<p>Medium-sized cyclic ethers are challenging synthetic targets due to enthalpic and entropic barriers. Methods for the stereoselective synthesis of &#945;,&#945;&#900;-disubstituted medium-sized cyclic ethers began to appear with the discovery of naturally-occurring, ladder-shaped polycyclic ethers, such as brevetoxin B, and monocyclic ethers, such as (+)-laurencin. Despite the progress made in this field, limitations remain including competing formation of smaller ring sizes and scarcity of catalytic methods. Our aim has been to develop stereoselective syntheses for 7- and 8-membered cyclic ethers which have potential for application in natural product synthesis. The C-O bond disconnection was selected for the methods described within because cyclization and stereoinduction could be achieved simultaneously. In the case of 7-membered cyclic ethers, an organocatalytic oxa-conjugate addition reaction promoted by the gem-disubstituent (Thorpe&#8722;Ingold) effect has been developed to stereoselectively provide &#945;,&#945;&#8242;-trans-oxepanes. A gold(I)-catalyzed alkoxylation reaction has also allowed access to &#945;,&#945;&#8242;-cis-oxocenes. This method has been probed for feasibility in the stereoselective synthesis of (+)-intricenyne, an 8-membered cyclic ether belonging to the C15 nonterpenoid acetogenin natural product class. These methods have the potential to become general and efficient routes to highly functionalized oxepanes and oxocenes.</p> / Dissertation

This Thesis describes the use of alkaline earth and rare earth complexes bearing phenolate ligands as catalysts in the amine-initiated, immortal ring opening polymerisation (ROP) of cyclic esters. Mechanistic elucidation was performed and two propagation pathways are presented. Chapter One introduces cyclic esters and catalytic routes to polyesters by ROP. Common techniques for polymer characterisation are described and an overview of relevant phenolate-supported ROP catalysts is given. Reversible chain transfer in ROP is also discussed. Chapter Two describes the synthesis and characterisation of zwitterionic Group 3 complexes bearing bis(phenolate)-amino ligands and the development of the amine-initiated, immortal ROP methodology using this class of catalyst. Detailed studies into the ROP of rac-lactide using amines and a zwitterionic yttrium complex are presented and the mechanism of amine-initiated, immortal ROP was derived. Chapter Three documents further amine-initiated, immortal ROP studies using a zwitterionic yttrium complex as the catalyst. The preparation of multiarm polymers is described and further investigations using the cyclic esters, ε-caprolactone and rac-β-butyrolactone are presented. Chapter Four describes the use of Group 2 and lanthanide phenolate complexes in the amine-initiated ROP of rac-lactide. Bulk polymerisation studies revealed the generality of the amine-initiated, immortal ROP methodology and an alternative propagation pathway was derived from mechanistic studies. Chapter Five details the synthesis and characterisation of Group 3 amide complexes supported by phenolate-amino ligands. Each complex was screened for ROP capability and amine co-initiators were employed. Chapter Six contains experimental details and characterisation data for the new complexes and polymer products described in this Thesis. CD Appendix contains crystallography .cif files, supporting information for each Chapter and spreadsheets containing polymerisation data.

The fragmentation of γ-silyloxy-β-hydroxy-α-diazoesters to provide tethered aldehyde ynoates was discovered and developed in Prof. Brewer's laboratory. This reaction is a Lewis acid mediated heterolytic cleavage of the Cβ-γ bond of a γ-silyloxy-β-hydroxy-α-diazocarbonyl functional group array contained in a ring compound. This dissertation describes a further study of this ring fragmentation reaction and application of this fragmentation to the preparation of synthetically useful organic molecules. The purpose of this dissertation work was three fold. The first objective was to extend this ring fragmentation reaction to the synthesis of tethered aldehyde ynones by fragmenting various γ-silyloxy-β-hydroxy-α-diazo ketone compounds. The second objective was to develop a new way to make medium size rings by fragmenting fused bicyclic γ-silyloxy-β-hydroxy-α-diazo ketones. The final goal was to use this reaction to make medium size ynolides by fragmentation of fused bicyclic γ-silyloxy-β-hydroxy-α-diazo esters to provide core structures for medium-size lactones which are synthetically challenging to make using other available methods.

A novel cryptand was synthesized which contained a 3,5-disubstituted-4- oxahexacyclo[5.4.1.02,6.03,10.05,9.08,11] dodecane "cage" moiety. In alkali metal picrate extraction experiments the cryptand exhibited high avidity towards Rb+ and Cs+, when compared with the corresponding model compound. A computational study of a series of cage-functionalized cryptands and their alkali metal-complexes was performed. The X-ray crystal structure of a K+-complexed bis-cage-annulated 20-crown-6 was obtained. The associated picrate anion was found to be intimately involved in stabilization of the host-guest complex. The interaction energy between the host-guest complex and picrate anion has been calculated, and the energy thereby obtained has been corrected for basis set superposition error.

The principal objective of this investigation is to develop a constitutive model to simulate the hysteresis behaviour of unbonded flexible risers. A new constitutive model for flexible risers is proposed and a procedure for the identification of the related input parameters is developed using a multi-scale approach. The constitutive model is formulated in the framework of an Euler-Bernoulli beam model, with the addition of suitable pressure terms to the generalised stresses to account for the internal and external pressures, and therefore can be efficiently used for large-scale analyses. The developed non-linear relationship between generalised stresses and strains in the beam is based on the analogy between frictional slipping between different layers of a flexible riser and frictional slipping between micro-planes of a continuum medium in nonassociative elasto-plasticity. Hence, a linear elastic relationship is used for the initial response in which no-slip occurs; an onset-slip function is introduced to define the ‘noslip’ domain, i.e. the set of generalised stresses for which no slip occurs; a nonassociative rule with linear kinematic hardening is used to model the full-slip phase. The results of several numerical simulations for a riser of small-length, obtained with a very detailed (small-scale) non-linear finite-element model, are used to identify the parameters of the constitutive law, bridging in this way the small scale of the detailed finite-element simulations with the large scale of the beam model. The effectiveness of the proposed method is validated by the satisfactory agreement between the results of various detailed finite-element simulations for a short riser, subject to internal and external uniform pressures and cyclic bending and tensile loadings, with those given by the proposed constitutive law. The merit of the present constitutive law lies in the capturing of many important aspects of risers structural response, including the energy dissipation due to frictional slip between layers and the hysteretic response. This privilege allows one to accurately study the cyclic behavior of unbonded flexible risers subject to axial tension, bending moment, internal and external pressures.

Numerous nickel titanium rotary file systems have been introduced in recent years. The recommended speed at which these files are to be operated varies widely between manufacturers. The purpose of this study was to test the number of rotations to fracture of NiTi rotary files when operated at different speeds and at different angles. Sixty size 25 Profile® nickel titanium rotary files were tested, thirty 0.04 and thirty 0.06 taper. Files were operated at speeds of 350 or 600 rpm and at angles of 25°, 28°, and 33.5°. The time to fracture was recorded and the numbers of rotations to fracture were calculated. The data was analyzed using a two-way ANOVA, and post hoc Tukey-Kramer multiple-comparison test with alpha = .001. The results show a significant difference in the number of rotations fracture according to taper and angle. Files of .06 taper fractured more readily than files of .04 taper as the speed increased, and the time to fracture decreased for both tapers as the angle increased. No significant difference was produced by the speed at which the files were operated. The number of rotations until fracture of NiTi rotary files is not related to the operational speed.

The purpose of this study was to examine the effect of immersion in sodium hypochlorite on nickel titanium rotary files and determine whether resistance to fracture was influenced by the immersion time. 70 ISO size 25 0.04 taper nickel titanium K3 and Profile rotary instruments were immersed in 5.25% sodium hypochlorite for 1, 5, 15, 30, 60, and 120 minutes (n=5 each). Resistance to fracture was tested by rotating files at a 30 degree angle at 350 rpm in a universal testing machine. Time to fracture was recorded and analyzed by a two-way ANOVA. Within both file types, there was a decreased time to fracture with increased immersion time in sodium hypochlorite. K3 files required an average of 327 rotations to fracture, while Profiles required 420 rotations to fracture. Profiles required significantly more rotations to failure than K3 files.

FABRICATION OF NANOPOROUS GOLD AND BIOLOGICAL APPLICATIONS By Badharinadh Uppalapati A Dissertation submitted in partial fulfillment of the requirements for the degree of Master of Science at Virginia Commonwealth University. Virginia Commonwealth University, 2014 Major Director: Maryanne M. Collinson, Professor, Department of Chemistry Fabrication of nanoporous gold electrodes by dealloying Au:Ag alloys has attracted much attention in sensing applications. In the first part of this work, the electrochemical response of the redox active molecule, potassium ferricyanide, in a solution of bovine serum albumin in buffer, serum or blood was studied using nanoporous gold and comparisons made to planar gold. Nanoporous gold electrodes with different surface areas and porosity were prepared by dealloying Au:Ag alloy in nitric acid for different dealloying times, specifically, 7.5, 10, 12.5, 20 minutes. Characterization was done using scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), energy dispersive X-ray spectroscopy (EDX), and cyclic voltammetry (CV). Using cyclic voltammetry, planar gold electrodes exposed to bovine serum albumin in buffer showed a decrease in Faradaic peak current and an increase in peak splitting for potassium ferricyanide. The time required for the peak Faradaic current to drop to one-half of its original value was 3 minutes. At nanoporous gold electrodes, however, no significant reduction in Faradaic peak current or increase in peak splitting was observed. Nanoporous gold electrodes having the smallest pore size and largest surface area showed ideal results to biofouling. These electrodes are believed to impede the mass transport of large biomolecules while allowing small redox molecules to exchange electrons effectively with the electrode. In the second part of this work, the open circuit potential (OCP) of biologic solutions (e.g., blood) was measured using nanoporous gold electrodes. Historically, the measurement of blood redox potential has been hindered due to significant fouling and surface passivation of the metal electrodes. As nanoporous gold electrodes retained electrochemical activity of redox probes like potassium ferricyanide in human serum and rabbit blood, they were used to measure the OCP of blood and plasma from various animals like pig, rabbit, rat, monkey and humans. Comparisons were made to planar gold electrodes. The OCP values at both the planar gold and nanoporous gold electrodes were different from each other and there was variability due to different constituents present in blood and plasma. The OCP of rabbit blood and crashed rabbit blood was measured and the values were found to be different from each other indicating that ORP helps in measuring the animal condition. Ascorbic acid was added to rabbit and sheep blood and OCP measured at the nanoporous electrodes. Addition of reducing agent to blood at different intervals and different concentrations showed a change in potential with concentration.

Lyme disease is the most common tick-borne disease in North America, with approximately 35,000 cases reported to the Centers for Disease Control in 2008. The genome of its causative agent, Borrelia burgdorferi, encodes for a set of genes involved in the metabolism and regulatory activities of the second messenger nucleotide, cyclic-di-GMP (c-di-GMP). Rrp1 is a response regulatory-diguanylate cyclase, and its regulatory capability is likely mediated via production of c-di-GMP, as it lacks a DNA-binding domain. One known class of c-di-GMP effector/binding proteins are those that harbor a PIlZ domain. The genome of B. burgdorferi strain 5A4 encodes for one chromosomally-carried PilZ domain, which we have designated PlzA. Additionally, certain B. burgdorferi strains encode for a second PilZ domain-containing protein (PlzB) which is plasmid-carried. Both PlzA and PlzB were found to bind specifically to c-di-GMP, and c-di-GMP binding by PlzA was found to be dependant upon arginine residues in the c-di-GMP binding region. Additionally, expression of PlzA was found to be upregulated by tick feeding and was constitutive in the mammalian host. We next constructed two deletion/allelic exchange mutants – one with the targeted deletion of PlzA, and on ethat replaced PlzA with PlzB in a strain lacking the plzB gene. Our studies demonstrated that ΔplzA was deficient in motility and was also non-infectious in the mouse model of B. burgdorferi infection. Additionally, this strain remained viable in larval Ixodes ticks. Also, B31-plzB KI was deficient in motility, as well as infectivity, demonstrating that PlzB is unable to complement for functions fo PlzA in vitro and in vivo and that it may play other roles in the biology of B. burgdorferi strains carrying the plzB gene. These studies represent the first identification of a c-di-GMP binding protein in any spirochete, but also represent the first demonstration of the importance of PilZ domain proteins in a spirochetal system. We additionally examined the effects of c-di-GMP synthesis and breakdown in the related bacterium, B. hermsii, a causative agent of tick-borne relapsing fever (TBRF). Deletion mutants in Rrp1 (B. hermsii’s sole diguanylate cyclase) and PdeA (B. hermsii’s only EAL domain-containing phosphodiesterase) were created. These strains were analyzed in order to determine: 1) the effect(s) of the losse of Rrp1/PdeA on intracellular spirochete c-di-GMP levels, and 2) the effects of Rrp1/PdeA on the establishment of murine infection and on gross motility/chemotaxis. It was demonstrated that c-di-GMP accumulates intracellularly in the cells lacking PdeA. Additionally, spirochetes were shown to chemotax towards N-acetyl-glucosamine (NAG) and they did not form soft agar swarms. In contrast, cells lacking Rrp1 did not accumulate detectable levels of c-di-GMP, demonstrated a reduced ability to chemotax towards NAG, and swarmed on soft agar in a fashion indistinguishable from wild type. Despite these differences in phenotype, both mutant strains display an attenuated murine infectivity. These results indicate that c-di-GMP is indeed important in the TBRF spirochete, B. hermsii and this vital second messenger plays key roles in virulence, motility, and chemotaxis. These studies also pave the way for future investigation of B. hermsii through use of targeted genetic manipulation.

Lyme disease is a tick-borne infection caused by Borrelia burgdorferi, B. garinii, and B. afzelii. These spirochetes experience environmental fluctuations as they are passed between mammalian and Ixodes tick hosts throughout their enzootic cycle. Recent studies have suggested cyclic diguanylate (c-di-GMP), a ubiquitous secondary messenger, is a key modulator of B. burgdorferi adaptive responses and may play a significant role in cycle progression. In this study, we examined the impact of the sole diguanylate cyclase (Rrp1), c-di-GMP binding proteins (PlzA and PlzB), and HD-GYP-containing phosphodiesterase (PdeB) in disease establishment of both murine and Ixodes tick systems. Strains harboring targeted gene deletions or plasmid-based constitutive gene expression constructs were generated. Rrp1 was required for tick colonization, yet overexpression abolished murine disease, thus implicating the requirement of finely regulated c-di-GMP levels for enzootic cycle progression. Deletion of rrp1 disrupted translational motion and swarming patterns by causing extended cell runs, eliminating stops/flexes, and reducing swarming capabilities. This was attributed to a defect in N-acetyl-D-glucosamine (NAG) metabolism and chemotaxis. NAG is a major source of nutrition for B. burgdorferi within the tick environment; therefore this defect would impede spirochete migration towards feeding ticks, as well as pathogen uptake and survival within the Ixodes vector. In contrast, the downstream c-di-GMP effector, PlzA, was critical for murine disease but nonessential for survival within ticks nor functionally complemented by PlzB. Deletion of plzA altered strain motility and swarming similarly to the rrp1 deletion mutant, yet had a distinct phenotype with significantly slower translational motion and no affect on NAG chemotaxis and metabolism. This indicates B. burgdorferi could possess alternate c-di-GMP effectors or Rrp1 could be directly influencing these cellular processes. Uniquely, PdeB did not abolish murine infection via needle inoculation, but wasrequired for natural transmission from ticks. This defect was linked to the decreased tick colonization efficiency upon pdeB deletion. Together, these analyses indicate that c-di-GMP signaling is an important virulence mechanism of Borrelia burgdorferi and demonstrate the complexity of this signaling pathway in an arthropod-borne pathogen. The data presented here additionally provide significant new insight into the gene regulatory mechanisms of the Lyme disease spirochetes.