PHD, Kent State University, 2025, College of Arts and Sciences / Department of Chemistry and Biochemistry

This dissertation discusses novel synthetic strategies for synthesizing complex natural products, such as cafestol and grayanane diterpenes. Ent-kaurene diterpenes, like cafestol, have a unique 6/6/6/5 tetracyclic skeleton, and their rearranged counterparts, grayanane diterpenes, feature a distinct 5/7/6/5 framework. These classes of diterpene show promising biological properties, including anti-inflammatory, anticancer, antidiabetic, and sodium channel-modulating activities, and are, therefore, of great interest to synthetic and medicinal chemists. This dissertation details the total synthesis of cafestol in 14 steps from the ent-kaurenoic acid extracted from the mature sunflower heads, and in 19 steps from the commercially available stevioside, with an overall yield of 1.59 %. It streamlines the synthesis process using stevioside or ent-kaurenoic acid, which already have all the carbon skeleton and only needs the introduction of a furan ring and some functional group manipulation. A novel one-pot synthesis of furan ring has been accomplished through oxa-Michael Wittig annulation reaction. This work also provides a synthetic pathway to grayanane diterpenes, with a novel discovery of rearranging the ent-kauranes' 6,6 rings system to the grayanane 5,7-rings system to produce a universal intermediate that opens the gate for the synthesis of numerous grayanane diterpenes. Besides, this work also investigates the in-silico studies of synthesized grayanane and other related natural products with the membrane protein of human voltage-gated sodium channels (PDB:6j8h). It provides the insight into the structural features that are associated with biological activity. Furthermore, this dissertation also explores the synthesis and lead optimization of 1,3,4-oxadiazole derivatives targeting methicillin-resistant Staphylococcus aureus (MRSA). The most promising lead compound has advanced into preclinical trials in beagles, demonstrating potential effective treatments in th (open full item for complete abstract)

Doctor of Philosophy, Miami University, 2025, Chemistry and Biochemistry

The continuing rise of antibiotic resistance (AR) is an emerging public health problem. This problem is further compounded by a reduction in the development of novel antibiotics to combat these persistent pathogens. This is due to the cost to bring the drug to market and the low likelihood of developing a successful drug. However, one alternative and promising approach is the application of bacteriophage (phage) viruses to selectively infect and kill pathogenic bacteria. These viruses are ubiquitous and innumerable, making them a compelling therapeutic alternative. However, this innumerable nature has a downside for therapy development. Current methods are limited in their throughput, generation of quantitative data regarding phage-host interactions, and are not amenable to multiplexing or parallel screening (i.e. scalability). Because phage therapy is personalized and requires screening collections of phages to identify efficacious phages to formulate into a therapeutic cocktail, new methods are required to address these gaps. Traditional screening methods include plaque and kinetic growth assays, but these assays are not representative of native conditions and are limited in multiplexing and scalability. Phage-layer Interferometry (PLI), engineered multiplex compatible reporter phages (MultiPhlex), and an indirect phage susceptibility assay called Phage-Assisted Droplet Sorting (PhADS) are aimed at addressing these current challenges to the field of phage therapy. PLI was developed to provide a standardized method for generating quantitative data regarding phage-host interactions. The development of MultiPhlex is ongoing but seeks to address the current inability to multiplex phage screening by developing fluorescently barcoded reporter phages with fluorogenic RNA aptamers. PhADS addresses the main short coming of MultiPhlex, not all phages are amenable to engineering, through indirect detection and provides a high throughput massively parallel phage screening sys (open full item for complete abstract)

Doctor of Philosophy (PhD), Wright State University, 2024, Biomedical Sciences PhD

DNA replication can be perturbed by various agents that slow or stall the replication forks, causing replication stress. If undetected, stressed forks may collapse, causing mutagenic DNA damage or cell death. In response to replication stress and DNA damage, the eukaryotic cell activates the DNA replication checkpoint (DRC) and DNA damage checkpoint (DDC) pathways to promote DNA synthesis, repair, and cell survival. The two cell cycle checkpoint pathways are controlled by the protein sensor kinases Rad3 (hATR/scMec1) and Tel1 (hATM/scTel1) in fission yeast, although Tel1 plays a minimal role in checkpoint functions. Rad3 and Tel1 belong to a family of phosphatidylinositol-3-kinase-related kinases (PIKKs), whose stability is regulated by the heterotrimeric TTT (Tel2-Tti1-Tti2) complex. The current model suggests that the TTT complex works with Hsp90 and R2TP complex in the co-translational maturation of all PIKKs for their proper folding and stability. We have previously reported a tel2-C307Y mutant with a moderately reduced Rad3 protein level (~60% of wild-type cells). This mutation eliminates Rad3 mediated signaling in the DRC pathway but moderately reduces signaling in the DDC pathway. This result suggests that Tel2 of the TTT complex may specifically regulate the DRC pathway. In this study, we investigated this possibility by taking a genetic approach to analyze the functions of Tti1, the largest subunit of the TTT complex. We randomly mutated the tti1 gene and integrated the mutations at the genomic locus by pop-in and pop-out recombination strategy. As a result, 100 primary tti1 mutants were successfully screened, based on their increased sensitivities to hydroxyurea (HU) which depletes cellular dNTPs and/or the DNA damaging agent methyl methanesulfonate (MMS). Preliminary characterization of the primary Tti1 mutants, based on their relative sensitivities to HU, MMS or both agents, led us to focus on a collection of 24 mutants. Among the 24 mutants, DNA seq (open full item for complete abstract)

Master of Science (MS), Wright State University, 2024, Biochemistry and Molecular Biology

Duchenne Muscular Dystrophy (DMD) is a devastating progressive muscular disorder caused by a mutation in the dystrophin gene affecting approximately 1 in 3,500 males. However, despite its prevalence, there is currently no cure for this disease. Mutations in dystrophin lead to enhanced inflammation, fibrosis, cell death, development, and decreased skeletal muscle function. Phosphatidylcholine, synthesized by choline, is a major phospholipid that functions in maintaining and synthesizing cell membranes and has previously been found to be decreased in DMD patients. This led us to hypothesize that the dystrophic phenotype could be improved through treatment with choline. In this study, we used concentrated choline to treat B10 wildtype and mdx mice for both short-term and long-term treatments. We found that compared to untreated groups, treatment with choline showed less inflammation, fewer macrophage markers, and reduced fibrosis development within the skeletal and cardiac tissue of mdx mice. We also found that within skeletal muscle, necroptotic protein markers were downregulated as a result of choline treatment. We further evaluated the effects of choline treatment on dystrophic skeletal and cardiac muscle function to explore the potential mechanism of action. This study determined whether choline could be a potential therapeutic agent for the treatment of DMD.

Doctor of Philosophy, Miami University, 2025, Chemistry and Biochemistry

Chloroplasts are organelles involved in photosynthesis and require thousands of proteins to function. More than 95% of these proteins are encoded by the nucleus and synthesized in the cytosol as precursor proteins. The chloroplasts' double-membrane architecture necessitates the implementation of complex protein import mechanisms to facilitate the transportation of nuclear-encoded proteins to the thylakoid. Proper routing and sorting of thylakoid-targeted proteins is essential for assembling the photosynthetic apparatus within the thylakoid membrane during chloroplast biogenesis. Once proteins reach the chloroplast stroma, they employ one of four protein sorting mechanisms to target the thylakoid membrane or lumen. The chloroplast Twin Arginine Translocation (cpTat) system is an essential and evolutionarily conserved system that transports fully folded proteins across the thylakoid membrane in chloroplasts, as well as the cytoplasmic membrane of eubacteria and archaebacteria. The cpTat system is distinct from other protein transport systems because it utilizes the proton motive force as the sole energy source to transport proteins across the thylakoid membrane by forming a transient pore. Two key components of the system, cpTatC and Hcf106, facilitate the binding of the substrate signal peptide and the recruitment of the pore-forming component Tha4. As the mechanical features of this system are still largely unknown, understanding how these components interact will provide important clues for solving the transport mechanism of the cpTat system. The core component of the system, cpTatC, has a much longer N-terminal soluble domain compared to the predicted lengths for bacterial and algal TatC proteins. This dissertation focuses on the functional characterization of the N-terminal extension of the cpTatC protein through Cysteine (Cys) scanning disulfide crosslinking assays. We observed close contact between the N-terminal extension of cpTatC and the precursor mature do (open full item for complete abstract)

Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, 2024, College of Arts and Sciences

Over the past 60 years, mass spectrometry (MS) has emerged as a premier analytical technique for fatty acid (FA) analysis due to its exceptional sensitivity and selectivity. Techniques combining chromatography with MS, such as liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS), have significantly enhanced the accuracy and robustness of FA quantification. Among FA metabolites, polyunsaturated fatty acids (PUFAs), particularly arachidonic acid-derived eicosanoids, play crucial roles in inflammation and disease pathology. Dysregulated eicosanoids are implicated in various diseases, underscoring the need for precise quantification in biological samples. LC-MS is the preferred method for eicosanoid analysis due to their low concentrations in biological matrices. The first part of this dissertation presents the development and validation of an LC-MS/MS method for quantifying 11-dehydrothromboxane B2 (11-DHTXB2) in human urine, a stable metabolite of thromboxane A2. Elevated urinary 11-DHTXB2 levels serve as a robust prognostic biomarker for atherosclerosis progression, offering the potential for monitoring disease progression and evaluating the efficacy of anti-atherogenic therapies. Short-chain fatty acids (SCFAs), produced by bacterial fermentation of undigested dietary components, are vital gut health biomarkers. Alterations in SCFA concentrations reflect gut microbial imbalances and are linked to inflammatory bowel disease (IBD), which arises from genetic, environmental, and microbial factors leading to an aberrant immune response. IBD-associated gut dysbiosis involves a reduction in SCFA-producing bacteria such as Bacteroides, Firmicutes, and Lactobacillus, resulting in significantly altered SCFA levels. GC-MS, widely regarded as the gold standard for analyzing volatile and semi-volatile compounds, excels in SCFA analysis due to its superior separation capabilities. The second part of this dissertation explores the use (open full item for complete abstract)

Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, 2024, College of Arts and Sciences

Phenolic acids are microbial products of non-absorbed fibers that are fermented by bacteria using indigestible components that reach the gastrointestinal (GI) tract. It has been shown that phenolic acid compounds significantly impact the human body, particularly their microbial metabolism and their ability to influence several aspects of gut homeostasis. For this reason, more attention should be placed on investigating these metabolites. Multiple factors may influence the biosynthesis of phenolic acids, including age, medications, and several environmental factors, but the production of gut metabolites is primarily affected by diet. In this research we aim to enhance the knowledge on the synergistic relationship between gut microbes, diets, and the biosynthesis of phenolic acids. Chapter I summarizes our current understanding of phenolic acid in the GI tract, fermentative pathways, etiology, and beneficial impacts of phenolic acids. In chapter II we aim to construct an in vitro fermentation model to find efficient ways to increase the generation of phenolic acids in the gut using different combinations of probiotics, prebiotics, and other factors, including fats, sugars, and amino acids. Chapter III discusses the development and complete validation of a high-throughput, fast, and reliable liquid chromatography-mass spectrometry method to quantify phenolic acids (ferulic acid, caffeic acid, and gallic acid) in biological samples. In conclusion, there are many studies that support the benefits of prebiotics on our health, but further investigations are needed to understand the interactions between prebiotics and the gut microbiota as they alter the biosynthesis of phenolic acids, which may be vital in enhancing health benefits, including reduced levels of oxidative stress, reduction of inflammation, enhancement of gut microbiota composition, improvement of metabolic health, and contribution to the prevention of chronic diseases such as obesity, diabetes, cardiovascula (open full item for complete abstract)

Master of Science (M.S.), University of Dayton, 2024, Materials Engineering

Rare earth elements (REEs) are essential to many modern-day technological applications. Due to their difficult and environmentally harmful refining methods, many of these REEs are imported to the U.S. from various other countries. With countries like China dominating the market, the U.S. supply chain is at risk. A potential solution to this issue would involve the use of proteins to extract these REEs in an environmentally sustainable manner. Custom proteins would be designed to extract specific REEs from their mixed metal ores through computer simulations, namely molecular dynamics. Currently the design process is stymied by the lack of working force fields for REEs within many molecular dynamics programs. This work seeks to address this issue by creating custom force fields designed around replicating basic experimental properties the REE ions have with water, counterions, and REE binding proteins. This is done utilizing a Bayesian optimization algorithm which can efficiently and accurately choose new parameters to test and verify for a wide variety of systems.

Doctor of Philosophy, Case Western Reserve University, 2025, Pathology

The COVID-19 pandemic claimed the lives of millions of people and affected communities worldwide. SARS-CoV-2, the virus that causes COVID-19, continues to be a global health concern, as does the inevitable threat of new viral outbreaks. We must therefore learn from this virus in the hope of better preparing for future pandemics. We investigated the SARS-CoV-2 accessory protein ORF3a and its roles in modulating host antiviral strategies, namely inflammatory signaling and autophagy regulation. ORF3a activates NF-κB signaling, which induces an inflammatory response in infected cells and can also prime certain cells for inflammasome assembly and subsequent cell death. We found that, unlike the homologous protein SARS-CoV ORF3a, SARS-CoV-2 ORF3a does not depend on its N-terminal TRAF-binding sequence to activate NF-κB. The ORF3a homologs thus affect NF-κB signaling through different mechanisms. Second, SARS-CoV- 2 ORF3a blocks autophagy by binding to the human protein VPS39, a member of the complex that facilitates membrane fusion between autophagic compartments. We discovered that the predicted β-propeller domain of VPS39 is critical to its interaction with ORF3a. Regulating autophagy is important for productive SARS-CoV-2 infection; disrupting the ORF3a:VPS39 interaction could therefore be a future strategy to hinder SARS-CoV-2 propagation.

Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, 2024, College of Arts and Sciences

The cell surface is coated with glycans with terminal sialic acids (Sias), a process known as sialylation that plays important roles in a variety of biological processes. Removal of Sias (desialylation) by sialidase, on the other hand, is often involved in a number of pathological pathways. Neu1, Neu2, Neu3, and Neu4 are the four isozymes of mammalian sialidase, which differ in subcellular localization and substrate specificity. Lipopolysaccharide (LPS) induces endogenous sialidase expression in macrophages, which could result in desialylation of the TLR4, thereby initiating the LPS/TLR4 signaling pathway. In this dissertation study, a variety of analytical methods have been applied to investigate desialylation and localization of sialidases such as Neu1 and Neu3. Higher expression of sialidases was confirmed on THP-1 monocytes and macrophages upon LPS stimulation. Furthermore, lectin blot analysis and flow cytometry showed desialylation of total proteins of LPS-stimulated THP-1 macrophages. Next, we investigated the impact of sialidase inhibitor 2,3-dehydro-2-deoxy-N-acetylneuraminic acid (DANA) on the regulation of LPS/TLR4 signaling pathways. As a result, DANA effectively mitigates the inflammatory responses induced by LPS in THP1 macrophages. Our findings demonstrate that the presence of LPS and the sialidase inhibitor DANA reduces both NF-κB phosphorylation and cytokine release. Finally, we investigated the secretion of Neu1 and Neu3 in the cell culture media of THP1 monocyte and macrophage upon stimulation with LPS. Surprisingly, sialidase levels significantly increased in the cell culture media, indicating sialidases secretion from THP-1 macrophages and monocytes. Furthermore, we investigated the secretion of exosomes from THP-1 macrophages to assess the presence of Neu1 and Neu3 in response to LPS stimulation. The exosome release from THP-1 macrophages is significantly increased by LPS treatment and Vacuolin-1 since it is inhibiting lysosomal exocytosis. (open full item for complete abstract)

Doctor of Philosophy, Case Western Reserve University, 2025, Biochemistry

Appropriate levels of gene expression are required for cellular fitness and survival. Messenger RNAs (mRNAs) play a central role in gene expression and thus must be tightly regulated. Despite most mRNAs undergoing degradation through the same decay pathway, mRNA half-lives are highly disparate and can vary by orders of magnitude. Some transcript-specific features, such as stable structures in the 5' UTR or regulatory sequences in the 3' UTR, can affect mRNA expression. However, these elements are not sufficient to explain the variation in mRNA stability observed transcriptome-wide. Our lab originally identified codon optimality as a global determinant of mRNA stability. Codon optimality refers to how efficiently each codon is translated and is dependent on the relative availability of charged cognate tRNAs. The most likely explanation for the observed link between codon optimality and mRNA stability is that ribosome decoding rates influence mRNA stability. However, a transcriptome-wide link between the contribution of codons to translation rate and to mRNA stability has not been previously established. Using ribosome profiling in Saccharomyces cerevisiae, we demonstrate that codon-level and transcript-level elongation rates indeed globally correlate with mRNA stability. Chemical modification of mRNA nucleosides provides an additional layer of regulation. Prior to our work, N4-acetylcytidine (ac4C) was an understudied modification solely found to exist in polyA-selected mRNA by HPLC/MS-MS. Through a collaborative effort, we identified more than 2,000 human mRNAs that are modified with ac4C, and our studies revealed novel roles for ac4C in promoting mRNA stability and translation. We also find that the acetylating enzyme N-acetyltransferase 10 (NAT10) associates with translating ribosomes, suggesting that it may monitor translation and co-translationally acetylate mRNA. Furthermore, ac4C displayed stronger base-pairing interactions with guanosine (open full item for complete abstract)

Doctor of Philosophy, The Ohio State University, 2024, Biophysics

Fragment-based quantum chemistry methods provide a way to circumvent the steep nonlinear scaling of electronic structure calculations, enabling the investigation of large molecular systems using high-level methods. First, we present calculations on enzyme models containing 500-600 atoms using the many-body expansion (MBE) and compare them to benchmarks where the entire enzyme-substrate complex is described at the same level of density functional theory (DFT). When amino acid fragments contain ionic side chains, the MBE exhibits oscillatory behavior under vacuum boundary conditions, but rapid convergence is restored using low-dielectric boundary conditions. This suggests that full-system gas-phase calculations are unsuitable as benchmarks for assessing errors in fragment-based approximations. A three-body protocol maintains sub-kcal/mol accuracy compared to supersystem calculations, as does a two-body approach combined with a low-cost full-system correction. In the next section, we use fragmentation to compute protein–ligand interaction energies in systems with several thousand atoms. Convergence tests using a minimal-basis semi-empirical method (HF-3c) indicate that two-body calculations, with single-residue fragments and simple hydrogen caps, are sufficient to reproduce interaction energies obtained using conventional supramolecular electronic structure calculations, to with 1 kcal/mol at about 1% of the cost. Additionally, we show that semi-empirical methods can be used as an alternative to DFT, to assess convergence of sequences of quantum mechanics (QM) models (of increasing size) generated by different automated protocols. Two-body calculations afford a low-cost way to construct a “QM-informed” enzyme model. This streamlined, user-friendly approach to building ligand binding-site models requires no prior information or manual adjustments, making it accessible and practical for a wide range of applications. For the latter parts of this work, we will be focusi (open full item for complete abstract)

PHD, Kent State University, 2024, College of Arts and Sciences / School of Biomedical Sciences

Wound healing is a very complex process requiring a well-orchestrated integration of multiple cellular and molecular events involving many players for antimicrobial activity and the promotion of new vascular formation. Acute wounds are healed by following the normal process of repair including inflammation, proliferation, and remodeling phases. If acute wounds fail to progress through the normal healing phases, it can develop into delayed healing or non-healing chronic wounds, particularly associated with the presence of bacterial biofilm (a community of bacteria encased in a protective matrix) and impaired angiogenesis. Conventional antibiotics frequently develop resistance and have limited efficacy against biofilm-associated wound infections. Approaches for promoting pro-angiogenic activity for wound healing have been relied on the use of bioactive molecules or growth factors, which have limitations in developing cost-effective treatment options. Additionally, thus far, each of the above aspects for antimicrobial and proangiogenic activities have been separately investigated to a great extent and an integrated approach to simultaneously addressing these three issues in a single drug delivery platform has yet to emerge. Wound scaffolds are biomaterial platforms designed to support tissue regeneration and enhance wound healing. In particular, nanoparticle-based scaffolds hold promise for treating chronic wounds due to their characteristics to exhibit higher reactivity due to high surface to volume ratio for improved cellular interactions, and easiness in surface functionalization, and controlled release of bioactive molecules. Additionally, their small size enables deep penetration into the wound bed. In this study, we propose to harness the unique characteristic of copper ion that can exhibit antibacterial and pro-angiogenic properties towards developing a cost-effective 33 scaffold for treating chronic wounds. Copper has emerged as an essenti (open full item for complete abstract)

Doctor of Philosophy, Miami University, 2024, Chemistry and Biochemistry

Pancreatic cancer remains a devastating disease with exceptionally poor prognoses due to inability to detect the disease when it is still surgically resectable and due to largely ineffective treatment modalities. Here, three engineered adenovirus vectors were developed to specifically target and suppress oncogenic activity of the HMGA1 architectural transcription factor that plays a crucial role in tumorigenesis, chemotherapy resistance and transformation of pancreatic cancer cells into pancreatic cancer stem cells. HMGA1 plays a similarly crucial role in many human cancers, including breast cancer. HMGA1 is an especially insidious oncoprotein because it is intrinsically disordered and therefore cannot be targeted by conventional small molecule drug therapy. Here, alternative adenoviral-mediated therapy approaches were designed and developed to suppress HMGA1 oncogenic activity in cancer cells and tested in MIA PaCa-2, PANC-1 and BxPC-3 human pancreatic cancer cell lines and in the ZR-75 breast cancer cell line. The engineered viruses characterized included: 1) a virus engineered to sequester overexpressed HMGA1 in cancer cells, 2) a virus engineered to express an artificial HMGA1 cis-antisense transcript and 3) a virus engineered to express an HMGA1-targeted shRNA transcript, the latter two viruses designed to suppress translation of HMGA1 mRNA into HMGA1 protein. Cancer cell characteristics after viral infection were evaluated using viability, toxicity, proliferation and wound healing assays and effects on HMGA1 mRNA transcript levels and HMGA1 protein levels were quantitatively evaluated. It was found that the virus designed to sequester HMGA1 proved more effective in suppressing HMGA1 oncogenic activity than the viruses designed to suppress HMGA1 translation.

Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, 2024, College of Arts and Sciences

Inflammatory bowel disease (IBD) is a non-contagious, chronic inflammation of the gastrointestinal (GI) tract classified into two subgroups, Crohn's Disease (CD) and Ulcerative Colitis (UC). IBD is a disease of the industrialized world, and its incidence and prevalence has increased worldwide. Short and branched chain fatty acids (SCFAs, BCFAs) produced by the gut microbiome are implicated with the immune systems inflammatory response. Chapter I summarizes our current understanding of SCFAs and BCFAs in the GI tract, fermentative pathways, etiology, inflammatory pathways relevant to the GI tract and beneficial impacts of SCFAs and BCFAs. Chapter II discusses the development and complete validation of a high throughput, fast and reliable gas chromatography-mass spectrometry (GC-MS) method with a simplified pre-treatment to quantify SCFAs and BCFAs in human stool. Chapter III summarizes a case-control study of 74 stool samples (21 healthy; 24 UC; 29 CD) measuring acetic, propionic, isobutyric, butyric, isovaleric, valeric, and caproic acid (μg/g stool) using the GC-MS method developed. Significant differences were observed for propionic, butyric and valeric acid (p < 0.05; all p values < 0.001) between healthy and IBD groups. Receiver operator curve (ROC) analysis resulted in area under the curve (AUC) value of 96% (95% CI: 0.89 – 0.98, p < 0.001). Significant differences were observed for propionic (p < 0.05; p = 0.018) and isobutyric acid (p < 0.05; p = 0.002) between UC and CD subgroups. ROC analysis resulted in AUC of 83% (95% CI: 0.66 – 0.92, p < 0.001). Acetic acid served as an endogenous, internal standard to normalize for watery stools because of its abundance and non-significant difference between groups. Chapter IV discusses a literature review of 11 published case-control studies quantifying SCFAs and BCFAs in stool between healthy, IBD, UC and CD subgroups. Valeric and butyric acid were increased in the stool of healthy groups when compared to IBD groups. (open full item for complete abstract)

Doctor of Philosophy in Clinical-Bioanalytical Chemistry, Cleveland State University, 2024, College of Arts and Sciences

Metabolomics is the study of small molecules (also termed as metabolites) in biological or botanical specimens using highly sophisticated analytical instruments, such as ultrahigh-performance liquid chromatography coupled with mass spectrometry (UHPLC-MS). The concept of computational chemistry is using software and algorithm-based analysis to pinpoint biomarkers by uncovering the molecular mechanisms that are helpful for treatment strategies. Integrating these two techniques (metabolomics and computational chemistry) enables a deeper understanding of the molecular mechanisms underlying diseases and therapeutic interventions, facilitating the identification of novel drug targets, biomarkers, and personalized treatment strategies. This integrative approach enhances our ability to unravel the complexity of biological networks and improve drug development and precision medicine. In the first half of the work, we developed a UHPLC-QTOF/MS-based untargeted metabolomics method to establish a biological model for allergic rhinitis by identifying, analyzing the regulation, and semiquantitative the key metabolites and exploring the impacted biological pathways. A computational chemistry approach was developed and applied to identify the potential therapeutic bioactive components of Astragalus radix, a traditional Chinese medicine, for allergic rhinitis treatment. In this, the protein targets were mined, and networks were constructed and analyzed to identify vital protein targets. Then, molecular docking simulations were conducted to identify the potential therapeutic components. Following this, the previously established metabolomics approach was used to investigate the therapeutic effects of the bioactive components on allergic rhinitis-induced human mast cells. In the second half of this work, we developed and validated a targeted metabolomic UHPLC-MS/MS analytical method for star anise samples. Building on our previous laboratory work, we quantified the three therapeuti (open full item for complete abstract)

Bachelor of Science, Marietta College, 2024, Chemistry

Parabens, a common antimicrobial agent used in a wide variety of industrial, cosmetic, and pharmaceutical products, have been used for many years due to their odorless and colorless properties. Structural modification of this traditional preservative may serve as a therapy for melanoma which has incidence and mortality rates that are predicted to increase by 78% and 73% respectively in the next two decades. In this study human melanoma cells and normal human epithelial cells were treated with 0.05-10mM paraben solution, consistent with LC50 values. Parabens were dissolved in ethanol or methanol solvent and added to complete DMEM for cell treatment. Methylparaben (methyl 4-hydroxybenzoate) and a paraben derivative, heptylparaben, were used. A colorimetric Caspase-3 microplate assay kit was used to assess the ability of these compounds to induce apoptosis. Previous research has shown parabens have induced apoptosis in human melanoma cells, while this study will assess parabens' ability to induce apoptosis in normal human epithelial HaCat cells versus human melanoma M624 cells and provide support for the continued study of paraben as a possible topical treatment for melanoma.

Doctor of Philosophy, Case Western Reserve University, 2024, Chemistry

Free radical-induced oxidation of lipids generates a vast array of molecules. One of these is a phospholipid ester of 8-oxo-4-hydroxyoct-2-enoic acid. A chemical mechanistic hypothesis suggested that conjugation of this a,b-unsaturated aldehyde with glutathione could lead to the formation of a molecule, which we named pseudo leukotriene (øLT)C, that bears structural and functional resemblance to leukotriene LTC4. Catabolism of LTC4 to a glycyl cysteine derivative LTD4 and of LTD4 to a cysteine derivative LTE4, suggested that øLTC would be catabolized to produce a glycyl cysteine derivative øLTD and then a cysteine derivative øLTE. Because LTE4 is acetylated in the kidneys and excreted in urine, the production of N-acetyl-øLTE in vivo was also postulated. To test these hypotheses, total syntheses of øLTs and N-acetyl-øLTE were performed. In ten steps, a divergent strategy generated each øLT from a single precursor, ethyl glutaryl chloride. Because two diastereomers were expected to be generated in the Michael addition of a thiol and the reduction of a ketone, each synthesis is expected to produce four enantiomers. Pure samples are especially important for testing the biological activities of øLTs. Therefore, HPLC methodology was developed that provided pure samples, as diastereomeric mixtures, of all synthetic øLTs. The ethyl ester intermediates øLTE-OEt could be separated into two pure diastereomers and a mixture of two other diastereomers, confirming that the syntheses are non-diastereoselective. Heavy isotope derivatives were also prepared and used to develop LC-MS/MS analyses to facilitate the detection, structural characterization, and quantitation of øLTs in vitro and in vivo. The Salomon group exploited these HPLC pure samples of øLTs to determine their production and biological activities in vitro and in vivo. In a pilot model study of Alzheimer's disease, I demonstrated that exposure of differentiated human neuronal SH-SY-5Y cells to the Ab1-42 peptid (open full item for complete abstract)

Doctor of Philosophy, The Ohio State University, 2024, Chemistry

Paraoxonase-1 (PON1) is an enzyme universally found in mammalian serum. It has been shown to hydrolyze a very broad range of substrates. The native substrates of this enzyme are likely to be lactones, while it also hydrolyzes aryl esters and organophosphates with lower efficiencies. PON1 also binds to high-density lipoproteins (HDL) in serum, which stabilizes the protein significantly and enhances its activities in vitro. It's also related to this enzyme's physiological function in mammalian serum. After being produced in liver cells, PON1 is transferred to HDL in serum and exerts anti-oxidative and anti-inflammatory functions. Loss of serum paraoxonase activities increases the risk of atherosclerosis development. Currently, due to the unstable nature of this protein, the structure information of native human PON1 is still not available. Several recombinant PON1 variants were created with higher stability and solubility. One variant along with several of its mutants (G2E6) was successfully crystalized and its structure was solved. Even though all current research is using G3C9 variant due to its better performance, a lot of information still can be exploited from G2E6 structures considering the high level of homology in their sequences. And thus, numerous mechanistic studies are performed based on this limited structural information. Based on all the available information, it is reasonable to believe that PON1 may adopt different mechanisms for different substrates. For lactonase and aryl esterase activities, key residues include the H115/H134 dyad, which have been hypothesized to serve as proton shuttlers and activate a water molecule for nucleophilic attack. This hypothesis is supported by a lot of mutagenesis and kinetics studies. But for paraoxonase activity, D269 is more likely to be the Lewis base that activates a water molecule for subsequent nucleophilic attack. Out of all the proposed mechanisms, the catalytic calcium ion is always seen as a Lewis acid t (open full item for complete abstract)

Master of Science in Engineering, Youngstown State University, 2024, Department of Civil/Environmental and Chemical Engineering

Wearable biosensors have become a valuable tool for their promising applications in personalized medicine. Cortisol is a biomarker for various diseases and plays a key role in metabolism, blood pressure regulation, and glucose levels. In this study, we fabricated an interdigitated laser-induced graphene (LIG) biosensor for the non-faradaic impedimetric detection of cortisol in sweat. A direct laser writing technique was used to produce the LIG. Gold nanoparticles (AuNPs) were electrochemically deposited onto the surface to enhance impedance response. A Self-Assembled Monolayer (SAM) was formed with on the AuNPs via 3-Mercaptopropionic acid (MPA) thiol chemistry. The carboxylic acid (-COOH) groups of the MPA were activated using EDC/NHS chemistry. Following activation, anti-cortisol antibodies were immobilized on the surface. Lastly, the LIG was incubated in the blocking agent bovine serum albumin (BSA) to avoid unwanted detection. Surface characterization of the LIG was performed at each step of modification by Electrochemical impedance spectroscopy (EIS) in a phosphate buffered saline (PBS) solution containing a 5 mM Fe(CN)3-/4- (1:1) redox couple. Further characterization of the modified LIG electrode was achieved through Fourier transform infrared (FT-IR), surfaced-enhanced Raman spectroscopy (SERS), and X-ray diffraction (XRD). The detection experiment using EIS was conducted in increasing concentrations of cortisol (0.1 pM-100 nM) in PBS. The ZMod decreased logarithmically (R2=0.97) with a 0.0085 nM limit of detection. Reproducibility was examined by percent change of ZMod at 100 nM and a 5.93%RSD (n=5) was observed. Additional analysis of sensor specificity and interference studies showed no substantial effect on detection. This research establishes the feasibility of using the gold nanoparticle decorated LIG electrode for flexible, wearable cortisol sensing devices, which would pave the way towards an end-user easy-to-handle biosensors as point-of-care diagno (open full item for complete abstract)