http://theses.ncl.ac.uk/jspui/handle/10443/78 2026-02-04T09:40:27Z http://theses.ncl.ac.uk/jspui/handle/10443/6323 Title: Towards the Determination of the Active Set of Elementary Flux Modes using Metabolic Flux Data Authors: Murphy, Koren Abstract: Vaccine development at lab scale through to large scale produc6on can take 10-15 years. With the outbreak of the SARS-CoV-2 disease, emphasis on fast vaccine produc6on was emphasised. However, the cells that are grown to produce an6gens have complex metabolic networks consis6ng of thousands of reac6ons, metabolites, and genes. There is liJle understanding of why a cell in the same environmental condi6ons may grow via one route over another. If this process was beJer analysed, process op6misa6on to increase biomass growth and reduce inhibi6ng metabolites could be performed. All routes that a cell can use during its life are collec6vely known as elementary flux modes. Genome networks are being constructed over 6me allowing for full reac6on stoichiometry to be known. However, genome networks do not have all the elementary flux modes iden6fied due to the combinatorial explosion that occurs when solving as there can be billions of possible routes. In this thesis, mixed integer linear programming has been presented to enumerate elementary flux modes as a future proof method towards genome scale solving. It is compared to publicly available tools and mixed integer linear programming methods throughout literature. The benefits of this method in the future for finding elementary flux modes are also discussed. Compression techniques and code parallelisa6on are examined and reduced solve 6mes presented. Alongside elementary flux mode enumera6on this thesis also applies flux analysis techniques as a method of finding biologically relevant elementary flux modes. Disadvantages of these techniques are highlighted whilst presen6ng an integrated form of metabolic flux analysis to alleviate some of the issues. The technique presented is proven to be a viable method for enumera6ng elementary flux modes with the integra6on of fluxes. E.coli can be modelled as the full genome network or a reduced set of reac6ons represen6ng the key areas of the network; this is known as the core network. E. coli fermenta6on data from GlaxoSmithKline was provided for this work, allowing for analysis techniques iden6fied and created in this work to be applied. However, this data was found to be underdetermined preven6ng aspects to flux analysis and elementary flux mode enumera6on to be performed. This thesis discusses the process data and es6mates specific growth and uptake rates for all III fermenta6ons in batch and fed batch opera6ons. This key data was missing and helpsin beJer understanding the opera6ons taking place in the fermenters. More importantly however areas where more data is required for flux analysis are presented along with the issues of data limita6on on finding the elementary flux modes even for the core network. Underdetermined flux analysis allowed for es6ma6ons on the number of possible elementary flux modes in batch and fed batch opera6ons, highligh6ng the reduc6on in feasible routes during fed batch due to the cell’s phase. Description: Ph. D. Thesis. 2024-01-01T00:00:00Z http://theses.ncl.ac.uk/jspui/handle/10443/6193 Title: Interactions between microplastics and the liquid/gaseous interface for environmental remediation Authors: Saczek, Joshua Abstract: In 2021, 2.5 million metric tons of plastic packaging waste was generated in the UK. The plasticene age has allowed for the development of numerous highly versatile materials useful throughout all aspects of society, but the consequence of our success has resulted in extensive plastic pollution. One end life cycle-point of plastic waste is into water, with almost 10 million tons entering oceans every year. These, once durable materials can now break down into smaller fragments, eventually resulting in an abundance of microplastics and even much smaller nanoplastics, with an estimated 5 trillion plastic particles within the world’s surface waters. Due to their prevalence in all aspects of our environment and recently found presence within human circulatory systems, these plastic pieces have become a focus of worldwide attention that require urgent remediation. As such their interactions with liquids, both bulk and droplets, and subsequent modes of removal are paramount to understanding how to bring this problem under control. The work within this thesis explored how microplastics interacted with water in two distinct forms, droplets and in bulk, and assessed how particle properties relate to various parameters. This work also considered how both scenarios could be effectively used to remove microplastics from water, be it in the form of liquid marbles for droplets or a bubble and oil system for removal from bulk water. Liquid marbles allow for various liquids to be encapsulated by hydrophobic particles, thus ensuring isolation from the external environment. They occur when a droplet, often water, is rolled through a powder bed causing the particles to decorate the exterior of the droplet. This unique structure allows for combinations of solid-liquid-gas reactions to occur, providing that stability is maintained throughout. This stability is regularly described in terms of lifetime and is one of their main drawbacks, often impacted by particle characteristics. The impact the coating material has on lifetime is mainly linked to thickness of coating layers and contact angle. Additionally other factors, such as internal and external environments play a major role in overall lifetime, making it possible to engineer a robust long-lived liquid marble. It is therefore useful to know factors which have a positive impact on lifetime and by using a reactor engineering approach it is possible to model and predict the overall lifetime of various coatings on liquid droplets. As such, this phenomenon has been applied to emerging applications, such as unconventional computing, cell mimicry and soft lithography, as well as within green and environmental applications, such as energy conversion, heavy metal recovery, CO2 sequestering, oil removal and microplastic capture. II Microplastics in the environment are more commonly found dispersed within bulk liquids rather than decorating a droplet, with various concentrations found within salt, fresh and potable water. Methods commonly employed to remove microplastics from water, such as membrane filtration, can achieve high rates of removal, however, polymer membranes are susceptible to mechanical and chemical failure due to backwashing and ageing. Another simple yet effective removal method of microplastics from water using oil was investigated and showed early promise, however, this technique is currently incorporated into systems that are unsuitable for scale up into a continuous removal method. Bubbles travel up through the water column attached to polymer particles, transporting them to the surface where they migrated into and subsequently trapped within an ‘environmentally friendly’ oil layer due to their hydrophobic and oleophilic nature, without a concern of secondary pollution. This waste to capture waste method was demonstrated to achieve high removal efficiencies of > 99.4 % of 6 of the most prevalent microplastic types (low-density polyethylene, polypropylene, polystyrene, nylon, poly(ethylene terephthalate) and polyvinyl chloride) and sizes. This bubble-oil method was adapted into a continuous system, where a reactor prototype was built and flow rates, liquid volume, liquid type, particle type and concentration variables were examined to optimise the removal process for vertical and horizontal systems. Description: PhD Thesis 2023-01-01T00:00:00Z http://theses.ncl.ac.uk/jspui/handle/10443/5959 Title: The optimization of the synthesis of Ni/Sb-SnO2 nanopowders for the electrochemical generation of ozone Authors: Eje, Ndubuisi Kennedy Abstract: Nickel and antimony co-doped tin oxide (Ni/Sb – SnO2) anodes have been shown to exhibit high selectivity for ozone generation at current efficiencies of ca. 50% and current densities of ca. 100 mA cm-2 . However, the synthesis involves dip coating Ti substrates in solutions containing the precursors, calcining at temperatures between 400 and 900 o C, and repeating these steps a number of times to produce ceramic coatings with geometric surface. The synthesis proved to be somewhat irreproducible, and the current densities that could be attained were low. The latter precluded the use of these anodes to treat “real” waters in the absence of added electrolyte: such a process would require a zero gap, polymer electrolyte membrane (Nafion) cell. Unfortunately, at 100 mA cm-2 there is a problem of poisoning polymer electrolyte membrane by cations such as Na+, Mg2+, and Ca2+ present in such waters, thereby rendering the ceramic anodes incapable of treating real waters. The postulate that forms the basis of the work reported in this report was that if high surface area, active and selective nanopowders of Ni/Sb – SnO2 could be synthesised then the current densities could be sufficient to prevent fouling of the polymer electrolyte membrane and allow the application of zero gap cells in water treatment. Preliminary work in Newcastle had suggested that hydrothermal method could be employed to produce active materials, in which the precursors salts were dissolved in deionized water and refluxed before the hydrothermal process and calcination. The second method explored in this thesis was the same as the first except that Sb-SnO2 (ATO) was synthesized, calcined, and then doped with Ni by mixing the powder with the appropriate weight percent of nickel chloride solution before calcination. It was hoped that this latter method would be more reproducible, as well as producing active and selective Ni/Sb – SnO2 nanopowders. The nanopowder samples prepared from the two routes were characterized using X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), Energy dispersive X-ray (EDX), and Brunauer Emmett-Teller to evaluate how much the changes in compositions and synthesis method affected some properties of the nanopowder samples. The XRD results revealed that increasing the Sb content of the nanopowder samples inhibited crystal growth and reduced the crystallite size, whereas increasing the amount of Ni at fixed iii Sb content and calcination temperature had no effect on the crystallite size. It also showed that the method of doping Ni had little or no effect on crystallite size. The BET analysis revealed that surface areas of the nanopowders from both routes increased as the Sb concentration increased. Cyclic voltammetry was employed to ascertain the onset potentials and potential ranges for redox processes. In general, the cyclic voltammograms of the ATO and the Ni/Sb – SnO2 electrodes in acidic electrolytes showed redox peaks. The responses of the ATO and the Ni/Sb – SnO2 anodes were similar; hence the peaks were attributed to the redox process associated with the Sb and Sn at the surface. The presence of the peaks in the cyclic voltammograms of these electrodes is most likely because the electrodes were prepared with high surface area powders which rendered the features visible in the voltammetry, unlike that of the ceramic electrodes geometric areas reported in the literature. Ozone activities and selectivity of the Ni/Sb – SnO2 nanopowders prepared by both routes were investigated in 0.5 M H2SO4 using a UV-Vis cuvette as the electrochemical cell. Ni/Sb – SnO2 anode was held flat on the bottom of the cuvette containing the electrolyte, and a 0.64 cm2 Pt/Ti mesh cathode inserted was vertically. The electrodes were placed against the opaque sides of the cuvette to avoid interfering with the light passing through the UV Vis spectrometer. On comparing the data obtained, it was found that the first doping route where the precursors salts were dissolved together before the hydrothermal process is more active in ozone generation. In general, it was observed that ozone activity decreased at higher Ni concentrations for nanopowders of both routes. XPS analysis of the nanopowders showed the presence of Sn at the oxidation state of +4 and Sb at the oxidation state of +3 and +5. Ni was not detected, probably because of the detection limit of the XPS or the location of the Ni in the lattice. On comparing the Sb/Sn atom ratio of the Ni/Sb – SnO2 nanopowders from the two routes, it was found that the second doping route had higher surface enrichment of Sb3+ than the Ni/Sb doping route, and this could be the reason for the observed high ozone current efficiency with the NATO anodes. The effects of ozone on the skin using the Phenion Skin Model were investigated. Three skin models were exposed to 0.3 ppm ozone in the exposure chamber for a week at 8hr each day, and three were kept as control. Haematoxylin and eosin (H&E) staining was employed to iv determine any changes to the thickness of the skin layers due to ozone exposure. In brief, it was found that ozone exposure increased the thickness of the epidermis and decreased the thickness of the dermis. The effects of ozone on the protein content of the skin models were also analysed using the Bradford assay, and it was observed that ozone caused the release of protein from the skin cells models. Description: PhD Thesis 2022-01-01T00:00:00Z http://theses.ncl.ac.uk/jspui/handle/10443/5636 Title: Mechanistic modelling of microscale chromatography Authors: Whitelock, Nick Abstract: Microscale chromatography as an experimental tool has shown much utility in process development due to reduced material consumption and ease of parallelisation which are of major benefit when compared to conventional lab-scale studies. Microscale columns are commonly used in early process development where the most impactful decisions, such as choice of unit operation, purification strategy, resin, and the choice of candidate are made with limited resources and knowledge. Understanding the behaviour of microscale chromatography and better applying the knowledge gained from microscale studies to large scale chromatography may allow faster, more efficient and more robust early process development, and therefore more effective processes once a bioprocess is fully developed and products commercialised. It is the overall aim of the project to develop a model to determine large scale mass transfer parameters describing a lab-scale chromatographic process from microscale data, and allow one to simulate and optimise large scale separations whilst enjoying the benefits of reduced resource consumption of the microscale domain. From the outset, characterisation ofthe differences between lab-scale columns operated on a conventional Fast Protein Liquid Chromatography (FPLC) system and microscale columns on a robotic Liquid Handling System (LHS) was performed. Determining the common metrics of column performance, HETP, asymmetry and experimentto-experiment or column-to-column variation between columns and experiments provides an understanding of some of the key differences between lab-scale and microscale column formats with regards to system, scale and data quality, as well as providing an opportunity to optimise the experimental design of microscale experiments. This was performed through evaluating methods of improving resolution, including fashioning rigs to use microscale columns on a conventional system, evaluating various tracer substances and evaluating a novel strategy of pre-filling collection plates. Investigations into ascertaining the dynamic binding capacity (DBC) of IgG to Protein A resin using microscale data has been performed with 3 microscale column volumes at several residence times using the high throughput system, and repeated at lab scale, with further work into understanding the effect of intermittent flow on resin: target interaction by mimicking the microscale operation on a larger system. This effort has led towards data used to calibrate a mechanistic model of chromatography at both lab scale and microscale with the intention of predicting lab scale behaviour. By correcting for scale, operational and flow effects, one may predict large scale performance through calibrating a model with microscale data, enabling better process understanding with reduced material consumption. Description: Ph. D. Thesis. 2022-01-01T00:00:00Z