Vaccines derived from non-pathogenic antigens often have greater safety profiles over live vaccines, whilst still eliciting a specific immune response. However, the decreased immunogenicity of these vaccines means that numerous doses have to be administered or an adjuvant has to be used. In this study gonadotrophin releasing hormone (GnRH), a fertility regulating self-peptide, was used as a model antigen applied to Stober silica nanoparticles (used as an adjuvant) in a vaccine formulation. It is proposed that an inorganic silica nanoparticle can act as a foreign carrier particle for adsorbed peptides, thus stimulating an immune response against these peptides in vivo. This work utilised molecular dynamics simulations of silica substrates and their interaction with peptides for optimal presentation of the peptide to the immune system for antibody production. The interactions at this step allowed the optimum design of the nanoparticle with surface peptides that were free to interact with the surrounding environment, thus having the potential to invoke a biological response. Quartz crystal microbalance and surface plasmon resonance measurements were also used in order to investigate the adsorption of the peptides onto a silica surface. Through running these experiments at various pH levels and ionic strengths the optimum conditions for peptide coverage of silica nanoparticles could be determined, enabling enhanced design of the silica nanoparticle-peptide system and thus providing invaluable data to inform immunisation studies. Following an immunological study in male BALB/c mice it was found that the use of silica as an adjuvant along with bovine serum albumin (BSA) as a carrier protein increased the immunogenicity of GnRH-I peptides in comparison to just BSA alone. It was observed that peptides adsorbed solely onto silica nanoparticles did not elicit a strong antibody response. However, this formulation caused a significant decline in testosterone production, suggesting that silica coated with native GnRH-I peptides could be useful in receptor blocking.

The effective combination of a bacterial Reverse Two-Hybrid System screening platform and a SICLOPPS randomised library has been previously used for the identification of cyclic peptide inhibitors of various protein-protein interactions. A more robust Reverse Two-Hybrid System was designed and constructed to utilise a fluorescent protein reporter as well as an antibiotic resistance gene and HIS3 to enable selection of an inhibitor. The functionality of the new system was tested with the HIF1[alpha]/HIF1[Beta] and p6/UEV heterodimeric interactions and the cyclic peptide inhibitors that had been previously identified for these interactions. Next, a split-intein with improved properties was used to construct a second generation SICLOPPS library, which was tested to show that it displayed more efficient splicing. The increased toxicity of the Nostoc punctiforme DnaE splitintein employed in the new library was reduced by the addition of the SsrA protein degradation tag to enable the identification of more varied cyclic peptide inhibitors in future SICLOPPS screens.

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The detection of prohibited performance-enhancing drugs in sports is often carried out using urine samples. The reason for this is that urine samples can be collected under non-sterile conditions and do not require the presence of a sanctioned medical officer. Improper storage of the urine samples from athletes can lead to microbial contamination, which can cause changes in the steroids profile, leading to false positive or false negative results for a particular athlete. To address this problem, a new analytical method was proposed that employs a fluorinated steroid as an internal standard and fluorine-19 nuclear magnetic resonance spectroscopy (19F-NMR) spectroscopy to identify both microbial and thermally-induced changes in the urine samples. In Chapter 2, synthesis of fluorinated steroids was carried out using method that involve the reaction of Selectfluor® with enolates/enols of steroids. A range of fluorinated steroids was prepared (2 novel F-steroids) in moderate yields and varying diastereoselectivities. Several synthesised steroids were recrystallized and crystals suitable for X-ray were obtained. In Chapter 3, selected fluorinated steroids were incubated with microorganisms such as Escherichia coli, Bacillus subtilis, Bacillus megaterium and Streptomyces griseus. Fluorinated steroids were transformed to various oxidised metabolites upon incubation with Streptomyces griseus. Escherichia coli did not produce any metabolites due to lack of cytochrome P450 enzymes. Incubation of Bacillus subtilis and Bacillus megaterium was not successful and therefore metabolites were not detected. In Chapter 4, hydroxy steroids were reacted with pentafluoropyridine (PFP) to form perfluoropyridine ethers in good yield. Several novel steroids were synthesised and the structures of 4 perfluoropyridine ethers were confirmed for the first time by X-ray structure. It was found that the hydroxy steroid PFP adducts have very similar 19F NMR spectra however they can be distinguished using this technique. This novel derivatisation technique could be potentially used for identification of hydroxy steroids in biological material by 19F NMR.

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Natural products have been the source of many of our successful drugs providing us with an unrivalled chemical diversity combined with drug-like properties. The search for bioactive compounds can be helped considerably by the phytotherapeutic knowledge held by indigenous communities. In this study solvent extracts from the bark of a Dalbergia species and used by a community in Borneo, will be investigated to isolate, identify and biochemically characterise compounds showing vasorelaxation. At the core of this study is the hypertensive model, which uses phenylephrine precontracted rat aortic rings as a bioassay to identify solvent extracts, fractions and sub-fractions that cause relaxation. These fractions are generated using chromatographic techniques and solvent systems developed specifically for this purpose. Structural elucidation of the isolated compounds was undertaken by studying extensive data from UV, MS, 1D and 2D 1H and 13C NMR spectra. This study also undertook the pharmacological characterisation of the isolated compounds using the same bioassay together with enzyme and receptor inhibitors to identify the signalling pathways involved.

Saudi Arabia has amongst the highest incidence of type 2 diabetes in the world, with nearly 20 percent of the adult population suffering from the condition. The prevalence of vitamin D deficiency in type 2 diabetics has been shown to be twice that of non-diabetic individuals. Recent studies have shown, over 95 percent of the Saudi adolescent population are vitamin D deficient. Pancreatic beta-cells express vitamin D receptors, as well as the vitamin-D3–activating enzyme 1-alpha-hydroxylase. Vitamin D plays a key role in both insulin production and glucose homeostasis. Supplementation with vitamin D could help tackle morbidity in diabetes by decreasing insulin resistance and reducing levels of advanced glycation endproducts (AGEs) which contribute to the onset and progression of diabetic complications. Despite vitamin D deficiency and diabetes being highly prevalent amongst the female Saudi population, the effect of vitamin D3 supplementation on improving diabetic outcomes is an area that is currently understudied. This was a double-blind randomized control study of 156 overweight Saudi females with poorly controlled type 2 diabetes mellitus recruited from Al-Noor Hospital in Saudi Arabia. Each subject was randomly allocated to either a placebo, 2000 IU/day or 4000 IU/day vitamin D3 intervention group for 16 weeks. Serum measurements were analysed using routine laboratory procedures. Oxidative stress biomarkers, including total antioxidant levels, were measured using a colorimetric method. AGEs were measured using an AGE reader at baseline and 16 weeks. Significant improvements in vitamin D status, HbA1c, LDL and total cholesterol were demonstrated after 16 weeks of supplementation (p < 0.001), (p = 0.001), (p = 0.028) and (p = 0.049) respectively. There were no statistically significant changes in HOMA-IR, AGE concentrations, fasting insulin, fasting glucose, HDL-cholesterol and triglyceride levels. This study suggests vitamin D may have a role in improving outcomes for type 2 diabetics and slowing the natural progression of the disease. Further research however is needed to determine the optimum dose, duration and form of delivery of supplementation in order to achieve these effects.