in mixed solvents of different composition have been compared using 32 experimental data sets. The solvents are binary mixtures of water-methanol, water-ethanol and methanol-ethanol, respectively. Mean percentage deviation (MPD), overall MPD (OMPD) and individual percentage deviation (IPD) have been considered as comparison criteria. The results showed that a reorganized solution model, namely the combined nearly ideal binary solvent/Redlich-Kister equation, is the most accurate model among other similar models concerning both correlation ability and prediction capability