Browsing by Author "Henry, Chuck, committee member"
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Item Open Access A rapid, point of need open cow test(Colorado State University. Libraries, 2023) Mendez, Jacy, author; Dandy, David, advisor; Henry, Chuck, committee member; Bailey, Travis, committee member; Hansen, Thomas, committee memberIn the dairy industry, maintaining non-pregnant (open) cows is expensive, and may require multiple rounds of artificial insemination (AI) for a cow to become pregnant. There is a need for early pregnancy detection in dairy cows, which allows the use of protocols such as prostaglandin F2-alpha (PGF) and gonadotropin releasing hormone (GnRH) to prepare a cow for another round of breeding via AI, with an emphasis on reduced time between each breeding attempt. The current gold standard method for confirming pregnancy in cows is a rectally-guided ultrasound at day 32 after AI. Interferon-tau (IFNT) is a biomarker that can be detected during days 7-28 of pregnancy in cattle, and is expressed by the cow conceptus. The goal of this work was to develop a cow-side test utilizing IFNT as the biomarker for early cattle pregnancy detection. A lateral flow assay (LFA) was chosen and investigated due to its simplicity and ease of use, but was later adapted to utilize the enzymatic oxidation of 3,3',5,5' – Tetramethylbenzidine to amplify the signal in the test line. C-reactive protein was used to develop protocols for aspects of device development involving nitrocellulose, including antibody striping, blocking, and nitrocellulose selection. These protocols were then utilized as optimization of the lateral flow assay was conducted. The resulting LFA has a limit of detection (LOD) of 10 μg/mL, with an LOD of 100 ng/mL in a half-strip format, with some limitations imposed by false positives. This work provides a novel method of detection for pregnancy in cattle and with further development, has the potential for use by dairy farmers in their respective industry.Item Open Access An efficient multi channel, 0.2 nJ/bit transmitter with tuning for process variation for biomedical telemetry in the MedRadio band of 401-457MHz(Colorado State University. Libraries, 2016) Gundla, Abhiram Reddy, author; Chen, Tom, advisor; Collins, George, committee member; Henry, Chuck, committee memberWith the increasing applications of bio-integrated telemetric systems, there is a growing demand for wireless transceivers in these systems to interface with the outside world. The use of wireless transceivers is desirable because they allow complete untethering of medical devices from patients. Applications of the medical devices that have transceivers may include, but not limited to, neuro-prosthetics for stimulation, sensing vital signs, wireless monitoring of neuro chemicals in the brain, wireless endoscopy, and remote medical diagnosis and therapy. The implantable medical devices to introduce impulses to the central nervous system to treat the diseases efficiently and/or to provide relief to pain are usually in the medical implantable communication services band of 401-406 MHz. The spectrum of 401-457 MHz band is called medical device radio communications service (MedRadio) band, was allocated by FCC on secondary basis. There exists various transmitter designs for the MedRadio band aimed at high energy efficiency (i.e. low energy per bit transmitted), as low as 0.16 nJ/bit. A few designs are targeted to work at high dc power transmission efficiency, as high as 22%. But, the existing designs fail to be truly MedRadio-standard complaint with short-comings either in terms of not using all the channels in the MedRadio band, low transmitter efficiency, or low output power emitted. The search for better designs of transmitters that can utilize all the channels with high transmission efficiency and high emitted output power continues. This thesis proposes an efficient multichannel transmitter circuit design in the MedRadio band at 401-457 MHz. The transmitter circuit consists of a multichannel phase locked loop (PLL) with rail to rail quadrature output voltage controlled oscillator (VCO), a low power digital synchronous programmable integer N-divider, bang bang Phase frequency detector (PFD), charge pump and a 3rd order loop filter, a passive mixer and a power amplifier (PA). The VCO of the transmitter is designed to account for process variation. The proposed transmitter uses quadrature phase shift keying (QPSK) modulation scheme to transmit data. The power consumption of the transmitter is 460 µW at the power supply voltage of 1.2 V, and consumes only 0.2 nJ of energy for every bit transmitted in the MedRadio band. The output power emitted by the power amplifier of the transmitter is -10.8 dBm. The transmitter is able to hop through all the 10 channels of 300 kHz bandwidth of each from 402 to 405 MHz, all the 4 channels of 6MHz bandwidth of each from 413 to 457 MHz. The overall global efficiency of the transmitter is 13.9 %. The proposed transmitter meets all the FCC requirements for the MedRadio band. This proposed work is implemented in a 180nm CMOS process. The proposed transmitter working in the MedRadio band consumes only 0.2 nJ/bit compared to 0.65 nJ/bit of the only other MedRadio-band compliant design. The transmitter energy consumption is low at 460 µW and efficiency is high at 13.9% when compared to mW energy consumption and single-digit efficiency achieved by existing designs.Item Open Access Anticancer potential of nitric oxide-based therapeutics for pediatric and adult cancers(Colorado State University. Libraries, 2021) Gordon, Jenna Leigh, author; Reynolds, Melissa, advisor; Henry, Chuck, committee member; Kennan, Alan, committee member; Brown, Mark, committee memberBased on 2015-2017 data, nearly 40% of men and women will be diagnosed with cancer at some point throughout their lives. As a worldwide pandemic, cancer presents a colossal challenge for researchers and clinicians to continually develop and implement new strategies to prevent, diagnose, and treat the many variations of this disease. Currently, treatment protocols are dominated by surgery, chemotherapy, and radiation therapy. Although valuable, these treatments are often ineffective and are limited to specific situations. Surgery is typically useful for early-stage cancer treatment while chemotherapy and radiation therapy are more common for late-stage treatment. Chemotherapy and radiation therapies are subject to drug resistance and all three produce patient side effects. Thus, a persistent need to develop drugs that are more effective, preferential (to neoplastic cells), and accessible remains. This work implements therapeutics that addresses those concerns while demonstrating efficacy within both pediatric and adult cancers. An evaluation of the anticancer potential of nitric oxide (NO) releasing S-nitrosothiol based anticancer therapeutics is presented herein. In the determination of clinical translatability of a drug, it is essential to understand the desired outcome and potential sources of error prior to execution of analyses and the corresponding methodologies and measurements. Thus, an in-depth analysis of indicators for therapeutic efficacy using tumor-derived cell lines and a detailed investigation of the protocol development and potential interferences of three common cellular viability assays is presented prior to the in vitro work detailed in this study. Specifically, this study involves the application of the NO releasing S-Nitrosothiol, S-Nitrosoglutathione (GSNO) in two variations to determine efficacy against pediatric neuroblastomas and adult breast cancers. Initially, two studies explore the application of GSNO in solution to multiple neuroblastoma cell lines of various origins to determine the potential of NO to act as an adjuvant therapeutic in the clinical management of the prevalent pediatric cancer neuroblastoma. These studies highlight the incredible impact of NO on clonogenic capacity as well as remarkable discriminatory characteristics between neoplastic and healthy cells. Further, the insight presented regarding the mechanism of action of NO on neuroblastomas expands the comprehension of NO-based anticancer therapeutics. Excitingly, when the same GSNO preparation is subsequently applied to more common adult breast cancers to determine if therapeutic efficacy is maintained, results display analogous consequences to those mentioned above. The final study in this dissertation will also explore another application of solution-phase GSNO to adult breast malignancies by combining it with a novel SMYD-3 inhibitor, termed Inhibitor-4 (by collaborators). Since Inhibitor-4 has been shown to similarly impact viability, clonogenic capacity, and apoptosis, this combination is expected to reveal a greater impact than each individual treatment. Overall, an analysis of the significance and feasibility of NO-based therapeutics, delivered via GSNO, is explored to determine their potential application in the clinical management of various cancers. Ultimately, this work expands the knowledge of the practicality, mechanism of action, and effectiveness of NO-based anticancer therapeutics in various cancers with a specific focus on its applicability in neuroblastomas, a malignancy where minimal focus has been placed on NO as a treatment option.Item Open Access Characterization of nanoparticles generated from drilling activities within a sub-surface mine using a novel sampler(Colorado State University. Libraries, 2020) Theisen, Daniel R., author; Brazile, William J., advisor; Tsai, Candace, advisor; Reynolds, Stephen J., committee member; Henry, Chuck, committee memberThis study employed nanoparticle sampling techniques to characterize the aerosol generated from a routine mining activity. A preliminary survey of the particle emission from the feed-leg drilling activity was conducted in the excavations of an experimental mine. The level of particulate exposure was sampled using a novel sampler for respirable and nanometer sized particles; and monitored by direct reading real time instruments. A NanoScan scanning mobility particle sizer (measurement range 10-420 nm) and an optical particle sizer (measurement range 0.3-10 µm) were used. Particulate morphological and structural examination of samples collected with the novel nanoparticle sampler and a thermophoretic sampler was conducted through transmission and scanning electron microscopy and x-ray dispersive analysis. Based on the real-time instrument data, the researchers found high concentrations (> 3.5 x 106 particles/cm3) of ultrafine/nanoparticles generated from the drilling activity. A large amount of submicron silica, spherical primary and agglomerated particles rich in carbon were discovered via analysis of particle sampler specimens with energy-dispersive x-ray spectroscopy. Many particle agglomerates contained primary particles less than 100 nm. Exposure to particles in the nanometer size from various sources within the mining environment has not been well characterized and may be associated with respiratory and systemic disease among miners.Item Open Access Determination of spatial distribution, dissipation, and efficacy of insecticides used for control of citrus greening disease(Colorado State University. Libraries, 2022) Rehberg, Rachelle Anne, author; Borch, Thomas, advisor; Henry, Chuck, committee member; Bailey, Travis, committee member; Trivedi, Pankaj, committee memberCitrus greening disease has devastated citrus production globally. While Florida growers explore management strategies, Asian citrus psyllids (ACP) continue spreading this detrimental disease. Determining the efficacy of insecticides applied in citrus groves is a necessity. In these field studies, the efficacies of foliar insecticide treatments to citrus trees were investigated with liquid chromatography tandem mass spectrometry. Insecticide spatial distribution, dissipation, degradation, and effectiveness at reducing ACP were quantified over time after commercial application at a field site in Florida. Citrus leaves, and sample discs attached to leaves, were collected at specific times and locations within individual citrus trees. ACP were inspected before and after treatments to quantify reductions associated with insecticide concentrations over time. We investigated several insecticides commonly used against ACP including malathion, imidacloprid, dimethoate, and one newer insecticide, afidopyropen. Our findings showed highly variable spatial distribution of insecticides throughout individual trees and rapid dissipation within 24 hours after application. Inadequate distribution to different sides of the leaf and tree canopy areas was observed for all aerial and ground spraying methods tested. Fast degradation rates were observed in sampling discs and citrus leaves with half-lives ranging from 0.6 to 4.0 hours while metabolite concentrations increased. Results showed faster dissipation rates during warmer months (July) and in younger-aged trees ground sprayed with the speed-sprayer. A wide range of insecticide efficacy was observed, with ACP reductions of 63 to 100%. When ACP remained after treatment, effectiveness decreased over time and ACP increased (e.g. from 6 to 172% after afidopyropen treatment). The observed variable spatial distribution, rapid insecticide dissipation, and inadequate efficacy allow remaining ACP or ACP from surrounding groves to continue spreading citrus greening disease, leaving citrus trees unprotected. For contact, or semi-systemic insecticides like afidopyropen, full coverage to both sides of the leaves and tree canopy is crucial to effectively manage ACP populations. ACP regeneration suggests lower metabolite toxicity or pest resistance development and reveals ineffective pest management. This research not only helps inform citrus growers of actual insecticide efficacy in the field, which may influence their pest and disease management strategies, but also provides better understanding of insecticide dissipation from citrus leaves, which assists those advancing predictive models for agricultural applications. Additionally, these results help inform insecticide manufacturers of their products' performance in field conditions which can be compared to laboratory studies. Lastly, this work reveals information on the fate of insecticides in the field which could be used to evaluate its impact on other species and the environment.Item Open Access Enabling and understanding low-temperature kinetic pathways in solid-state metathesis reactions(Colorado State University. Libraries, 2020) Todd, Paul Kendrick, author; Neilson, James, advisor; Finke, Richard, committee member; Prieto, Amy, committee member; Henry, Chuck, committee member; Ma, Kaka, committee memberFor the kinetic pathway to influence the outcome of a solid-state reaction, diffusion barriers must be lowered or circumvented through low-temperature chemistry. Traditional ceramic synthesis use high temperatures to overcome diffusion, yet they result in the thermodynamically stable product. If the desired product lies higher in energy, they are unattainable at such temperatures. Extrinsic parameters, like pressure, can be used to change the stability of products (kinetic trapping), yet require extreme conditions. Another strategy involves kinetically controlling the energy barriers of the reaction to select for a given product. Here, we use solid-state metathesis reactions to understand and control kinetic pathways in the formation of complex oxides and binary metal sulfides. Through simple changes to precursor composition, three unique polymorphs of yttrium manganese oxide are synthesized, two of which are metastable phases. Using in situ diagnostics, the reaction pathways are characterized to identity intermediates and the temperature regimes at which they react. Using this information we identify why different polymorphs form using different precursors. Additionally, small functional organosilicon molecules are shown to catalyze the formation of iron(II) sulfide using metathesis reactions. Here we show that the Si-O functional group stabilizes intermediate species along the pathway to avoid forming more stable intermediates. The result is higher yields of FeS2 at lower temperatures and times. The included chapters will hopefully better inform future solid-state chemists when exploring new composition spaces and reaction pathways.Item Open Access Evaluating the effects of fire on carbon and nitrogen biogeochemistry in forested ecosystems(Colorado State University. Libraries, 2023) Roth, Holly, author; Borch, Thomas, advisor; Henry, Chuck, committee member; Reynolds, Melissa, committee member; Prenni, Jessica, committee member; Wilkins, Mike, committee memberForests provide ecosystem services (e.g., carbon storage, nutrient processing, and water filtration) valued at ~$5 trillion per year which are vulnerable to disturbances such as wildfire. Although fires are a natural component of healthy forests, climate change has begun to increase the size, frequency, and severity of wildfires outside of their historic range. Expected increases in burn severity have implications for carbon (C) and nitrogen (N) cycling, with the potential to shift forests from C sinks to C sources due to long delays in tree re-establishment. There is great interest in resolving changes to soil organic matter (SOM) composition, especially organic nitrogen, to predict how forests respond to wildfires. Therefore, the purpose of the work included in this dissertation was to improve nitrogen analysis in fire-impacted forest systems and apply these methods to soil and water samples. In the following work, a suite of advanced analytical approaches were used to determine the molecular composition of SOM, which was evaluated for the impacts of severe wildfires on microbially-mediated SOM processing and water quality in fire-impacted watersheds. Field-based soil and water samples were collected from subalpine forests in the Colorado Rocky Mountains and investigated for shifts in the water-soluble and solid fractions of SOM in lodgepole pine-dominated forests and their influence on microbial processing and water quality was determined. The objectives of this study were to leverage ultrahigh mass spectrometry to improve N analysis in fire-impacted systems (Objective 1), determine the post-fire changes to surface water C and N chemistry in reducing conditions (Objective 2) and to characterize how fire severity influences SOM composition along soil burn severity gradients (Objective 3). Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) currently achieves the highest mass resolving power in the world, which allows for the study of complex mixtures with tens of thousands of compounds that are separated by the mass of an electron across a wide molecular weight range. The most widely used FT-ICR MS analytical approach uses negative-ion mode electrospray ionization (-ESI) to selectively ionize highly abundant carboxylic acids in SOM. The application of this approach has allowed for rigorous analysis of C composition; however, -ESI FT-ICR MS vastly underestimates N-dense species which are formed during combustion. The biases associated with ionization are propagated in chemical property calculations that are determined by elemental compositions and which must be fully understood for proper use in C and N cycling models. We compared traditional -ESI with positive-ion mode electrospray ionization (+ESI) of burned soil extracts and found that +ESI increased compositional coverage by 87%, including nearly 10,000 additional N species (Objective 1). We applied our +ESI FT-ICR MS findings on a burn severity gradient (low, moderate, and high severity) to evaluate the compositional changes to SOM with increasing severity, with a specific focus on organic nitrogen. We collected soils from burned lodgepole pine forests along the Colorado-Wyoming border from two depths to characterize changes to organic N chemistry. Since organic N is the most abundant form of soil N in conifer forests, a better understanding of post-fire organic N will help address a critical gap in our understanding of fire severity-induced changes in the molecular composition of soil organic nitrogen. Nuclear magnetic resonance spectroscopy and FT-ICR MS analysis showed that N content and aromaticity of water-extractable SOM (0-5 cm depth) increased with burn severity, while minimal changes to the 5-10 cm depth were observed. Heterocyclic N species are generally higher in toxicity compared to their non-nitrogenated counterparts, which prompted soil toxicity measurements. We used Microtox ® to determine that soil toxicity increased with increasing burn severity, which may be partly attributed to newly formed N-species (Objective 2). In conjunction with increased fire activity, North American beaver (C. canadensis) populations have steadily increased since the early 1900s. The ponds that beavers create slow or impound hydrologic and elemental fluxes, increase soil saturation, and have a high potential to transform redox active elements (e.g., oxygen, nitrogen, sulfur, and metals). While surface water runoff composition has been studied in many environments, the effects of reducing conditions (i.e., beaver ponds) on these products are not well known. We collected surface water and sediment samples to investigate the impact of beaver ponds on the chemical properties and molecular composition of dissolved forms of C and N, and the microbial functional potential encoded within these environments from a combination of FT-ICR MS and metagenomics. We found that N-containing compounds and aromaticity increased in the surface water of burned beaver ponds, and that C/N and O/C ratios decreased. Microbial communities within the ponds did not have the capacity to process aromatic species, but they did have the potential for anaerobic metabolism and the potential to respire on microbial necromass (Objective 3). Fires burn heterogeneously across the landscape, and overstory vegetation likely plays a large role both in the way fires burn and how soils recover post-fire. Site factors such as soil type affect the interactions of SOM with abiotic soil components and can have cascading effects on soil C storage, including SOM partitioning between particulate organic matter (POM) and mineral associated organic matter (MAOM). POM is generally considered to have a faster turnover time than MAOM, which is physically protected from microbial degradation. Soil under two common trees in Colorado (lodgepole pine and aspen) are known to differ in SOM quantity and composition, including their relative proportions of POM and MAOM but post-fire products in these soils are relatively uncharacterized. To determine the differences in post-fire SOM between aspen and pine soils, we collected soils from under aspen and pine stands and burned them in open-air pyrocosms to minimize environmental variables which confound field-based studies. We concluded that fire influenced the dissolved fraction of the soils, with higher concentrations of dissolved organic carbon, dissolved total nitrogen, ammonium-N, and nitrate-N in burned aspen soil extracts. To determine the implications for less bioavailable carbon fractions, we will determine %C and %N in soils that have only been dried and sieved, as well as separated into POM and MAOM. We will also characterize the dissolved fractions using FT-ICR MS and NMR to evaluate differences in soil functional groups. Complementary microbiome analyses will be performed to determine the implications of shifts in soil functionality for microbial processing and C and N sequestration. The novel application of +ESI in this dissertation allowed for the identification of increasingly N-dense species at high burn severities which were not previously observed in field samples. N-dense species are enriched under reducing conditions as they are unable to be processed by local microbial communities. In total, these findings contribute to our understanding of newly formed organic C and N species in soils, with implications for microbial activity in fire-affected watersheds.Item Open Access Fundamental studies of reverse micellear aggreagates by multinuclear and multidimensional NMR spectroscopy(Colorado State University. Libraries, 2012) Sedgwick, Myles, author; Levinger, Nancy, advisor; Crans, Debbie, advisor; Henry, Chuck, committee member; Roess, Deborah, committee member; Van Orden, Alan, committee memberSelf-assembled reverse micellar aggregates using cationic, anionic and non-ionic surfactants have been investigated by multinuclear and multidimensional NMR. By utilizing 51V NMR chemical shifts and line widths of decavanadate, the local proton concentration and characteristics of the reverse micellar environment are measured. There is a distinct environmental change on the interior of the reverse micelle depending on the surfactant used. 51V NMR signals for decavanadate inside an Igepal CO-520, non-ionic surfactant, reverse micelle display sharp signals indicating the decavanadate experiences water like environment. Conversely, 51V NMR signals for decavanadate inside an Igepal CO-610/430 mixed reverse micelle show significant broadening of the decavanadate signal indicating that the environment inside the reverse micelle in which the decavanadate resides is more viscous. These data provide a description in that the water pool of non-ionic surfactants can be compared. Time resolved anisotropy decays, ultrafast time-resolved transient absorption, and 2D NMR spectroscopy have been used to study proton transfer reactions in the interiors of Igepal-CO 520, CTAB and AOT reverse micelles. For ѡ0 = 10 reverse micelles formed with anionic AOT surfactant, the HPTS proton transfer dynamics are similar to dynamics in bulk aqueous solution, and the corresponding 1H 2D NOESY NMR spectra display no cross peaks between HPTS and AOT consistent with the HPTS residing, well-hydrated by water, in the interior of the reverse micelle water pool. In contrast, ultrafast transient absorption experiments show no evidence for HPTS photoinduced proton transfer reaction in reverse micelles formed with the cationic CTAB surfactant. In CTAB reverse micelles, clear cross peaks between HPTS and CTAB in the 2D NMR spectra show that HPTS embeds in the interface. Similar behavior is observed for HPTS in Igepal reverse micelles as in CTAB reverse micelles and we interpret the slowed dynamics in the same manner. The 2D NMR spectra for HPTS in Igepal-CO 520 reverse micelles shows interaction that imply the HPTS molecule is rested near the interface inside the reverse micelle. Dynamic light scattering (DLS) and 1H NMR spectroscopic experiments suggest that the assembly of the reverse micellar aggregates depends on non-polar solvent and co-surfactant used. Two different self-assembled particles form in the AOT/cholesterol /water in cyclohexane, where in the similar system of AOT/cholesterol /water in 1-octanol there is only one particle present. In microemulsions employing 1-octanol as the continuous medium, AOT reverse micelles form in a dispersed solution of cholesterol in 1-octanol. Although the size distribution of self-assembled particles is well-known for many different systems, evidence for simultaneous formation of two distinctly sized particles in solution that are chemically different is unprecedented. By utilizing optical spectroscopic techniques, 2D NMR, and DLS, the structure of the non-ionic reverse micelles have been characterized. The impact of adding cholesterol, a biologically relevant molecule, has on the structure of the reverse micellar solutions has also been shown.Item Open Access Investigation into Mycobacterium bovis phenolic glycolipid as a potential biomarker of bovine tuberculosis in urine(Colorado State University. Libraries, 2019) Morphet, Stephanie Marie Little Thunder, author; Belisle, John, advisor; Crick, Dean, committee member; Wilusz, Carol, committee member; Henry, Chuck, committee memberMycobacterium bovis, the etiological agent of bovine tuberculosis (bTB) is reported to cause disease in man and animal alike on every continent aside from Antarctica. Although coordinated efforts have been made for over a century in the US to cease transmission of this pathogen, outbreaks still occur. It has been posited that the failure to eliminate transmission of this pathogen is partially due to the diagnostic in use, which lacks critical sensitivity and specificity. To address this gap, we investigated a potential new method of identifying infected animals that is through the detection of a pathogen-derived biomarker. M. bovis phenolic glycolipid (PGL) is a species-specific, highly abundant, and unique glycolipid that comprises up to 2.5% of the dry cell mass. Coupling an LC-ESI-TOF-MS, method with a solid phase extraction, we successfully detected PGL derived from the urine of naturally-infected cattle. With this knowledge, we aimed to generate a detector of PGL that could be applied in a rapid and field friendly diagnostic platform. Using phage display technology, we selected M13 bacteriophage capable of binding M. bovis PGL with specificity that differentiated between M. bovis PGL and M. Canetti PGL, as well as between total lipid fractions of various species of Mycobacterium and other lipids with similar biochemical properties to PGL. These M. bovis PGL specific phage were able to differentiate between unspiked cattle urine and urine spiked with PGL. Lastly, we assessed the relative stability of PGL, specifically contrasted to another highly abundant mycobacterial glycolipid, phosphatidylinositol mannoside (PIM). We found that PGL was the more stable molecule when testing thermal and chemical stability, as well as when treated with protease K. However, when treating these glycolipids with whole cell lysates derived from fresh bovine organs, accurate detection on an LC-ESI-TOF-MS platform was lost. Further studies will be required to probe the stability of these molecules in vivo. Overall, this potential methodology to assess for infection status may be beneficial in improving the control of M. bovis if further developed, as this new approach has the potential to be more specific and sensitive than the currently used diagnostic.Item Open Access Investigation of surface interactions of pyrazinamide and pyrazinoic acid with synthetic and natural lipid membrane model systems(Colorado State University. Libraries, 2022) Gasparovic, Nathaniel, author; Crans, Debbie C., advisor; Henry, Chuck, committee member; Ryan, Elizabeth, committee memberPyrazinamide (PZA) is a pro-drug used in the treatment of tuberculosis. Upon administration of the drug, it is converted to its active form of pyrazinoic acid (POA) by the tuberculosis bacterium; this is believed to be the biologically active form of the drug which exerts anti-tubercular activity. However, it is generally accepted that both compounds interact with and transverse the membrane, with POA potentially functioning as a protonophore and lowering the intracellular mycobacterial pH. To investigate the interactions of PZA and POA in model membranes, we employed Langmuir monolayers to investigate the potential membrane-disrupting effects of PZA and POA on a model membrane. At physiological pH, neither PZA nor POA disrupted the membrane, although a difference in compressibility was observed. At acidic pH, POA became more disruptive but only at high, non-physiological concentrations. 1H NMR spectroscopy of a microemulsion system was used to investigate the location of PZA and POA in the interface in different protonation states. The neutral POA species was found to preferentially reside in the interface while the charged species remained in the interfacial water. Finally, the effects of PZA and charged POA on the bilayer in liposomes were investigated. A leakage assay on fluorophore-filled liposomes showed that PZA and POA do not induce leakage in the membrane at physiological conditions.Item Open Access Low-noise, low-power transimpedance amplifier for integrated electrochemical biosensor applications(Colorado State University. Libraries, 2014) Wilson, William, author; Chen, Tom, advisor; Pezeshki, Ali, committee member; Henry, Chuck, committee memberBiosensor devices have found an increasingly broad range of applications including clinical, biological, and even pharmaceutical research and testing. These devices are useful for detecting chemical compounds in solutions and tissues. Current visual or optical methods include fluorescence and bio/chemiluminescence based detection. These methods involve adding luminescent dyes or fluorescent tags to cells or tissue samples to track movement in response to a stimulus. These methods often harm living tissue and interfere with natural cell movement and function. Electrochemical biosensing methods may be used without adding potentially harmful dyes or chemicals to living tissues. Electrochemical sensing may be used, on the condition that the desired analyte is electrochemically active, and with the assumption that other compounds present are not electrochemically active at the reduction or oxidation potential of the desired analyte. A wide range of analytes can be selectively detected by specifically setting the potential of the solution using a potentiostat. The resulting small-magnitude current must then be converted to a measurable voltage and read using a low-noise transimpedance amplifier. To provide spatial resolution on the intra-cellular level, a large number of electrodes must be used. To measure electrochemical signals in parallel, each electrode requires a minimum of a transimpedance amplifier, as well as other supporting circuitry. Low power consumption is a requirement for the circuitry to avoid generating large amounts of heat, and small size is necessary to limit silicon area. This thesis proposes the design of a low-noise, low-power transimpedance amplifier for application in integrated electrochemical biosensor devices. The final proposed design achieves a 5MΩ transimpedance gain with 981aA/√Hz input inferred noise, 8.06µW at 0.9V power supply, and occupies a silicon area of 0.0074mm2 in a commercial 0.18µm CMOS process. This thesis also explores the development of a multi-channel electrochemical measurement system.Item Open Access Measuring dissolution rates and interfacial energetics of monolayer molybdenum disulfide electrodes in electrochemical systems(Colorado State University. Libraries, 2023) Toole, Justin R., author; Sambur, Justin, advisor; Henry, Chuck, committee member; Ackerson, Chris, committee member; Field, Stuart, committee memberMeeting carbon zero goals within the next few decades requires advances in energy conversion efficiency, and hydrogen fuel is believed to be a key part of the solution. Photoelectrochemical (PEC) devices can contribute to a renewable-based energy portfolio by directly producing storable chemical fuels. The electrode is a key component that determines what is thermodynamically and kinetically possible for a given PEC device. Unfortunately, semiconductor electrode efficiency can come at the cost of chemical stability. Also, the energetic description of an ultra-thin semiconductor electrode at the liquid interface is unclear. Here, we studied molybdenum disulfide (MoS2), a promising two-dimensional (2D) semiconductor, to improve understanding of interfacial energetics and electron transfer. The overarching hypothesis of this work is: if we quantitatively measure band energies of this 2D material, then we improve understanding of electron transfer efficiency and rates for involved chemical reactions. Knowledge from this research informs new ways to reduce solar energy conversion losses and may improve control over chemical reactions. Our experimental approach is to make in situ optical measurements while changing two key variables: (1) the electrode applied voltage (E), and (2) the liquid redox electrolyte environment (E0'). This thesis is organized into six chapters. Chapter 1 motivates semiconductor photoelectrochemistry as a viable approach for solar energy and chemical fuel production. Following the chronology of key scientific advances over the past few decades, Chapter 2 delves deeper into the established principles of semiconductor photoelectrochemistry, the unique properties of monolayer MoS2, and the current state of the field for making in situ optical measurements in an electrochemical cell. This chapter concludes with open questions that are addressed in Chapters 3 – 5. In Chapter 3, the stability of MoS2 is tested by literally pushing the semiconductor to its anodic decomposition limit. The crucial results are identification of the MoS2 dissolution onset potential (ED) and its thickness-dependent dissolution rates. Additional insights pertain to the long-term stability differences between monolayer and multilayer material. Chapter 4 includes the most noteworthy results wherein we develop a method to quantitatively measure the electronic band gap of monolayer MoS2 using a relatively simple optical setup. For the first time, we use an all-optical approach and many-body theory to report an abrupt change in potential-dependent band gap energies of monolayer MoS2 under electrochemical conditions. Chapter 5 summarizes preliminary work investigating how redox couples in the electrolyte may tune the optical signature of a monolayer MoS2 electrode. Finally, Chapter 6 concludes the thesis with suggestions for subsequent investigations available based on the expertise and resources within the Sambur group at Colorado State University.Item Open Access Phosphorus ligand-coupling reactions for the functionalization of pyridine and other azines(Colorado State University. Libraries, 2021) Nottingham, Kyle G., author; McNally, Andrew, advisor; Paton, Robert, committee member; Henry, Chuck, committee member; Cohen, Robert, committee memberPyridines and related azines are ubiquitous in pharmaceuticals, agrochemicals, and materials. The discovery and development of new purpose-built molecules is contingent on our ability to modify these motifs. Described herein are the development of methods that selectively functionalize pyridine and diazine scaffolds through phosphorus ligand-coupling. Novel phosphine reagents were designed and leveraged to construct C–C, C–O, and C–N bonds on azines from their C–H precursors. Chapter one introduces the history of phosphorus ligand-coupling and defines the reactivity explored throughout this thesis. Both seminal and contemporary examples of phosphorus ligand-coupling reactions are also discussed to provide context for this work. Chapter two focuses on a method to incorporate fluoroalkyl groups onto azines and pharmaceuticals using phosphorus ligand-coupling. This method offers a complementary alternative to widely used radical addition approaches which often produce regiomeric product mixtures on azines. Chapter three presents the investigation of a phosphorus-mediated alkenylation reaction on pyridines and quinolines. Examination of the reaction of pyridylphosphines with alkyne acceptors uncovered divergent reaction pathways from alkenylphosphonium salts. Mechanistic studies provide an explanation for the origin of selectivity obtained in these reactions. Lastly, chapter four expands upon one of these reaction pathways and describes the development of a method for the direct conversion of pyridines into pyridones and aminopyridines.Item Open Access Protein engineering therapeutic strategies and tools(Colorado State University. Libraries, 2019) Ta, Angeline Ngoc, author; Snow, Christopher, advisor; Henry, Chuck, committee member; Kennan, Alan, committee member; Stasevich, Tim, committee memberProteins have become an important tool for research development and therapeutics. Proteins complement the use of small molecules as well as overcome challenges that small molecules cannot. The contrasting difference of their diverse functional and structural properties allows for complex processes like molecular recognition and catalysis. Through loops, turns, helixes, and sheets, these structural motifs provide a protein with shape and electrostatics to achieve a particular function. Overall, I describe here two examples of functional proteins where the protein's complex structure plays an important role in the development of new strategies and tools for therapeutics. The first part of this dissertation shows the effects of increased antibody recruitment on targeted cell death through the use of an immunotherapeutic cocktail of cell surface HER2 receptor binding proteins. The second part of this dissertation describes the use of a protein's chiral environment to develop a new artificial metalloenzyme that selectively catalyses synthesis of the most common N-heterocycle found in FDA approved pharmaceuticals.Item Open Access Rotational motion and organization studies of cell membrane proteins(Colorado State University. Libraries, 2016) Zhang, Dongmei, author; Barisas, B. George, advisor; Van Orden, Alan, committee member; Henry, Chuck, committee member; Roess, Deborah A., committee member; Crans, Debbie, committee memberCell membranes are dynamic structures with complex organization. The complexity of the cell membrane arises from intrinsic membrane structure, membrane microdomains within the plasma membrane and the membrane cytoskeleton. Plasma membrane receptors are integral membrane proteins with diverse structures and functions which bind specific ligands to trigger cellular responses. Due to compartmentalization of the plasma membrane and the formation of membrane microdomains, receptors are distributed non-homogeneously in the cell membrane bilayer. Both lateral and rotational diffusion of membrane receptors reflects different kinds of intermolecular interactions within the plasma membrane environment. Understanding protein diffusion within the membrane is very important to further understanding biomolecular interactions in vivo during complex biological processes including receptor-mediated signaling. Rotational diffusion depends linearly on the in-membrane volume of the rotating proteins. Relative to lateral diffusion, rotational diffusion is a more sensitive probe of an individual molecule’s size and local environment. We have used asymmetric quantum dots (QD) to conduct imaging measurements of individual 2H3 cell Type I Fcε receptor rotation on timescales down to 10 msec per frame. We have also used time-tagged single photon counting measurements of individual QD to examine μsec timescales, although rapid timescales are limited by QD emission rates. In both approaches, decays of time-autocorrelation functions (TACF) for fluorescence polarization fluctuations extend into the millisecond timescale, as implied by time-resolved phosphorescence anisotropy results. Depending on instrumental parameters used in data analysis, polarization fluctuation TACFs can contain a contribution from the intensity fluctuation TACF arising from QD blinking. Such QD blinking feed-through is extremely sensitive to these analysis parameters which effectively change slightly from one measurement to another. We discuss approaches based on the necessary statistical independence of polarization and intensity fluctuations to guarantee removal of a blinking-based component from rotation measurements. Imaging results demonstrate a range of rotational behavior among individual molecules. Such slow motions, not observable previously, may occur with large signaling complexes, which are important targets of study in cell biology. These slow motions appear to be a property of the membrane itself, not of the receptor state. Our results may indicate that individual mesoscale membrane regions rotate or librate with respect to the overall cell surface. The luteinizing hormone receptor (LHR) is a seven transmembrane domain receptor and a member of the GPCR family. It is located on luteal cells, granulosa and theca cells in females. Understanding how these protein receptors function on the plasma membrane will lead to better understanding of mammalian reproduction. LHR becomes aggregated upon binding hCG when receptors are expressed at physiological numbers. Binding of hormone to LHR leads to activation of adenylate cyclase (AC) and an increase in intracellular cyclic AMP (cAMP). ICUE3 is an Epac-based cAMP sensor with two fluorophores, cyan fluorescent protein (CFP) and the YFP variant, cpVenus, and a membrane-targeting motif which can be palmitoylated. Upon binding cAMP, ICUE3 undergoes a conformational change that separates CFP and YFP, significantly reducing FRET and thus increasing the ratio of CFP to YFP fluorescence upon excitation with an arc lamp or 405nm laser source. Hence we have investigated hLHR signal transduction using the cyclic AMP reporter probe, ICUE3. A dual wavelength emission ratio (CFP/YFP) imaging method was used to detect a conformational change in ICUE3 upon binding cAMP. This technique is useful in understanding the sequence of intercellular events following hormone binding to receptor and in particular, the time course involved in signal transduction in a single cell. Our data suggested that CHO cells expressing ICUE3 and directly treated with different concentrations of cAMP with saponin can provide a dose-dependent relationship for changes in intracellular cAMP levels. Forskolin (50μM) causes maximal activation of the intracellular cAMP and an increase in the CFP/YFP emission ratio. In CHO cells expressing both ICUE3 and hLHR-mCherry, the CFP/YFP ratio increased in cells treated with forskolin and in hCG- treated cells. In flow cytometry studies, similar results were obtained when CHO cells expressed < 60k LHR-mCherry per cell. Our results indicate that ICUE3 can provide real time information on intracellular cAMP levels, and the ICUE3 is a reliable cAMP reporter can be used to examine various aspects of LH receptor-mediated signaling.Item Open Access Site-selective functionalization of azines and polyazines via heterocyclic phosphonium salts(Colorado State University. Libraries, 2020) Dolewski, Ryan D., author; McNally, Andrew, advisor; Paton, Robert, committee member; Henry, Chuck, committee member; Kanatous, Shane, committee memberPyridine and diazines are frequently found in FDA approved drugs, biologically active compounds, agrochemicals, and materials. Given the importance of these structural motifs, direct methods that selectively functionalize pyridine and diazine scaffolds have been developed. These methods and their associated challenges are discussed in chapter one. In chapter two, a strategy to directly and selectively functionalize pyridines and diazines via heterocyclic phosphonium salts is presented. The process is broadly applicable for pyridines and diazines and the late-stage functionalization of pharmaceuticals. Four reaction manifolds are amenable to transforming heterocyclic phosphonium salts into valuable derivatives. In chapter three, inherent factors that control site-selectivity in polyazine systems are described along with mechanistically driven approaches for site-selective switching, where the phosphonium ion can be predictably installed at other positions in a polyazine system. The fourth chapter focuses on a new strategy to selectively alkylate pyridines via a traceless dearomatized phosphonium salt intermediate. Preliminary studies show this protocol is amenable to building-block pyridines, drug-like fragments and pharmaceuticals. A late-stage methylation strategy is also presented.Item Open Access The dynamic nature of ligand layers on gold nanoclusters(Colorado State University. Libraries, 2020) Hosier, Christopher Allen, author; Ackerson, Christopher J., advisor; Kennan, Alan J., committee member; Henry, Chuck, committee member; Kipper, Matthew, committee memberGold nanoclusters have been heavily investigated over the last few decades for their potential use in sensing, imaging, energy conversion, and catalytic applications. The development of methodology that allows for controlled functionalization of the surface ligand layer in these compounds is of particular interest due to the role of ligands in determining a large number of cluster properties. One of the fundamental ways of tailoring the ligand layer is the use of ligand exchange reactions. Despite the synthetic utility that ligand exchange reactions afford, a significant number of unanswered challenges currently limits the scope and control that can be obtained with these reactions. While a large variety of ligand types have been used to protect nanocluster surfaces, the majority of reported ligand exchange reactions revolve around chalcogenate-for-chalcogenate exchange. Site-selectivity in these reactions is limited to kinetic phenomenon, and the role of intercluster exchange largely remains a mystery. Additionally, recent works suggest that changes in ligand orientation can impact bulk material properties. In this thesis, we seek to address these challenges by reporting new exchange methodology, probing the evolution of exchanged ligand layers over time, investigating the stability of ligand layers in reaction conditions, and exploring the impact of ligand orientation on nanocluster behavior and reactivity. By addressing these questions and challenges, we seek to move closer to the goal of developing methodology that can be easily and reliably used to tailor gold nanoclusters for directed applications.