Grain oriented steels containing 3 % silicon are widely used as stacks of thin laminations in transformers and other electrical devices. Phosphate coatings are applied to these laminations in order in insulate between the sheets and hold them under tension, reducing the electrical energy losses and making the transformers more efficient. This thesis explores the properties of aluminium and magnesium phosphate, two commonly used coating materials. Using model phosphate coatings it has been shown that an excess of phosphoric acid is required to cause the condensation reaction that produces the metaphosphate, thought to be the cause of the increased tension imparted on the steel. The addition of chromium oxide was found to prevent this transformation by reacting with the excess acid to form chromium pyrophosphate, which lead to a more stable coating which imparted a greater tension upon the steel substrate. XPS has been shown to be a useful technique for the characterisation of model phosphate coatings, however it cannot be used to quantitatively analyse the systems (unlike other phosphate systems) possibly due to the higher number of phases present within the samples.

The design of a photoswitchable homeodomain artificial transcription factor (PATF), modelled on an engrailed homeodomain, for the purpose of controlling DNA binding affinity and controlling the transcription process in cells using light has been investigated. This study was conducted using a 3,3’-bis(sulfo)- 4,4’bis(chloroacetamino)azobenzene crosslinker, alkylated between two cysteine residues with different spacings (i, i+4, i, i+7 and i, i+11) and either a rigid or flexible linker domain. In previous studies, basic leucine zipper transcription activators have been photocontrolled in living cells by incorporating a photoswitchable azobenzene crosslinker. Circular dichroism spectroscopy showed the conformation of crosslinked PATF (XLPATF) peptides (i, i+11 spacing) containing rigid and flexible linkers could be controlled reversibly by light. Fluorescence anisotropy experiments using labelled DNA confirmed the in vitro DNA binding affinity of PATF was considerably higher with the crosslinker in the trans (ground state) configuration than in the cis (photoexcited state) configuration. Further studies of peptides with i, i+4 and i, i+7 spacings with a semirigid and rigid linker domains showed increased binding affinity with the crosslinker in the cis configuration. Initiation of transcription was investigated by an in vitro transcription assay to measure the ability of PATF molecules to moderate the production of RNA by irradiation with UV light. PATF molecules with i, i+11 spacings showed increased transcriptional activation with the crosslinker in the ground state configuration and i, i+4 and i, i+7 spacings resulted in increased transcription activation with the crosslinker in the excited state conformation. Control of 50% of transcriptional activity was achieved for i, i+11 v spacings, and PATFs with a rigid linker domain were more effective switches than those with flexible linkers. Using i, i+4 and i, i+7 spacings in PATFs resulted in a lower degree of control but, as anticipated, transcriptional activation was increased after irradiation.

The main focus of the work contained within this thesis was to explore the possible use of novel organoboron chemistry in generating quaternary carbon centres, with the ultimate aim of developing new routes for the asymmetric synthesis of chiral quaternary carbon centres. Chapter 1 contains a general introduction to the literature of organoboron chemistry, focusing particularly on organoboron reactions that contain 1, 2 - boron to carbon migrations. Chapters 2 and 3 contain attempts at generating tertiary alkyl groups bonded to boron in an asymmetric fashion. Chapter 2 specifically deals with attempts at designing a chiral version of the dichloromethyl methyl ether (DCME) reaction, giving disappointing results most probably due to the instability of the chiral analogue of DCME. Chapter 3 focuses on attempts at designing a chiral version of the cyanidation reaction. Although the generation of tertiary alkyl groups next to boron was shown to be possible when using imidoyl chlorides as the acylating reagent for relatively unhindered trialkylboranes, alkene side-products predominated when using hindered trialkylboranes – which meant that the reaction was unlikely to become an efficient method for the asymmetric generation of tertiary alkyl groups bonded to boron. Chapter 4 contains a detailed investigation into the migration of tertiary alkyl groups in the reaction between tertiary-alkyl boronic esters and (bromomethyl)lithium. As well as gaining a better understanding of the factors at work in this potentially highly important reaction, the reaction was applied to a number of highly hindered boronic esters. Chapter 5 deals with attempts at tertiary alkyl group migration using 3-chloro-1-lithiopropyne, which were in general disappointing due to the apparent instability of the intermediate ‘ate’ complex. Chapter 6 contains novel work on the migration of alkyl groups using the previously unreported 1-lithio-3-chloropropene organolithium reagent. The reaction was successfully applied to the migration of several alkyl groups (including tertiary), giving the corresponding 3-alkylprop-1-en-3-ols in good to excellent yields.

Dihydrofolate reductase (DHFR) is the enzyme that catalyses the reduction of 7,8-dihydrofolate (DHF) to 5,6,7,8-tetrahydrofolate (THF) in the presence of the cofactor reduced nicotinamide adenine dinucleotide phosphate (NADPH). The DHFR catalysed reaction has often been used to study enzymatic tunnelling and the contribution of protein dynamics to catalysis. To gain a better understanding of such phenomena and to investigate the key elements of structural adaptation in DHFR, in this thesis the hydride transfer reaction of DHFR from Moritella profunda (MpDHFR), a cold adapted enzyme, was studied and compared to the mesophilic and extensively studied enzyme from Escherichia coli (EcDHFR) and the thermophilic enzyme from Thermotoga maritima (TmDHFR). Chapter 1 gives a brief introduction to the thesis. Description of the materials and methods used in evaluating this work is reported in Chapter 2. In Chapter 3, the steady state and pre-steady state temperature dependences of the kinetic isotope effect (KIE) for the MpDHFR catalysed reaction was elevated, compared to data obtained for the mesophilic and the thermophilic DHFR homologues and the results interpreted according to the environmentally coupled tunnelling model. The work presented in Chapters 4 and 5 has investigated the role of dynamics during catalysis by DHFR using site directed mutagenesis. In Chapter 4, mutations were created in the GH loop for both EcDHFR and MpDHFR to elucidate the role of the occluded conformation during catalysis by DHFR. In Chapter 5, different MpDHFR and EcDHFR variants in the highly mobile M20 loop were generated and their temperature dependences of KIE were studied in addition to studying the two variants MpDHFR-G123V and MpDHFR-D124N in the catalytically important FG loop. The results obtained suggest that MpDHFR does not undergo the dynamical loop movements that have been recognized previously for EcDHFR in spite of following the same catalytic cycle. Further findings were found which contradict the current models that relate protein dynamics to catalysis efficiency, thus modifying these models has become essential. Chapter 6 has focused on studying the effect of different denaturants/salt concentrations on MpDHFR chemical step. Finally, a summary of the work presented in this thesis and future guidelines are provided in Chapter 7.

EDITED ABSTRACT. This thesis is divided into two parts. The first part describes the production of a small stereodiverse library of 2-substituted piperidines. Novel chiral titanium alkylidene reagents ii alkylidenated resin-bound esters i to give acid-labile resin-bound enol ethers iii. These were cleaved to give amino ketones iv. The switch in the nature of the resin from acid-stable to acid-labile is key to the purity of the amino ketones iv, as during cleavage only the acid-sensitive enol ethers iii are cleaved, leaving the unreacted esters i on the resin. The amino ketonse iv were cyclized using TMSC1 to give cyclic iminium salts v. Diastereoselective reduction of the iminium salts v with NaBH(OAc)[sub]3 gave piperidines vi which, after cleavage of the chiral protecting group gave the desired enantiomerically-enriched, 2-substituted piperidines vii. [Illustrated] The piperidines vii were produced in good overall yield, high purity, and good to excellent stereochemical purity. By switching the enantiomer of the phenylethylamine chiral protecting group used, either enantiomer of the desired piperidine could be produced at will. The second part of the thesis describes a solution-phase route to 2,6-syn substituted piperidin-4-ones xii inspired by the Petasis-Ferrier rearrangement. Imino esters x derived from [Beta]-amino acids viii were methylenated using the Petasis reagent, dimethyltitanoce, to give imino enol ethers xi containing nucleophilic and electrophilic functionality in the same molecule. The mild microwave conditions used for the methylenation geve the enol ethers xi in minutes. Potentially, the reaction takes advantage of selective heating of the polar Petasis reagent in a non polar solvent system so that the rate determining decomposition of the Petasis reagent is accelerated without affecting any sensitive substrate. Acidic conditions activated the imine and induced cyclization to give the desired 2,6-syn piperidin-4-ones xii in good yield and excellent diastereoselectivity. A small library of piperidinones was produced to demonstrate the method. [Illustrated]

The aim of this work was to synthesise phosphorous-containing polymers with controlled molecular weights and low PDIs. Phosphorus functionality was introduced both by polymerising a phosphorus-containing monomer and by the reaction of functional polymers with phosphorous compounds in a postpolymerisation approach. Cu(0)-mediated SET-LRP was studied to ascertain if it could be used for the polymerisation of ethylene glycol containing monomers. When using copper wire as the source of metal, significant periods of inactivity or ‘induction’ time were observed at the beginning of the polymerisation. By altering the polymerisation temperature and the surface area of the copper, their affect on the ‘induction’ time was studied. SET-LRP was used to polymerise DEGMEMA, PEGA454, PEGMA475 and PEGMA1100, serving as model compounds for subsequent polymerisations of phosphorus-containing monomers. ATRP and SET-LRP were used to synthesise homopolymers of DEMOEP and copolymers of DEMOEP with both DEGMEMA and PEGMA475. ATRP resulted in well defined polymers with low PDIs, but SET-LRP of DEMOEP and DEMOEP/DEGMEMA exhibited high termination and high PDIs. By replacing DEGMEMA with PEGMA475, polymer synthesis by SET-LRP was much improved; termination was low, molecular weight growth was linear with respect to conversion and the final polymers had narrow molecular weight distributions. The control of these polymers by SET-LRP equalled those of ATRP. Deprotection chemistry was used to attempt the removal of the ethyl groups of the pendant phosphate groups in DEMOEP-containing polymers. Three different systems were used and their affect on the polymers was studied. Using a post-polymerisation modification strategy, reactive succinimidyl-ester containing polymers were modified with the amino-functional phosphate, AMPA. In a similar fashion, the reaction of P-H bonds residues with amines and aldehydes was also studied for the modification of reactive polymers. These ‘Kabachnik-Fields’ reactions were performed on small molecule model amines, commercially available amino-functional polymers and finally, a water-soluble succinimidyl-ester containing copolymer synthesised by ATRP.

Microstereolithography is a rapidly developing technique for the generation of 3D structures by the building up of 2D layers and has shown great potential for the development of scaffolds for clinical applications. Significant research has been carried out on the development of functionalised biocompatible polymers specifically for this technique. With this technique there is potential for the fabrication of a patient/injury specific scaffold, by scanning the injured site, developing a 3D model and converting it into 2D layers for fabrication. The construction of 3D scaffolds requires control of the curing depth of the resin to prevent loss of 3D control by through polymerisation into the monomer. This control is present in commercial resins, however, a method using biocompatible materials has not been reported upon. This work demonstrates how natural products, specifically paprika extracts, can be used to control the penetration of UV light into resin, and so control the thickness of layers generated. The behaviour of these resin compositions upon exposure to light has been determined, and equations governing curing depth have been determined. The microporosity of cell growth scaffolds is an important factor in the reactivity of the material, and this is a scale which cannot be controlled in some μSL apparatus due to the lack of resolution. Microporosity has been obtained by the use of monomer soluble, nonfunctionalised polymers. This has allowed the development of scaffolds with increased surface area and so potentially higher reactivity than those without porogens. Material reactivity is also important for scaffolds, specifically in hard tissue replacement, where the differentiation of mesenchymal stem cells to osteoblast cells is guided by calcium phosphates. The development of photopolymerisable resins containing hydroxyapatite in suitably stable suspensions is reported upon. These materials can be easily polymerised, and is has been shown that adult stem cells adhere to, and proliferate, on them.

Single walled carbon nanotubes (SWNTs) have unique structural and electronic properties which drive their use in many different fields of modern technology. In electrochemistry, SWNTs have been shown to have outstanding electrochemical characteristic enabling their application in trace level electroanalysis amongst other areas. The SWNT electrochemical activity has been described as originating solely from sidewall structural defects, oxidised open ends and post-growth metal nanoparticles (NPs). However, recent studies have demonstrated the electrochemical activity of pristine and defect-free SWNTs, grown on insulating substrates using catalysed chemical vapour deposition (cCVD) method, suggesting sidewalls support electron transfer (ET). In this thesis cCVD is employed to grow SWNTs of different geometrical arrangements on single crystal stable temperature (ST)-cut quartz substrates for the development of novel nanoelectrodes (NEs) and optically transparent electrodes (OTE). Majority of the work concentrates on cCVD and electrochemical studies of horizontally aligned SWNTs on ST-cut quartz. The quality of SWNT horizontal arrangement is shown to strongly depend on the crystallographic structure of the ST-cut quartz substrate. Perfectly aligned SWNTs are utilised as (i) one dimensional (1D) templates for electrodeposition of silver NPs, and (ii) NEs for the fundamental assessment of SWNT electrochemical behaviour. Formation of uniform silver nanowires (NWs) on most of the SWNTs is possible at high deposition overpotentials and times. Uniformity of the NWs indicates high structural quality of the horizontally aligned SWNTs. Nernstian behaviour of SWNT NEs for simple outer-sphere redox couples is shown in localised voltammetric measurements with microcapillaries, 25 - 50 μm in diameter serving as probes, filled with the redox active electrolyte solution and the reference electrode (RE). For the first time measurements at single SWNTs in the positive and negative potential window are performed, revealing a strong dependence of SWNT electrochemical activity on SWNT structural and electronic properties. Finally, careful tuning of the cCVD parameters enables growth of interconnected SWNT networks on ST-cut quartz, with complete surface coverage and no sign of alignment. These ultrathin mats are utilised as novel OT disk ultramicroelectrodes (OT-UMEs) with complete transparency in the ultraviolet and visible (UV-Vis) range, metallic behaviour and improved electrochemical usability in comparison to conventional solid UMEs.

The advent of electrospray ionisation (ESI) has been a major factor driving the revolution which has taken place in the field of biological mass spectrometry over the past twenty years. In this thesis, efforts to improve electrospray ionisation in two respects are detailed. One is the transmission of ions from ambient pressure into the high vacuum of a mass spectrometer. The other is control and influence over making and breaking of bonds, in particular long and weak bonds, during this journey into the mass spectrometer. These improvements to electrospray ionisation have been sought through the development, theoretical analysis and application of the ion conveyor, which is an electrodynamic ion-transport device. The ion conveyor as reported here is a device consisting of stacked ring-electrodes intended for operation within the first and second differential-pumping regions of a mass spectrometer with an electrospray ionisation source. It is demonstrated how the ion conveyor achieves aspects of ion trapping and ion transport through the application of one or more radiofrequency waves to the stacked ring-electrodes. The operating principles of the device are detailed, and related technologies are described. Theoretical simulations have been used to explore for explanations for the behaviour of the device and to make predictions as to the optimum performance of the device. The first experiments with an ion conveyor and subsequent experiments with two evolved designs are described. The operation of the device has been investigated with three different types of mass spectrometer, each with its own design of electrospray ionisation source. These designs are described in detail, and the responses of the ion conveyor to various operating parameters have been characterised. A major finding was that the transmission of the ion conveyor remains significantly high in the absence of all applied potentials. This discovery reveals and emphasises the importance of mechanical forces to ion transport through the ion conveyor. A wide variety of analytes have been used in experiments to characterise the device. The mass spectra of vancomycin hydrochloride presented are of particular note, because they exhibit unusually intense peaks corresponding to the doubly charged dimer-species. The dimer of vancomycin plays the central role in long-standing theories of anti-bacterial action, but has not been observed in previous studies of vancomycin by mass spectrometry. The experimental results support the view that the ion conveyor is a useful technique for the effective transport of ions through the differential-pumping stages of an electrospray ionisation source. The results suggests that the device could be developed through appropriate manipulation of ion-neutral collisions in the higher-pressure regions to preserve and transmit delicate non-covalently bound species and facilitate their accurate measurement by means of mass spectrometry.

No description available.