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Chemistry

Research Presentation Video

Watch Jasper Werby '14 discuss his research project.

Differences in Random Walks Between One Jump and Two Jump Particles on Catalytic Surfaces

Raul Sun Han Chang (2016); Student Collaborator(s): Albert Kakkis (2016); David Sharfi (2016); Dion Boyd (2016); Vince Morgan (2015); Nick Alvarez (2016); Oluwamayowa Ige (2016); Noam Hurwitz (2017); Charles Minkah-Premo (2016); Mentor(s): Roberto Garza-Lopez

Abstract: Computational random walk calculations using Markovian chains can be used to predict chemical properties such as kinetics, diffusion, reaction rates, and overall dynamics of a given chemical system. We study the efficiency of diffusion-controlled reactions of the type A+B->C on different families of 2-dimensional and 3-dimensional lattices with reaction centers located at unique sites. We calculate numerically-exact values for the absorption time (or mean walk length) of a particle performing a nearest -neighbor and second-nearest neighbor random walk on finite, nth generation Sierpinski and triangular lattices of non-uniform connectivity, with a deep trap at different locations of the lattice (Garza-López, et al., J Phys. Chem. B 1999). We obtain results that are analyzed in terms of size of the system N, valency of the active sites, v, dimensionality of the lattice, d, and boundary conditions.
Funding Provided by: Pomona College SURP (RS, CM); Sontag Fellowship (AK,OI); Howard Hughes Medical Institute (DB); Pomona College Department of Chemistry (NH, VM)

Remote Sensing of Chlorophyll Fluorescence

Allison Wallingford (2014); Additional Collaborator(s): Tom Pongetti (Jet Propulsion Laboratory, Caltech); Stanley Sander (Jet Propulsion Laboratory, Caltech); Mentor(s): Fred Grieman

Abstract: We are attempting to improve techniques for measuring the intensity of chlorophyll fluorescence from remote targets. Remote sensing techniques measure fluorescence in the visible region from chlorophyll α by looking at added light in gaps in solar radiation called “Fraunhofer lines.” Many studies have used remote sensing of fluorescence from broad expanses of vegetation to obtain an estimate of gross primary production (GPP) from that area, and thus CO2 usage. Ultimately, fluorescence sensing projects aim to monitor the biosphere’s carbon uptake. We take spectra using a Fourier transform spectrometer (FTS) at the California Laboratory for Remote Sensing (CLARS) on Mt. Wilson. The detector takes spectra from various targets throughout the day. In order to determine quantitatively the amount of fluorescence, the spectra are fitted using a model written in Matlab validated by Frankenberg et al. (Geophys. Res. Let., Vol. 38, Iss. 3). The program fits the solar lines using optimal estimation using a simple forward model for continuum radiation and fractional depths of Fraunhofer lines. The diurnal variations of the fluorescence from the targets are an indication of the accuracy of the spectrometer. We have settled on a window of 13270-13400 cm-1 to best fit the spectra and improved the program by adding additional terms to the baseline polynomial. We determined that a Spectrolon® panel is an unsuitable control target, as it likely reflects fluorescence from surrounding vegetation.
Funding Provided by: Pomona College Department of Chemistry

Matrix-Isolated Infrared Spectroscopy of Simple Criegee Intermediates CH2OO and CH3CHOO

Sarah Buchhorn (2014); Additional Collaborator(s): Xu Zhang (Jet Propulsion Laboratory, Caltech); Mentor(s): Frederick Grieman

Abstract: Carbonyl oxides formed during ozonolysis of alkenes known as Criegee intermediates play key roles in the formation of several atmospheric compounds including the hydroxyl radical. These Criegee intermediates, proposed for decades, have only recently been directly observed. In this study we combine the technique of cryogenic matrix isolation with Fourier-transform infrared spectroscopy to study formaldehyde oxide and acetaldehyde oxide produced via reaction of oxygen with haloalkyl radicals from the pyrolysis of CH2IBr and CH3CHI2, respectively. Four bands are tentatively assigned to vibrational modes of acetaldehyde oxide. Future work will include using spectral math processes to further analyze current spectra, and gathering spectra of CH2OO precursor mixtures with reaction partners such as SO2 to aid in identification of formaldehyde oxide vibrational modes.
Funding Provided by: Pomona College Department of Chemistry

Hydrogel-mediated Delivery of Stimuli Responsive Liposomes Against Bacterial Infection

Jieming Li (2015); Student Collaborator(s): Drew Vecchio (2015 University of Michigan -Ann Arbor); Additional Collaborator(s): Liangfang Zhang (UCSD); Victoria Fu (2013 UCSD); Mentor(s): Malkiat Johal

Abstract: Liposomes are a well-established antimicrobial platform. They have innate antimicrobial properties through fusion with bacterial membranes as well as the ability to deliver antibiotics. Stimuli responsive liposomes have been created by adsorbing charged gold nanoparticles onto liposomes. In neutral conditions, the gold nanoparticles prevent liposomes from fusing with one another or bacterial membranes. However, in acidic environments, the gold stabilizers detach and fusion resumes. Since infectious lesions on human skin are typically acidic, this pH-sensitive liposome can be used to treat various skin diseases like staph or acne. Although promising, the application of responsive liposomes against skin infections is limited by the ineffective retention of liposomes at the infection site. To address this, a polyacrylamide hydrogel loaded with gold stabilized liposomes was developed. Varying the cross-linker concentrations resulted in tunable rheological properties and release kinetics. Released liposomes were approximately 100 nm in size, close to the size of liposomes prior to encapsulation. Liposomes released from hydrogels showed pH-responsive fusion with bacterial membranes, indicating the preservation of antibacterial activity. When applied onto the skin of mice, the formulation did not induce inflammation and caused no skin toxicity. Overall, using hydrogels to deliver stimuli responsive liposomes is potentially an effective and safe strategy against skin infections.
Funding Provided by: Paul K. Richter and Evelyn E. Cook Richter Memorial Fund

Effect of Substrate Charge Density on the Layer of Bound Water in Solid Supported Lipid Bilayers

Marco Lobba (2013); Student Collaborator(s): Juliah Shay Kim (2016); Mentor(s): Malkiat Johal

Abstract: Solid supported lipid bilayers provide a convenient system for mimicking biological membranes. Unfortunately, these simple systems come with limitations that prevent them from modeling important biological functions. One drawback of solid supported lipid bilayers has been the inability to use them to model cellular hydration due to the small size and lack of control over the Layer of Hydration (LoH) between the bilayer and the surface. Using a combination of Quartz Crystal Microbalance with Dissipation monitoring (QCM-D) and Dual Polarization Interferometry, we show that it is possible to tune the LoH by varying the electrostatic interaction between the bilayer and its solid support. We found that the thickness of the LoH was dependent upon the pH and varied from 12.7 Å at pH 1.6 to 9.6 Å at pH 10. This work presents an important step towards developing solid supported biomembrane models that can be used to probe changes in cellular hydration in response to well controlled biological phenomena. This work is now a manuscript that will be submitted for publication in the coming weeks.
Funding Provided by: Rose Hills Foundation (ML); Pomona College Department of Chemistry (JK)

The Effects of Glycation on HSA’s Affinity for Hemin: A QCM-D Study

Alison Mercer-Smith (2015); Student Collaborator(s): Gabriella Heller (2014); Additional Collaborator(s): Lewis Johnson; Mentor(s): Malkiat Johal; Kevin Sea; Matthew Sazinsky

Abstract: Non-enzymatic glycosylation is the process by which sugars covalently bond to proteins, potentially altering their structure and function. We investigated the change in affinity of human serum albumin (HSA) for hemin, a small iron containing molecule, after the protein was incubated for two weeks with three sugars: glucose, fructose, and glyoxal. The formation of protein-heme complexes was measured using a quartz crystal microbalance with dissipation monitoring (QCM-D). We hypothesize that as HSA's exposure time to sugar increases, less hemin will bind to HSA. A decrease in the binding affinity for hemin can negatively impact HSA's ability to transport hemin, which can even lead to free hemin in the blood. Free hemin may lead to higher rates of bacterial infection as some bacteria may use it as a micronutrient.
Funding Provided by: Kenneth T. and Eileen L. Norris Foundation (AM); Howard Hughes Medical Institute (GH)

Bivalent and Trivalent Cation Effects on Supported Lipid Bilayer Structure: A QCM-D Study

Hannah Wayment-Steele (2015); Additional Collaborator(s): Angelika Kunze (Chalmers University of Technology); Sofia Svedhem (Chalmers University of Technology); Mentor(s): Malkiat Johal

Abstract: Studying the interactions of lipid membranes and ions is of critical importance for understanding membrane processes such as lipid packing, membrane fusion, and signal transduction. In this work, we investigate cation-mediated membrane phenomena using the Quartz Crystal Microbalance with Dissipation (QCM-D) technique, which monitors changes in mass and viscoelastic properties in real time. “Bilayer slip,” the removal of bivalent ions causing the bilayer to detach from the surface, is compared for Ca2+ and Mg2+ ions. We observe Mg2+ to be removed significantly faster than Ca2+ ions. Binding of Ca2+ and Mg2+ to the lipid membrane is compared to binding of Al3+. We find the affinity for Al3+ ions to be significantly stronger than for the bivalent cations. Furthermore, we show that Al3+-mediated liposome fusion can be used for the formation of lipid multilayers.
Funding Provided by: Sherman Fairchild Foundation

Characterization of DANPY binding to biological substrates

Conner Kummerlowe (2016); Student Collaborator(s): Juliah Shay Kim (2016); Additional Collaborator(s): Bruce Robinson (University of Washington); Rose Ann Cattolico (University of Washington); Mentor(s): Lewis Johnson

Abstract: DANPY-1 is a fluorescent dye with high affinity for DNA that has potential applications as an alternative option for commonly used carcinogenic dyes. Using a Quartz Crystal Microbalance (QCM) we characterized DANPY binding to biological substrates and compared this to the binding of the dyes DAST and Ethidium Bromide. DANPY proved to bind irreversibly to double stranded DNA, single stranded DNA, and RNA and reversibly to the electrostatic control polystyrene sulfonate. These results indicate that DANPY may be an effective cell stain for nucleic acids due to its irreversible binding mode. QCM studies on DANPY binding to Bovine Serum Albumin, anionic proteins, lipid membranes, and other biological substrates will be carried out to further understand DANPY binding to biological substrates and evaluate its efficacy for cellular staining.
Funding Provided by: Pomona College Department of Chemistry

The Vc1 riboswitch can serve as a biosensor for cyclic diguanylate in Vibrio cholera

Abraham Cass (2014); Mentor(s): Jane Liu

Abstract: Cyclic diguanylate (c-di-GMP) is an important second messenger in Vibrio cholerae, and interacts with several types of riboswitches in the V. cholerae genome. We investigated the interaction between c-di-GMP and the c-di-GMP riboswitch Vc1 to determine whether Vc1 could serve as a biosensor for the second messenger. Three V. cholerae strains were used, each containing two plasmids. The first plasmid allowed for the over-and under-expression of c-di-GMP by expressing, respectively, either a protein that synthesizes c-di-GMP (VCA0956, a diguanylate cyclase) or one which degrades the molecule (VieA, a phosphodiesterase). An empty vector was also used for a control. The second plasmid contained, in all three strains, the Vc1 riboswitch fused to the 5’-end of the lacZ coding sequence. Activity levels of β-galactosidase (LacZ) were analyzed to determine how the riboswitch responded to varying intracellular levels of c-di-GMP. Preliminary results show that the riboswitch functions in an ‘ON’ mechanism; the interaction between c-di-GMP and the riboswitch upregulates synthesis of the β-galactosidase reporter protein. HPLC experiments will further confirm the relative amount of c-di-GMP in each strain. Ultimately, this technology would be used for cost-effective water quality control in the field, as well as investigation into the environmental factors that help determine V. cholerae biofilm and virulence pathways.
Funding Provided by: National Science Foundation #CBET-1258307

Early Detection of Breast Cancer using Single Molecule Array Technology

Erick Velasquez (2016); Additional Collaborator(s): Stephanie Schubert (Tufts University); David Walt (Tufts University); Mentor(s): Jane Liu

Abstract: Breast cancer is the second most diagnosed cancer among women of the United Sates. Current diagnostic methods, such as mammograms and biopsies, have greatly improved early detection of breast cancer; however, these techniques are associated with high numbers of false positives, false negatives, and in the case of biopsies, are extremely invasive. Our overall goal is to implement a simple blood test that will measure a fingerprint of protein biomarkers associated with breast cancer to aid in earlier detection of the disease as well as improve therapeutic efficacy in a non-invasive manner. Since the majority of vital proteins associated with breast cancer within blood exist at levels below the detectable limits of standard methods, i.e. ELISA, we are utilizing the ultra-sensitive Single Molecule Array (SiMoA) technology developed in our laboratory that is capable of quantifying proteins down to sub-femtomolar levels. Here we describe the progress made on developing assays for monocyte chemotactic protein-1 (MCP1) and prostate specific antigen (PSA). With calculated limits of detection (LOD) of 61 fg/mL (7 fM) and 27 ag/mL, the assays developed for MCP1 and PSA, respectively, were 150 and 8000 times more sensitive than our bulk bead based ELISAs. These promising results suggest that we can use these assays for validation of our mouse models.
Funding Provided by: Linares Family SURP for Chemistry

Synthesis of the C-C Analog of Malformin A1

Claire Brickson (2014); Mentor(s): Daniel O'Leary

Abstract: The bicyclic pentapeptide malformin A1 (cyclo-D-cysteinyl-D-cysteinyl-L-valyl-D-leucyl-L¬isoleucyl), a metabolite of the fungus Aspergillus niger, demonstrates antibiotic activity and induces severe curvatures in bean plants and corn roots. Malformin’s uncommon oxidized Cys-Cys eight-membered ring is likely responsible for the peptide’s biological activity. We synthesized the C-C analog of malformin A1 using solid-phase peptide synthesis (SPPS), replacing the cysteine residues present in the natural product with allylglycine (Agy) residues. The eight-membered ring was formed using ruthenium-based ring closing metathesis (RCM), a unique application of olefin metathesis. To make intramolecular RCM possible, we alkylated the Agy-Agy amide bond with a 2,4¬dimethoxybenzyl (DMB) protecting group, causing the pendent olefins to adopt the cis conformation over trans. The subsequent challenging secondary amine peptide coupling was achieved via the acyl chloride both in situ and in two separate steps. As yet RCM with Grubbs first generation catalyst has been unsuccessful, although traces of the product have been observed. We will attempt to build the pentapeptide on resin after generating the protected allylgycine dipeptide in solution, after which cleavage and cyclization should generate the C-C analog of malformin A1.
Funding Provided by: Pomona College Department of Chemistry

Optimization of Shewanella oneidensis FeoB expression and purification – Measurement of the G-Domain activity

Lazaros Marios Konstantinos Chalkias (2016); Student Collaborator(s): Benjamin Smith (2013); Mentor(s): Matthew Sazinsky

Abstract: Identifying the differences in iron acquisition between eukaryotic cells and anaerobic bacteria presents an exciting opportunity to target harmful bacterial infection without affecting the human host. While eukaryotes primarily rely on Ferric Iron (Fe+3), bacteria has been shown to have higher affinity for the reduced Ferrous Iron (Fe+2) in anaerobic conditions such as the human gastrointestinal track. Previous studies have identified the ferrous iron enzyme system (Feo) as the major regulator of iron levels in bacteria such as Escherichia coli and Salmonella enterica. Our work focuses on the key component of the system, FeoB, a guanine nucleotide binding protein (G-protein) with an integral and a cytosolic domain, which dictates the differences in iron acquisition between the bacteria and humans. To this date, we have optimized the expression of Shewanella oneidensis FeoB in E. coli in aerobic conditions, simplifying the process and producing amounts of protein beyond those of the available literature. Protein purification has also been achieved by nickel affinity chromatography. For the purposes of this project, the quantification methods of FeoB were investigated and the activity of the G-Domain was compared in the full-length enzyme and cytosolic areas. Primary results indicate a significant increase in the rate of GTP hydrolysis when the full-length enzyme is present.
Funding Provided by: National Institute of Health #R15¬GMO93308

Exploring the Structure, Substrate Kinetics, and Mechanism of phCoADR

Berniece Chen (2015); Mentor(s): Matthew Sazinsky; Kevin Sea

Abstract: Coenzyme A disulfide reductase (CoADR) in Pyrococcus horikoshii (ph) is a FAD and NAD(P)H¬dependent homodimer that reduces disulfide bonds in various substrates. Previous studies focused on wildtype phCoADR, analyzing its structure and substrate specificity, and found that CoA disulfide is not a substrate at 50°C, possibly because its large site is not accommodated by the active site. In this project, four specific mutations, Y65A, Y66A, P67G and H367G, were made to the active site in order to remove the side chains blocking the site, possibly allowing access to CoA disulfide. This quad mutant was crystalized to 3.7 Å and we found little impact on the backbone structure. We also characterized the substrate specificity of the enzyme and found the enzyme actively used CoA disulfide as a substrate. An aerobic titration of NADH into the oxidized enzyme showed the reduction of FAD by NADH, but the FAD was reduced only after one-half equivalent of NADH was added. We hypothesized the first one-half equivalent of NADH reduced a disulfide bond between the substrate and the active site cysteine 48, so we made the quad mutant with an additional C48S mutation. FAD in the quad-C48S enzyme was immediately reduced by anaerobic addition of NADH, thus supporting our hypothesis. Future projects include making a heterodimer, with one wild type and one C48S monomer, in order to explore why only one of the two active site cysteines appeared to be reduced in the anaerobic titration.
Funding Provided by: Pomona College Department of Chemistry -Robbins Fund

An Activity Assay on Fatty Acid 2-Hydroxylase in Pichia Pastoris

Roger Sheu (2014); Student Collaborator(s): Kimberly Elizabeth Ona Ayala (2016); Mentor(s): Matthew Sazinsky

Abstract: Fatty Acid 2-Hydroxylase (FA2H) is a diiron membrane protein that plays a role in hydroxylating sphingolipids, converting fatty acids (C16 to C26) into 2¬hydroxy fatty acids. 2-hydroxy sphingolipids are prevalent in the brain, as such,mutations in FA2H have been linked with leukodystrophy and neurodegeneration. Pichia pastoris, a methyltropic yeast commonly used in experimentations for its high level of inducible expression, and mammals possess FA2H. The goal of our experiments is to conduct an in vitro activity assay on FA2H embedded in Pichia membrane. An electron transfer system consisting of NADPH and cytochrome P-450 reductase is needed to carry out the FA2H reaction. Known concentrations of C16 and 2-hydroxy C16 fatty acid methyl esters are run with Gas Chromatography-Mass Spectrometry (GC-MS) and interpolating these results allows us quantify the rate of the FA2H enzyme.Toluene is a standard in GCMS to ensure that trials are consistent.Hexanes had difficulty dissolving fatty acid methyl esters, so methanol is being used in its place. Coomassie Blue staining, Western blotting, and Fast Protein Liquid Chromatography are methods used in identifying and purifying FA2H.Growing E. coli cells with the human cytochrome b5 domain of FA2H is also in progress. The diiron complex consists of an 8-histidine motif; since the diiron complex's structure is currently unknown, purifying and analyzing it with X-ray crystallography could reveal some answers about FA2H's mechanism.
Funding Provided by: Rose Hills Foundation (RS); Pomona College Department of Chemistry (KO)

Expression and characterization of L-ferritin-binding protein, Scara5

Gabriella Heller (2014); Student Collaborator(s): Erica Storm (2013); Mentor(s): Matthew Sazinsky

Abstract: Investigating the interaction between the iron transport protein L-ferritin and its receptor, Scavenger Receptor Class A Type 5 (Scara5), is an important avenue for understanding the relationship between iron regulation and disease. Although ferritin is a ubiquitous and well-understood iron storage protein, the receptor of its L-chain variant, Scara5 is comparatively unknown since is identification in 2009. Isolation of purified recombinant products should provide a foundation for the characterization of Scara5 and its interaction with ferritin. In this study, the scavenger receptor cysteine-rich domain of Scara5 was expressed in sHuffle E. coli cells under various conditions and purified using Fast Protein Liquid Chromatography. Purity was confirmed with polyacrylamide gel electrophoresis. Pure protein will be used to crystallize Scara5 and investigate Scara5-ferritin binding kinetics.
Funding Provided by: Howard Hughes Medical Institute

Substituent Effect on Radical Scavenging Activity of Catecholic Compounds: A QSAR model

Kevin Guan (2015); Mentor(s): Cynthia Selassie

Abstract: The objective of our investigation was to determine quantitatively the substituent effect on the radical scavenging activity level of catecholic compounds. The basic catechol moiety is ortho¬benzenediol, or 1,2-benzenediol, and we examined specifically catecholic compounds with substituents at the 4-position. To assess the extent of radical-scavenging activity, we utilized the radical compound 2,2-diphenyl-1-picrylhydrazyl (DPPH) and observed via spectrophotometer the neutralization rate of multiple dilutions of each catechol. Afterward, we generated a Quantitative Structure Activity Relationship model relating each catechol’s activity level with each corresponding substituent’s σpara+ constant, a positive charge/radical variant of the Hammett electronic constant that represents the extent of a para substituent’s electron donating or withdrawing properties. The best fit equation was found to be: LOG(1/C) = -0.119(0.067) σpara+ + 3.710(0.040), indicating that there is a small but significant relationship wherein the more electron donating substituents correlated with higher radical scavenging activity by the catechol. Other constants, such as the σ, π, ClogP and MR constants, were considered but yielded no significant correlation to activity of the catechols.
Funding Provided by: Pomona College Department of Chemistry

Measuring the Hydration of BSA-Ligand complexes Using the Quartz Crystal Microbalance and Dual Polarization Interferometer

Jung (James) Kang (2014); Student Collaborator(s): Noah Stanton (2015); Mentor(s): Cynthia Selassie; Malkiat Johal; Lewis Johnson

Abstract: To determine the number water molecules involved in protein-ligand complexes, we determined the mass of the complex on silicon layered surfaces with the dual polarization interferometer (DPI) and quartz-crystal microbalance with dissipation monitoring (QCM¬D). The DPI and QCM-D have a difference in hydration sensitivity which allows us to determine the degree of hydration from binding drug molecules to a rigid bovine serum albumin layer. With this difference in mass, we can determine the number of water molecules involved in ligand-protein complexes. Various ligands with different hydrophobicities were examined in this manner so that we could find a relationship between the ligands' physicochemical properties and their hydration with bovine serum albumin.
Funding Provided by: Pomona College Department of Molecular Biology -Stellar Fellowship

Synthesis of 4,6-diamino-1,2-dihydro-2,2-dimethyl-1¬(3’-(4”-hydroxyacetamide-anilinomethyl)phenyl)-s¬triazine as a potential Plasmodium Falciparum DHFR/HDAC inhibitor

Briton Lee (2015); Mentor(s): Cynthia Selassie

Abstract withheld upon request.
Funding Provided by: National Institute of Health; Pomona College Department of Chemistry

Synthesis of a potential benzoxazine inhibitor of dihydrofolate reductase from P. falciparum

Alex Samuels (2015); Mentor(s): Cynthia Selassie

Abstract: Plasmodium falciparum is a microbe that causes malaria, a devastating disease that kills approximately 1.5-2.7 million people every year. A key enzyme in this parasite's biological processes is thymidylate sythase-dihydrofolate reductase, which both catalyzes the reduction of dihydrofolic acid to tetrahydrofolic acid and generates thymidine monophosphate. The goal of this project is to synthesize a new antimalarial antifolate, 8-isopropyl-6¬(2',4'-diamino-5'-pyrimidylmethyl)-2H-1,4-benzoxazin-3(4H)-one, which inhibits the dihydrofolate reductase constituent of the enzyme, potentially resulting in the inactivation of P. falciparum. The aforementioned molecule was synthesized through a series of reactions beginning with 2,4-diamino-5-cyanopyrimidine, which was reduced to obtain 2,4-diamino-5-carboxyaldehyde, and again to yield 2,4-diamino-5-hydroxymethyl pyrimidine. This compound was then coupled with 2¬isopropylphenol, forming 2,4-diamino-5-(3'-isopropyl¬4'hydroxybenzyl) pyrimidine. It will then be treated with bromo-acetamide to make 2,4-diamino-5-(3'-isopropyl¬4'carboxamidomethoxybenzyl) pyrimidine, which will then be nitrated, reduced and cyclized to form the desired product.
Funding Provided by: Sherman Fairchild Foundation

Synthesis of 4,6-diamino-1,2-dihydro-2,2-dimethyl-1¬(3’-(3’’-X-anilinomethyl)phenyl)-s-triazines as potential Plasmodium falciparum DHFR inhibitors

Makeda Tekle-Smith (2014); Mentor(s): Cynthia Selassie

Abstract withheld upon request.
Funding Provided by: Howard Hughes Medical Institute

Design and Synthesis of novel bifunctional antimalaria drugs

Jasper Werby (2014); Mentor(s): Cynthia Selassie

Abstract: Malaria annually kills around one million people, mainly in the developing world. The rapid life cycle of the malaria parasite and the subsequent development of resistance have rendered most currently used drugs as ineffective. This project focuses on the development and synthesis of a new bifunctional, antimalarial drug that targets multiple inhibitory sites at the same time. Falcipain 2 and Dihydrofolate reductase (dhfr) are enzymes that are critical to the growth of the plasmodium. A critical analysis of the current literature has led to the design and synthesis of 4-((2,4-diaminopyrimidin-5¬yl)methyl)phenyl4-(3-(5-ethyl-1,3,4-thiadiazol-2-yl)-3¬oxopropyl)benzenesulfonate that incorporates structural features necessary for binding to Falcipain 2 and dhfr. The multistep synthesis has involved the synthesis of two separate pharmacophores that are eventually connected to each other. Further work will be done to create other analogues that optimize the antimalarial activity and bioavailability of these compounds as well as to test the cytotoxicity and the in vitro activity of these novel antimalarial drugs.
Funding Provided by: Corwin Hansch and Bruce Telzer Fund

Identifying Bacterial VOC Biomarkers using SPME/GC¬MS

Alexandra Antonoplis (2014); Additional Collaborator(s): Ilya Tolstorukov (Keck Graduate Institute of Applied Life Sciences); Angelika Niemz (Keck Graduate Institute of Applied Life Sciences); Mentor(s): Charles Taylor

Abstract: Solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS) facilitates identification of volatile organic compounds (VOCs) produced by infectious bacteria as metabolic waste. A SPME/GC-MS analysis of VOCs emitted in vitro by planktonic Escherichia coli demonstrated the organism’s production of indole, an odorous compound with a mothball-like scent. Sampling the E. coli headspace in glass vials capped with Teflon-lined rubber septa promoted indole detection by SPME/GC-MS, as polypropylene sampling vessels leak confounding VOCs. Additionally, sampling the headspace above E. coli growth media (after sterile-filtration to remove E. coli cells) allowed VOC snapshots to be taken at particular points in culture growth without the challenges of maintaining a growing culture. Future work will use SPME/GC-MS to detect VOC biomarkers for other pathogens, such as Pseudomonas aeruginosa, in both a planktonic and biofilm growth state.
Funding Provided by: Rose Hills Foundation

Characterizing Trace Element Profiles in Coffee Samples Using Wavelength-Dispersive X-Ray Fluorescence

Laura Barry (2014); Mentor(s): Charles Taylor

Abstract: Coffee is consumed daily by approximately one-third of the world’s population. The quality of coffee, while a subjective measure, is believed to depend on geographic origin, making correct identification important. Existing literature on trace element profiles of coffee is limited. Wavelength-dispersive X-ray fluorescence (WD-XRF) provides a wealth of elemental data in a short amount of time with minimal sample preparation. WX-XRF spectroscopy was used to obtain a trace element profile of coffee samples of known origins and principle component analysis (PCA) was performed on the data. One pre-ground sample and fourteen whole bean roasted coffee samples of specified origin were obtained from Trader Joe’s in 2011 and 2013. Seven whole bean samples, both roasted and green beans, were obtained from Herkimer Coffee. Minimal difference was found between trace element profiles of the same Trader Joe’s coffee samples of different years. There were also few differences seen between the roasted and green beans from Herkimer Coffee. By combining the Trader Joe’s and Herkimer Coffee data we were able to show that by using WD-XRF spectroscopy and PCA, coffee samples can be separated by origin.
Funding Provided by: Pomona College Department of Chemistry

Developing Evanescent-Wave Raman Spectroscopy to Detect VOCs of Pulmonary Diseases

Constance Wu (2014); Student Collaborator(s): Allison Boden (2014); Mentor(s): Charles Taylor

Abstract: Ventilator assisted pneumonia (VAP) and tuberculosis (TB) are treatable diseases, but current diagnostic methods and tools are often invasive, inaccurate, and inefficient. While gas chromatography-mass spectrometry (GCMS) has been used previously for analysis and characterization of volatile organic compounds (VOCs), this method is expensive and non-portable. This project aims to develop a non-invasive, portable, easy-to-use Raman-based system for the analysis of VOC biomarkers that could be used to rapidly diagnose many different types of infections. We have modified a commercial Raman scattering system to enable us to perform novel evanescent-wave excitation scattering measurements. Although Raman spectroscopy typically lacks adequate sensitivity for trace analysis, theory predicts that the geometry used should enhance sensitivity of the system tremendously, thus allowing its use for trace analysis. The results obtained show good agreement with theory.
Funding Provided by: Howard Hughes Medical Institute

Untangling the Important Variables in Vanadium Oxide Chemical Vapor Deposition

Kyle Roskamp (2014); Mentor(s): Chuck Taylor

Abstract: Vanadium oxides have a wide variety of applications but their utility depends strongly on the oxidation state and morphology of the material. Chemical vapor deposition has been demonstrated to enable precise control over morphology and chemical composition but these results are highly variable, especially at lower deposition temperatures. This summer, I will grow thin films of vanadium oxides using two molecular precursors (vanadium (V) oxytriisopropoxide and vanadium (V) oxytrinitrate) with the goal of identifying conditions for preparing thin films with strong texturing. Samples will be analyzed by field emission scanning electron microscopy (including energy-dispersive X-ray analysis) and X-ray diffraction. Kinetics will be modeled to provide additional insight into the deposition process.
Funding Provided by: Pomona College SURP

Synthesizing Antimicrobial Peptides via Solid Phase Peptide Synthesis: An Exploratory Lab Exercise

Cristina Saldaña (2015); Mentor(s): Thomas Vasquez; Jane Liu

Abstract: Antibiotics have been one of our main weapons against pathogenic microorganisms. However, their overuse has given rise to resistant strains that are becoming serious problem. Thus, finding novel antibiotics is of upmost importance. Many peptides have been shown to exhibit antimicrobial activity that may be promising alternatives to current antibiotics. With this in mind I set out to create several antimicrobial peptides through solid phase peptide synthesis (SPPS). The goal of this research was to identify potential new antibiotics that are effective and less prone to leading to resistance. I synthesized several peptides of varying lengths and sequences using both ¬D-and L-amino acids. Other reagents used included DCM, DMF, piperidine, and coupling agents HBTU and DIEA. Of the thirteen peptides I synthesized, the ones that exhibited antimicrobial activity were hexapeptides. Thus, in the future, I plan to synthesize peptides that are at least six amino acids long and test them on a variety of bacteria. A second goal of this research was to develop a teaching laboratory that would expose students to SPPS and the analytical tools that are involved, namely LCMS. To do this, I created a protocol that can be used in an organic chemistry teaching lab setting. I anticipate that through this laboratory exercise, students will get a better understanding of the chemistry behind peptide bond formation as well as the implications antimicrobial peptides have on today’s antibiotic crisis.
Funding Provided by: Pomona College Department of Chemistry

Research at Pomona