The primary aim of this work was to provide an alternative method of synthesis of the non-steroidal anti-inflammatory drug copper indomethacin (Cu-Indo) and to produce alternative forms of the drug to increase its marketability. Dense gases as anti-solvents were used to achieve these aims. The study involved the synthesis, purification, micronisation and co-precipitation of Cu-Indo with polyvinylpyrrolidone (PVP) using dense carbon dioxide as an anti-solvent. Initially the volumetric and solubility behaviours of the solvent???anti-solvent systems were investigated to determine the optimum processing conditions. The solubility of Cu-Indo in an expanded solution was found to be a complex function of the solvent and other solutes. Copper indomethacin was successfully synthesised and purified in a single vessel using dense carbon dioxide as an anti-solvent. Drug yields of 98 % and purities near 100 % were achieved at optimum conditions with the advantages of less residual solvent in the drug, less solvent waste, reduced processing time and increased yields over the conventional synthesis process. Copper indomethacin was produced in a variety of morphologies and particle sizes using dense carbon dioxide as an anti-solvent. An investigation of the effect of process parameters on the particle characteristics showed that solute concentration was the dominant variable. Spherical particles with diameters less than 8 mm were obtained at optimum conditions. The immediate benefit of micronising Cu-Indo was demonstrated with an eight fold increase in dissolution rate when compared to the conventionally produced drug. Polyvinylpyrrolidone was successfully co-precipitated with Cu-Indo using dense carbon dioxide as an anti-solvent. The PVP???Cu-Indo co-precipitates were found to increase the solubility of the drug in ethanol with a 36 fold solubility enhancement at optimum conditions. The use of dense carbon dioxide as anti-solvent in this work demonstrates the potential of the GAS and ASES processes in the pharmaceutical industry. Copper indomethacin was synthesised, purified and micronised in a single vessel at a substantial saving in terms of time and solvent usage. The micronisation of Cu-Indo and the formation of the PVP???Cu-Indo co-precipitate provided alternative forms of the drug substantially increasing its marketability.

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

published_or_final_version / Dentistry / Doctoral / Doctor of Philosophy

Atherosclerosis is the major cause of cardiovascular diseases, which constitute the top ten leading causes of death worldwide. Atherosclerotic plaque development initiates from the inflamed endothelium under an atherogenic environment – chronic low grade inflammation, hypercholesterolemia, endotoxemia, etc. The principal cause of such inflammation has yet to be defined – with growing evidence that microbial stimulants like lipopolysaccharide (LPS) and peptidoglycan (PGN), which can activate toll-like receptors (TLRs) and nuclear factor-kappaB (NFκB) signaling might be the plausible origins. The gastrointestinal tract is suggested to be the major site for absorption and translocation of such stimulants, where gut microbiota have been associated with systemic inflammation and is essential in generating atherogenic substances. Since probiotics have the potential to induce systemic anti-inflammatory effects and fortify gut barrier to reduce bacterial translocation, evaluation of whether probiotics can help reduce atherogenesis was done by feeding the disease model, ApoE-/- mice with high fat diet alone, with telmisartan (1 or 5 mg/kg/day, positive controls) or with probiotics VSL#3 (2.8 x 1011 CFU/day) and/or Lactobacillus rhamnosus strain GG (LGG) (1x108 CFU/day), or the combination of which for 12 weeks. All treatments reduced lesion size significantly; with some treatments reduced plasma endotoxin, cholesterol and various proinflammatory biomarkers. The gut microbiota assessed with PCR-amplified 16S rRNA gene sequences using 454 pyrosequencing and thereafter correlation studies revealed that at least 20 bacterial families that were significantly altered by high fat diet in apolipoprotein E knockout (ApoE-/-)mice correlated with atherosclerotic plaque size and related biomarkers including cholesterol, adipocyte fatty acidbinding protein (A-FABP), etc. Probiotics showed potential in improving atherogenic environment by immunomodulation and induction or inhibition of growth of bacteria correlated with the atherosclerotic plaque and biomarkers. The atherosclerotic condition was also improved by telmisartan, which correlated with the altered gut microbiota. The newly identified atherosclerosis-related gut bacteria will require further exploration into their properties and mechanisms, which will eventually lead to the potential of developing probiotics for the treatment or prevention of atherosclerosis, and thus may be used as an affordable and non-invasive alternative that brings health benefits worldwide. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

In dentistry, repair and restoration of tooth structure to regain its mechanical properties is the ultimate aim of caries management. Currently, preventive therapies are preferred to operative interference. New preventive approaches are necessary to treat dental caries. Dentine is composed of an organic matrix, consisting of type 1. Degradation of collagen matrix is the main reason for breakdown of resin-dentine bonds and development of secondary caries over time. Hence, preservation of collagen fibrils is essential for preventive and reparative procedures in minimum intervention dentistry. Proanthocyanidin (PA) is a natural collagen cross-linker, which can be obtained from grape seeds, pine bark or elm tree. It has a high affinity to proline-rich proteins, like collagen. Cross-linking of the collagen fibrils increases their mechanical properties and their resistance to breakdown by proteolytic enzymes. Thus, in the first part of the study, with the aim of discovering other possible natural dentine collagen cross-linkers, the effect of two smaller molecular size flavonoids, naringin and quercetin, on the mechanical properties of demineralized dentine was compared to PA. Demineralized dentine treated with PA showed the greatest increase in mechanical properties, followed by quercetin and naringin. The protease inhibitory effect of PA was evaluated by examining its actions on soluble and collagen-bound matrix metalloproteinases and cysteine cathepsins. Proanthocyanidin, even at low concentration of 1%, exhibited excellent inhibitory effects on soluble and matrix-bound proteases. Secondly, PA was incorporated in a dental adhesive to facilitate its application in clinical situations. The effect of PA incorporation on durability of resin-dentine bond was evaluated. Up to 2% of PA could be added to dental adhesive with no adverse effects on immediate resin-dentine bond strength. However, the bond strengths of PA-incorporated adhesives dropped significantly following ageing. This could be attributed to the free radical scavenging effect of PA, which might also have interfered with polymerization of dental adhesive. The mechanical properties of PA-incorporated adhesive were evaluated and it was shown that up to 1% PA could be incorporated into an adhesive resin. With the addition of higher concentration of PA, the mechanical properties of the adhesive resin were reduced with increased solubility. Proanthocyanidin release from the cured resin showed an initial burst for 48 hours and was stabilized after five days. Finally, the remineralization potential of PA on artificial root caries was also compared with quercetin and naringin. All three flavonoids showed remineralization potential; however, their effects were inferior to fluoride. Proanthocyanidin formed a precipitate band on the superficial layer of carious lesion, preventing further mineral deposition. Subsequently, PA was incorporated in a CPP-ACFP (casein phosphoproteins amorphous calcium fluorophosphates) containing-paste to increase the mineral uptake in subsurface layer of caries lesion. Simultaneous application of PA and CPP-ACFP was shown to have a favourable outcome on mineral deposition in root caries lesion. / published_or_final_version / Dentistry / Doctoral / Doctor of Philosophy

Titanium and its alloys have been widely used as materials in biomedicine and in particular, for dental implants, and one main reason is their unique ability to osseointegrate with the host bone. This phenomenon forms a strong bone-to-implant bonding. The rate and quality of osseointegration of a titanium dental implant depends heavily upon its surface properties. Over time, various surface treatment methods have been developed to further improve osseointegration of titanium-based biomaterials. This PhD thesis focuses on promising surface treatment methodology: surface charge modification. A negative charge has been known to promote the osseointegration of titanium biomaterials. Before the current work there was no practical approach to induce negative charge on titanium surfaces. The goal was to find such a practical technique. The current investigations revealed that sandblasting, a commonly used surface treatment method, can generate a negative charge on the surface of titanium, and also titanium alloy and stainless steel micro-implants, due to the triboelectric effect. Stainless steel doesn’t osseointegrate. The basic methodology of these studies was to sandblast titanium and measure the amount and polarity of the static charge on titanium surfaces. The effects of several important parameters were evaluated, including the sandblasting material, the size of the blasting grits, and the shape of the titanium material. Statistical analysis was performed on the results of the static charge measurements. In addition, the contamination of the titanium sandblasting was also evaluated. The results of the current studies suggested that sandblasting could generate static charges on titanium and stainless steel surfaces. However, such charges gradually dissipate into the atmosphere. The amount, polarity, and the decay rate of the charge depend on many factors tested in the experiments. These results might explain the beneficial of sandblasting on the osseointegration of titanium implants. Osseointegration has been traditionally attributed to its roughening effects. Moreover, the current studies may potentially lead to improved sandblasting techniques, becoming in mind that more research should be carried out. / published_or_final_version / Dentistry / Doctoral / Doctor of Philosophy

Gold complexes have a long history of being used as therapeutic agents, especially in applications against immune diseases such as rheumatoid arthritis. In 1979, an oral gold(I) drug – auranofin (AuRF, brand name as Ridaura®) – was demonstrated to exhibit anticancer properties. Since then, a considerable number of gold(I) complexes have been reported to show remarkable anticancer activities, but the understanding of their mechanism of actions is limited. In the present study, AuRF and several other anticancer gold(I)-phosphine complexes including AuPEt ([Au(triethylphosphine)Cl]) were demonstrated to induce autophagy – a cellular catabolic process of macromolecules and organelles through lysosomal degradation. The induced autophagy involved the accumulation of autophagosomes, which was mediated by the enhancement of autophagy initiation rather than by the blockage of autophagosomes maturation. Moreover, the AuRF and AuPEt induced autophagy was demonstrated to have a pro-survival effect for the cancer cells. To better explore the mechanism of actions of AuRF and other anticancer gold(I) complexes, a subcellular fractionation-based proteomic approach has been developed and optimized. This approach combined the use of subcellular fractionation, protein extraction, HPLC-LTQ-Orbitrap mass spectrometry, and bottom-up protein identification and quantification. By using this approach, the proteome coverage was increased, the complexities of the sub-proteomes were reduced, and the low-abundant organelle proteins were enriched. The nuclear sub-proteomes of AuRF-treated or AuPEt-treated cells were analyzed to identify the significantly regulated transcription regulators and the signaling pathways involved. The analysis delineates the possible AuRF-activated anticancer pathways involving up-regulation of the tumor suppressor cyclin-dependent kinase inhibitor 2A (〖p14〗^ARF), inhibition of the E2F transcription activity, blocking of the translocation of E3 ubiquitin-protein ligase (MDM2) from nucleus to cytoplasm and induction of the tumor suppressor p53. Furthermore, the KeyNode-based pathway analysis was applied to analyze the whole proteomes obtained from merging the sub-proteomes. Alongside the p53 pathway and E2F network, the regulation of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR, the rate-limiting enzyme of cholesterol biosynthesis) is one of the most up-regulated pathways of AuRF treatment. AuRF also showed significant inhibition to HMGCR activity in vitro with an IC50 value at the micromolar level. The effects of AuRF and AuPEt on the high mobility group box-1 protein (HMGB1), which exhibits distinct functions dependent on its cellular locations, were investigated. Treatment of cells with AuRF or AuPEt resulted in down-regulation of nuclear HMGB1, which is associated with p53-dependent cytotoxicities. The cytoplasmic HMGB1, which can induce autophagy, was found to be up-regulated. The levels of secreted HMGB1, which exhibits pro-inflammatory properties, were reduced, possibly contributing to anti-rheumatoid arthritis actions of AuRF. Collectively, the pharmacological and proteomic analyses in this research of AuRF and other anticancer gold(I) complexes supplement the current knowledge of their mechanism of actions. / published_or_final_version / Chemistry / Doctoral / Doctor of Philosophy

Background: Thalassaemia is a genetic disorder disease, one of the most clinically relevant haemoglobinopathies in paediatric population. It interferes with the synthesis of haemoglobin chain. For the sake of maintaining the serum haemoglobin at a normal level, regular blood cell transfusion is required to the patients with thalassaemia. In general, patients with thalassaemia are often diagnosed at an early age and need to take a life-long iron chelating therapy to prevent the multi-organ failure caused by iron-overload. The safety issue is considered a very importance aspect in the treatment among paediatric population and young people. In the past decades, numerous randomised controlled trials (RCTs) and meta-analysis regarding the efficacy and safety of iron chelating agents including deferoxamine, deferiprone, deferasirox, in reducing iron accumulation among patients with thalassaemia had been published, yet limited meta-analysis reveal the same issue among paediatric population and young adults. Further evidence and understanding are therefore needed to confirm whether or not the iron chelating agents are safe among young patients with thalassaemia. Objective: To conduct a meta-analysis to evaluate the safety of iron chelating agents in paediatric population and young adults with thalassaemia. Methods: Literature search was carried out in PubMed, EMBASE, BIOSIS Previews, Science Citation Index Expanded and Cochrane Library databases. This meta-analysis of observational studies was conducted following the PRISMA and MOOSE statements. Selection Criteria All prospective uncontrolled cohort studies were eligible to include. Articles were assessed according to the age range of its participants and the quality of the reported adverse effects. All enrolled studies should record countable cases of adverse effects of paediatric population and young patients up to 25 years of age, diagnosed with alpha/beta thalassaemia or sickle cell disease and under the treatment of iron chelating agents. Data Collection and analysis Two reviewers independently retrieved the data and conducted the quality assessment for each of the included studies. Agency for healthcare research and quality assessment tool was used to evaluate the general risk of bias, whilst the quality of harms was assessed with the Mcharm question criterions. Meta-analysis was carried out for detecting the entire proportion value of six adverse effects including liver abnormality, renal abnormality, rash, abdominal pain/discomfort, nausea and neutropenia, and the forest plot was generated accordingly. The I2 was estimated to assess the methodological quality of each outcome. Random or fixed model was used for the analysis. The sensitivity analysis was conducted to assess the robust of the result. Results A total of 8199 articles were identified in the initial database search. After removal of duplications, update from other sources and exclusion based on the exclusion criteria, 25 full articles were retrieved and 14 uncontrolled cohort studies were included in this review. Eight hundred and fifty-four patients up to 25 years of age were included in the analysis. The general quality of the studies was moderate while the quality of adverse effects was low to moderate. Out of the 14 included studies, nine were under the deferasirox treatment; five for deferiprone therapy. No study included deferoxamine. Most adverse effects were observed among the paediatric patients under deferasirox treatment. The meta-analysis of pooled proportion under deferasirox were 17.23% (95%CI, 8.78-25.68%) for liver abnormality, 11.58% (95%CI, 5.91-17.25%) for renal abnormality, 5.41% (95%CI, 3.23-7.58%) for rash, 11.03% (95%CI, 1.83-20.22%) for abdominal pain/discomfort and 5.77% (95%CI, 1,50-10.03%) for nausea. Only one study reported case of neutropenia in the patients under deferasirox, whereas more cases were recorded within the paediatric patients with deferiprone, estimated proportion of 5.98% (95%CI, 2.79-9.16%). However, the meta-analysis of estimated proportion for liver abnormality in paediatric patients with deferiprone was 10.08% (95%CI, 2.67-17.49%), abdominal pain/discomfort was 4.31% (95%CI, 1.65-6.98%). Only one study reported case of renal abnormality, rash and nausea respectively in the patients with deferasirox, which a meta-analysis could not be conducted. Conclusions Among paediatric population and young patients with thalassaemia disease, most drug-related adverse effects were liver injury among patients under both deferasirox and deferiprone. For patients under deferasirox, the proportions of the risk of abdominal pain/discomfort and renal abnormality were in a secondary high-level, whereas the proportions of the risk of rash and nausea were comparatively very low. Few adverse effects were detected among young patients with deferiprone. In addition, the proportion of liver abnormality and abdominal pain/discomfort were lower for deferiprone than deferasirox. Further investigation is needed to assess the safety and efficacy between different dosage of iron chelating agents and the risk of other adverse effects among paediatric population, which are necessary to guide the clinical practice in the treatment of paediatric patients with thalassaemia. / published_or_final_version / Pharmacology and Pharmacy / Master / Master of Medical Sciences

Not available / text

M.Tech. Drama. Tshwane University of Technology / When the healthy resonant frequency is out of balance, physical and emotional health is affected. The imbalance can be restored by the use of sound waves to once again create a healthy balance. Taking into account that the actor-in-training needs to focus on several elements on a physical, mental and emotional level all at once, it can be argued that s/he needs to be centred and keep all physical, mental and emotional aspects in balance. Therefore, well-being is certainly needed for optimal functioning of the actor and this is often archived through relaxation. This study hypothesizes that Sound Therapy will be beneficial as means of achieving a sense of well-being in the actor.