A product we consume daily, such as honey, may contain heavy metals like copper (Cu). Copper can get into our environment and food from urban sources such as tires, roofing materials, and generators. We therefore expect food products from more urbanized areas to show a higher concentration of copper. To test this hypothesis we evaluated copper levels in raw honey from different regions with different levels of urbanization. The honeys we tested were divided into three different categories according to their source: rural, suburban, and urban. The samples were digested by reflux with HNO₃ and H₂O₂ then it was diluted and filtered in preparation for analysis by Atomic Absorption Spectroscopy. The results suggest that the honey with the highest concentration of Cu was from an urban area, and the honey from a rural setting had the least amount of Cu. A one-way ANOVA analysis confirmed a statistically significant difference in the copper levels in raw honey from the regions (p < 0.01). The concentration of Cu in the samples of honey we analyzed ranged from 0.2 ug/g to 0.4 ug/g which is well within the recommended upper limit of 300 ug/g. Since bees collect pollen and honey from plants within roughly one mile of their hive, evaluation of the levels of copper and other metals in honey can provide a snapshot of the background levels of exposure in that area.

As our world progresses through technological advancements, much of our planet regresses as an effect of climate change, highlighting a need for underutilized resources to be brought to the forefront of industry. One avenue for transforming abundant resources into useful chemicals for generating sources like fuel is the catalytic upgrading of biomass derived molecules. However, catalysts traditionally used for these reactions are not stable to contaminants in biomass mixtures, such as water or organic acids. For biomass derived molecules to serve as precursors for biofuel and other related energy sources, more stable and efficient catalysts are needed. Our group has recently shown that a bifunctional acid–base MOF with co-localized acidic and basic sites outperforms a MOF with randomly dispersed acid–base sites for the aldol condensation reaction. To further demonstrate the importance of having the acid and base groups co-localized, I synthesized and tested three control frameworks for comparison: (1) a framework with no functionality, (2) a framework with only acidic sites, and (3) a framework with only basic sites. I then tested stability and recyclability of the bifunctional acid–base frameworks by conducting recycling experiments. I resubjected the same sample to reaction conditions for a total of 5 cycles. After each cycle, I used 1H NMR to quantify the conversion of starting material to ensure that there were no changes in catalytic activity. Lastly, I used powder X-ray diffraction (PXRD) to ensure that the catalysts maintained their crystalline structure after 5 cycles. Here I show that metal–organic frameworks (MOFs), a class of porous crystalline solids, can be used as efficient and recyclable catalysts for the aldol condensation, an important reaction for biomass conversion. Overall, this work illustrates the stability and reusability of metal organic frameworks as catalysts and thus their potential for utility in biomass upgrading reactions.

The kidney is composed of thousands of filtration units called nephrons. Within each nephron lies a tuft of capillaries, the glomerulus, that filters from the blood through a filtration barrier. Over time this filtration barrier thickens, ultimately causing decreased blood filtration. A main marker of this age-related degradation are telomeres which comprise the ends of chromosomes and protect the coding DNA from degradation. If telomeres become too short, the coding region of DNA will begin to degrade. To combat this, telomere shortening signals for cells to enter a state of permanent cell cycle arrest, senescence, which prevents replication of cells with degraded DNA. Accurately quantifying telomere length will enable the development of correlations between cell lineage and structural changes within the kidney. I hypothesize that Expansion Microscopy (ExM) and quantitative-Fluorescent In-Situ Hybridization (Q-FISH) will allow me to determine the relationship between physiological changes in the filtration barrier and single-cell telomere length. ExM enables a superresolution cellular view by embedding a tissue sample in a swellable hydrogel, achieving four-fold isotropic expansion. This technique confers greater resolution of Q-FISH signal versus traditional confocal microscopy. To determine telomere length, I developed custom analysis scripts to quantify Q-FISH signal brightness. Preliminary results indicate an increased brightness of younger mice compared to their aged counterparts. Additionally, to receive a base-pair output I compared the Q-FISH signal to the signal of a DNA region of known length, Major Satellites, determining young telomeres to have an average base-pair length of 30 kb. I am validating these results in collaboration with the Miller Lab using next-generation sequencing techniques. Future work includes concurrent application of general physiology stains to identify and measure the glomerular filtration barrier physiology. Results from this method will allow for a wealth of information regarding the relationship of single-cell telomere length and glomerular structural health.

Epigenetic factors, including histone marks, change the patterns of gene expression without altering the DNA sequence. Variations in such marks are known to account for the ability of stem cells to differentiate into various cell types, but a preliminary experiment conducted by a former member of the Vaughan Group, Dr. Marcus Woodworth, has revealed that even in a phenotypically homogeneous, terminally differentiated cell population, the presence of H3K27me3, a repressive histone mark, varies on the HOXC gene of human retinal pigment epithelium (RPE1) cells at single-cell, single-loci level. My role is to evaluate the two possible origins of such heterogeneity: inheritance (histone mark varies due to events that happened during differentiation, or the random drift after differentiation, and the variations are kept within each lineage of cells), and multiple mark co-repression (one histone mark varies, but summing its effect with another histone mark that perform a similar function lead to the observed functional homogeneity), and to validate that such pattern exists among a broader range of genes. To achieve these ends, I profile selected histone marks (H3K27me3 and H3K9me3) on genes that experience different types of regulations during differentiation (HOXC, GAPDH and SIX6), using imaging-based methods, including the time-lapse imaging of live cells to map out lineages, and expansion microscopy (ExM) to capture fluorescently labeled histone marks at single-loci level. If the hypothesized origins are true, a significant difference in the number of histone mark clusters around the genes of interest would be observed between cells of different lineages, and complementary variation patterns would be observed between H3K27me3 and H3K9me3. The study reveals the nuances of histone mark dynamics on the single-cell, single-loci level, and optimizes an imaging-based method that has the potential for multiplexing at high spatial resolution, thereby providing a powerful tool for further studies on epigenetics.

Glomeruli are the basic filtration unit of the kidney. The current understanding of its physiology is limited by the partial or 2D analysis of its structural components. The Vaughan Group uses optical super-resolution microscopy in combination with advanced chemical labeling techniques and powerful data analysis approaches to perform high-resolution 3D reconstruction of the whole mouse glomeruli. Overall, the work has the potential to provide a novel understanding of the glomerular structures and how they are altered in aged and diseased conditions. The labeling of the overall morphology of the glomeruli is achieved by chemically labeling the distribution of abundant macromolecules (carbohydrates, amine, and DNA) using Fluorescence Labeling of Abundant Reactive Entities (FLARE). Though we could visualize the general physiology of the sample with FLARE, incorporating specific targeting of molecules with FLARE is still challenging. My role is to optimize the FLARE protocol to add the capability of labeling the distribution of specific molecules using immunolabeling. The most challenging part is that all the fluorophores labeled prior to FLARE will be bleached out by the strong oxidation step while labeling carbohydrates. I am focusing on exploring possible workarounds to incorporate immunostaining with FLARE. The only way to bypass the bleaching fluorophores is to label dyes after the FLARE. However, the FLARE involves the gelation part, and the gel makes antibodies which are linked to fluorophores hard to get into the sample. So, instead of using regular secondary antibodies, I use biotin and then link to the streptavidin dye, which is smaller in size and easier to enter the sample. With this optimization working, we could incorporate whatever target of interest with high resolution on top of three general stains provided by FLARE, giving us an extra degree of information for our 3D reconstructions of glomeruli.

The four letters in DNA (ATGC) construct the basis of life as we know it. Unnatural base pairing xenonucleic acids (ubp XNAs) are synthetic nucleic acids that can be used orthogonally to the 4-letter code. XNAs have the potential to revolutionize a myriad of biotechnologies, but commercial sources of XNA nucleotides are limited and expensive. Here, we fill one step of an enzymatic cascade required to sustainably produce XNA nucleotides. Nucleoside phosphorylases (NPs) are enzymes that catalyze the reversible phosphorolysis of nucleosides to their base and sugar components. We purified and assayed promiscuous NPs from two thermophiles, Geobacillus thermoglucosidasius (GtNP) and Thermus thermophilus (TtNP). Using a combination of mass spectrometry and fluorescence assays, we show that these phosphorylases have activity on a subset of three XNA substrates (B, Sⁿ, and P). This enzymatic pathway allows us to synthesize non-standard nucleotides in a cost-efficient manner and provides a crucial tool for the biosynthesis of XNAs.

Exclusionary zoning laws — which limit population densities and land uses in specific neighborhoods — are a typical feature of American municipal land use regulation. An extensive body of evidence links traffic-related air pollutant (TRAP) exposure to adverse health effects. Using zoning data and a model of TRAP levels in cities across the Seattle metropolitan area, I hypothesize that TRAP exposure will be greater on average in zones where higher-density housing is an allowed use, and lower on average in zones reserved for lower-density housing. I used the software package QGIS to spatially join zoning and air pollution data and used the software package R to perform correlation analyses between zone types (classified by maximum population density) and three common TRAPs (NO₂, black carbon, and ultra-fine particles.) This research highlights the public health implications of normative policy regimes like exclusionary zoning. These results can assist elected officials and planners in pursuing a more geographically distributive approach to increasing housing supply in the Seattle area, in order to minimize the TRAP exposure burden – and associated adverse health effects – faced by residents.

The synthesis of key organic molecules often requires toxic, expensive, non-reusable reduction agents and extreme conditions. In recent years, electrochemistry has emerged as a sustainable alternative to standard methods, but this approach is often hindered by high energy barriers for electron transfer to the substrate. Electrocatalysts address this challenge by shuttling charge between the electrode and dissolved substrates, accessing lower transfer barriers, and reducing the overall energy needed. Current electrocatalysts, however, cannot stabilize reactive intermediates, which often leads to harmful side reactions and degradation of the electrode. We hypothesize that redox-active supramolecular cages can address this limitation by both shuttling charge and providing unique microenvironments capable of stabilizing intermediates. Previously, we synthesized two tetrahedral supramolecular cages that incorporate redox-active perylene diimide (PDI) and pyromellitic diimide (PMDI) motifs. Using cyclic voltammetry, we then showed that both cages can lower the voltages required for the electroreduction of vicinal dihalides to alkenes, indicating electrocatalysis. To better understand these results, I used density-functional theory (DFT) calculations to obtain computer models of the PDI and PMDI cages. These DFT-optimized structures revealed significant differences in charge density between redox centers due to electron-donating functional groups, which may explain why the PMDI cage lowered the substrate reduction voltages more than the PDI cage. With these models, I have also studied the shape and volume of the cages’ internal cavities, thereby providing information about substrate compatibility. I am conducting additional DFT analysis to understand how modifications to the ligand motifs may alter the electrocatalytic behavior. By continuing to investigate supramolecular cages for reductive electrocatalysis, I aim to contribute to the development of low-waste synthetic strategies for the production of alkenes and other commercially significant organic compounds.

Vulcanized rubber, the main component of tires, is prized for its chemical durability and thermal stability. These properties, however, make disposal difficult and contribute to the increasing problem of polymer waste. To broaden the applications of end-of-life tires, we developed a method to chemically upcycle polybutadiene, a primary component of vulcanized rubber, via selenium-mediated allylic amination. We hypothesized that functionalizing the backbone of crosslinked polybutadiene with sulfonamide groups—without breaking their double bonds—would result in favorable thermal properties, creating a new life for the crosslinked polybutadiene. We used infrared spectroscopy and scanning electron microscopy with energy dispersive X-ray spectroscopy to confirm the aminated crosslinked polybutadiene’s molecular structure and differential scanning calorimetry and thermogravimetric analysis to measure its changes in thermal properties. Our research has future implications for the reduction of tire waste and reprocessing of other end-of-life crosslinked polymers.

The catalytic ability of an industrial heterogeneous catalyst is determined by the interactions between the active sites, which are often transition metals, and the support. Insights into the interplay between the active sites and support during catalysis are difficult to gain because of the inherent complexity of heterogeneous surfaces. Alternatively, molecular catalysts are well-defined, and can be studied by a range of spectroscopic characterization techniques. To model multi-active site dynamics on a molecular scale, the Velian group has developed a system involving a cobalt selenide cluster with amido phosphine ligands that are used to tether transition metals that act as catalytically active sites onto the cluster surface. My project is probing the tri-metalated clusters’ (M₃Co₆Se₈L₆; M = Cr, Mn, Fe, Co, Cu, Zn; L = PPh₂N^-Tol, Ph = phenyl, Tol = 4-tolyl) ability to catalyze intramolecular carbon-hydrogen (C-H) amination. Previous work has shown that these clusters are remarkable catalysts for carbodiimide formation, but we have yet to compare reactivity among the tri-metalated clusters. I probed the transformation of aliphatic azides to pyrrolidines, a class of 5-membered-N-heterocycles with. This study seeks to understand how the reactivity of the clusters change as edge metal identity changes, and the role of the three active sites during catalysis. A substrate scope has shown how the steric and electronic profile of the azide affects the capability of the clusters for this reaction. This research provides insights into metal-support interactions that are important for heterogeneous catalysis. Development of next generation catalysts that can perform complex transformations benefits from the information these studies provide.

Solar energy is a promising form of renewable energy that will play a major role in reducing carbon emissions. Perovskite-based solar cells have attracted significant attention due to their high power conversion efficiency (PCE), which reached 26.1% this year, surpassing commercial silicon’s (23.3%). High PCE, low cost of materials, and ability to be solution processed make perovskite solar cells a prime candidate to replace silicon. However, efficiencies are still below the theoretical limit and these materials suffer from limited operational stability. To tackle these problems, scientists have focused on minimizing active layer and interfacial defects which act as barriers for charge extraction in a solar cell, lowering the device efficiencies. Defects also electronically dope the perovskite layer, changing the recombination kinetics in the sample. The goal of this project is to quantify how the electronic doping and defect concentration of the perovskite sample is affected by surface passivation treatements via fluence dependent photoluminescence (PL) and time resolved photoluminescence (TRPL) spectroscopy. By solving the kinetic equations at the basis of charge recombination, we can extract the rate constants that correspond to different charge recombination pathways. We pioneer a global fitting analysis to simultaneously fit TRPL and PL measurements for robust determination of these kinetic constants that are subsequently used to determine the doping density of films before and after passivation. We show that the electronic doping density is higher than previously reported in literature, and that this doping is reduced with a surface passivation treatment. We collaborate with the University of Arizona to correlate our measured electronic doping density to electrochemically measured defect densities on the same samples. This work will provide an implementable tool to quantitatively assess electronic doping and defect density values for various perovskite compositions which will be useful for optimizing future solar cell devices.

Although immunotherapy with T-cells is successful in treating non-solid cancers, targeting solid cancer tumors remains a challenge. Unlike T-cells, bacteria can colonize solid tumors and thrive in a hypoxic tumor microenvironment (TME). This means that bacteria could be used to treat solid tumors that T-cells cannot reach. I propose to develop new bacterial immunotherapies that can be used as an alternative treatment method to fight solid cancer tumors. Specifically, I aim to engineer E. coli that secretes therapeutic payloads upon sensing the TME. My research focused on comparing secretion efficiencies of signal peptides and secretion tags. Signal peptides are short sequences that transport cargoes, such as therapeutic proteins, to the periplasm. Similarly, secretion tags are small secreted proteins that can transport a partner cargo fused to them to the extracellular medium. From literature, I selected the signal peptide PelB and the secretion tag YebF. I compared their efficiencies in secreting the cargo human interleukin 2 (h-IL2), an immunostimulatory cytokine. To this end, I expressed h-IL2 with either the genes for the signal peptide or secretion tag fused to the N-terminus and a detection tag on the C-terminus, in an E. coli expression strain. I induced the expression of cargoes, after which I isolated the proteins that were secreted into the extracellular medium. I detected the proteins through quantitative Western blot analysis. I concluded from my experimental data that the cargoes were secreted at a higher concentration with YebF than with PelB. I plan to repeat this experiment with another secretion tag, OsmY. The next step is to use the secretion system with the highest secretion yield to secrete a variety of potential immunomodulatory cargoes. I plan to evaluate their effects on immune signaling and their ability to eliminate tumor cells.

The nitrile imine produced by photolysis of 2,5 dimethyltetrazole undergoes a cross-linking reaction with the amide group in peptide-tetrazole conjugates and tetrapeptide-nucleotide complexes. In our work, we synthesized various peptide conjugates furnished with 2,5-diphenyltetrazole phototag. Upon laser pulses at 250 nm, nitrile imine intermediates can be generated by loss of N2 from tetrazoles. These nitrile imines can then crosslink with other parts of the molecule that contains amide groups. These crosslinking reactions are quite effective, achieving about 50% conversion with just two laser pulses at about 2 mJ. We could detect the formation of crosslinked products by tandem mass spectrometry. The UVPD-CID-MS3 spectra of these conjugates showed unique fragments including internal fragments of peptide sequence, indicating possible crosslinking. Moreover, we can confirm the structures and compositions of these crosslinked products using UV–Vis action spectroscopy and cyclic ion mobility mass spectrometry (c-IMS). By comparing experimental and calculated data, we confirmed the presence of nitrile imines and certain crosslinked products. We also explore thermal chemistry when nitrogen gas is lost from the peptide-tetrazole conjugates, and it seems to be a mildly energy-consuming process. The extra energy from breaking down tetrazoles is likely driving the reaction towards forming crosslinked structures involving peptide amide groups. Digging into the mechanism of this reaction, we found the proton transfer as the initial step, followed by a series of steps like cycloaddition and breaking of certain chemical bonds. Interestingly, other reactive groups, like cysteine thiol, do not interfere with this process. Within the complex of peptide conjugate and 2′-deoxycytidylguanosine, the intermolecular crosslinking efficiency is over 80%. The CID-MS3 and optimized structure showed the nitrile imine selectively targets guanine. In particular, the discovered reactivity of peptide amide groups toward nitrile imines appears promising as it provides potential clues to cross-link structure elucidation and conformational analysis.

As global demand for renewable energy grows, new avenues emerge to design cost-effective routes for enhancing energy storage and capture, such as high-capacity batteries and next-generation solar cells. This study focuses on characterizing the underlying dynamics of model energy storage and conversion materials in response to oscillating stimuli. The near-term goal is to develop a generalizable method of doing this that may be adapted to a wide array of different materials. The long-term goal is to apply this method to understand the fundamental chemical dynamics involved in the function of these materials, ultimately accelerating progress in improving ion storage media for batteries and enhancing photovoltaic efficiency. The first target is to selectively detect vibrational signatures associated with electron accumulation in response to an applied potential in semiconducting nanoparticles, with applications as anode materials for alkali metal ion-based batteries. I aim to do this by developing a phase-sensitive detection method using lock-in amplification, which allows measurement of small spectral signals that would otherwise be undetectable due to noise. To hit this target, a well-studied, reversible ferrocene/ferrocenium system was subjected to alternating current (AC) electrochemical modulation and probed using visible light. The electrical signals induced in these materials were analyzed at characteristic frequencies. By monitoring the spectral fingerprints of each ferrocene and ferrocenium, I will extend the application of these spectroelectrochemical methods to the model titanium(IV) oxide (TiO₂) electrode. Moving forward, this method to probe reaction dynamics may be applied to analyze the stabilizing effect of performance increasing modifications on anode materials within alkali metal-based batteries.

Black phosphorus (bP), an allotrope of phosphorus, is a 2D Van der Waals material composed of corrugated layers of phosphorus atoms. Few-layer bP is a semiconductor with interesting physical properties, including relatively high carrier mobility and layer-dependent band gap. These properties may be harnessed for applications including nitrogen fixation photocatalysts, thin film transistors, and sensing devices. One limitation that must be overcome before bP can be used in devices is its degradation into phosphoric acid when exposed to oxygen, water, and/or light. Finding passivation methods is crucial for the future use of bP in electronics or photochemistry. As each passivation treatment changes bP’s electronic properties, it is important to find protection methods that are compatible with each use case. To investigate possible methods to reduce surface degradation, we exfoliate bP in solution and treat it with a passivation candidate. We then use ultraviolet-visible light (UV-Vis) spectroscopy to track the amount of unoxidized bP that remains in solution during ambient exposure. Since bP absorbs strongly across the UV-vis region, while the decomposition products, phosphorus oxides, do not, UV-vis is an ideal method for measuring degradation. Possible treatments include attaching alkoxy or thiolate groups via peroxides or disulfides to bP edges to protect the particularly reactive dangling bonds, treatment with radical scavengers such as butylated hydroxytoluene, or noncovalent protection with Tetracyanoquinodimethane (TCNQ).

In this pilot project, we conducted think-aloud interviews with undergraduate students who had recently completed CHEM 142 as they worked through a set of multiple-choice, general-chemistry questions in order to investigate the levels of cognition they exhibited during problem solving. In collaboration with members of my research group, I designed recruitment materials and selected eight undergraduate interviewees from a pool of around 50 volunteers, with attention to diverse identities— including race, gender, sexuality, disability, prior courses in science/math, and parent’s educational background—as such identities and experiences may influence what problem-solving skills one has acquired. I co-developed the interview format, and conducted two of eight interviews. Participants were asked to narrate their approach while solving four multiple-choice CHEM 142 questions, then to critique the questions in terms of relative difficulty. The questions we used were previously characterized according to Marzano and Kendall’s New Taxonomy of Educational Objectives by a group of chemistry educators from our research group. I developed and refined a modified version of Marzano’s Taxonomy inspired by Teodorescu’s work in introductory physics (Teodorescu et. al., 2013), and am using it to code the interviews according to cognitive level exhibited by the interviewees. My preliminary results suggest that students approach a particular question in a variety of ways, engaging different levels of cognition. Students who display mastery of a concept tend to utilize lower cognitive levels, apparently due to familiarity with the concepts. Students who seem less familiar with or state they are struggling with a topic tend to employ more cognitively demanding strategies, whether or not they arrive at the correct answer. I will also discuss a comparison of the cognitive level exhibited by students to the level predicted by previous group members for each question, and any observations unique to different student identities.
 

With the increasing use of lithium-ion batteries, the trajectory of the modern world’s energy needs calls for an improvement in their safety and functionality. Current lithium-ion batteries use a lithium salt dissolved in organic carbonates, which results in a liquid electrolyte with high lithium dissociation and fast conductivity. However, the use of organic solvents makes these electrolytes flammable and prone to combustion in the event of a dendritic formation causing a short. One approach to potentially improving these electrolytes is a solid single-ion polymer electrolyte where the anion is part of the polymer chain to potentially allow higher conductivity and lithium transference numbers. Our approach is to synthesize single-ion copolymers with varying weight percentages of a single-ion monomer (SIM) to control the ion concentration and a monomer with an oligomeric ethylene oxide sidechain (ONDI-12) to lower the glass transition temperature. The goal of my work is to identify the ratio of the two monomers that gives optimal conductivity, improving the potential use of our copolymer as a solid single-ion copolymer electrolyte in lithium-ion batteries. Initial electrochemical impedance spectroscopy measurements of our previous copolymers indicate that lower ion concentrations and lower glass transition temperatures resulted in increased conductivity with 20 wt.% SIM. Building from this work, I synthesized the single-ion monomer (SIM) with an attached anion incorporated into the structure to facilitate lithium cation motion. I anticipate copolymerizing this SIM with ONDI-12 at lower SIM percentages using ring-opening metathesis polymerization (ROMP) and subsequently measuring their conductivity. Identifying the SIM to ONDI-12 ratio that optimizes conductivity will improve our understanding of these materials and potentially advance future polymer electrolyte design.

Since their introduction to clean energy applications, organic-inorganic lead halide perovskites have received great attention for their potential to create highly efficient, manufacturable and cheap solar cell devices. To make effective perovskite solar cells, charge transport layers are used to remove electrons and holes from the bulk perovskite semiconductor, increasing current, voltage and power conversion efficiency. Phosphonic acid self-assembled monolayers (SAMs) are a common hole transport layer. The phosphonic acid binds to the transparent conductive oxide electrode while an organic head group forms the SAM/perovskite interface. This head group is key for charge transfer and voltage characteristics, but the structure-function relationship is still poorly understood. My project investigates the role that deposition techniques and electronic structure play in the optimization of this SAM/perovskite interface. Expanding from the standard two step spincoating SAM/perovskite deposition method, I explored whether the codeposition of the two layers or the addition of a SAM solvent wash step produced an improved interface. I also fabricated films using several different SAM compositions to test for performance trends and improvements compared to the current field standard SAM, Me-4PACz. I collected photoluminescence lifetimes, quantum yields and solar simulation measurements to evaluate film performance. Preliminary data shows that neither the washing step nor codeposition add any performance benefit, but the single step codeposition achieves a more streamlined manufacturing method. Two of the new experimental SAMs performed comparably to Me-4PACz. These results encourage codeposition of the SAM/perovskite interface as the most efficient method to create high quality devices and show promising alternatives to the industry standard Me-4PACz SAM.

Nickel phosphide (Ni2P) nanoparticles have been gaining attention due to their ability to catalyze various clean energy-relevant reactions, e.g., the hydrogen evolution and carbon dioxide reduction reactions; however, Ni2P has been known to corrode in aqueous electrolytes. Studies have indicated that nickel phosphide alloys have shown a small amount of oxidation to nickel phosphate at oxidizing potentials or complete dissolution at more aggressively oxidizing potentials. However, understanding of the speciation and kinetics of oxidation is limited. Therefore, we aim to understand the corrosion mechanism of Ni2P in neutral buffered electrolyte; which we hypothesize to show significant conversion to nickel phosphate at oxidizing potentials. First, I synthesized colloidal Ni2P nanoparticles from NiCl2 and tris(diethylamino)phosphine [P(NEt2)3] in oleylamine. Techniques such as powder X-ray diffraction, transmission electron microscopy, and thermogravimetric analysis confirmed the formation of uniform 5 nm diameter nanoparticles. Next, we probed the electrochemical corrosion of Ni2P through anodic polarization and operando X-ray absorption spectroscopy (XAS). In order to prevent premature oxidation of Ni2P, all procedures were performed in an air-free environment which posed many challenges for the preparation of the electrochemical cells, especially the operando XAS cell. Finally, we found that Ni2P nanoparticles corrode upwards of 0.4 V vs RHE and can no longer be restored when anodically polarized beyond 0.6 V vs RHE. Future experiments will probe the corrosion of Ni2P in acidic and basic electrolytes. This study aims to understand how Ni2P can be used industrially to replace rare and expensive metals, such as platinum, as electrocatalysts to electrify the petrochemical industry and reduce greenhouse gas emissions.

Upconversion (UC) is a non-linear optical process where a material absorbs two lower energy photons and subsequently emits one of higher energy. Currently, inorganic UC materials used in lasers and photovoltaics are primarily lanthanide-based. However, a few transition metals also exhibit UC, such as Re4+ , Os4+, Ti2+, Ni2+, and Mo3+, and due to their high oscillator strengths, d-d transitions, and a strong ligand field dependency, offer the potential for greater tunability and efficiency in upconverting optoelectronics than their than their lanthanide counterparts. The goal of this work is to increase Re4+’s PLQY by isovalently doping low-phonon vacancy-ordered double perovskites (A2BX6 : A = Cs+, NH4+; B = Ti4+, Zr4+; X = Cl-, Br-) with rhenium to minimize non-radiative decay that can occur through defects and lattice vibrations. This has been attempted via schlenck line synthesis of the host lattice and coprecipation and ion-exchange doping procedures. While [ReX6]2- has previously demonstrated near-IR to visible upconversion in the bulk, this work aims to characterize its upconversion mechanism on the nanoscale with variable temperature and time-resolved photoluminescence. If made successfully, the colloidal stability of Re4+:Cs2TiBr6 nanocrystals would allow for new post-synthetic processing avenues including electrohydrodynamic inkjet printing and core-shelling, and new applications in flexible electronics.

In a recent breakthrough, bullvalene, renowned for its “shape-shifting” molecular nature with over 1.2 million degenerate isomers, has been successfully integrated into polymer backbones. This integration addresses challenges in solubility and thermal properties crucial for tailoring polymers used in manufacturing diverse products ranging from phone screens to organic solar cells. This project aims to deepen our understanding of the interplay between fluxionality and thermal properties by examining the thermal stability of small molecule bullvalene models. Through extrapolating insights for bullvalene-substituted polymers, our research seeks to contribute to the advancement of the development of advanced materials suited for varying thermal conditions. We synthesized small molecule bullvalenes to mimic polymer chains, subjecting them to diimide reduction to suppress fluxionality before comparison with their fluxional counterparts. Their thermal properties were characterized using Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry (DSC). Key findings reveal a decrease in glass transition temperature upon reduction of bullvalene, highlighting the impact of fluxionality on thermal stability. Future work will delve deeper into exploring the thermal properties of small molecule models, providing insights into polymer behavior. We anticipate bullvalene as an internal plasticizer capable of modulating rigidity, solubility, and thermal properties within different classes of polymers, thus enabling a more efficient and cost-effective large-scale industrial production of a wide array of polymeric materials.

In recent years, there has been a stark decline in bee colony populations. Many studies believe neonicotinoids to be a significant contributor to this problem. Neonicotinoids are insecticides that are often used in farming or agricultural settings. One of the neonicotinoids suspected to play a role in the bee colony decline is Imidacloprid. Imidacloprid is believed to directly negatively affect the navigational skills of bees. It interrupts their homing abilities, making it difficult for them to relocate their hive after coming in contact with Imidacloprid. Previous research projects have been conducted to study the affect of Imidacloprid on bees, but the proportion of Imidacloprid to the bee itself is not realistic of what they would be affected by in a veritable agricultural setting. Here we demonstrate an extraction protocol based on sample clean-up with both ion-exchange and reversed phase separations prior to targeted gas chromatography-mass spectrometry (GC-MS) analysis. This protocol is similar to the QuEChERS (Sigma, St. Louis, IL) method, however, the extraction protocol presented here shows how capturing and concentrating the Imidacloprid while washing away the other matrix components can help improve sensitivity and selectivity for Imidacloprid. Our targeted detection strategy involves selected ion monitoring (SIM) to decrease limits of detection. Initial findings show that C-18 separation is optimized by adding a centrifugation step prior to SPE (solid phase extraction). As we continue to develop a targeted detection strategy for Imidacloprid, the hope is that the strategy will be used in future farming and agricultural environments to determine if the level of neonicotinoids being used in specific fields is detrimental to the bee colony population in the area. 

Recent progress in synthetic biology has focused on utilizing probiotics as therapeutic production factories in the gastrointestinal environment to treat GI-related diseases. Although oral administration of probiotics is a convenient method for patients, a key challenge lies in the poor survival rate of probiotics in gastric and intestinal areas. Engineered living materials (ELMs), which are comprised of genetically engineered microbes embedded in a polymer matrix, present a novel formulation for orally-administered probiotics. Herein, we developed ELMs containing probiotics in a protein-based polymer matrix, aiming to enhance their viability in the GI tract. The ELMs’ photocurable polymer matrix allows us to 3D print our formulation into oral tablets. To form our protein-based polymer matrix, we functionalized bovine serum albumin with polyethylene glycol diacrylate. We then added a photoinitiator and E. coli Nissle genetically engineered to produce tryptamine (an anti-inflammatory agent) and subsequently photopolymerized this resin to 3D print probiotic tablets. We placed these tablets through a simulated gastrointestinal tract and observed cell escape using optical density measurements and cell viability through live/dead staining and fluorescence imaging. Liquid-chromatography mass-spectrometry was used to quantify the extent of therapeutic bioproduction in vitro by our ELMs over time. Overall, we found that the ELMs successfully delivered viable probiotic cells able to perform in situ therapeutic bioproduction. Furthermore, we observed that encapsulation of probiotics in ELMs yielded a higher survival rate of cells in the GI tract, suggesting that our polymer matrix formulation protected cells and allowed for extended proliferation and colonization in the colon. These findings are also supported by our observations that ELMs produced significantly higher amounts of tryptamine in the GI tract compared with non-ELM, free cells. The findings from our study can be applied to further development of orally-administered probiotic therapeutics, and show promise for future directions in drug delivery.

Lck is a lymphocyte specific tyrosine kinase involved in T cell activation in response to T cell receptor (TCR) mediated signaling. T cell activation is essential for the adaptive immune response, as it results in the proliferation of T cells after the detection of a peptide presented on a Major Histocompatibility Complex (MHC) and the production of cytokines necessary for immune response coordination. Lck activity is dependent on its global conformation, which is dynamically regulated via phosphorylation on its activation loop and C-terminus tail. Upon TCR engagement, active Lck phosphorylates the CD3ζ chains of the TCR complex, transducing the intracellular signaling events that activates T cells. Because Lck activity is dependent on its global conformation, we sought to map the conformational changes in Lck upon TCR simulation, as well as identify cysteine-reactive fragments that target and stabilize Lck in its conformational extremes. Lck has few endogenous cysteines, so we performed a yeast-growth-based deep mutational scan (DMS) of Lck–in which we utilized Lck’s toxicity to yeast to calculate the activity scores of ~5,000 Lck mutants–and identified 109 solvent-exposed, wild-type-like cysteine mutants of Lck. Expressing these wild-type-like cysteine mutants in T cells, and utilizing competition-based mass spectrometry, we can quantify changes in electrophilic reactivity of the cysteine side chains in the wild-type-like cysteine mutants upon T cell receptor (TCR) stimulation. Thus far, I have identified six wild-type-like cysteine mutants of Lck that are quantifiable using mass spectrometry and exhibit reactivity to our set of cysteine-reactive fragments, some of which show differential reactivity upon TCR simulation and fragment selectivity. Currently, I am using these mutants to map the dynamics of a hyperactive mutant of Lck. These quantifications provide insight into changes in the conformational flexibility of Lck, accessibility of the mutated residue sites, and intramolecular protein-protein interactions of Lck upon TCR stimulation.
 

Soluble factor signaling between immune cells and fibroblasts is critical in regulating biological processes. However, it is often dysregulated in diseases and leads to physiological changes, including airway inflammation in asthma and allergies. One immune cell type that can be attributed to airway inflammation is eosinophils (EOS). When activated by interleukin-3 and heat-aggregated immunoglobulin G, EOS release certain soluble factors associated with the activation of lung fibroblasts. To investigate the interactions between human lung fibroblasts (HLFs) and EOS, we used the open microfluidic coculture device. This device has two chambers, in which two types of cells can be cocultured in the shared media while being physically separated by a half wall. We found that HLFs in coculture with activated EOS had the highest levels of proinflammatory gene expressions and proinflammatory cytokines. However, the exact mediators responsible for promoting these biological processes are still uncertain. We hypothesize that EOS secrete a protein, transforming growth factor alpha (TGFa), to be consumed by HLFs, triggering proinflammatory responses of HLFs. The goal of this study was to elucidate the roles of TGFa in airway inflammation. HLF-EOS cocultures are seeded in the microfluidic coculture device, then TGFa and their respective cellular receptors are neutralized using antibodies. Then, reverse transcription quantitative-polymerase chain reactions are used to quantify gene expression levels relevant to proinflammatory responses of HLFs, in addition to multiplex immunoassays to analyze the secreted soluble factors from both cell types. We anticipate that HLF-EOS cocultures treated with neutralizing antibodies have lower expression levels of proinflammatory genes than cocultures without antibodies. Findings from this study will help us better understand the key regulators that promote proinflammatory behaviors of HLFs in airway inflammation.

Gastrointestinal symptoms are a common comorbidity of autism spectrum disorder (ASD), and individuals with the disorder tend to have a distinct gut microbial community composition and circulating metabolomes. My work in the Rajakovich Group focuses on a gut-derived metabolite, 4-ethylphenolsulfate (4-EPS), found in higher abundance in ASD mouse models and children with ASD. 4-Ethylphenol (4-EP), its precursor, is produced by gut microbiota before host-mediated sulfation, but the microbial biosynthetic pathway is unknown. A proposed metabolic pathway suggests the microbial stepwise conversion of plant-derived complex polysaccharides to 4-EP. My project goal is to identify a gut microbial enzyme responsible for the first step of this proposed pathway: a hydroxycinnamoyl esterase. I used literature searches and bioinformatics tools to identify characterized bacterial cinnamoyl esterases and candidate enzymes. I designed plasmids for two candidate enzymes (both from E. faecium, known to colonize the gut) and one characterized esterase (from L. plantarum). Currently, I am working on expressing the proteins in E. coli cells and purifying them by affinity chromatography. Once purified, I will assess the enzymes for their anticipated cinnamoyl esterase activity by incubating them with dietary hydroxycinnamic acid esters and detecting products with high-performance liquid chromatography (HPLC) and UV/Vis spectroscopy. Since the candidate enzymes are homologs of confirmed esterases and have conserved catalytic motifs, I hypothesize that they will have hydrolytic activity. If correct, I will see consumption of the substrate (no detection) and detect the anticipated products. Positive results from these assays would complement ongoing work by the lab to identify other E. faecium enzymes in this proposed pathway. Though it is debated if 4-EPS is causal to the disorder or simply a biomarker, elucidating its biosynthetic pathway and studying the biochemistry of gut microbes will contribute to detangling the gut’s role in ASD.

Eutrophication fuels toxic algal blooms that can harm biodiversity and human health. Phosphate is often the limiting nutrient in freshwater ecosystems and, when in excess, causes eutrophication. Our study compares urban lakes prone to algal blooms to rural lakes with fewer anthropogenic sources of pollution to better correlate nutrient dynamics of pacific northwest lakes to population density. We collected sediment and water samples from two lakes in the Seattle area and two lakes in a more remote setting, then measured phosphate uptake and release over time using ion chromatography. To determine the potential for the sediments to uptake phosphate, we placed air-dried sediment samples into a phosphate solution and measured the concentration over time. We also put the sediment in distilled water and measured the phosphate released from the sediment over a 24 hour time period. These two data sets allow us to quantify the capacity of sediment to store and release phosphate into the surrounding environment. Our research shows sediments from urban lakes release more phosphate and have a reduced ability to uptake nutrients from the water. This suggests that the lake will continue to eutrophy whereas the lakes more removed from human activity have a better ability to mitigate excess phosphates. This model for assessing the ability of sediment to store phosphate allows prediction of future eutrophication events.

We have observed that conventional respiratory pathogen sampling methods, such as pharyngeal swabs, elicit unpleasant experiences for both adults and children. Particularly for pediatric patients, having a non-invasive, enjoyable sampling approach is crucial to facilitate prompt diagnosis and treatment. In prior research, we introduced a novel saliva sampling device, the CandyCollect. This lollipop-inspired device, with its isomalt candy coating, is specially produced and surface-treated for capturing pathogens from saliva, providing a pleasant sampling experience to child patients. Clinical studies approved by multiple Institutional Review Boards (IRB) revealed an average candy dissolving time for CandyCollect (with a mass of 0.90g~1.10g) of 3.51 minutes, with a minimum of 1.25 minutes. To compete with original sampling methods which take up to 10 seconds, it is desirable for CandyCollect to have a shorter sampling time around 15–20 seconds. Therefore, this study aims to decrease the dissolving time by introducing a new CandyCollect recipe and design. For the new candy recipe, we replaced isomalt with a mix of glucose and sucrose. Additionally, baking soda (sodium bicarbonate) was added to increase the candy’s contact area with the tongue. An ongoing experiment will assess if baking soda affects PCR results for pathogen samples, and this modified recipe will be employed in a new clinical study. Concurrently, we have modified the CandyCollect design by placing the candy on the same side as the spiral, with a small candy reservoir beneath the spiral to decrease the mass to 0.06g-0.1g. To validate the efficiency of this new design, we plan to conduct another clinical study recruiting 30 younger participants and obtaining feedback about the new design. This collaborative study will provide valuable insights towards achieving a faster dissolving time, ultimately enhancing the viability of CandyCollect as an improved and more efficient replacement for conventional sampling methods.

Traditional methods of sampling for respiratory illnesses can be uncomfortable and possibly act as a deterrent for children and adults. The CandyCollect is a lollipop-inspired device developed to aid in the testing for respiratory infections. Fabricated by a computer numerically controlled (CNC) mill, the device contains a plasma-treated microfluidic channel that collects pathogens from saliva; the addition of strawberry-flavored isomalt candy on the head of the stick aids in the resemblance, feel, and taste of a traditional lollipop. In previous remote human subjects studies, approved by the Institutional Review Board (IRB), we demonstrated the ability of CandyCollects to detect commensal bacteria, Streptococcus mutans and Staphylococcus aureus, in healthy adults using qPCR and detect infections in adults displaying symptoms of respiratory infections. From these and IRB-exempt device improvement studies, we aimed to shorten the dissolve time of the candy to make it comparable to the sampling time of nasal and oral swabs. In order to investigate the usability and accuracy of the CandyCollect for children, our team launched the Parent-Child Dyad Study. The IRB-approved study recruited parents with children displaying symptoms of respiratory illness via online forums. The 40 eligible dyads, consisting of children aged 5-15 and their parents, received a kit including a nasal swab, two mouth swabs, and three CandyCollects. After using the CandyCollects, the children and parents completed their respective surveys asking them about their comfort and experience with the different testing methods. As with all remote sampling methods, limitations included potential biases towards nasal and/or mouth swabs. However, the CandyCollect is no more susceptible than currently employed methods. User feedback data demonstrates children preferring the CandyCollect over mouth and nasal swabs, reporting better taste and comfort. The parent survey results also show preference for the CandyCollect. Here we will present the results of their user feedback.

Cell co-culture systems are used to study intracellular interactions by culturing distinct cell-type populations within a shared environment. This in vitro method is more representative of the highly complex and diverse processes that occur in organisms, allowing accurate insight into mechanisms of cell signaling pathways, disease, drug interactions, etc. Existing designs can be two- or three-dimensional, with or without cell-cell contact, and use systems like microfluidics, solid supports, or transwells to control contact. Methods that physically partition individual cell-type populations often allow soluble factor transmission by using permeable material or flooding the compartments so the solvent is shared. However, this is less representative of the human body, where physical partitions do not divide different cell types. Thus, there is a need for a co-culture system with a partition allowing initial separation, that can later be removed to allow interaction without a physical barrier. We are developing this system by utilizing open microfluidic gel patterning techniques to test if a removable partition can be formed with enzyme-degradable polyethylene glycol (PEG). I first use computer-aided design to engineer a rail scaffold outlining two compartments, and fabricate these devices using 3D printing. I pipette PEG into an inlet in the rail, flowing along the scaffold channel due to spontaneous capillary flow and patterning the insert. The PEG polymerizes to form the insert with two distinct cell chambers on the well’s bottom surface, and the cells are seeded into their corresponding compartments. After the cell culture period is complete, sortase (SrtA) is added to completely degrade the PEG insert, allowing the cell populations to interact. We expect the PEG inserts to polymerize similarly to agarose, and leave no residue in the well once degraded. Future work will include utilization of this device for experiments using functionalized beads to monitor soluble factor signaling in co-cultures.

Biological tissues are a group of cells that have similar structure and that function together as a unit. In between these cells is the extracellular matrix (ECM), which provides structural support for resident cells. The makeup of the ECM consists of fibrous proteins, such as collagen, that are interlocked and cross-linked, following a nonlinear stress/strain curve and is considered viscoelastic. The dominant mechanism behind this response is the presence of largely elastic, spring-like straightening/uncrimping of fibrils. This can be thought of like applying a force to springs in parallel. Overall, this mechanism allows the ECM fibrils to align and elongate significantly under small loads, thereby aligning the cells. This, in turn, affects the overall tissue structure and its mechanical properties. We have developed a method for patterning a cell-infused collagen mixture as a three-dimensional tissue, and subsequently stretching it, in order to observe how the cells develop in a strained environment. Specifically, we have engineered two devices that fit within a 6-well plate: the tissue is patterned on the first device, and then transferred to the second for stretching. During each phase, the suspended tissue is incubated for a period of time in order to facilitate cell development and hydrogel gelling. Once the tissue has been stretched for a certain period of time, it is then removed from the device and imaged. Our modular design designates strain as an known and adjustable value, allowing us to relate it to the internal stresses of the tissue via Hooke's Law. We are able to identify the quantitative conditions that promote tissue alignment and maturation within the suspended tissue.

Substitution reactions are frequently utilized in organic chemistry due to their capacity to enable the interconversion of functional groups among a wide array of molecules to generate a variety of new distinct compounds. Our research focused on optimizing and designing a general reaction procedure between acid chlorides and alcohols. Benzoyl chloride and 1-pentanol were used as our model. High yields (92 – 97%) were achieved relatively quickly (2 h) when the acid chloride was added in excess. Lower temperatures (0oC) and gradual addition of the acid chloride to the reaction mixture also increased yields. Future studies will focus on the versatility of these reaction parameters on electron-donating, withdrawing, and sterically hindered acid chlorides.

The modification of tRNA plays a significant role in the efficiency and accuracy of translation during protein synthesis. A modification that plays a direct role in reading cognate codons of mRNA in E. coli is the 5-oxyacetic acid methyl ester (mcmo5) modification. This modification occurs on the uracil base at position 34 (U34). The biosynthetic pathway of this modification is initiated via a hydroxylation reaction. Previous in vivo studies demonstrate the enzyme TrhP, tRNA hydroxylation protein, performs this hydroxylation reaction in anaerobic conditions. No in vitro work has been done to study this enzyme and its mechanistic function. TrhP is known to coordinate an iron-sulfur cluster, a metallic cofactor known to contribute to a variety of critical cellular processes, however, the necessity of an iron-sulfur cluster for a hydroxylation reaction is unique to this newly discovered protein family. The goal of this research project is to spectroscopically characterize TrhP’s iron-sulfur cluster to understand the importance of the FeS cluster. Site-directed mutagenesis is utilized to study the coordination of the iron-sulfur cluster. Changes to iron-sulfur cluster coordination are monitored via UVVIS, electron paramagnetic resonance (EPR), and colorimetric assays. These experiments determine how the loss of cysteine, a known iron-sulfur cluster ligand, impacts the iron-sulfur cluster coordination. Coordination of a [2Fe2S] cluster by 4 conserved cysteines is expected, and UVVIS data agrees with that hypothesis. Colorimetric assays show the cysteine to alanine mutants contain less iron than wild-type TrhP, indicating each cysteine has a significant role in cluster binding. Learning more about the specific coordination will establish the site of cluster-binding within TrhP and shed light on the cluster’s role in TrhP’s stability, geometry, and redox properties which all contribute to the enzyme’s modification activity.