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<p> We investigated the local electronic properties of the triangular-lattice materials NaxCO2 (x = 0.3, 0.72) and the superconductor Na0.3Co02-1.3H20 by 59Co and 170 Nuclear Magnetic Resonance(NMR). For Na0.72Co02 , 59Co NMR line shape shows clearly that there are two types of Co sites - Co(A) site and Co(B) site. The electronic character of Co(A) site is close to that of the less magnetic Co+3-like ion with spin rv 0, while the electronic character of Co(B) site is close to that of the strongly magnetic Co+4-like ions with spin ~ 1/2. The temperature dependence of the Knight shifts suggests that the Co(A) and Co(B) sites are electronically coupled, which is not consistent with simple phase separation. The local Co electronic environments propagate to the adjacent 0 layers through p-d hybridization. Therefore, there are two types of oxygen sites, O(A) site and O(B) site. We introduced a different route to do K vs x plot analysis for the Co sites and determined that for the Co sites, Karbitat(A) is 1.816 % and Kspin(A) is about rv 0.2 %; Karbitat(B) is 4.0255 % and Kspin(B) is at least 1.5 %. For Na0 .3Co02 , 59Co NMR line shape shows that the Co valence is averaged out in this material. There are two types of oxygen sites, 0( C) site and O(D) site, presumably because of the nearest neighbor Na+ sites. The constant behavior of Knight shifts below 100 K for both the Co and 0 sites suggests the emergence of a low temperature canonical Fermi-liquid behavior. For the superconductor Na0.3CoO2-1.3H20, both 59Co and 170 NMR line shapes show that there is only one type of Co site and oxygen site. The Knight shifts of 59Co and 170 are temperature independent below 100 K down to Tc. Combined with our spin-lattice relaxation 1/T 1 T measurements, we can rule out the possibility of ferromagnetic scenario of spin excitations above Tc. </p> / Thesis / Master of Science (MSc)

<p> The formation of soluble amyloid oligomers by polypeptide chains is the main pathogenic mechanism underlying several neurodegenerative disorders including some of the most common debilitating and aging-related illnesses such as Alzheimer's and Parkinson's diseases. However, the molecular basis of polypeptide oligomerization and amyloid formation is currently not fully understood. In this thesis the focus will be on the early steps of oligomer formation that precede the nucleation of amyloid fibrils, that are still reversible. The reversibility of these initial self-association equilibria makes them an attractive target for therapeutic intervention in the treatment of amyloid diseases. Specifically three general questions will be addressed: (a) What are the residues within a given polypeptide chain that mediate self-recognition? (b) What are the driving forces for self-association? (c) Is self-recognition coupled with conformation changes? </p> <p> The objective of this thesis is to provide initial responses to these key questions using as prototypical system the Ap (12-28) peptide, which has been previously proposed as a model for the initial self-association events that are linked to Alzheimer's disease. Given the flexibility of this peptide the main tool for its investigation will be Nuclear Magnetic Resonance (NMR) spectroscopy. Specifically, both classical (i.e., TOCSY and NOESY) and more novel (i.e. saturation transfer difference and off-resonance relaxation) NMR experiments were used to probe the soluble oligomers through the comparative analysis of samples with different monomer/oligomer distributions. The combined analysis of this integrated set of experiments reveals that while the residues in the central hydrophobic core (CHC) drive self-recognition, stable oligomers require a conformational change towards more folded structures that affects residues well outside the CHC. The conformational change occurring upon self-association thus effectively couples CHC and non-CHC residues. This model may also explain why mutations outside the CHC (i.e. E22, D23) can affect significantly the kinetics of self-association. </p> <p> / Thesis / Master of Science (MSc)

A novel partial agonist of the exchange protein activated by cAMP isoform 1 (EPAC1), I942, was recently discovered and shown to reduce the guanine exchange factor activity of cAMP-bound EPAC1 to approximately 10% relative to cAMP activation. However, the inhibition mechanism of I942 remains unknown. Here, we utilize NMR spectroscopy to probe the inhibitory I942 - EPAC1 interactions at atomic resolution. The EPAC1 - I942 interface was mapped through intermolecular NOEs measured by 15N and 13C filtered NOESY-HSQC experiment. Intermolecular NOE mapping combined with other protein NMR methods, such as saturation transfer difference, transfer Nuclear Overhauser Effect spectroscopy and chemical shift mapping, we revealed that I942 interacts with the phosphate binding cassette (PBC) and base binding region (BBR) of the EPAC1 cyclic nucleotide binding (CNB) domain, similar to cAMP. The PBC controls the conformation of the hinge region, and subsequently, allosterically shifts the hinge region between its active/inactive states. Molecular dynamics simulation based on the NMR spectroscopy data revealed that EPAC1-CNB adopts an intermediate conformation between its inactive and active states, which explains the partial agonist nature of I942. / Thesis / Master of Science (MSc) / The exchange protein activated by cAMP (EPAC) is a receptor for the classical secondary messenger cAMP. EPAC is present in multiple human systems and plays a pivotal role in the development of a wide range of diseases. In this study, we aim to establish the inhibition mechanism of a novel small molecule EPAC inhibitor/partial agonist I942 using NMR spectroscopy with the goal of achieving a better understanding of EPAC inhibition and paving the way for new small molecule EPAC inhibitors that can potentially treat EPAC-related diseases such as heart failure and diabetes.

Mass spectrometry was used in conjunction with numerous 1-D and 2-D NMR techniques to determine the structures, devoid of stereochemistry, of five different compounds isolated from the extracts of Eunicea succinea, Eunicea tourneforti, and an unidentified species isolated from the Eunicea genus by Professor Meledath Govindan, of the University of the Virgin Islands. Three of the compounds were then identified as the known compounds eunicin, 12,13-bisepieupalmerin, and 7(S),8(S)-epoxy-1(S),11(R)-dolabella-3E,12(18) -dien-13-one by comparison of their spectroscopic data and optical rotations with those published in the literature. Optical rotations could not be measured accurately for the other two compounds because of small sample sizes; therefore, another method had to be found to elucidate the stereochemistry of these two structures. To solve this problem, molecular modeling and NOESY were employed. Comparison of the NOESY interactions to the thermodynamically available conformations of several possible stereoisomers, calculated by molecular modeling, proved to be a useful technique. One of the remaining two structures was identified as the known stereoisomer euniolide. The stereochemistry of the one remaining structure could not be assigned because sample size was not large enough to obtain a clean NOESY spectra. Finally, based on published synthetic work by Corey and Kania, the absolute stereochemistry of the dolabellane was revised to 7(R),8(R)-epoxy-1(R),11(S)- dolabella-3E,12(18)-dien-13-one. / Master of Science

Ion transport of electrolytes determines the performance of many electroactive devices, from fuel cells to batteries to soft mechanical actuators. This dissertation aims to address some fundamental issues regarding ion transport of ion dense electrolytes using electrophoretic NMR and NMR diffusometry. I first describe the design and fabrication of the first instrumentation capable of reliable ENMR on highly ion-dense electrolytes such as ionic liquids and electrolytes for zinc-air batteries. I design a new electrophoretic NMR sample cell using parallel capillaries to investigate the electrophoretic mobilities of pure ionic liquids. It shows the first study of a highly ion-dense electrolyte with electrophoretic NMR. Then I employ NMR diffusometry and electrophoretic NMR to investigate ion association of pure ionic liquids. Then I use electrophoretic NMR technique to investigate the electrophoretic mobilities of electrolytes for zinc-air batteries. For Zn2+ salt added dicyanamide (dca) based ionic liquids, I investigate the effects of Zn2+ salt on chemical shift of dca and ion motion. The combination of mobilities measurements and diffusion measurements provides some new insight of ion aggregation. We explore ion transport of ionic liquids inside the ionic polymer Nafion as a function of hydration level. When ionic liquids diffuse inside ionic polymers, isolated anions diffuse faster (e 4X) than cations at high hydration whereas ion associations result in substantially faster cation diffusion (d 3X) at low hydration inside membranes, revealing prevalent anionic aggregates. Finally, we compare diffusion activation energy measurements in a hydrated perfluorosulfonate ionomer and aqueous solutions of triflic acid, which provides insight into water transport dynamics on sub-nm lengthscales. And we explore the physical meaning of activation energy, characterizing local intermolecular interactions that occur on the pre-diffusional (~ 1 ps) timescale. / Ph. D.

Polymeric methylenebis(phenylisocyanate) (pMDI) has increasingly been used in the wood particulate composite industry. Wood composites, especially oriented strand board (OSB) are made with many variations of wood species. Little research has been done to investigate how pMDI adhesion has been affected by species. The present is divided into two parts. First, mode I fracture mechanics and surface free energy analysis was performed to investigate differences in adhesion between southern yellow pine and yellow-poplar bonded with pMDI. Secondly, an improvement in the synthesis of 13C, 15N labeled pMDI is discussed. Mode I fracture results show that pMDI adhesion was affected differently by southern yellow pine compared yellow poplar. The shear energy release rate was significantly higher in pine/pMDI composites than in yellow poplar/pMDI composites. The total surface energy of southern yellow pine was shown to be significantly greater than yellow poplar. The free energy of adhesion (DG) of the pine/pMDI and the poplar/pMDI was investigated. The DG indicated that the pine/pMDI system would take more energy to separate compared to the poplar/pMDI system. Lastly, a double-labeled 13C, 15N pMDI adhesive was successfully synthesized to produce Solid-State NMR composites. / Master of Science

<p dir="ltr">Biologic drugs are a key defense against many health issues. In many cases, biologic drugs are not stable in the solution state and must be lyophilized. Lyophilization in the presence of excipients increases the stability of the drug by interactions with the excipients through hydrogen bonding, which will lower the local mobility of the drug. Key threats to stability include: inhomogeneity of the drug substance and excipients, high mobility, and crystallization. Solid-state nuclear magnetic resonance spectroscopy was used to identify crystallization, assess homogeneity, and measure the local mobility of lyophilized protein and mRNA/LNP systems. </p><p dir="ltr">The impact of disaccharide type and concentration on protein stability was explored. Human serum albumin (HSA) was lyophilized with disaccharides (sucrose and/or trehalose) in different relative concentrations, and solid-state nuclear magnetic resonance spectroscopy (ssNMR) ¹H T₁ and ¹H T_1rho relaxation times were measured to determine the homogeneity of the lyophilized systems on 20-50 and 1-3 nm domains, and measure local mobility with ¹H T₁ relaxation times. HSA/sucrose systems had longer ¹H T₁ relaxation times and were slightly more stable than trehalose systems in almost all cases shown. HSA/sucrose/trehalose systems have ¹H T₁ relaxation times between the HSA/sucrose and HSA/trehalose systems and did not result in a more stable system compared to binary systems. Phase separation was evident in a sample containing relative concentrations of 10% HSA and 90% trehalose, suggesting trehalose crystallization during lyophilization. Under these stability conditions, a ¹H T₁ relaxation time below 1.5 s correlated with an unstable sample, regardless of disaccharide(s) used.</p><p dir="ltr">The effect of mannitol on protein stability was studied. Human serum albumin was lyophilized in binary systems with mannitol, and in ternary systems with sucrose or trehalose and mannitol. The monomer content of the HSA was monitored over 36 weeks of storage at 50 C. The amount of mannitol in the system dictated the ability of mannitol to crystallize, and the polymorph that mannitol crystallized into. In HSA/mannitol systems, mannitol crystallization caused inhomogeneity of the matrix, determined by ¹H T_1rho relaxation times. Adding a disaccharide to the matrix, however, increased the homogeneity of the matrix. Addition of mannitol to a HSA/disaccharide matrix resulted in less stability at similar HSA:disaccharide ratios.</p><p dir="ltr">The impact of storage temperature on protein stability was investigated. Human serum albumin was lyophilized with sucrose or trehalose in histidine, phosphate, or citrate buffer. ¹H T₁ relaxation times were measured by ssNMR and were used to assess the homogeneity and mobility of the samples after zero, six, and twelve months at different temperatures. The mobility of the samples decreased after 6 and 12 months storage at elevated temperatures, consistent with structural relaxation of the amorphous disaccharide matrix. Formulations with sucrose had lower mobility and greater stability than formulations with trehalose.</p><p dir="ltr">The effect of an RF-assisted lyophilization method on homogeneity, mobility, stability, and moisture content was explored. This method, utilizing 18 GHz microwave frequency to accelerate the lyophilization cycle, resulted in equivalent or better stability for attenuated live virus or protein formulations, respectively. ssNMR showed comparable amounts of homogeneity in the formulations, however mobility of the samples produced by RF-assisted lyophilization was slightly higher.</p><p dir="ltr">A lyophilized mRNA/LNP formulation was prepared. Disaccharide type, disaccharide concentration, and freezing rate were found to alter critical quality attributes of the system. When mRNA/LNP formulations were stored at 4 C, solution formulations outperformed lyophilized formulations for at least 6 months. When mRNA/LNP formulations were stored at room temperature, solution formulations were superior for the first three months, however lyophilized formulations outperformed solution formulations after 6 months, with less growth in particle size and less loss of encapsulation efficiency. ssNMR was used to assess the interactions between the formulation components.</p>