Browsing by Author "Crick, Dean, committee member"

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Open Access
Antibacterial growth effects and speciation of several vanadium salts and complexes
(Colorado State University. Libraries, 2023) Arhouma, Zeyad Kamal, author; Crans, Debbie C., advisor; Crick, Dean, committee member; Roess, Deborah, committee member; Jackson, Mary, committee member
Vanadium (V) is a first-row transition metal ion that acts as a phosphatase inhibitor with a wide variety of biochemical and physiological functions. The ability of vanadium to form stable polyoxovanadates (POVs) and organometallic complexes has attracted attention for studying the properties and effects of these compounds in various biological systems. In my research, I used the bacterium species Mycobacterium smegmatis (M. smeg), which has undergone reclassification and is now classified as Mycolicibacterium smegmatis. Despite the taxonomic change, both the previous and current classifications use the same abbreviation, M. smeg. I also carried out some studies in Mycobacterium tuberculosis (M. tb). In my work, I explored the properties of several types of vanadium compounds including salts, oxometalates, and coordination complexes to investigate how they impact cellular growth. The first chapter of this dissertation focuses on determining the growth inhibitory effects of decavanadate (V10) and rapidly exchanging oxovanadates on the growth of two mycobacterial species: M. tb and M. smeg. Speciation analysis, utilizing 51V NMR spectroscopy, was employed to document that one specific oxometalate exhibits greater potency as a growth inhibitor for these mycobacterial species compared to other oxovanadates, indicating selectivity in its cellular interaction. Oxometalates have been involved in numerous applications in biological and medical studies, including their ability in addressing the phase-problem in X-ray crystallography of the ribosome. This study investigated the effect of different vanadate salts on the growth of M. smeg and M. tb, highlighting the critical role of speciation in the observed growth inhibition. Specifically, the large orange-colored sodium decavanadate (V10O286−) anion was found to be a stronger growth inhibitor for these bacteria compared to the colorless oxovanadate derived from sodium metavanadate. The 51V NMR spectroscopy and speciation calculations were employed to monitor the vanadium(V) speciation in the growth media and its conversion among species under growth conditions. Our results show that the decavanadate was 200-20 times more potent in inhibiting growth dependent the consideration of molecules or total vanadium content. The findings presented in this work are particularly important in the context of the numerous applications of polyoxometalates in biological and medical studies. The second chapter focuses on investigating the inhibitory effects of two monosubstituted decavanadates (V10): monoplatino(IV)nonavanadate(V) ([H2PtIVV9O28]5−, V9Pt), and by MoIV in monomolybdo(VI)nonavanadate(V) ([MoVIV9O28]5−,V9Mo) on the growth of M. smeg. The inhibitory effects of V9Pt and V9Mo were examined against the growth of M. smeg with EC50 values of 0.0048 mM and 0.015 mM, respectively. These values were compared to the reported inhibitory value of decavanadate ([V10O28]6−/[HV10O28]5−, V10) on M. smeg (EC50 = 0.0037 mM). Time-dependent 51V NMR spectroscopic studies were carried out for all three polyanions in aqueous solution, biological medium (7H9), and heated and non-heated supernatant. These studies aimed to evaluate their stability in their respective media, monitor their hydrolysis over time to form different oxovanadates, and calculate the corresponding EC50 values. The results presented in this study indicate that the two related derived decavanadate derivatives (V9Pt and V9Mo) and V10 exhibited greater potency as growth inhibitors of M. smeg, compared to monomeric vanadate (V1). The spectroscopic characterization conducted in the growth medium led to the conclusion that both the decavanadate structure and its properties play significant roles in their growth effects. In the third chapter in this dissertation, we investigated the growth effects of an anticarcinogenic non-toxic Schiff base oxidovanadium(V) complex (N-(salicylideneaminato)-N'-(2-hydroxyethyl) ethane-1,2-diamine) coordinated to the 3,5-di-tert-butylcatecholato ligand on a representative bacterium, M. smeg.. In addition, we synthesized a series of the Schiff base V-complexes based on previously reported methods and examined the effect of complexes as well as the free catecholates on the bacterial growth. To determine the inhibition activity of these complexes on M. smeg., the biological studies were complemented by spectroscopic studies using UV-Vis spectrophotometry and NMR spectroscopy. These spectroscopic studies determine which complexes remained intact under biologically relevant conditions. In this work, we examine (1) the growth effects of Schiff base oxidovanadium complexes coordinated to a catechol, (2) the growth effects of the respective free catecholates on M. smeg., and (3) the effects of the scaffold. These studies allowed us to demonstrate that some metal coordination complex exhibited greater potency than the ligand alone under biological conditions, whereas others showed greater effects of the free catecholate ligand and in one case the effects were similar of complex and catecholate ligand. The findings from these studies revealed that the observed effects of the Schiff base V-catecholate complex were influenced by the properties of the catechol, including toxicity, hydrophobicity, and steric factors. Finally, the fourth chapter presents preliminary research data on the antimicrobial effects of two pseudospherical mixed-valence polyoxovanadates (MV-POVs), namely K(NH4)4[H6PVIV2VV12O42]·11H2O (V14) and (Me4N)6[VIV8VV7O36(Cl)] (V15) on the growth of M. smeg. These MV-POVs showed complex effects on cell growth, as many of these systems are not stable under biological conditions. To investigate the vanadium(V) speciation in aqueous solutions and growth media, as well as to monitor any conversion among species under growth conditions, 51V NMR spectroscopy was employed. The 51V NMR spectra revealed some hydrolysis and more extensive oxidation of vanadium(IV) in V14 compared to V15 in both aqueous solutions and media. The studies show that both MV-POVs are effective growth inhibitors. The combined findings from the studies described in all the chapters of this dissertation indicate that the stability of the vanadium compound and its structure plays a significant role in the ability of the vanadium complexes to inhibit bacterial growth. These studies highlight the importance of speciation in the biological activity of vanadium complexes.
Open Access
Characterization and quantification of urinary metabolic biomarkers for early response to anti-tuberculosis treatment
(Colorado State University. Libraries, 2016) Fitzgerald, Bryna, author; Belisle, John, advisor; Crick, Dean, committee member; Dobos, Karen, committee member; Cohen, Robert, committee member
Development of new anti-tuberculosis (TB) therapies remains a major priority to combat this infectious disease and to prevent continued transmission of the causative agent Mycobacterium tuberculosis (Mtb). However, newly developed therapies require large, lengthy clinical trials to determine the number of treatment failures and relapses for evaluation of treatment efficacy. Biomarkers for the prediction of treatment outcome in TB patients at early time points would facilitate movement of new therapies through clinical trials. Previously, liquid chromatography-mass spectrometry (LC-MS) based metabolomics experiments identified potential biomarkers for early response to anti-TB treatment. The research presented in this dissertation involves experiments needed for the progression of these compounds towards a clinically useful biosignature. A major impediment to metabolomics-based biomarker discovery is metabolite identification, as approximately 50% of detectable products do not match structures in existing databases. In concordance with this, several of the potential small molecule biomarkers of anti-TB treatment response lacked structural identification. This research resulted in the structural characterization of three of these compounds as a core 1 O-glycosylated SerLeu peptide, N-acetylisoputreanine, and N1, N12-diacetylspermine. Both the core 1 O-glycosylated SerLeu peptide and N-acetylisoputreanine are novel compounds that had not been previously detected in human urine. Characterization of these compounds indicated a potential alteration of polyamine catabolism and the complement and coagulation pathways during anti-TB treatment. Another key aspect in biomarker discovery is defining the processes involved in formation of potential biomarkers. In order to determine whether these compounds were formed by processes upregulated during active disease, the abundances of these compounds were assessed in active TB patients and household contacts as well as in Mtb infected and uninfected Balb/c mice. The core 1 O-glycosylated SerLeu peptide and N1, N12-diacetylspermine were increased in the urine of index patients demonstrating a potential link between Mtb infection, associated disease pathology, and the formation of these compounds. N-acetylisoputreanine, however, was not increased in TB patient urine or infected mouse tissue indicating that this compound may be formed due to off target drug interactions. These experiments not only provided insights into the mechanisms behind alteration of these compounds during anti-TB treatment, but also highlighted those compounds that may be better biomarkers for anti-TB treatment response. Assessment of these compounds using an independent set of patient samples is needed to validate them as biomarkers for early anti-TB treatment response. Unlike the untargeted experiments used for discovery of potential biomarkers, validation typically employs targeted assays. This research describes the development of a targeted multiple reaction monitoring (MRM) assay which enabled accurate and precise quantitation of compounds previously detected in an untargeted metabolomics experiment. This targeted assay will be used for validation of these compounds in a larger set of patient samples representing a variety of different treatment outcomes. Overall these experiments confirmed the identity of three metabolites that decrease with anti-TB treatment response. Two of these metabolites are novel compounds and their characterization adds to metabolite databases expanding the number of metabolites available to other metabolomics researchers. Assessment of these compounds in samples representative of active TB disease confirmed two of them as promising biomarkers for anti-TB treatment response and highlighted another as a potential result of unintended drug effects. The development of a MRM assay for the quantification of these compounds enables their validation and confirmation as biomarkers of anti-TB treatment response. The work presented in this dissertation describes the advancement of metabolites identified during biomarker discovery towards application in clinical trials.
Open Access
Elucidation of the biogenesis of the paraherquamides, malbrancheamides, citrinalins, and brevianamides
(Colorado State University. Libraries, 2019) Klas, Kimberly R., author; Williams, Robert M., advisor; Shi, Yian, committee member; Chen, Eugene, committee member; Crick, Dean, committee member
Various fungi of the genera Aspergillus, Penicillium and Malbranchea produce prenylated indole alkaloids that possess a bicyclo[2.2.2]diazaoctane ring system and a variety of biological activities such as insecticidal, cytotoxic, anthelmintic, and antibacterial properties. After the discovery of distinct enantiomers of the natural alkaloids Stephacidin A, Notoamide B and their corresponding diastereomers, from Aspergillus protuberus MF297-2, Aspergillus amoenus NRRL 35660 and Aspergillus taichungensis, the structurally diverse metabolites became of particular biosynthetic interest. The bicyclo[2.2.2]diazaoctane core of the divergent natural metabolites may be enzymatically derived via a putative intramolecular hetero-Diels-Alder cycloaddition. We completed the total synthesis of ZwtP and MeZwtP, unveiling the role of a newly discovered Diels-Alderase. We are also undergoing further synthetic efforts to access other novel natural products, as well as further understand additional unprecedented transformations in nature.
Open Access
Extreme ultraviolet laser ablation mass spectrometer for molecular imaging at the nanoscale
(Colorado State University. Libraries, 2018) Kuznetsov, Ilya, author; Menoni, Carmen, advisor; Rocca, Jorge, committee member; Crick, Dean, committee member; Bernstein, Elliot, committee member; Krapf, Diego, committee member
The demand for high-precision analytical instrumentation in modern science and technology is exploding. The quality of questions to be answered sets the requirements for a given piece of technology. The type of analytical instrumentation that enables users to unambiguously identify, quantify, and map the chemical structure of a solid is imaging mass spectrometry (IMS). Most common commercially available instruments include desorption electrospray ionization (DESI), matrix-assisted laser desorption and ionization (MALDI), and secondary ion mass spectrometry (SIMS) as well as their derivatives. Each of these methods possesses a set of capabilities that define its use for one or another research task. None of them, however, enables scientists to map a solid's molecular composition in three dimensions at the nanoscale. We have developed an extreme ultraviolet laser ablation time-of-flight mass spectrometer (EUV TOF) that relies on sample probing by a 46.9 nm wavelength laser. In this work, the unique interaction of EUV light with matter was experimentally assessed and compared to SIMS TOF. It was found that the spatial resolution can be as small as 80 nm in molecular and atomic analysis in organic and inorganic materials respectively. Depth resolution is as high as 20 nm as measured on an organic bilayer. Sensitivity of the EUV TOF reaches ~0.02 amol, which is estimated to be 20× better than that of SIMS TOF in the sample of the amino acid alanine. Sensitivity in other units—sample utilization efficiency (SUE)—was found to be similar to SIMS TOF when assessed by means of detecting trace actinides in a glass matrix. It was shown that it can be further improved by means of post-ablation ionization (PI) with a secondary UV laser source. Using vacuum ultraviolet (VUV) laser light can increase the mass range of molecular detection. For instance, an intact cholesterol molecule was first detected by EUV TOF operating in VUV PI mode. This approach opens a range of opportunities to use the technique for biological studies. EUV TOF is capable to image chemical composition. This capability is demonstrated by imaging the 3D nanoscale spatial distribution of low mass fragments in a single mycobacterium. With additional instrumental modifications, it will be possible to achieve sub-cellular imaging of the molecular structure of a single microorganism without the need for using externally applied ionization-promoting matrix. Such capabilities may help to steer the development of new drugs in pharmacology and identify the signature isotope pattern of the miniscule bits of material examined by nuclear scientists.
Open Access
Extreme ultraviolet time-of-flight mass spectrometry for the characterization of actinides
(Colorado State University. Libraries, 2017) Green, Tyler, author; Menoni, Carmen, advisor; Crick, Dean, committee member; Marconi, Mario, committee member
Many scientific disciplines, as well as several technologies, have a great demand for analytical tools capable of assessing chemical composition and imaging chemical content that are highly sensitive and have sub-micron spatial resolution. Laser ablation mass spectrometry meets both of these requirements and as such is broadly used for chemical analyses and chemical mapping of atomic and molecular species on solid samples. This thesis describes results on the characterization of Extreme Ultraviolet Time-of-Flight Laser Ablation Mass Spectrometry (EUV TOF) for analysis of trace actinides and for high lateral resolution imaging of inorganic samples. We demonstrate that this technique, with a nominal sensitivity around 50 ppm and a lateral resolution of 80 nm, is already competitive with other well-established mass spectrometry techniques that focus on analytical isotopic imaging. The thesis also details the diagnosis and determination of causes of noise that compromise sensitivity in the mass spectra. These results enable EUV TOF to properly orient itself in the field of mass spectrometry and allow multiple disciplines to conduct high resolution compositional mapping quickly and with minimal isobaric interferences.
Open Access
Flavivirus control of lipid metabolism: implications for virion formation, function and pathogenesis
(Colorado State University. Libraries, 2018) Gullberg, Rebekah, author; Perera, Rushika, advisor; Crick, Dean, committee member; Di Pietro, Santiago, committee member; Geiss, Brian, committee member; Wilusz, Jeff, committee member
Dengue viruses (DENV) are the most aggressive arthropod-born viruses worldwide with no currently available antivirals. There is a clear need to understand host viral interactions that can be exploited for therapeutic options. DENV are members of the Flaviviridae family with a positive sense single-stranded RNA genome surrounded by a virally encoded capsid protein, a host cell derived lipid envelope and an icosahedral shell of virally encoded glycoproteins. Its genome is replicated in virally–induced invaginations in the endoplasmic reticulum of the host cell that consistently develop in a time-dependent manner. These invaginations display a highly curved architecture and seem to increase the membrane contact sites within the ER and its vicinity. Functionally, these membranes condense the replication machinery, provide a scaffold to coordinate replication, and hide the viral double stranded RNA intermediate from the host cellular defenses. It has been shown that fatty acid synthesis is increased during infection to provide substrates for this membrane expansion. To identify further changes to cellular metabolism, we have profiled the metabolome of DENV serotype 2 (DENV2) infected Human Hepatoma cells (Huh7) cells at key time-points in virus replication. We have found time-dependent changes in cellular essential fatty acid metabolism. Furthermore, we have interrogated a library of siRNAs directed at the unsaturated fatty acid biosynthesis pathway to determine key enzymes involved in viral replication. We have identified that stearoyl Co-A desaturase 1 (SCD1), the rate-limiting enzyme responsible for converting stearic to oleic acid, is critical for viral replication, maturation and infectious particle formation. Finally, we have profiled the serum metabolome of acute-phase patients with dengue diseases, chikungunya virus infection, or an unknown febrile illness to identify metabolic changes with potential use as prognostic biomarkers. Hypothesis: Since dengue viruses are enveloped viruses, lipid metabolites in the human host are a critical resource hijacked by these viruses for their replicative advantage. Important metabolites will be altered during infection in a time dependent manner and can be quantified and correlated directly to their role in viral genome replication and infectious particle assembly and release. These metabolic changes could also be identified in human bio-fluids and could function as early biomarkers of disease manifestation.
Open Access
Investigating biosynthetic pathways of the Aspergillus genus through biomimetic total synthesis of secondary metabolites
(Colorado State University. Libraries, 2022) Benson, Brooke, author; Williams, Robert M., advisor; Kennan, Alan J., advisor; Paton, Robert, committee member; Crans, Debbie, committee member; Crick, Dean, committee member
The prenylated indole alkaloids are a class of secondary metabolites containing a unique bicyclo[2.2.2]diazaoctane core and a wide range of biological activity. This complex structure has prompted extensive investigation into the biochemical synthesis of these compounds. Currently, three disparate biochemical strategies are known to be used by producing fungi to construct the bicyclic core: (1) NADPH-dependent bifunctional reductase/Diels-Alderase-mediation in formation of the monooxopiperazines; (2) brevianamide assembly through cofactor-independent pinacolase resulting in spontaneous intramolecular Diels-Alder (IMDA) generation of the bicyclo[2.2.2]diazaoctane core; (3) Diels-Alderase mediated enantiodivergent generation of the dioxopiperazines via cytochrome P450 oxidation to achiral azadienes and successive enzyme-mediated stereoselective IMDA reaction. This work aimed to employ biomimetic total synthesis to aid in elucidation of the biosynthetic pathways in the Aspergillus genus, which utilizes the third strategy. This author reports the first total syntheses of 6-epi-Notoamides T10-12 and Notoamide T2, as well as an improved total synthesis of 6-epi-Notoamide T. Also reported are synthetic efforts towards 6-epi-Notoamide T9, Notoamide TI, and Citrinalin C.
Open Access
Investigation into Mycobacterium bovis phenolic glycolipid as a potential biomarker of bovine tuberculosis in urine
(Colorado State University. Libraries, 2019) Morphet, Stephanie Marie Little Thunder, author; Belisle, John, advisor; Crick, Dean, committee member; Wilusz, Carol, committee member; Henry, Chuck, committee member
Mycobacterium bovis, the etiological agent of bovine tuberculosis (bTB) is reported to cause disease in man and animal alike on every continent aside from Antarctica. Although coordinated efforts have been made for over a century in the US to cease transmission of this pathogen, outbreaks still occur. It has been posited that the failure to eliminate transmission of this pathogen is partially due to the diagnostic in use, which lacks critical sensitivity and specificity. To address this gap, we investigated a potential new method of identifying infected animals that is through the detection of a pathogen-derived biomarker. M. bovis phenolic glycolipid (PGL) is a species-specific, highly abundant, and unique glycolipid that comprises up to 2.5% of the dry cell mass. Coupling an LC-ESI-TOF-MS, method with a solid phase extraction, we successfully detected PGL derived from the urine of naturally-infected cattle. With this knowledge, we aimed to generate a detector of PGL that could be applied in a rapid and field friendly diagnostic platform. Using phage display technology, we selected M13 bacteriophage capable of binding M. bovis PGL with specificity that differentiated between M. bovis PGL and M. Canetti PGL, as well as between total lipid fractions of various species of Mycobacterium and other lipids with similar biochemical properties to PGL. These M. bovis PGL specific phage were able to differentiate between unspiked cattle urine and urine spiked with PGL. Lastly, we assessed the relative stability of PGL, specifically contrasted to another highly abundant mycobacterial glycolipid, phosphatidylinositol mannoside (PIM). We found that PGL was the more stable molecule when testing thermal and chemical stability, as well as when treated with protease K. However, when treating these glycolipids with whole cell lysates derived from fresh bovine organs, accurate detection on an LC-ESI-TOF-MS platform was lost. Further studies will be required to probe the stability of these molecules in vivo. Overall, this potential methodology to assess for infection status may be beneficial in improving the control of M. bovis if further developed, as this new approach has the potential to be more specific and sensitive than the currently used diagnostic.
Open Access
The C3HeB/FeJ mice as a novel preclinical mouse model for Mycobacterium tuberculosis: an analysis of the host pathogenesis and the in vivo environment of the necrotic granuloma
(Colorado State University. Libraries, 2014) Driver, Emily R., author; Lenaerts, Anne, advisor; Basaraba, Randall, committee member; Crick, Dean, committee member; Gustafson, Daniel, committee member
Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis (TB), infects and kills millions each year. Actively infected patients exhibit a heterogeneity of disease lesions in the lungs, which make it difficult for current treatment regimens to effectively cure. Efforts are underway to develop novel pre-clinical drugs, and this requires a thorough re-evaluation of the current animal models used to test these formulations. A new mouse model using C3HeB/FeJ mice, deficient at the sst-1 locus develops a similar heterogeneity of pulmonary lesions to human disease when aerosol infected with MTB. The studies presented here have found the C3HeB/FeJ mice develop 3 types of lesions: type I) necrotic granulomas delineated with a collagen rim, type II) necrotic alveolitis, and type III) cellular inflammation. Type I lesions, consist of an accumulation of amphophilic cellular debris admixed with numerous extracellular bacilli. At the periphery of the necrotic core on the inside of the collagen rim is a cuff with foamy macrophages of variable size, which contain very large numbers of intracellular bacilli. Type II lesions, the alveolar septal walls remain mostly intact, while the alveolar spaces are filled with high numbers of dying neutrophils, intact cells, extracellular bacteria and debris, creating a honeycomb-like necrosis, which is only seen at very late stage in immunocompetent mouse models. Type III lesions, are disorganized and have a mix of lymphocytes, macrophages (epitheloid and foamy), and a small number of neutrophils with low numbers of intracellular bacilli. The necrotic encapsulated type I granuloma creates an environment that promotes hypoxia, creates a physical barrier, and causes physiological changes to the various bacterial populations. The Kramnik mouse model has higher populations of resistant bacteria when drug treated, such as isoniazid (INH), rifampin (RIF), and pyrazinamide (PZA). These drugs are less effective when administered as monotherapies in C3HeB/FeJ mice. The adaptations the MTB undertake in the core of the type I granuloma while under stress not only make drug treatment less effective, but also make standard acid-fast staining techniques, such as auramine-rhodamine less efficient at detecting bacilli. While, Sybr Gold, a novel acid-fast stain that binds to nucleic acid, is capable of staining bacilli throughout infection and in all lesion types. The discrepancy in staining ability of the two acid-fast stains is indicative of the physical target for auramine-rhodamine no longer being present or available for staining in the bacilli in the core of the necrotic granuloma. The knowledge on histopathology then leads us further to modify the granuloma in the Kramnik mouse model. Firstly, using collagen disrupters we aimed to improve treatment efficacy. Secondly, by using clinical TB strains the goal was to advance disease to potentiate cavitary disease and gain a greater understanding of strain relevance to treatment. Thirdly, by taking advantage of the structural similarity of the necrotic granuloma and solid cancerous tumors allow for the translation of therapy technologies, such as liposomal nanoparticles to be exploited for treatment of MTB. Preliminary findings suggest that the use nanomedicines in TB therapy may be an effective method of drastically reducing treatment as well as potential issues. The C3HeB/FeJ mouse strain is a highly relevant disease model that can be used for determining the efficacy of novel pre-clinical drugs and drug regimens, to gain a better understanding of disease pathogenesis, to understand the specific immunological events of disease, and to explore alternatives to standard therapy.
Open Access
The formation and reactivity of α,β-unsaturated platinum carbenes: new approaches to heterocycle synthesis
(Colorado State University. Libraries, 2014) Allegretti, Paul Andrew, author; Ferreira, Eric, advisor; Rovis, Tomislav, committee member; Kennan, Alan, committee member; Elliott, C. Michael, committee member; Crick, Dean, committee member
The use of transition metal carbenes in organic synthesis has enabled chemists to promote an incredible variety of bond-forming reactions. These intermediates are traditionally accessed via the decomposition of diazo species. Alternative methods for accessing metal carbene intermediates from functional groups beyond diazo species could present many practical advantages and would complement the existing methods. Presented herein are our efforts to develop a novel method of carbene generation from the reactions of alkynes and π-acidic transition metals, and their use in the synthesis of different heterocycles. Our initial investigations focused on the ability to generate substituted furan products from the intramolecular addition of alcohols into alkynes containing propargylic methyl ethers. We then extended this reaction manifold to the synthesis of complementary positional isomers of isoxazoles, adding both propargylic N-hydroxycarbamates and propargylic N-Boc amino ethers into the activated alkyne. Key to this transformation was the inclusion of Brønsted acids to facilitate activation and elimination of the methyl ether leaving group. While studying the synthesis of furans and isoxazoles it was observed that the nature of the carbene could be modulated, leading to selective silicon or hydrogen migration. Also observed was the addition of nucleophiles into the β-position of the carbene species. This last mode of reactivity led to the discovery of a new way to synthesize vicinal bisheterocycles using alkyne activation conditions.
Open Access
The synthesis of the pentacyclic carbon framework of the PF1270 family of natural products
(Colorado State University. Libraries, 2014) Sanchez, Michelle A., author; Williams, Robert M., advisor; Kennan, Alan, committee member; Rovis, Tomislav, committee member; Levinger, Nancy, committee member; Crick, Dean, committee member
The PF1270 family of natural products contains novel indole alkaloids that display interesting biological activity; the synthesis of these natural products and their analogs could lead to the discovery of novel therapeutics. Discussed herein is the synthesis of the complete pentacyclic carbon framework of the PF1270s, accomplished through a key intermolecular Diels-Alder reaction. Other highlights of the synthesis include an acid catalyzed opening lactim ether at a late stage, and a particularly difficult decarboxylation promoted by diphenylphosphoryl azide.
Open Access
The use of model membrane techniques for the analysis of interactions, conformation and redox properties of menaquinones and other small molecules
(Colorado State University. Libraries, 2021) Doucette, Kaitlin A., author; Crans, Debbie, advisor; Crick, Dean, committee member; Roess, Deborah, committee member; Menoni, Carmen, committee member
This thesis explores the use of model membranes to solve complex problems in determining the placement, conformation, and electrochemical properties of hydrophobic compounds as they interact with a model membrane. Menaquinone, an electron transporter commonly found in Gram-positive and Gram-negative obligate anaerobes, consists of a naphthoquinone head group and isoprene tail of variable length and saturation. Chapter two shows the use of liposomal model membranes to solubilize menaquinone analogues of variable length and saturation for aqueous electrochemical studies characterizing half-wave potentials, reversibility, and diffusion coefficients to examine its redox properties in connection to its role as an electron transporter. This work shows a distinct odd-even effect with respect to the isoprene chain length of the compound and its electrochemical properties. The exploration of this project is continued in chapter three, in which the conformation and placement of menaquinone-2 is determined in the context of a phosphatidylcholine liposome using 1D and 2D 1H NMR. Finally, chapter four explores the use of a reverse micellar model membrane for determining the placement of glycine and short glycine peptides to explore its placement near a membrane with regards to its role as a neurotransmitter and to explore its potential role in antimicrobial peptides. The experiments contained herein show that model membranes are a useful tool for the study of hydrophobic compounds and molecules commonly found within a cellular membrane.