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This thesis describes synthetic efforts towards two natural products matrine and pinnaic acid. The strategy towards matrine focused on effecting an overall, but not necessarily concerted [4+2] cycloaddition in which a 1,4-dihydropyridine supplies the dienophile and aN-vinyl pyrldone is activated to act as the diene component. Attempts to progress an expedient route to pinnaic acid that starts with a chiral pyrrolidine cata lysed conjugate addit ion between propionaldehyde and I - nitrocyclopentene is discussed. Chapter 1 is an introduction to previous syntheses of matrine and introduces background literature examples of azadienes or dihydropyridines being used in [4+2] cycloadditions, as well as methods for constructing enamides required for the target. Chapter 2 describes our progress towards the synthesis of matrine. Significant contributions include the use of organotrifluoroborates as air and moisturestable alternatives to boronic acids in the synthesis of N-vinyl pyridones. Zincke reaction and subsequent dithionite reduction gave a high-yield ing route to install the l ,4-dihydropyridines required for the target precursors. Several studies to initiate intramolecular bond formation between these functional groups are reported. Chapter 3 provides an introduction to previous syntheses of pinnaic acid. Examples of transformations required for our proposed route are introduced including organocatalysed 1,4-addit ions of carbon nucieophiles to nitroolefins, an important stereochemistry generating step, and the vinylogous aldol reaction, which are both important components of the planned route. Chapter 4 highlights progress towards the synthesis of pinnaic acid. Significant contributions include the improvement of the diastereoselectivity of our key conjugate addition step via reduction of the in situ generated aldehyde. A panel of organocatalysts was screened for this conjugate addition step, with chiral HPLC studies confirming modest gains in e.e. when trans-4-hydroxy- L-proline was employed. This route was used to gain step-efficient access to an intermediate in Heathcock's synthesis of pinnaic acid. Further studies toward a step efficient route to pinnaic acid are described including a Friedel-Crafts acylation between a protected butynol species and chloroacetyl chloride that was demonstrated on a large scale, albeit with a slight preference for the undesired (E)-isomer. Conversion of this material to phosphonate and sulphone species, as well as its use in a model Wittig reaction, was also demonstrated.

The research presented in this thesis involves the development of a variety of synthetic routes which have enabled the preparation of a library of novel polymeric materials, primarily block copolymers containing a polyferrocenylsilane metalloblock. A large emphasis has been placed on the subsequent investigations into the self-assembly of these materials, where they have been found to form a variety of interesting nanostructures, making them candidates for use in a number of different scientific fields. An introduction to the principal areas of research of this thesis is given in Chapter 1. Block copolymer (BCP) self-assembly in the bulk is discussed, highlighting the interest in studying BCP systems exhibiting greater complexity. This is followed by a similar discussion concerning block copolymer self-assembly in the solution phase. The synthesis and selfassembly of BCPs containing polyferrocenylsilane is introduced. The final sections of the introduction are dedicated to block copolymer lithography and bit-patterned media. Miktoarm star terpolymers represent a class of BCPs which are comprised of three chemically distinct polymeric blocks joined at a single junction point. Previous studies into their bulk self-assembly have shown them to self-assemble into a large number of complex morphologies, including a range of Archimedean tiling patterns which aren't obtainable through the self-assembly of their linear triblock terpolymer counterparts. These materials could be utilised for block copolymer lithography, facilitating the fabrication of features ~xhibiting greater complexity than simpler diblock copolymer. In Chapter 2, the synthesis of a range of polystyrene-arm-polyisoprene-arm-poly(ferrocenylethylmethylsilane) miktoarm star terpolymers ()..t-SIF) is discussed; two synthetic routes were pursued, a chlorosilane route and a core-first route. In total, 5 j.l-SIFs were prepared. Their bulk self-assembly was investigated by transmission electron microscopy (TEM) and three different morphologies were obtained - two Archimedean tiling patterns ([8.8.4] and [12.6.4]) and a lamellae + cylinder morphology. Additionally, the thin film self-assembly and pattern transfer of one of the [8.8.4] Archimedean-tiling-pattern-forming j.l-SIFs is presented, demonstrating the applicability of these materials for use in block copolymer lithography. In addition to the structures formed by the self-assembly of block copolymers in the bulk and thin film, a wide variety of colloidally stable nanoscale architectures have been shown to be accessible through the solution self-assembly of block copolymers. The solution self-assembly of two j.l-SIF terpolymers is described in Chapter 3, one bearing a crystallisable poly(ferrocenyldimethylsilane) block (j.l-SIFc) and the other bearing an amorphous, non-crystallisable poly(ferrocenylethylmethylsilane) block (j.l-SIFa). The selfassembly of these materials was studied by TEM, atomic force microscopy (AFM) and dynamic light scattering (DLS), whereupon it was shown that the solvent and the 1 crystallisable nature of the PFS block have a profound influence on the micelle architectures formed. Spheres bearing phase-mixed cores and distorted spheres bearing phase-separated cores were obtained for j.l-SIFa in different solvents. For j.l-SIFc, a transition from distorted spherical micelles bearing a phase-separated core to elongated fibres bearing a phaseseparated core was observed. Additionally, cylindrical micelles with a phase-separated corona were obtained for Il-SIFc in another solvent. Finally, successful seeded growth studies with the crystallisable j.l-SIFc terpolymer were demonstrated, yielding B-A-B block co-micelles.

Natural product synthesis is a highly regarded and essential aspect of organic chemistry. Its challenging nature requires a wide spectrum of reactions to be performed with a broad set of skills being developed throughout each project. This thesis describes studies towards the total synthesis of nominine, an indole diterpenoid which exhibits antiinsectant properties. Synthesis of this natural product could provide entry into this family of indole diterpenoids allowing several natural products to be accessed. Chapter 1 introduces this natural product and its proposed synthetic route. Two total syntheses, by Bonjoch and Nicolaou published after this project had ceased, are also presented. Chapter 2 discusses the methods of constructing key intermediate enone 1.1 and describes its successful efficient synthesis on a multigram scale. The synthesis of natural products dehydrofukinone and fukinone using this key intermediate follows in chapter 3. Chapter 4 details the challenges and successes of the investigations towards the total synthesis of nominine. Future work for its completion and other possible related indole diterpenoids are described in chapter 5. A conclusion to the project and a chapter containing experimental details conclude this thesis.

This thesis focuses on our progress towards the total synthesis of N-methylwelwitindolinone C isothiocyanate 7, commonly called welwistatin. This major alkaloid of the welwitindolinone family, which was isolated from Hapalosiphon welwitschii in 1994, represents a particularly attractive target due to its interesting biological activities (MDR reversing agent) and its challenging structure. Welwistatin possesses a complex bicyclo[4.3.1]decane ring system consisting of four stereogenic centres, three quaternary carbons and two unusual reactive functionalities: the isothiocyanate bridgehead and a vinyl chloride group. Inspired by the synthetic challenge of this complex architecture, the Simpkins group reported an expedient four-step synthesis of its core structure in 2005. Three years later, our group had investigated the reactivity at the bridgehead enolate positions. Taking inspiration from this previous work, we successfully synthesised bridgehead alkene 126. The other features present on welwistatin 7 were then investigated and enone 198 was obtained. Facing some difficulties on the model system 88, we turned our attention towards the synthesis of tetracycle 170, possessing the gem-dimethyl at the position C\(_{16}\), and its subsequent functionalisation.

Polyphosphate biosynthesis and the stringent response play an important role in the virulence of pathogenic bacteria. Our objective is to validate these pathways as an antimicrobial target and to identify inhibitors of the key enzymes polyphosphate kinase (PPK), (p)ppGpp synthetase I (RelA) and (p)ppGpp synthetase/hydrolase II (SpoT). The role of polyphosphate and (p)ppGpp metabolism in Francisella virulence has been explored with deletion mutants. These exhibited defects for intracellular growth in macrophages and were attenuated in mice, indicating a key role for the FtPPK, FtRelA and FtSpoT in the virulence of Francisella. The development of three in vitro activity assays will enable the discovery of PPK inhibitors. Ion-pairing HPLC analysis has been used to measure substrate kinetics, providing evidence that FtPPK belongs to the PPK2 superfamily with little preference between substrates (KM: ADP - 369 μM; GDP - 624 μM) as displayed by other PPK2 enzymes. 31P NMR spectroscopy has been used to monitor the overall time course of the PPK reaction. To facilitate high-throughput screening, a coupled luminescence based activity assay has been developed in a 96-well plate format. These assays can also be applied to the discovery of inhibitors for FtRelA and FtSpoT. Understanding the structural basis of inhibitor action requires a crystal structure of the target enzyme. For FtPPK, a crystallisation screen has identified conditions for obtaining suitable crystals and data has been collected to 2.1 Å resolution. Future studies will use the high throughput assay to identify PPK inhibitors; NMR and HPLC assays to characterise the mode of action and crystal structures of PPK:inhibitor complexes will identify the precise molecular interactions.

The study of interactions within complex formulations is essential for the understanding of stability in colloidal dispersions and in particular the use of adsorbed polymers. Instability of dispersions caused by the displacement of the stabiliser is a well-known problem in industry, especially when two particles compete for the same stabiliser. Most experiments in this thesis involve the adsorption of poly(vinyl pyrrolidone), PVP, onto silica. Although a particle size dependency was observed, all measured silica diameters gave an NMR solvent relaxation rate enhancement upon PVP adsorption. Competition between PVP and poly(ethylene oxide), PEO, was investigated. When there is excess polymer, the PVP will replace the PEO at the silica/water interface. Small-angle neutron scattering measurements were used to complement the NMR data and provide detailed information on the structure of the polymer layer. The competition between particles for polymer adsorption was studied; silica and PVP were used, in competition with either alumina-modified silica or titanium dioxide. The ability for the solvent relaxation NMR technique to measure a population weighted average of the relaxation environments allowed predictions to be made for the relaxation rate in a number of polymer adsorption scenarios. These were compared to the experimental data to reveal that, in both situations, the PVP preferentially adsorbed onto silica. An experiment using SANS indicated that polymer competition may also be studied with the use of contrast-matching methods. Finally, the use of solvent relaxation NMR to study the kinetics of processes such as pigment sedimentation, polymer adsorption and competition was investigated. For the PVP/PEO system, it appeared that the adsorption of the polymers to Si02 was very rapid after sample agitation and displacement occurred in a timescale of minute

This work describes the controlled organocatalytic ring-opening polymerisation (ROP) of cyclic carbonates with pendant groups for the preparation of functional aliphatic poly(carbonate)s and poly(estercarbonate) s. Their subsequent post-polymerisation functionalisations were studied yielding a range of functional aliphatic poly(carbonate)s. Chapter 1 reviews the preparation of cyclic carbonates with pendant functionalities, their ring-opening polymerisation and the postpolymerisation modifications of the resulting poly(carbonate)s. The properties and some applications of functional poly(carbonate)s are also discussed. Chapter 2, 3 and 4 describe the controlled ring-opening polymerisation of allyl- functional cyclic carbonate MAC (5-Methyl-5-allyloxycarbonyl-1,3- dioxan-2-one) and propargyl- functional cyclic carbonate MPC (5-Methyl-5- propargyloxycarbonyl-1,3-dioxan-2-one) using organic catalysts. Successful functionalisation of allyl-functional poly(carbonate)s was achieved via radical addition of thiol-containing molecules to the pendant allyl esters (Chapter 2), while functionalisation of propargyl-functional poly(carbonate)s was realised via the Huisgen 1,3-dipolar cycloaddition of azides to the pendant propargyl groups (Chapter 3). In addition, the copolymerisation of MAC and MPC and the subsequent orthogonal functionalisation of a copolymer was investigated (Chapter 4). Chapter 5 describes the copolymerisation of MAC (and MPC) with stereopure lactide, with resulting copolymers with opposite chiralities being succesfully applied in stereocomplexation. Chapters 6 summarises the results obtained in Chapters 2, 3, 4 and 5 whilst Chapter 7 provides the experimental methodologies.

Verdazyl Radicals as Mediators in Living Radical Polymerisation: The aim of this work was to investigate verdazyl radicals as an alternative to nitroxides as mediators in stable free radical polymerisation. Verdazyl radicals and their unimolecular initiators were synthesised and utilised in the polymerisation of styrene and n-butyl acrylate. Varying degrees of success was observed in the polymerisations depending on the structure of the verdazyl radical. The polymerisation of methyl methacrylate and the copolymerisation of styrene and methyl methacrylate were also investigated. Correlations between observed molecular weight and theoretical molecular weight were poor but may be improved by optimisation of the reaction conditions. Electron paramagnetic resonance was used to elucidate the radical structure as well as to confirm the living nature of the polymerisation technique. Electron paramagnetic resonance was also utilised to provide an insight into radical stability and reactivity in the various reactions undertaken. Dopamine End-Functionalised Polymers for Application as Friction Modifiers: The aim of this work was to synthesise oil soluble dopamine end-functionalised polymers for mechanical testing to determine if the polymers can reduce friction by film formation at a surface. A dopamine based initiator was synthesised and used in Cu(I) and Cu(0) mediated polymerisations with little success and the dopamine catecholic end-group could not be identified as the polymer end-group. To enable a successful living polymerisation, the catechol groups on dopamine required protection. Complete deprotection of the catechol group can be achieved post polymerisation. The polymerisation of lauryl methacrylate was achieved using a polymerisation method designed for the long chain, non-polar molecule which utilised Cu(I)Cl. The polymerisations were scaled up to obtain a baseline, protected dopamine and deprotected dopamine polymers for mechanical testing. A reduction in friction and wear observed for the deprotected dopamine polymer, however, corrosion was also observed and may have affected the results.

The size of the prenylated indole alkaloid family sharing the unique bicyclo[2.2.2]diazaoctane core ring system has grown steadily over the last decades. Due to the complex structures, and in some cases the potent biological activity, these molecules have been adopted as challenging targets for several synthetic groups. Chapter One gives an introduction to these alkaloids, their origin, biological activity and biosynthetic studies. Our strategies to deliver 5,6- and 6,6-fused diketopiperazines (DKPs) and monoketopiperazines (MKPs) are discussed in Chapter Two. Radical cyclisation of a phenylselenyl DKP 108 (Scheme 2.10) and a bromo MKP 140 (Scheme 2.22) gave mixtures of exo and endo products. An alternatizve thio-mediated radical approach allowed access to exo products (Table 2.2 and Scheme 2.28). Previous experiments on the established cationic cyclisation showed that the pyran ring present in the stephacidins is particularly sensitive to the presence of acid. These results prompted us to explore alternative approaches in which the formation of the pyran ring occurs after the key-step (Scheme 3.19 and 3.20). Our progress towards stephacidin A and previous syntheses are discussed in Chapter Three. Synthesis of a sulfide DKP 218 (Scheme 4.22) allowed access to indoline products via radical approach (Scheme 4.27). An oxidative radical approach as well as a cationic approach is also discussed in Chapter Four. In comparison with our new strategy there is also a review of all previously described approaches to assemble the bicyclo[2.2.2]diazaoctane framework. Among the different approaches to access the core structure of these natural products the radical cyclisation approach appeared to be the most efficient. Based on this strategy, synthesis of indolines 348/349 which is structural related to avrainvillamide is discussed in Chapter Five (Scheme 5.10). Our rapid radical methodology allows synthesis of these indolines in 6 steps and 28% overall yield.

Peptide nucleic acids (PNAs) were used as encoding tags to enable the analysis of peptide libraries by PNA/DNA hybridisation onto DNA microarrays. This allowed entire peptide libraries to be organised and sorted in a two dimensional format whereby all library members could be interrogated and analysed on a one-byone basis. In this thesis, PNA-encoded peptide libraries, generated by split-and-mix library synthesis, were screened for a variety of functions. Peptide sequences identified from the screening of a PNA-encoded library were analysed in detail as the first specific substrates for chymopapain. A new PNAencoded library consisting of D-amino acids was synthesised and screened with a number of proteases in attempts to identify novel/unusual substrates. PNA-encoded libraries were also used in the screening of peptide libraries for other activities. Thus substrates for catalyst-free Hüisgen cycloaddition were identified following the reaction between an alkyne modified peptide library and azidofluorescein, while cell-penetrating peptides were identified by hybridization of an internalized encoded library onto a DNA microarray.

The steady-state and time-resolved fluorescence of 2-aminopurine (2AP) have been used to monitor base dynamics and base stacking interactions in DNA single strands and dinucleotides, and to investigate the interactions between DNA and a polymerase, Pfu-Pol. A new class of base analogues has also been investigated using a combination of experiment and quantum chemical computation. In recent years, 2AP has been widely used as a fluorescent probe to study conformational changes and inter-bases interactions in duplex DNA, but the conformational behaviour of DNA in single strands has been far less investigated. In the present work, six 2AP–labelled single strands have been studied by steady-state and time-resolved fluorescence measurements. Single strands were found to show similar conformational heterogeneity (manifested by 4-exponential fluorescence decays) to duplex DNA, but highly stacked conformations, in which 2AP is rapidly quenched by inter-base charge transfer, are less populated in single strands, whereas imperfectly stacked (weakly quenched) conformations are more highly populated. The effect of base pairing in constraining base mobility is evident. To further investigate the influence of base stacking interaction on DNA conformation and the mechanism of inter-base quenching of 2AP, the time-resolved fluorescence of 2AP-containing dinucleotides was measured. The fluorescence decay of 2AP-containing dinucleotides in PBS buffer at room temperature is also multiexponential and the shortest lifetime varies with the identity of the natural base partner, in a manner consistent with quenching by inter-base electron transfer. When the dinucleotides are frozen to 77 K, the quenching of 2AP is almost eliminated, demonstrating the importance of thermal fluctuations of the bases in facilitating inter-base quenching at room temperature. In the frozen dinucleotides, an additional decay component with a lifetime significantly longer than unquenched 2AP is also observed, suggesting the formation of a new, delocalised, inter-base excited-state. Archaeal family-B DNA polymerases bind tightly to uracil and stall replication when they encounter this base in template strands, four bases ahead of the primer-template junction. If the polymerase progresses further towards the uracil, the 3′-5′ proof reading exonuclease becomes stimulated, trimming the primer and re-setting uracil to the +4 position. Uracil sensing prevents copying of the deaminated base and the introduction of mutations into the genome. Time-resolved fluorescence of 2AP has been used to investigate the role played by unwinding of primer-templates in this mechanism. 2AP-labelled primer-templates (2AP positioned next to the terminal 3′ base of the primer strand), with a misincorpated uracil at the +2 position (U+2) or +4 position (U+4) from the replication fork in the complementary template strand, were investigated in complex with the polymerase Pfu-Pol. For the U+2 primer-template, the fluorescence decay parameters show clear evidence for a decrease in the amount of double-stranded DNA on polymerase binding, manifested by marked weakening of inter-base stacking and a large transfer of population from highly stacked to poorly stacked conformations. In contrast, for the U+4 primer-template only a small perturbation to inter-base stacking is seen, together with the persistence of a high population of strongly stacked states. A new class of base analogues with selenium replacing oxygen at the 4 position of thymine and the 6 position of guanine has been investigated experimentally and computationally. These base analogues are interesting because they show a large shift (>80 nm) in their absorption spectrum compared with the natural bases, taking their absorption into the visible region, with minimal change in molecular structure. The potential of two examples of these analogues, 4-selenium thymine-3’-phosphate and 6-selenium-2’-deoxyguanosine-3-phosphate as luminescent probes has been investigated. However, they prove to have very low emission quantum yields for both fluorescence and phosphorescence. The effect of selenium-substitution on the structural and photophysical properties of the bases has been studied by various ab initio computational methods. It has been found that replacement of oxygen by selenium does not affect the ground state structure but changes the structure of the first excited-state from buckled to nearly planar. The shift in the absorption spectrum on introduction of selenium is successfully predicted by the calculations; the red-shifted absorption band of selenium-substituted thymine is due to a new electronic transition that is not present in the natural base, whereas that of selenium-substituted guanine is from red-shifting of a guanine-like transition.

The three-dimensional structure of a protein, formed as a result of amino-acid sequences folding into compact domains, is regarded as a key factor in its biological function. How and why proteins fold into specific topologies, remain the key focus of scientific research in the field of biophysics. By stripping down complex reactions down to the most basic elements, biophysicists aim to develop simplified models for biological phenomena such as antibody discrimination, viral fusion or self-assembly. Focusing on small model peptide systems, rather than the full proteins from which they were derived, was hoped to result in accurate structural measurements and provide a more transparent comparison between simulation and experiment. The aim of this research was therefore to investigate how accurate these models were when compared against experiment. Furthermore, while breaking down the complex biological phenomena into simple models, there was also a conscious effort to ensure that the models were representative of real biological systems, and a major focus was therefore aimed at determining whether any meaningful biomedical insight may be extrapolated from such models. Peptides found in hormones (human chorionic gonadotropin, luteinizing hormone), viruses (HIV) and amyloid diseases (transthyretin) were selected in order to probe a variety of questions in relation to the aforementioned biological phenomena. Namely, how the primary sequence influenced the three-dimensional structure (and thus its biological function), how its environment could influence such a confirmation, and how these systems aggregated. This doctoral study has made use of a combination of computer simulations and experimental techniques to investigate a selection of biologically relevant peptides; utilising classical atomistic molecular dynamics (MD) simulations to characterise the free-energy landscapes of the chosen peptides, and compare these findings with the secondary structure content predicted by spectroscopic methods such as circular dichroism and infrared spectroscopy. The peptide systems studied within, were found to be characterised by rugged free-energy landscapes unlike their protein counterparts (defined by singular, deep minima). Furthermore, these landscapes were found to be highly plastic and sensitive to changes in the local environment.

The incorporation of a propargyl group to a natural product target allows for a streamlined approach to the investigation of structure activity relationships (SARs) and target identification in forward chemical genetics programmes using a ‘click’-based approach. To this end, an efficient synthesis of O-propargylated tyrosine derivatives was designed, and these have been used in the construction of peptide motifs both (a) derived from phage display libraries and (b) found in natural products. The L-tyrosine derivative Y* (compound I, X=H, R=H) was incorporated into a peptide sequence, PTTIYY, which is known to prevent the inhibition of p53 by the AG-2 protein. Y* has been included as both the terminal and the internal tyrosine in the peptide sequence. ELISA assays were carried out to determine how the binding of PTTIYY* and PTTIY*Y to AG-2 compared to that of the un-marked PTTIYY sequence. The results of these assays allowed new conclusions to be drawn regarding the important binding features of the peptide and possible sites for further optimisation of the AG-2 binding properties of this peptide through ‘click’ functionalisation of the modified tyrosine. The binding of the peptides incorporating Y* was also assessed using MCF-7 breast cancer cell lysate, known to contain the AG-2 protein. These results confirmed those seen for the purified AG-2 ELISA. The related bromo-D-tyrosine derivative (compound I, X=Br, R=Me) has been prepared and employed towards the synthesis of a bisebromoamide derivative. Bisebromoamide is a newly discovered polypeptide, and a promising anti-cancer agent. The bisebromoamide derivative contains a thiazole unit (Tzl), two N-methylated amino acids, and an oxopropyl pyrrolidine (Opp) moiety, which is unique to bisebromoamide in natural products. The activity of this bisebromoamide derivative will be investigated via ‘click’-based affinity chromatography using a new supported linker recently developed within the Hulme group.

This project consists of three parts. In the first part the synthesis of two mycolic acid diastereomers (I and II) was achieved, thereby completing the full range of the cisdicyclopropane a-mycolic acid diastereomers. The second part comprised the development of a new method for the synthesis of the a-alkyl-B-hydroxy fragment of mycolic acids; two novel routes that involved fewer steps ted using L-malic as starting material. The first route which was via diethyl malate was achieved in fewer steps without a significant reduction in percentage yield. The second route via dimethyl malate was also completed successfiilly. Finally, a model cord facto III of a B-hydroxy carboxylic acid without an a-alkyl chain was prepared.

Work was undertaken towards the synthesis of the promising antibiotic lactonamycin (iii). Following the work of Parsons et al. it was proposed that cyclisation of the ene-diyne (i) would give access to advanced pentacyclic intermediate (ii) and that from this a total synthesis of lactonamycin would be achieved (scheme I). Scheme I : Proposed Parsons, Board, Waters cyclisation to form the pentacycle (iii)(For image refer to pdf). A synthesis towards the cyclisation precursor (i) was carried out and a route to the key tetrasubstituted phthalide (v) established. Further chemistry was proposed to complete the synthesis of lactonamycin (scheme II). Scheme II : Formation of a fully substituted benzolactone.(For image refer to pdf). During attempts to introduce the β-bromoallyl group of key intermediate (v) using a high temperature Claisen rearrangement it was established that the benzodioxin (vii) underwent thermolysis to generate the reactive quinone methide intermediate (viii) and that in the presence of a nucleophilic solvent the adduct (ix) was formed (scheme III). Model studies showed the reaction to be both general and high-yielding. Scheme III : Novel quinone methide methodology. (For image refer to pdf).

Surface tethered bottle-brush co-polymers are prepared by ATRP grafting of the macroinitiator brush backbone onto plasmachemical deposited poly(vinylbenzyl chloride) initiator nanofilms, followed by ATRP growth of the side chains (bristles). Lateral force scanning probe microscopy demonstrates that poly(glycidyl methacrylate)-graft-poly(sodium 4-styrenesulfonate) bottle-brush decorated surfaces give rise to an enhancement in lubrication. Patterned polymer brushes are fabricated using molecular scratchcard lithography, where a functional top nanolayer (acting as a resist) is selectively removed using a scanning probe tip to expose underlying ATRP initiator sites. The lateral spreading of grafted polymer brush patterns across the adjacent functional resist surface is reversibly actuated by solvent exposure. Macroporous poly(vinylbenzyl chloride) scaffolds are used for ATRP initiation to generate polymer brushes and thereby actuate pore size. These functionalised macroporous scaffolds are fabricated by a decoupled two-step approach comprising plasmachemical deposition of the host material followed by spontaneous emulsion formation using amphiphilic species. Finally, charge nanopatterning onto polymer film surfaces is accomplished by using an SPM probe tip to create localised corona discharge electrification. The efficacy of surface charging is shown to correlate strongly to the polymer substrate hydrophilicity. Localised plasma generation using a scanning probe microscope tip is then demonstrated to actuate the movement of ATRP surface grafted polyelectrolyte and polyzwitterionic brushes. The raising or retraction of polymer brushes can be controlled by varying the SPM tip polarity.

Viridenomycin is a very important antibiotic, which showed strong inhibitory activity against gram-positive bacteria and an active agent against B16 melanoma. In this work we confirmed the asymmetric and stereoselective synthesis of tert-butyl(1R,3Z,5E,7E,10RS)-10-hydroxy-1-phenylundeca-3,5,7-trienylcarbamate as the key building block towards the synthesis of the southern polyene section of viridenomycin. In this synthesis, a series of reactions were carried out, including Heck-Mizoroki couplings, Suzuki-Myuara couplings, deprotection of the silyl ethers, selective oxidation and enol-acetylation. In addition, the full explanation for regeoselectivity of hydroboration with catecholborane on terminal alkynes was introduced. Also, our efficient iododeboroation methodology was applied to the alkenyl hindered boronate ester and led to the terminal (E)-alkenyl isomer which was the only isomer obtained. Therefore, this iododeboronation will be a very important method for practical use to prepare highly stereoselective terminal (E)-alkenyl iodide. Moreover, selective cleavage of tert-butoxycarbonyl (Boc) for the deprotection of the corresponding triene was studied. In addition, two mild cleavage protecting groups (TFAA, phthalimide) are studied. In conclusion, we have demonstrated an efficient methodology and a flexible approach to the synthesis of conjugated (Z,E,E)-triene core of viridenomycin.

A series of amino-acid derived bis-urea gelators were synthesised, some of which show strong gelation in a wide variety of solvents. The gels were probed at the molecular, microscopic and macroscopic levels to gain insights into the gelation behaviour observed. Mixtures of different gelators also result in gels, some of which show different fibre morphologies and X-ray powder patterns to the pure gelators. The series was extended to include fluorescent 1- and 2-pyrenylalanine derived gelators and the fluorescence behaviour of the gels in solution, the gel state, in mixed gels and with the addition of anions was investigated. Tetrabutylammonium-acetate was found to disrupt urea-hydrogen bonding leading to the break-down of the gels, a process which was followed by NMR spectroscopy, rheometry and fluorescence spectroscopy. Two gelators were used to template the formation of porous polymers which SEM and gas adsorption studies show reflect the different fibre morphologies observed in the gels. The use of supramolecular gels as a medium for controlling the crystallisation of pharmaceutical compounds was developed. Proof of principle for the growth of a wide range of pharmaceutical compounds from a variety of different gels was demonstrated. The supramolecular nature of the gels was exploited by using anions to break down the gels in order to recover the crystals. Comparison of crystals grown from gels with those grown from solution highlighted a number of differences in crystal form, habit and stability. Gelators which mimic the chemical functionality of the drug compound being crystallised were synthesised. A study investigating the crystallisation of 5-methyl-2-[(2-nitrophenyl)amino]-3-thiophenecarbonitrile (ROY) showed some differences in polymorphism when crystallised from a gel designed to mimic ROY compared to generic gels. A placement with the particle science group at GSK explored the potential of supramolecular gels for use in early stage screening of pharmaceutical compounds in an industrial setting.

Previous atomic-level simulations have been shown to provide invaluable insight into the adsorption behaviour of amphiphilic polymers at a water-air interface. Neutron reflectivity profiles generated from these simulations showed good agreement with experiment, particularly at low surface concentrations. Unfortunately, previous detailed atomistic simulations have failed to produce adequate results at high surface concentrations due to crowded configurations, which could not relax within the simulation times available. To tackle this problem, a coarse-graining (CG) technique, where the structure of the simulated molecule is simplified to a chain of beads, has been employed in this study. This provides for the simulation of larger time and length scales allowing for a more detailed study of the capture of polymer chains by a surface and the structure of surface layers. The work presented in this thesis involves development of coarse-grained models for water and for poly(ethylene oxide) (PEO)/water systems, with the aim of reproducing the properties of key importance for the bulk and liquid/vapour interfacial states. These models are then used in the coarse-grained simulation studies of di-and trifluoro dendritic end-capped PEO at an air-water interface; the amphiphilic polymers that have been studied recently by neutron reflectivity experiments. It is shown in this study that simulation of very large polymer chains comparable to that used in real experiments, is achievable using coarse-grained molecular dynamics. Neutron reflectivity profiles generated from simulations of di- and trifluoro dendritic end-capped PEO materials at low polymer concentrations are in good agreement with experiment data. Simulations at high polymer concentrations showed no evidence of a stretched brush structure, in accordance with experimental findings. It is shown from these simulations that there are polymers adsorbed to the interface by a combination of fluorocarbon ends and ethylene oxide segments,resulting in a rather at layer structure. At high surface concentrations of polymers, it proved possible to see the formation of polymer micelles in bulk water. The process of micelle capture by the surface and incorporation of the micelle contents into the surface, were also observed.

An ongoing challenge in polymer science is the preparation of materials with specific surface properties that differ from the bulk, whilst retaining the advantageous mechanical properties of the bulk polymer. We have explored the use of multi-end functionalised polymer additives, which undergo rapid spontaneous adsorption to a surface or interface, as an efficient method of modifying surface properties. These materials are of potential use in tailoring the hydrophobicity of the gas diffusion layer (GDL) in a polymer electrolyte membrane fuel cell (PEMFC), and hence optimising fuel cell efficiency. In this research, reversible addition-fragmentation transfer (RAFT) polymerisation has been used to synthesise a range of well-defined, low molecular weight, end-functionalised poly N-vinyl pyrrolidone (PVP) polymer additives bearing aryl-ether end groups containing up to three, low surface energy, C8F17 fluoroalkyl chains. Polymer end-functionality is introduced via the design of functionalised RAFT chain transfer agents (CTAs). Three novel CTAs have been made in addition to their corresponding end-functionalised PVP additives, in a range of molecular weights. Thin films have been prepared comprised of polymer blends of unfunctionalised PVP and varying percentages of each end-functionalised PVP additive, and these films have been analysed by contact angle measurements, ion beam analysis and atomic force microscopy in order to investigate effects of additive concentration, additive molecular weight, matrix molecular weight, design of functional group and annealing. We have shown that modest amounts of additive (<2.5%) can render the surfaces of bulk PVP hydrophobic and lipophobic.

Molecularly imprinted polymers (MIPs) are synthetic receptors containing binding sites selective for a particular analyte molecule. They have been investigated for applications in many of the same analytical techniques where biological antibodies are currently used, such as in binding assays and sensors. MIPs are made using a template which interacts with a functional monomer: after polymerisation the template is extracted to leave the vacant binding sites. In this project, two new amine functional monomers for molecular imprinting were synthesised;4-vinylbenzylarnine (4-VBA) and diethylvinylbenzyl amine (DEVBA). A thorough study of the molecular imprinting of bisphenol A (BPA) template using DEVBA as a monomer has been described. This MIP showed selective binding towards BPA in the presence of structural analogues. These results were in agreement with the NMR titrations data which suggest a hydrogen bond formation between the BPA and DEVBA during imprinting. The new monomers were also used to prepare MIPs for enantioseparation of ibuprofen (IBP). NMR titrations revealed that DEVBA formed strong complexes with the template (S-IBP). However, enantioseparation of IBP was not achieved on either MIP. In chapter 3, MIPs were prepared for the sulphonamide antibiotic sulfamethazine (SMZ). These polymers were successfully used as stationary phases in HPLC for the separation of SMZ and related structures, both in organic and aqueous mobile phases. Two novel derivatives of SMZ were synthesised in chapter 4. Dansylmethazine (DMZ) was designed as a fluorescent analogue of 5MZ and anthraquinonesulfamethazine (AqSMZ) as a dummy template or electroactive analogue. The optical properties of these novel compounds were investigated. A set of AqSMZ-MIPs were prepared and investigated as HPLC stationary phases, in comparison with the MIPs prepared using SMZ as a template. Unlike SMZMIPs, the AqSMZ-MIPs showed very poor recognition of SMZ and its derivatives. This could be due to limited accessibility to the imprinted sites under the investigated conditions due to polymer swelling. The SMZ-MIPs and AqSMZ-MIPs were investigated for use in a competitive binding assay for SMZ using DMZ as a fluorescent probe. Using a MIP based on AqSMZ and glycerol dimethacrylate (GDMA) as cross-linker, some limited evidence for selective binding of the fluorescent probe was obtained. The poor performance of II - the MIP in a competitive assay, however, suggested the need for more optimisation in order to develop a useful sulphonamides assay.

Amphidinolides T are challenging natural targets for the total synthesis community because of their unique structural architecture. This thesis describes the investigations of the establishment of a novel and convenient route to synthesise four members of the natural product family, amphidinolides T1 and T3-5, from a common late-stage intermediate. The key transformations towards the synthetic pathway are ring formation reactions. The trans-tetrahydrofuran ring is forged efficiently via diastereoselective [2,3]-sigmatropic rearrangement of an oxonium ylide from diazo-generated metal carbenoids. Coupling of a side chain is achieved using a sequence of esterification and RCM reactions. Alternatively, a one-pot RCM followed by hydrogenation is described to shorten the synthetic route.

Nakadomarin A is a polycyclic marine alkaloid of the manzamine family. It was first isolated from the marine sponge Haliclona sp. collected off Manzamo, (Okinawa, Japan) and was characterised by Kobayashi and co-workers in 1997. The structure of this unprecedented compound was elucidated by exhaustive spectroscopic studies which showed that the natural product has a unique hexacyclic skeleton. In terms of biological activity, this natural product displays cytotoxicity against murine lymphoma L1210 cells (IC50 1.3 μg/mL), inhibitory activity against cyclin-dependent kinase 4 (IC50 9.9 μg/mL), anti-microbial activity against the fungus Trichophyton mentagrophytes (MIC 23 μg/mL) and anti-bacterial activity against the Gram-positive bacterium Corynebacterium xerosis. The complex structure of nakadomarin A combined with its biological activities has made it a highly attractive synthetic target. The work described herein displays the most recent contribution to this field from our research group.

Surfactant activity is a property more commonly associated with small molecules than biological macromolecules. However, the significant advantages of improved biodegradability and biocompatibility that could be presented by natural surfactant proteins has elevated interest in a group of only a few proteins where intrinsic surfactant activity appears to be the primary function. Two examples of this group, ranaspumin-2 (Rsn-2) from the foam nests of the tungara frog and latherin, the surface active protein component of horse sweat, appear to be different from other surfactant proteins in the form of their activity. However, the exact molecular basis of this activity is poorly understood. This thesis describes work to rationalise surface activity and related properties in these unusual proteins. The properties of Rsn-2 and latherin including surface activity, interaction with lipid membranes and behaviour in solution were investigated to provide further insight into the characteristics that distinguish the surfactant proteins from both conventional surfactants and other proteins. A second protein component of the foam nests, ranaspumin-1 (Rsn-1), of previously unknown function was also found to be highly surface active and is proposed to function in a similar manner to Rsn-2. A model whereby Rsn-2 functions via a clamshell-like opening was tested through a combination of specialised NMR techniques and site-directed mutagenesis. The results identified features associated with surfactant activity, all of which were consistent with the model. The potential of Rsn-2 as a recombinant fusion partner for the production of functional surfactants or foams was proven by construction of a fluorescent conjugate. Solution state NMR was used to determine the structure of latherin. Information on the dynamic processes taking place in the molecule were derived by analysis of NMR relaxation data. The structure revealed is a super roll fold, similar to a single domain of the BPI-like proteins. A model is proposed whereby latherin recognises the air-water interface via three dynamic, hydrophobic loops at one end of its long cylindrical structure and then unfolds to expose its hydrophobic core at the air-water interface.

Biosynthesis and folding of proteins entering the secretory pathway of eukaryotic cells is coordinated at the endoplasmic reticulum (ER). Any terminally misfolded proteins are sorted for the endoplasmic reticulum associated degradation (ERAD) pathway for destruction. Eeyarestatin I (ESI) is a potent inhibitor of certain stages of protein biogenesis at the ER including protein translocation across the ER and deubiquitination, a late stage of the ERAD process. Here several structurally related Eeyarestatins (ES) were synthesised based on key structural requirements shown in ESI and the structural activity relationship of these compounds both in vivo and in vitro was investigated. Experiments conducted included: cytotoxicity to determine toxicity of ES compounds at several concentrations; cell microscopy to identify vacuole formation frequently exhibited by ESI; SDS page electrophoresis and western blotting for ubiquitin to ascertain the extent of accumulation of ubiquitinated protein and indicator of inhibition of deubiquitination; a translocation block assay looking to determine the inhibitory effect the ES compounds have on translocation of selected proteins across the ER membrane; and finally a secretion assay to determine the ES compound's ability to block overall secretion of proteins from cells. Whilst none of the 'new' analogues seem to be equipotent to ESI in all inhibitory activities, inhibitory activity of compounds ES40, ES24 Click meta and ES24 Click para appear to be more specific towards DUBs, with ES40 being the most active.

The aim of this project was to develop a cascade approach towards perhydropyrrolo-[1,2-a]-quinolines and to apply this to the asymmetric synthesis of the gephyrotoxin alkoids. Chapters Two and Three outline the development of a synthetic route towards a range of cascade precursors, whilst Chapter Four outlines investigations into the enamine-Michael cascade. Central to understanding the cascade process was the discovery that the major product of the enamine-Michael cascade was the unusual tricyclic hydroquinium salt. This can subsequently be engaged in a diastereoselective inter- or intramolecular reduction to afford either a trans-perhydro-[1,2-a]-quinoline or a tetracyclic aminal in high overall yield depending on the C1 oxygen substituent.

In this thesis we show the generation and characteriation of specific GPI-anchored cell surface markers. The creation of these constructs has also allowed the quantification of levels of GPI linked protein shedding into the cell-culture media. Experiments presented in the thesis also show that the addition of a GPI anchor to Growth hormone (GH) appears to reduce expression in the media and that these molecules retain the GPI moiety. We speculated that anchoring cytokine ligands to the cell surface might also modulate receptor mediated signalling. To test the effect of GPI modification on cytokine ligands we fused the GH, and Ob mRNA sequences to the GPI anchor signal sequence of the naturally GPI-anchored protein, Thy-l. To our surprise we found that endogenously expressed GPI anchored cytokines acted as potent non-competitive cytokine receptor antagonists. Furthermore, we demonstrate that GPI anchored GH from a stable CHO cell-line can be purified and reinserted into mammalian cell membranes where they are able to induce receptor mediated signalling. Mammalian cell expression technologies are reliant on methods for obtaining high-yielding stable clones. This thesis describes the development of a high-throughput screening procedure based on the direct pull-down and enrichment of stably transfected cells using magnetic activared cell sorting (MACS). Results presented in this thesis demonstrate that it may be possible to use this procedure to isolate transfected cells from a heterogeneous population and to preferentially recover those rare cell populations which express higher levels of recombinant biopharmaceutical. Thus, GPI anchored markers may be used to facilitating facile cloning and identify population heterogeniety.

Bio-based thermoset composites were prepared from epoxidised linseed oil (ELO) using bio-derived crosslinkers. The use of Pripol 1009 (a dimerised fatty acid derived from natural oils and fats) as a crosslinker yielded homogenous transparent films. The inclusion of catalysts, in particular, 4-dimethylaminopyridine (DMAP), demonstrated a significant improvement in the mechanical properties of the resins. An infrared spectroscopic study coupled with modulated differential scanning calorimetry revealed the epoxide ring opening, followed by etherification occurred during the curing process. The optimum DMAP catalyst loading was 0.5-1 % with respect to the total resin weight. The optimised formulation consisting of ELO, Pripol and DMAP were subsequently combined with starch or modified starch in order to improve the resin properties. Normal corn starch, high amylose corn starch and their acid hydrolysed derivatives were included in the formulation. The addition of starch improved the mechanical properties of the films with high amylose starch yielding a film with the most desirable properties. Expansion of high amylose corn starch (gelatinisation and retrogradation) yielded a high surface area material. The formulation with 20% wt. of gelatinised starch yielded a film with 227% improvement in tensile strength and 166% enhancement in Young’s modulus, compared to those with no added starch. Moreover, expanded starch granules uniformly dispersed in the polymer matrix, resulting in a complete disappearance of phase separation. This was attributed to better interfacial adhesion of porous expanded starch and the polymer matrix. Thermal analysis revealed retardation in the cure process in the presence of starch, however the hydroxyl groups of starch were likely to enhance the extent of curing, as indicated by the higher total enthalpy of reaction. Furthermore, these bio-based composites demonstrated excellent thermal stability. Esterification of expanded starch dramatically decreased the water uptake of the resins however, the mechanical properties were compromised, owing to low thermal stability of the esterified starch.

The central idea in Fragment Based Ligand Discovery (FBLD) is to identify small, low molecular weight compounds (MW < 250) that bind to a particular protein active site. Hits can be used to efficiently design larger compounds with the desired affinity and selectivity. Three approaches to FBLD are described in this thesis. The first topic is the development and assessment of different chemoinformatics procedures to select those fragments that maximally represent the chemical features of a larger compound library. Such a fragment library could be of great value in the so-called “SAR by Catalogue" approach, where the initial stage of fragment growth is by selecting existing compounds that contain sub-structures of the hit fragments. Five schemes implemented in the Pipeline Pilot software are described. The second project was to develop improved approaches to processing Thermal Shift Analysis (TSA) data. The shift in melting temperature can indicate that a ligand binds and thus stabilises a protein. A program, MTSA, has been written which allows more straightforward processing of the experimental data than existing available software. However, detailed analysis of fragment screening data highlighted difficulties in defining the melting temperature and suggest that TSA is not sufficiently reliable for routine screening use. Finally, a number of proteins were assessed experimentally for suitability for FBLD: N-myristoyl transferase (NMT), the bacterial homologue of a GlcNAcase enzyme (BtGH84) and the model system hen egg white lysozyme (HEWL). It was not possible to produce suitable NMT material due to the inherent instability of the protein produced in York. The screening results of HEWL with a new Surface Plasmon Resonance (SPR) assay, a cell based activity assay and TSA were inconsistent and difficult to interpret. However, BtGH84 was suitable for screening by both TSA and SPR. The resulting fragment hits are suitable starting points for further evolution.

Analogues of Antibacterial Natural Products Elizabeth Anne Heaviside, St Catherine’s College, University of Oxford DPhil Thesis, Trinity Term 2012 This thesis is concerned with the synthesis and biological evaluation of structural mimics for the natural products 16-methyloxazolomycin and lemonomycin which display potent biological activity including antibacterial and antitumour activity. Chapter 1 explores methods and approaches to the discovery of new antibacterial drugs and the challenges faced in this respect. It also gives an overview of the properties of the natural products investigated in the following chapters and summarises previous synthetic approaches to these molecules published in the scientific literature. Chapter 2 describes the work carried out towards the synthesis of the diazabicyclo[3.2.1]octane unit of the tetrahydroisoquinoline antitumour antibiotic lemonomycin. The intended retrosynthesis of the natural product led to a 2,5-disubstituted pyrrolidine bearing a 1ʹ-amino functional group; a series of routes were explored for the synthesis of this unit. Using (S)-pyroglutamic acid, strategies using Eschenmoser and thiolactim ether coupling reactions were investigated. A sequence based on the formation of a pyrrolidine ring from the cyclisation of an appropriately substituted oxime ether derived from L-phenylalanine was then implemented but a competing Beckmann rearrangement/Grob fragmentation prevented access to the desired heterocycle. Preliminary investigations were also carried out on the modification of cyclic imines derived from oxime ethers which did not undergo Beckmann rearrangement. Chapter 3 describes the synthesis of a library of densely functionalised tetramic acid and pyroglutamate mimics for the right-hand fragment of 16-methyloxazolomycin, and their coupling with a gem-dimethylamide unit mimicking the middle fragment of the natural product. Tetramates were accessed through the Dieckmann cyclisation of N-acyloxazolidines and were derivatised with various alkyl halides. The pyroglutamates were accessed via the highly diastereoselective aldol cyclisation of N-acyloxazolidines formed by the amide coupling of a threonine derived oxazolidine and β-keto-acids. A series of β-keto-acids were synthesised through the acylation and subsequent ring-opening/decarboxylation reaction of Meldrum’s acid. The formation of right-hand/middle fragment adducts was explored using cycloaddition, alkylation and Sonogashira chemistry before a Wittig protocol led to the formation of adducts (E)- and (Z)- 402 and 403. Biological evaluation of the compounds synthesised in this chapter was carried out using both broth and hole-plate bioassays and active compounds were identified. Of particular note was that the Wittig adducts displayed a higher level of activity against Gram-negative E. coli than either the pyroglutamate or amide motifs alone.

Dynamic covalent chemistry encompasses reversible bond forming reactions which proceed under equilibrium control, where the position of the equilibria are sensitive to changes in environment, and which are often able to undergo component exchange. These virtues provide polymeric nanoparticles incorporating dynamic covalent bonds with the ability to reconfigure or change their structural properties in response to stimuli. In Chapter 1 we critically discuss and evaluate the current state of the art whereby polymer chemists have exploited dynamic covalent bonds within responsive and adaptive polymeric nanoparticles. Chapter 2 describes the synthesis and study of a chemoresponsive polymeric micelle. In this work, aldehyde and alkoxyamine endfunctionalized polymers are shown to link together through a single oxime bond and then self-assemble into micellar aggregates. The chemoresponsive nature of these micellar aggregates is expressed when their disassembly is triggered through the addition of a small molecule alkoxyamine. Chemoresponsive core cross-linked star and nanogel nanoparticles which contain multiple imine cross-links are presented in Chapter 3. These imine linkages are utilized to facilitate the self-assembly process of the nanoparticles, which display chemoresponsive disassembly upon the addition of a small molecule amine. Chapter 4 describes the preparation of core cross-linked star polymers which are both pH-responsive and thermoresponsive. The pH-responsive nature is imparted through the pH-responsiveness of multiple imine linkages, and their thermoresponsive nature arises on account of the thermoresponsive polymer chains contained within their cores. In Chapter 5 nanoparticles possessing pH-responsive imine and redox-responsive disulfide cross-links have been developed where the simultaneous application of both low pH and a reducing agent is required to trigger their disassembly. It is shown that the application of either low pH or a reducing agent does not trigger disassembly. The research presented throughout this dissertation confirms the great potential of dynamic covalent chemistry in the development of stimuliresponsive polymeric nanoparticles.

Environmental pressures are driving detergent formulations to become ever-more so- phisticated blends of stabilisers; probing the interactions occurring in complex systems of hydrosoluble polymers with surfactants and their adsorption onto particles is key for the understanding of the mechanism of colloidal stability. In this thesis, the polymer-polymer and polymer-surfactant interactions have been investigated both in bulk and at the solid/liquid interface. The main techniques used were solvent relaxation NMR, photon correlation spectroscopy, small angle neutron scattering, diffusion NMR and optical reflectometry. The majority of experiments involved polyvinyl pyrrolidone, PVP, adsorbed on silica. Addition of sodium dodecyl sulfate, SDS, decreased the adsorbed amount but increased the layer thickness, leaving the remaining adsorbed polymer in an extended conformation caused by repulsive interactions amongst adsorbed surfactant aggregates and the silica interface. The introduction of a nonionic alcohol ethoxylate surfactant to this system prevented the desorption of the polymer layer as it formed mixed micelles with SDS reducing the total charges per micelle and lowering repulsions between the surface and the surfactant micelles. The effect of a non-adsorbing polymer, sodium polystyrene sulfonate, aPSS, on the stability of silica particles was also investigated, showing that addition of the polyelec- trolyte to the silica dispersion modifies the interparticles interactions. Small roughly spherical clusters were formed, followed by larger, less spherical aggregates resulting in the formation of a dense gel network. Exploring the PVP /N aPSS system showed that these polymers interact in bulk forming NaPSS-rich complexes; interfacial investigations revealed that adding aPSS to PVP previously adsorbed on silica led to surface complexes formation. Finally exposing PVP to sodium polyacrylate revealed some polymer interactions at high pH. These interactions were favoured by increasing molecular weights; adding this polymer to PVP adsorbed onto silica lead to a small decrease in adsorbed amount and some particle aggregation.

This thesis describes concise asymmetric syntheses of prostaglandin PGF2u (7 steps) and the anti glaucoma drug latanoprost. A one-pot proline-catalysed aldol reaction of succinaldehyde and subsequent intramolecular aldol reaction and dehydration reaction gave a bicyclic enal in excellent enantioselectivity on multi-gram scale. This bicyclic enal was converted to PGF2u in five further steps. The lower side-chain was introduced by a conjugate addition reaction and the enolate trapped as a silyl enol ether. The C-II alcohol was installed by a reductive ozonolysis of the silyl enol ether. Both reactions proceeded with excellent selectivity which allowed a gram scale synthesis of PGF 2u to be completed by deprotection and a Wittig reaction.

Clay platelets and silica nanoparticles are used as Pickering stabilizers in the fabrication of hybrid armored polymer particles through a Pickering emulsion polymerization process. A variety of hydrophobic comonomers (i.e., styrene-co- (n-butyl acrylate) (Sty:BA), methyl methacrylate-co-(n-butyl acrylate) (MMA:BA)), styrene-co-(2-ethyl hexyl acrylate) (Sty:2-EHA), vinyl acetate (VAc) and vinyl pivalate (VPiv) are used as organic film forming components. Polymerization kinetics and particle size distributions were examined as a function of monomer conversion. Additionally, key mechanistic features of the polymerization process by quantitatively analyzing the concentration of silica nanoparticles in the water phase during monomer conversion by disc centrifugation are unraveled. It is also showed the crucial role of Laponite clay discs in the particle formation (nucleation) of the Pickering emulsion polymerization process. Increasing amounts of clay nanodiscs leads to smaller average particles sizes, but broader particle size distributions. Polymer films of poly(styrene-co-n-butyl acrylate) armored with Laponite clay were studied as a function of clay amount. Improvements in mechanical, thermal and surface topography provided by clay platelets are reported. In addition, advantages are shown in use of hybrid polymer particles in comparison with simple blend mixtures of polymer particles plus inorganic particles. Humidity properties of poly(styrene-co-n-butyl acrylate) films as a function of clay content are investigated. It is demonstrated that the presence of Laponite clay improves the water storage capacity of polymer films. Also water barrier properties are improved when clay platelets are applied. Finally, a versatile two step Pickering emulsion polymerization for the fabrication of core-shell particles armored with Laponite clay XLS is developed. The obtained particles contain a "hard" core and a "soft" shell armored with clay. The different in the refractive indexes between the core and shell makes these core-shell particles interesting for possible use as colloidal crystals.

The formation of nucleated clusters of β-Lactoglobulin (β-Lg) under alcohol-induced denaturation has been studied by applying spectroscopic and scattering techniques. The concentration dependence of the cluster formation process of β-Lg in 50% (v/v) buffer-ethanol solutions was study at pH 7 and 22°C. Near UV circular dichroism results indicated that the tertiary structure of the native protein was completely lost upon addition of ethanol. Changes in the secondary structure were characterised by a drastic increase of the α-helical content (and decrease of β-sheets content) immediately after ethanol addition. The percentages of these two types of secondary structure evolved further during the 10 days of incubation at room temperature attained an equilibrium state which differs from the secondary structure found in the native protein: while dimeric β-Lg in its native state has a majority content of β-sheets, clusters presented a majority content of α-helix. Similar results on the evolution of the secondary structure of the protein upon ethanol addition were found with attenuated total reflection infrared spectroscopy (ATR-IR). ThT-binding fluorescence spectroscopy revealed that the kinetics of structural rearrangement follows a nucleation dependent growth mechanism for the lowest protein concentration studied here (0.5 mg/ml and 1 mg/ml) where a lag phase, an exponential growth, and a plateau phase could be clearly identified and quantified. For higher protein concentrations (from 4 to 40 mg/ml) the structural transition kinetics of the protein was much faster and no lag phase was observed. CD and fluorescence results suggest that the high amount of β-helix structure formed immediately after ethanol addition for the lowest concentrated samples acts as an activation barrier and slows down the nucleation process leading to a distinct lag phage. Kinetic of cluster growth was monitored by dynamic light scattering, indicating that clusters grew rapidly after ethanol addition reaching a constant hydrodynamic radius shortly after the organic solvent was present in solution. Correlation with spectroscopic results revealed that structural changes were still taking place even after the cluster size attained its equilibrium value for high concentrations of protein (10 and 40 mg/ml). DLS results were in good agreement ii with those obtained by small angle X-ray scattering (SAXS) and were consistent with compact sphere-like structure of clusters. Our results indicate that clusters formed in the presence of 50% (v/v) of ethanol corresponded to a thermodynamically controlled intermediate on the β-Lg aggregation pathway. However, the internal structure of these clusters must contain suitable β-sheet arrangements with enhanced binding of the ThT dye to explain the fluorescence intensity increase found when the clusters were formed.

This thesis is concerned with the total syntheses of the natural products aspidospermidine and aspidofractinine. These targets are noteworthy not only for the biological activity displayed within the class but also for their interesting molecular architecture. Herein, two routes towards the natural products are presented. Key features of the first route, a unified approach to both targets via the indolinyl radical, include the elegant construction of the core ABDE ring system, a mild lactam reduction and synthesis of the key cyclic imine. The second route features a highly efficient Stille coupling and a Wittig olefination. Attempts to effect a critical radical cyclisation reaction are also discussed. The chemistry of the C2 indolinyl radical is investigated, in particular the influence of the C3 indolinyl substitution upon the radical pathway followed. A discussion of the study and its findings is presented in Chapter 4. A review of synthetic approaches to these natural products since 2007 is presented in Chapter 1, in addition to details on their isolation, characterisation and biosynthesis, and an overview of their biological activity. Experimental procedures and characterisation data are provided in Chapter 6.

Triplex-forming oligonucleotides (TFOs) bind to the major groove of the DNA duplex via the Hoogsteen interactions to generate triple helices. Potential applications of triplex technology are in regulation of gene expression, site-directed gene-knockout, mutation correction and as tools in molecular biotechnology. The presence of 2’-modified nucleosides in therapeutic oligonucleotides inhibits enzymatic degradation in vivo. Therefore such sugar modifications have the potential to improve the biological activity of TFOs. We have synthesized the phosphoramidite monomers of six 2’-modified nucleosides from D-ribose via 1-O-methyl-3,5-di-O-benzyl-α-D-ribofuranoside. Three of these are N-linked nucleosides: 2’-O-methoxyethyl-5-propargylamino-uridine (MEPU), 2’-O methoxyethyl-5-methyl-cytidine (MOE-5-MeC) and 2’-O-aminoethylthymidine (AE-T); and three are C-linked nucleosides: 3-methyl-2-amino-pyrdine-2’-O-methyl-ribonucleoside (Me-MAP), 3-methyl-2- amino-pyrdine-2’-O-methoxyethyl-ribonucleoside (MOE-MAP) and 3-methyl-2- amino-pyrdine-2’-O-aminoethyl-ribonucleoside (AE-MAP). These monomers were incorporated into a number of oligonucleotides, on which the biophysical and biochemical studies have been performed. TFOs containing the N-nucleoside (MEPU) showed high duplex affinity and strong nuclease resistance. Studies on two C-nucleosides (Me-MAP and MOE-MAP) revealed that their triplex stability was determined by the sequence context. The incorporation of Me-MAP and MOE-MAP into oligonucleotides renders them much more resistant to the degradation by serum nucleobases compared to their 2’-deoxy derivative (dMAP) and 2’-deoxycytidine (dC). AE-MAP is a promising triplex stabilizer, which not only shows the highest triplex stabilization, but also displays an impressive resistance to enzymatic degradation

This thesis describes the development and improvement of an assay for the detection and discrimination of DNA sequences based on surface enhanced Raman spectroscopy (SERS) and electrochemically driven denaturation. A DNA helix can be immobilised at a nano-structured gold electrode and then denatured into its constituent components by driving the potential at the surface cathodic (electrochemical melting). This denaturation is typically monitored by observing changes in the SERS spectra of a reporter molecule attached to one of the two strands that comprises the helix. The ease of denaturation (defined as the potential required to denature half of the DNA at the electrode surface) was found to be directly related to the thermodynamic stability of the duplex. In addition to dsDNA structure, the effect of varying environmental conditions during an electrochemical melting experiment (pH and ionic strength) was explored. A number of possible mechanisms for electrochemical melting have been ruled-out, including electrostatic repulsion from the electrode surface. It was found that peptide nucleic acid (an uncharged analogue of DNA) could be denatured electrochemically. Local pH changes were also ruled-out as a possible mechanism through experiments in which the surface pH and electrochemical melting were monitored simultaneously. A method for detecting surface immobilised DNA without the need for the attachment of a synthetic label is described. This presents a significant milestone towards the development of a point-of care electrochemical melting assay, because no synthetic pre-treatment of the sample is required prior to analysis.

A variety of supramolecular porphyrin arrays on DNA or single walled carbon nanotube scaffolds are presented herein. A novel porphyrin modified nucleoside with multiple degrees of freedom about the linking moiety has been synthesised. Oligonucleotide strands containing the novel ‘flexible’ porphyrin modified nucleoside or a previously published ‘rigid’ linked porphyrin modified nucleoside were synthesised. The resulting systems were analysed by photospectrometric techniques. Stable B form duplexes were observed in all cases, with the porphyrin modifications imparting a stabilising effect on the duplexes, the degree of stabilisation the novel porphryin monomer provides is of a similar level to that of the rigid linked monomer. Excitonic coupling of the porphyrins is observed; the different monomers incorporated into DNA show different effects in the circular dichroism spectra, which may be explained through the increased conformational freedom of the ‘flexible’ linker. The synthesis of a novel anthraquinone modified nucleoside is presented. Modified DNA strands containing both porphyrin modifications and anthraquinone modifications were synthesised and analysed electrochemically. Cyclic voltammetry has shown that the inclusion of multiple porphyrin modifications increase the electron transfer rate to the anthraquinone redox marker. The synthesese of a novel ferrocenyl modified nucleotide, a novel naphthalene diimide modified nucleotide and an alternative synthesis route for a ruthenium tris-bipyridyl nucleoside are presented. Homo- and hetero-porphyrin single walled carbon nanotube adducts have been prepared with neutral, tetra-anionic and tetra-cationic porphyrins. Significantly elevated loading levels have been observed for the mixed charge species which forms a 1:1 salt on the nanotube surfaces

This thesis describes the development of a novel synthetic route towards the Securinega alkaloids, securinine 1 and virosecurinine 2. The key reaction in this innovative approach is the l,3-dipolar cycloaddition between pyridinium stabilised ylides 169 and γ,δ-unsaturated butenolides 142, to form spirocyclic dihydropyridines 168 as single diastereoisomers. The Introduction chapter introduces the Securinega alkaloids and describes previous syntheses of securinine 1. This section also reports on developments in the field of l,3-dipolar cycloaddition chemistry, with particular emphasis on the cycloadditions of pyridinium stabilised ylides 169. Cycloadditions of γ,δ -unsaturated butenolides 142 with azomethine and other ylides are also described. The Results and Discussion section describes investigations which were conducted towards the total synthesis of securinine 1 and virosecurinine 2. A number of routes to γ,δ -unsaturated butenolides 191 were evaluated and are discussed. The optimum route was found to be a three step procedure from propargyl alcohols 193, via Johnson-Claisen rearrangement to 13- allenic esters, which were hydrolysed and then cyclised under transition metal catalysis. γ,δ -unsaturated butenolides 191 were then employed in l,3-dipolar cycloaddition reactions with pyridinium stabilised ylides 169. Dihydropyridines 168 formed in the cycloaddition step were found to be unstable and as such, strategies for the stabilisation of these compounds are discussed. Reduction afforded stable intermediates 229 prompting investigation to deliver intermediates that could be used to carry out the remaining cyclisation required for the synthesis of tetracyclic structures. Attempts at a variety of such ring closing reactions are described, the most advanced intermediates being committed to an intramolecular Heck reaction. Finally, expansion of the scope of the key cycloaddition reaction is described and opportunities for further developments are discussed. The Experimental section describes all procedures which were used to synthesise compounds disclosed in this thesis. Full spectroscopic data and characterisation is also provided. The Appendices include some relevant XRD, NMR and MS data for clarity.

A novel version of the CSA (Chemical Shift Anisotropy) amplification experiment which results in large amplification factors is introduced. Large xa (up to 48) are achieved by sequences which are efficient in terms of the number of π pulses and total duration compared to a modification due to Orr et al. (2005), and greater flexibility in terms of the choice of amplification factor is possible than in our most recent version. Furthermore, the incorporation of XiX decoupling ensures the overall sensitivity of the experiment is optimal. This advantage has been proved by extracting the CSA tensors for a novel vinylphosphonate-linked nucleotide. The application of CSA amplification experiment to six nucleosides is also discussed. The measured principal tensor values are compared with those calculated using the recently developed first-principles methods. Throughout this work, the NMR parameters of all nucleosides are presented. Finally, high-resolution multi-nuclear solid-state NMR experiments are used to study some novel vinyl phosphonate-linked oligo-nucleotides.

The asperparalines belong to a large family of prenylated indole alkaloids possessing a characteristic bicyclo[2.2.2]diazaoctane core structure. More than 70 examples are known to have been isolated from various diverse sources. Chapter 1 discusses the isolation and structure of these compounds along with an introduction to their biosynthesis and biological activities. A combination of unique structural features and intriguing biological profiles has inspired a large body of work regarding the chemical synthesis of this alkaloid family. Chapter 2 gives an overview of the most important strategies for the construction of the central bicyclo[2.2.2]diazaoctane and presents these in the context of several total syntheses. Chapter 3 details an approach to the bicyclo[2.2.2]diazaoctane core using a free radical cascade comprised of a 1,6-hydrogen atom translocation followed by 6-exo-trig and 5-exotrig cyclisations. An initial model system is presented consisting of a series of DKPs synthesised from a propargylated proline derived from a modification of Seebach’s procedure for the self-reproduction of chirality. Addition of a sulfur radical to the acetylene triggers the desired reaction in good to excellent yield and favours the asperparaline stereochemistry. In Chapter 4 the application of this strategy to the synthesis of an advanced asperparaline core structure is presented. While construction of the key cyclisation intermediate proved difficult, the radical cascade proceeds in modest yield to furnish a bicyclo[2.2.2]diazaoctane possessing the key structural features of the asperparalines.

This thesis describes the synthesis, characterization and study of a range of oligonucleotides that were modified by the inclusion of a non-nucleosidic anthracene unit. This involved the synthesis of different stereoisomers of a non-nucleosidic backbone attached to DNA via a carbon chain linker. These monomers were then coupled to short (15-mer) oligonucleotide chains using solid state DNA synthesis and purified using RP-HPLC. Two systems were studied: a duplex with anthracene functioning as a non-bonding base opposite of a natural nucleobase and a wedge type system. The probes were then examined by a variety of physical techniques that included UV melting point studies, fluorescence and CD spectroscopy in order to understand the effect of varying base pairs and introducing mismatches. Certain duplexes were then studied in greater depth using transient absorption spectroscopy, to probe the formation of electron transfer intermediates and by high field NMR to look at the effect of intercalation between base pairs. In order to aid explanation of the acquired data, molecular modeling studies were undertaken using AMBER and the University of Birmingham Bluebear cluster to produce solvated, molecular dynamics optimized structures. The formation of excimers in bis-anthracene systems as potential precursors to photo-ligated DNA duplexes was also studied.

Using supramolecular principles, we have been designing luminescent lanthanide complexes with a defined hairpin bis-interlacator in order to obtain luminescent probes able to recognise DNA. The complexes are comprised of Platinum(II) terpyridine, which acts as a DNA recognition site and is brought together with a "remote" luminescent lanthanide unit. All the synthetic approaches were based on the accessibility of the lanthanide-platinum complexes by the self-assembly of different components in a one pot reaction. Thus, we have been able to isolate a water soluble heterometallic complex based on thiophenal linkage named [LnPt\(_2\)]Cl\(_2\). The complex has a relatively weak lanthanide luminescence, which increases upon addition of DNA. Photophysical and DNA binding properties of the lanthanide-platinum complex were investigated by UV-vis absorption, luminescent studies and circular and linear dichroism. Oligonucleotides of twelve bases were also used to investigate the intercalation [LnPt\(_2\)]Cl\(_2\) and the mono-intercalator AATP used as control compound. Using bidimensional NMR techniques, we investigated the binding site for [LnPt\(_2\)]Cl\(_2\) and AATP upon interaction with Dickerson-Drew sequence. The sulphur lanthanide-platinum linkage in [LnPt\(_2\)]Cl\(_2\) was replaced with an acetylide one in order to introduce new photophysical features. Thus the self-assembly procedures based on DTPA-bis(amido-acetylide) and a platinum(II) terpyridine led us to isolate a new lanthanide-platinum complex named [LnC\(\equiv\)CPt\(_2\)] (CH\(_3\)SO\(_3\))\(_2\). The photophysical properties and the DNA binding properties toward interaction with CT-DNA were investigated. The complex named LnC\(\equiv\)CPt\(_2\)](CH\(_3\)S)\(_3\))\(_2\) exhibited a relatively strong lanthanide luminescence that increased upon addition of DNA. The bi-functional metal complex [EuLPt](PF\(_6\)) (where Pt=platinum-2,2':6'2"-terpyridine and L=assymmetric DTPA bisamide ligand with a thiopheno pendant arm and a quinoline moiety) was synthesised and the interaction of [EuLPt](PF\(_6\)) with CT-DNA was examined by luminescence spectroscopy, linear and circular dichroism studies and thermal denaturation studies. The [EuLPt](PF\(_6\)) retained the ability to increase its luminescence upon the addition of CT-DNA. The binding properties of the complexes were tested toward interaction with plasmid DNA by gel electrophoresis and properties such as the unwinding angle were measured. The bis-intercalators [LnPt\(_2\)]Cl\(_2\) and [LnC\(\equiv\)CPt\(_2\)](CH\(_3\)SO\(_3\))\(_2\) showed the ability to uncoil DNA almost as well as cisplatin and at low concentrations, while almost double the amount of mono-intercalators, such as [EuLPt](PF\(_6\)) is required to observe the same uncoiling effect.