Applications of camphor-derived chiral auxiliaries in the asymmetric synthesis of α-amino acids and other systems
- Authors: Matjila, Joseph Moemise
- Date: 1998-04
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/191501 , vital:45106
- Description: A viable synthetic route to camphor-derived imino lactones as precursors for the asymmetric synthesis of a-amino acids has been established. Several synthetic strategies have been investigated and the required regioisomeric imino lactones were finally obtained via the step-wise condensation of JV-(carbobenzyloxy)glycine with the a-ketols, 3-exo-hydroxycamphor and 2-exo- hydroxy-3-bomanone. Enolates of the camphor imino lactones, generated using potassium tert- butoxide, were reacted with a range of alkyl halides. Dialkylation was observed using the 2-imino lactone, while the regioisomeric 3-imino lactone derivative gave monoalkylated products with diastereoselectivities, shown by NMR spectroscopy, to range from 43% d.e. for the methylated product to > 99% d.e. for larger alkyl groups. The expected preference for endo-alkylation is supported by NMR (chemical shift, coupling constant and NOE) data and was confirmed by acidic hydrolysis of the pentylated 3-imino lactone to afford the known acid. Computer modelling, with the software package HYPERCHEM®, was used to explore the conformational properties of the alkylated products and their enolate precursors. Exploratory work on the enantiomeric beneficiation of racemic amino acids, using alkylated imino lactone derivatives, revealed preferential exo-protonation of the enolate intermediates. Asymmetric Baylis-Hillman reactions between a novel camphor-derived acrylic ester and a range of aldehydes afforded the corresponding 2-(hydroxyalkyl)acrylates in up to 59% d.e., the observed stereoselectivities being sensitive to both steric and electronic factors. Attempts to prepare imino lactone derivatives from ketopinic acid, although unsuccessful, led to the isolation of two novel W-(carbobenzyloxy)glycinates, whose structures were established by 1- and 2-D NMR spectroscopy. Attempts to prepare "BINAP" analogues from dibomyl ether's also proved unsuccessful, but the investigation led to the discovery of a third, novel dibomyl ether. , Thesis (PhD) -- Faculty of Science, Chemistry, 1998
- Full Text:
- Date Issued: 1998-04
- Authors: Matjila, Joseph Moemise
- Date: 1998-04
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/191501 , vital:45106
- Description: A viable synthetic route to camphor-derived imino lactones as precursors for the asymmetric synthesis of a-amino acids has been established. Several synthetic strategies have been investigated and the required regioisomeric imino lactones were finally obtained via the step-wise condensation of JV-(carbobenzyloxy)glycine with the a-ketols, 3-exo-hydroxycamphor and 2-exo- hydroxy-3-bomanone. Enolates of the camphor imino lactones, generated using potassium tert- butoxide, were reacted with a range of alkyl halides. Dialkylation was observed using the 2-imino lactone, while the regioisomeric 3-imino lactone derivative gave monoalkylated products with diastereoselectivities, shown by NMR spectroscopy, to range from 43% d.e. for the methylated product to > 99% d.e. for larger alkyl groups. The expected preference for endo-alkylation is supported by NMR (chemical shift, coupling constant and NOE) data and was confirmed by acidic hydrolysis of the pentylated 3-imino lactone to afford the known acid. Computer modelling, with the software package HYPERCHEM®, was used to explore the conformational properties of the alkylated products and their enolate precursors. Exploratory work on the enantiomeric beneficiation of racemic amino acids, using alkylated imino lactone derivatives, revealed preferential exo-protonation of the enolate intermediates. Asymmetric Baylis-Hillman reactions between a novel camphor-derived acrylic ester and a range of aldehydes afforded the corresponding 2-(hydroxyalkyl)acrylates in up to 59% d.e., the observed stereoselectivities being sensitive to both steric and electronic factors. Attempts to prepare imino lactone derivatives from ketopinic acid, although unsuccessful, led to the isolation of two novel W-(carbobenzyloxy)glycinates, whose structures were established by 1- and 2-D NMR spectroscopy. Attempts to prepare "BINAP" analogues from dibomyl ether's also proved unsuccessful, but the investigation led to the discovery of a third, novel dibomyl ether. , Thesis (PhD) -- Faculty of Science, Chemistry, 1998
- Full Text:
- Date Issued: 1998-04
Reactor development and process optimisation for the bioremediation of phenolic wastewaters by trametes species
- Authors: Ryan, Daniel Reginald
- Date: 2004-04
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/191487 , vital:45103
- Description: In many service industries, the source of a company’s value has shifted from capital to knowledge and ideas, the quality of which is dependent on its employees (Wooldridge, 2006). In fact, human resources can be considered part of factor conditions which can positively impact on a firm’s competitive context. This impact can ultimately translate into improved financial results (Porter and Kramer, 2002). There is therefore a growing interest in ways to attract and retain talent. According to the managers of many big companies, well communicated corporate responsibility practices can improve staff attraction as well as retention rates by improving morale (CSRwire, 2002). To explore this, a small, creative company in Johannesburg which engages in charity work was selected as a case study, with the goal being to understand whether their culture of good deeds has a positive impact on staff wellbeing. While the owner of the company actively attempts to make the company an enjoyable place to work at, he appears to have initiated the philanthropic activities in a true spirit of giving, rather than with the motive of engaging staff in order to make more money. Nevertheless, the researcher’s investigative stance is that of an enlightened egoist, and the study focuses on the business case of giving being beneficial to the giver (ultimately the company) in the long term, as well as to the recipient. While the danger of suggesting that philanthropy could be instrumentalised is acknowledged (Morton, 2004), the investigation explores the possibility because such evidence could persuade other companies to become more socially concerned. Through a qualitative approach involving interviews, observation and analysis of video footage, it becomes apparent that there is clearly value for the staff in the charity work they do. Unfortunately the multiple initiatives undertaken to keep staff morale high at the company make it impossible to establish a clear link between the philanthropy and overall wellbeing, but as the study was conducted in the phenomenological paradigm the main concern was with understanding the experience of participants. However, an unexpected finding was that the employees derive great satisfaction from using their professional skills for charity work rather than just donating money to the charity. They feel that their skills uniquely position them to make significant changes to the lives of others, which gives them a sense of pride and achievement that they don’t necessarily experience in their ordinary activities at work. On the basis of this, it is recommended that companies look to involve staff with projects that require their specific expertise when evaluating philanthropic initiatives. , Thesis (PhD) -- Faculty of Science, Biochemistry, Microbiology and Biotechnology, 2004
- Full Text:
- Date Issued: 2004-04
- Authors: Ryan, Daniel Reginald
- Date: 2004-04
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/191487 , vital:45103
- Description: In many service industries, the source of a company’s value has shifted from capital to knowledge and ideas, the quality of which is dependent on its employees (Wooldridge, 2006). In fact, human resources can be considered part of factor conditions which can positively impact on a firm’s competitive context. This impact can ultimately translate into improved financial results (Porter and Kramer, 2002). There is therefore a growing interest in ways to attract and retain talent. According to the managers of many big companies, well communicated corporate responsibility practices can improve staff attraction as well as retention rates by improving morale (CSRwire, 2002). To explore this, a small, creative company in Johannesburg which engages in charity work was selected as a case study, with the goal being to understand whether their culture of good deeds has a positive impact on staff wellbeing. While the owner of the company actively attempts to make the company an enjoyable place to work at, he appears to have initiated the philanthropic activities in a true spirit of giving, rather than with the motive of engaging staff in order to make more money. Nevertheless, the researcher’s investigative stance is that of an enlightened egoist, and the study focuses on the business case of giving being beneficial to the giver (ultimately the company) in the long term, as well as to the recipient. While the danger of suggesting that philanthropy could be instrumentalised is acknowledged (Morton, 2004), the investigation explores the possibility because such evidence could persuade other companies to become more socially concerned. Through a qualitative approach involving interviews, observation and analysis of video footage, it becomes apparent that there is clearly value for the staff in the charity work they do. Unfortunately the multiple initiatives undertaken to keep staff morale high at the company make it impossible to establish a clear link between the philanthropy and overall wellbeing, but as the study was conducted in the phenomenological paradigm the main concern was with understanding the experience of participants. However, an unexpected finding was that the employees derive great satisfaction from using their professional skills for charity work rather than just donating money to the charity. They feel that their skills uniquely position them to make significant changes to the lives of others, which gives them a sense of pride and achievement that they don’t necessarily experience in their ordinary activities at work. On the basis of this, it is recommended that companies look to involve staff with projects that require their specific expertise when evaluating philanthropic initiatives. , Thesis (PhD) -- Faculty of Science, Biochemistry, Microbiology and Biotechnology, 2004
- Full Text:
- Date Issued: 2004-04
Synthesis and evaluation of novel heterocycles as potential HIV-1 enzyme inhibitors
- Ngnie Tuemgnie, Gaëlle Tatiana
- Authors: Ngnie Tuemgnie, Gaëlle Tatiana
- Date: 2014
- Subjects: Heterocyclic compounds , Enzyme inhibitors , Organic compounds , Green chemistry , Coumarins , HIV (Viruses) Enzymes
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194293 , vital:45440 , DOI https://doi.org/10.21504/10962/194293
- Description: This project has focussed on the synthesis and the evaluation of organic compounds as potential HIV-1 enzyme inhibitors, by making use of green chemistry (microwave assisted synthesis and click chemistry), palladium catalyzed reactions (Heck and Sonogashira coupling), Baylis Hillman methodology and aldol condensation. These compounds were synthesized in good yields and fully characterised by spectroscopic techniques. Biological assay data revealed that some of the compounds possess high inhibitory activity and their effective inhibitory concentration was as good as those of drugs in clinical use. These potential drug molecules were identified by preliminary investigations carried out by molecular modelling where a trend of their inhibitory activity against different enzymes was anticipated. Benzotriazole-AZT conjugates generated by 1,3-dipolar cycloaddition of anthranilic acid derivatives with AZT showed good inhibitory activity in silico against both HIV-1 protease (PR) and HIV-1 reverse transcriptase (RT) enzymes. Still in line with our dual action strategy, cinnamate ester-AZT conjugates were synthesized in three steps starting from benzaldehyde derivatives with a click reaction at the final step. These compounds also showed some inhibitory activity against HIV-1 RT enzyme (88%). In addition, the cinnamoyl fragment attached to AZT appeared to improve the activity of AZT against HIV-1 RT. Peptide chemistry involving carbonyl diimidazole as a coupling reagent between cinnamic acid derivatives and protected amino acids was used to prepare substituted amino acid derivatives which appeared to be very active against the integrase (IN) enzyme (88%). Commercially available coumarin was iodinated and derivatized through palladium catalyzed Heck and Sonogashira reactions with activated alkenes and a terminal alkyne respectively to afford novel coumarin derivatives in good yields. Optimization studies on the Heck reaction with regards to the phosphine ligand, the palladium catalyst and the solvent were carried out to afford novel formyl substituted cinnamate esters with nonaflyl salicylaldehyde derivatives. , Thesis (PhD) -- Faculty of Science, Chemistry, 2014
- Full Text:
- Date Issued: 2014
- Authors: Ngnie Tuemgnie, Gaëlle Tatiana
- Date: 2014
- Subjects: Heterocyclic compounds , Enzyme inhibitors , Organic compounds , Green chemistry , Coumarins , HIV (Viruses) Enzymes
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194293 , vital:45440 , DOI https://doi.org/10.21504/10962/194293
- Description: This project has focussed on the synthesis and the evaluation of organic compounds as potential HIV-1 enzyme inhibitors, by making use of green chemistry (microwave assisted synthesis and click chemistry), palladium catalyzed reactions (Heck and Sonogashira coupling), Baylis Hillman methodology and aldol condensation. These compounds were synthesized in good yields and fully characterised by spectroscopic techniques. Biological assay data revealed that some of the compounds possess high inhibitory activity and their effective inhibitory concentration was as good as those of drugs in clinical use. These potential drug molecules were identified by preliminary investigations carried out by molecular modelling where a trend of their inhibitory activity against different enzymes was anticipated. Benzotriazole-AZT conjugates generated by 1,3-dipolar cycloaddition of anthranilic acid derivatives with AZT showed good inhibitory activity in silico against both HIV-1 protease (PR) and HIV-1 reverse transcriptase (RT) enzymes. Still in line with our dual action strategy, cinnamate ester-AZT conjugates were synthesized in three steps starting from benzaldehyde derivatives with a click reaction at the final step. These compounds also showed some inhibitory activity against HIV-1 RT enzyme (88%). In addition, the cinnamoyl fragment attached to AZT appeared to improve the activity of AZT against HIV-1 RT. Peptide chemistry involving carbonyl diimidazole as a coupling reagent between cinnamic acid derivatives and protected amino acids was used to prepare substituted amino acid derivatives which appeared to be very active against the integrase (IN) enzyme (88%). Commercially available coumarin was iodinated and derivatized through palladium catalyzed Heck and Sonogashira reactions with activated alkenes and a terminal alkyne respectively to afford novel coumarin derivatives in good yields. Optimization studies on the Heck reaction with regards to the phosphine ligand, the palladium catalyst and the solvent were carried out to afford novel formyl substituted cinnamate esters with nonaflyl salicylaldehyde derivatives. , Thesis (PhD) -- Faculty of Science, Chemistry, 2014
- Full Text:
- Date Issued: 2014
Understanding the replication biology of Providence virus: elucidating the function of non-structural proteins
- Authors: Nakayinga, Ritah
- Date: 2014
- Subjects: Insects Viruses , Viruses Reproduction , Tombusviridae , RNA viruses , RNA polymerases
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193930 , vital:45408
- Description: Tetraviruses are non-enveloped, small insect RNA viruses with a single stranded positive RNA genome that is either monopartite or bipartite. Providence virus (PrV) is the only member of the three tetravirus families with a viral replicase similar to the replicases of tombusviruses and umbraviruses. The principle aim of this thesis was to study PrV replication, focusing on subcellular localization and potential interactions between PrV replication proteins. The first objective of this study was to generate an anti-p104 antibody that does not cross-react with p40. Expression of the C-terminal portion of p104 in E. coli resulted in no detectable protein. Further expression in an insect cell based expression system resulted in the production of an insoluble protein. Attempts to improve protein solubility with a range of solubilization treatments were unsuccessful. Bioinformatic analysis was used to detect an antigenic region at the C-terminus of p104 and the peptide was used to raise anti-p104 antibodies. These antibodies did not detect native protein by western blot detection however they were used for immunoprecipitation. The establishment of the subcellular localization of PrV required two approaches; immunofluorescence in persistently infected Helicoverpa zea MG8 cells using antip40 and anti-dsRNA antibodies and the expression of EGFP-replicase fusion protein in Spodoptera frugiperda Sf9 cells. Replication of PrV was found to take place in cytosolic punctate structures. Co-immunoprecipitation experiments revealed that p40 self-interacts and interacts with p104. Bioinformatic analysis of PrV p104 suggests that the RdRp is similar to viral RdRps of the carmo-like supergroup II. Potential RNA binding regions are present within p104. A potential p40 interaction domain that shares hydrophilic and surface exposed properties with the TBSV p33 interaction domain is present. A putative arginine-rich region and disordered C-terminal region is present in p130. In conclusion, PrV p104 is the viral replicase. The resemblance of the expression strategy and putative functional domains with tombusviruses and umbraviruses suggest that PrV replication is related to the replication system of the tombusviruses and umbraviruses. This has led to propose that tetravirus replication strategies are diverse and raises questions on the origin and evolution of PrV. , Thesis (PhD) -- Faculty of Science, Biochemistry, Microbiology and Biotechnology, 2014
- Full Text:
- Date Issued: 2014
- Authors: Nakayinga, Ritah
- Date: 2014
- Subjects: Insects Viruses , Viruses Reproduction , Tombusviridae , RNA viruses , RNA polymerases
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193930 , vital:45408
- Description: Tetraviruses are non-enveloped, small insect RNA viruses with a single stranded positive RNA genome that is either monopartite or bipartite. Providence virus (PrV) is the only member of the three tetravirus families with a viral replicase similar to the replicases of tombusviruses and umbraviruses. The principle aim of this thesis was to study PrV replication, focusing on subcellular localization and potential interactions between PrV replication proteins. The first objective of this study was to generate an anti-p104 antibody that does not cross-react with p40. Expression of the C-terminal portion of p104 in E. coli resulted in no detectable protein. Further expression in an insect cell based expression system resulted in the production of an insoluble protein. Attempts to improve protein solubility with a range of solubilization treatments were unsuccessful. Bioinformatic analysis was used to detect an antigenic region at the C-terminus of p104 and the peptide was used to raise anti-p104 antibodies. These antibodies did not detect native protein by western blot detection however they were used for immunoprecipitation. The establishment of the subcellular localization of PrV required two approaches; immunofluorescence in persistently infected Helicoverpa zea MG8 cells using antip40 and anti-dsRNA antibodies and the expression of EGFP-replicase fusion protein in Spodoptera frugiperda Sf9 cells. Replication of PrV was found to take place in cytosolic punctate structures. Co-immunoprecipitation experiments revealed that p40 self-interacts and interacts with p104. Bioinformatic analysis of PrV p104 suggests that the RdRp is similar to viral RdRps of the carmo-like supergroup II. Potential RNA binding regions are present within p104. A potential p40 interaction domain that shares hydrophilic and surface exposed properties with the TBSV p33 interaction domain is present. A putative arginine-rich region and disordered C-terminal region is present in p130. In conclusion, PrV p104 is the viral replicase. The resemblance of the expression strategy and putative functional domains with tombusviruses and umbraviruses suggest that PrV replication is related to the replication system of the tombusviruses and umbraviruses. This has led to propose that tetravirus replication strategies are diverse and raises questions on the origin and evolution of PrV. , Thesis (PhD) -- Faculty of Science, Biochemistry, Microbiology and Biotechnology, 2014
- Full Text:
- Date Issued: 2014
Establishment of human OCT4 as a putative HSP90 client protein: a case for HSP90 chaperoning pluripotency
- Authors: Sterrenberg, Jason Neville
- Date: 2015
- Subjects: Induced pluripotent stem cells , Heat shock proteins , Stem cells , Transcription factors , Molecular chaperones
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194010 , vital:45415 , 10.21504/10962/194010
- Description: The therapeutic potential of stem cells is already being harnessed in clinical trails. Of even greater therapeutic potential has been the discovery of mechanisms to reprogram differentiated cells into a pluripotent stem cell-like state known as induced pluripotent stem cells (iPSCs). Stem cell nature is governed and maintained by a hierarchy of transcription factors, the apex of which is OCT4. Although much research has elucidated the transcriptional regulation of OCT4, OCT4 regulated gene expression profiles and OCT4 transcriptional activation mechanisms in both stem cell biology and cellular reprogramming to iPSCs, the fundamental biochemistry surrounding the OCT4 transcription factor remains largely unknown. In order to analyze the biochemical relationship between HSP90 and human OCT4 we developed an exogenous active human OCT4 expression model with human OCT4 under transcriptional control of a constitutive promoter. We identified the direct interaction between HSP90 and human OCT4 despite the fact that the proteins predominantly display differential subcellular localizations. We show that HSP90 inhibition resulted in degradation of human OCT4 via the ubiquitin proteasome degradation pathway. As human OCT4 and HSP90 did not interact in the nucleus, we suggest that HSP90 functions in the cytoplasmic stabilization of human OCT4. Our analysis suggests HSP90 inhibition inhibits the transcriptional activity of human OCT4 dimers without affecting monomeric OCT4 activity. Additionally our data suggests that the HSP90 and human OCT4 complex is modulated by phosphorylation events either promoting or abrogating the interaction between HSP90 and human OCT4. Our data suggest that human OCT4 displays the characteristics describing HSP90 client proteins, therefore we identify human OCT4 as a putative HSP90 client protein. The regulation of the transcription factor OCT4 by HSP90 provides fundamental insights into the complex biochemistry of stem cell biology. This may also be suggestive that HSP90 not only regulates stem cell biology by maintaining routine cellular homeostasis but additionally through the direct regulation of pluripotency factors. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2015
- Full Text:
- Date Issued: 2015
- Authors: Sterrenberg, Jason Neville
- Date: 2015
- Subjects: Induced pluripotent stem cells , Heat shock proteins , Stem cells , Transcription factors , Molecular chaperones
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194010 , vital:45415 , 10.21504/10962/194010
- Description: The therapeutic potential of stem cells is already being harnessed in clinical trails. Of even greater therapeutic potential has been the discovery of mechanisms to reprogram differentiated cells into a pluripotent stem cell-like state known as induced pluripotent stem cells (iPSCs). Stem cell nature is governed and maintained by a hierarchy of transcription factors, the apex of which is OCT4. Although much research has elucidated the transcriptional regulation of OCT4, OCT4 regulated gene expression profiles and OCT4 transcriptional activation mechanisms in both stem cell biology and cellular reprogramming to iPSCs, the fundamental biochemistry surrounding the OCT4 transcription factor remains largely unknown. In order to analyze the biochemical relationship between HSP90 and human OCT4 we developed an exogenous active human OCT4 expression model with human OCT4 under transcriptional control of a constitutive promoter. We identified the direct interaction between HSP90 and human OCT4 despite the fact that the proteins predominantly display differential subcellular localizations. We show that HSP90 inhibition resulted in degradation of human OCT4 via the ubiquitin proteasome degradation pathway. As human OCT4 and HSP90 did not interact in the nucleus, we suggest that HSP90 functions in the cytoplasmic stabilization of human OCT4. Our analysis suggests HSP90 inhibition inhibits the transcriptional activity of human OCT4 dimers without affecting monomeric OCT4 activity. Additionally our data suggests that the HSP90 and human OCT4 complex is modulated by phosphorylation events either promoting or abrogating the interaction between HSP90 and human OCT4. Our data suggest that human OCT4 displays the characteristics describing HSP90 client proteins, therefore we identify human OCT4 as a putative HSP90 client protein. The regulation of the transcription factor OCT4 by HSP90 provides fundamental insights into the complex biochemistry of stem cell biology. This may also be suggestive that HSP90 not only regulates stem cell biology by maintaining routine cellular homeostasis but additionally through the direct regulation of pluripotency factors. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2015
- Full Text:
- Date Issued: 2015
Modification and application of the decentralised wastewater treatment technology for greywater treatment to reduce water needs
- Authors: Ngqwala, Nosiphiwe Patience
- Date: 2015
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193509 , vital:45338
- Description: Water is a scarce resource that is being acknowledged as a limiting factor to further social- economic growth and development. Demand for freshwater is increasing with corresponding increases in human population, industrial and agricultural activities. Alternative sources, such as greywater and rainwater are often polluted. Though greywater can be used for non-potable purposes, such as irrigation, it still requires some measures of treatment to improve its quality. To improve on greywater quality to facilitate its reuse, decentralised wastewater treatment technologies carry a great potential as complementary and alternative means of wastewater management particularly in peri-urban areas. Five research goals are addressed in this thesis: (i) to monitor the performance of Fly Ash Lime Filter Tower (FLFT) in the treatment of greywater; (ii) to modify the Fly Ash Lime Filter Tower in the treatment of greywater in order to reduce the pH of the greywater, and improved on the reduction of chemical oxygen demand (COD) and coliform counts; (iii) to investigate the potential of the reuse of greywater for irrigation; (iv) to undertake a techno analysis of the FLFT system for commercial use; and (v) to evaluate the use of hydrogen-sulphide (H2S) test kit to monitor faecal contamination of various water sources using a multidisciplinary approach. The modification of the FLFT indicated good treatment efficiency, reducing the concentrations of COD, chlorides, nitrates, ammonia and sulphate by 82.6%, 60.4%, 72.9%, 60.5%, and 53.9%, respectively; while the average pH was at 8.3. Greywater contains nutrients that are beneficial to the growth of most plants. Growth variables included biomass, stem height, number of leaves and number of vegetables harvested. Soil analysis showed no effects of the treated greywater on soil physico-chemical and microbial quality with bulk density 2.0g/cm3, average pH 7.4, total phosphorus 0.16mg/L 8, faecal coliform 0.3 CFU/100 ml. The tomatoes had high biomass and dry weight (150 g; 33g) than beetroot (35 g; 15 g). Crops irrigated with greywater significantly grew faster compared with those irrigated with tap water. The community approach highlighted the value of knowledge management in greywater reuse. It highlighted the importance of creating an institutional knowledge in water management using the H2S kit. The techno-economic analysis was used to evaluate key factors and the activities that are relevant to develop a sustainable FLFT in order to gain insights into the possibility of developing, and incorporating a business model framework to support decision making in value creation and value capturing during the research and the scaling up of the system. By this, a long term perspective to accomplish sustainable FLFT service businesses can be achieved. , Thesis (PhD) -- Faculty of Pharmacy, Pharmacy, 2015
- Full Text:
- Date Issued: 2015
- Authors: Ngqwala, Nosiphiwe Patience
- Date: 2015
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193509 , vital:45338
- Description: Water is a scarce resource that is being acknowledged as a limiting factor to further social- economic growth and development. Demand for freshwater is increasing with corresponding increases in human population, industrial and agricultural activities. Alternative sources, such as greywater and rainwater are often polluted. Though greywater can be used for non-potable purposes, such as irrigation, it still requires some measures of treatment to improve its quality. To improve on greywater quality to facilitate its reuse, decentralised wastewater treatment technologies carry a great potential as complementary and alternative means of wastewater management particularly in peri-urban areas. Five research goals are addressed in this thesis: (i) to monitor the performance of Fly Ash Lime Filter Tower (FLFT) in the treatment of greywater; (ii) to modify the Fly Ash Lime Filter Tower in the treatment of greywater in order to reduce the pH of the greywater, and improved on the reduction of chemical oxygen demand (COD) and coliform counts; (iii) to investigate the potential of the reuse of greywater for irrigation; (iv) to undertake a techno analysis of the FLFT system for commercial use; and (v) to evaluate the use of hydrogen-sulphide (H2S) test kit to monitor faecal contamination of various water sources using a multidisciplinary approach. The modification of the FLFT indicated good treatment efficiency, reducing the concentrations of COD, chlorides, nitrates, ammonia and sulphate by 82.6%, 60.4%, 72.9%, 60.5%, and 53.9%, respectively; while the average pH was at 8.3. Greywater contains nutrients that are beneficial to the growth of most plants. Growth variables included biomass, stem height, number of leaves and number of vegetables harvested. Soil analysis showed no effects of the treated greywater on soil physico-chemical and microbial quality with bulk density 2.0g/cm3, average pH 7.4, total phosphorus 0.16mg/L 8, faecal coliform 0.3 CFU/100 ml. The tomatoes had high biomass and dry weight (150 g; 33g) than beetroot (35 g; 15 g). Crops irrigated with greywater significantly grew faster compared with those irrigated with tap water. The community approach highlighted the value of knowledge management in greywater reuse. It highlighted the importance of creating an institutional knowledge in water management using the H2S kit. The techno-economic analysis was used to evaluate key factors and the activities that are relevant to develop a sustainable FLFT in order to gain insights into the possibility of developing, and incorporating a business model framework to support decision making in value creation and value capturing during the research and the scaling up of the system. By this, a long term perspective to accomplish sustainable FLFT service businesses can be achieved. , Thesis (PhD) -- Faculty of Pharmacy, Pharmacy, 2015
- Full Text:
- Date Issued: 2015
The design and evaluation of targeted patient-centred health information to improve knowledge and behavioural outcomes in tuberculosis patients with limited literacy
- Authors: Patel, Sonal
- Date: 2015
- Subjects: Tuberculosis Patients , Health literacy , Patient education , Communication in medicine , Picture-writing
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194071 , vital:45420 , DOI 10.21504/10962/194071
- Description: South Africa carries a significant TB burden as evidenced in the 2013 statistics which report 450 000 new active TB cases and 890 000 TB-related mortalities. For successful treatment outcomes, 90% adherence is necessary, but many patients prematurely discontinue treatment due to poor knowledge and understanding of their complex TB medicines. Patient education is pivotal in improving knowledge, health literacy and behavioural outcomes such as health information seeking, self-efficacy and adherence. In the under-resourced South African healthcare system, time and capacity to adequately counsel patients are limited. The value of written medicine information (WMI) to supplement the verbal information provided by healthcare professionals (HCPs) has been widely investigated but minimal South African research is available. Current WMI distributed in South Africa is mainly generated by pharmaceutical manufacturers and is complex, incomprehensible and undesirable to patients. TB-related WMI focuses mainly on the disease, with little information relating to TB medicines and their use. The overall aim of this project was to improve patient knowledge about their TB medicines through the use of a simple illustrated patient information leaflet (PIL). Objectives to achieve this aim included: investigation of the medicine information seeking behaviour (MISB) of long term patients attending public health sector facilities; the development and validation of a medicine literacy test (MLT) to identify patients with limited health literacy requiring additional support and counselling; the development and evaluation of a patient-centred illustrated PIL for first-line TB treatment; the assessment of self-efficacy and adherence using modified versions of the HIV Treatment Adherence Self-Efficacy Scale (HIV-ASES) and Morisky 8-item Medicine Adherence Scale (MMAS-8), respectively, and the investigation of the impact of the PIL on patient knowledge and these health-related behaviours. Six focus group discussions (FGDs) conducted in 34 isiXhosa-speaking patients with limited formal education taking long-term treatment explored themes related to information needs, information-seeking practices and awareness of and ability to utilize information sources. Codes were analysed and potential themes and subthemes were identified and refined. The findings of this study reflected a passive, disempowered patient due to both patient-related and systemic healthcare factors. Poor awareness of information sources, lack of health-related knowledge, stigma and lack of awareness of the importance of appropriate medicine-related knowledge contributed to a lack of information-seeking practice. Patients neither asked questions nor were encouraged to do so. All expressed an unmet need for information and a desire for receiving relevant, appropriate, written medicine-related information. Feedback from this phase of the study was used to inform the development of the targeted patientcentred PIL. A double-sided A4 PIL containing information about TB medicines was designed giving careful consideration to content, format and layout features. Twenty five pictograms were designed through a rigorous, iterative design process and were included in the PIL that was evaluated in a randomised control trial (RCT) conducted amongst 120 TB patients attending a high burden TB clinic in South Africa. Interviews were conducted in either isiXhosa or Afrikaans via a trained interpreter. Patients were randomly allocated to either a control (standard care) or an experimental group (standard care plus brief counselling using the PIL). Two interviews were conducted using a prepared questionnaire; one at baseline followed by a 4-week follow-up. Baseline data included demographics, medicine literacy test, health information sources, knowledge of TB medicines, self-reported adherence and self-efficacy. Data collected at the 4-week follow-up interview included TB knowledge, self-reported adherence, self-efficacy, opinion of TB medicine information and interpretation of pictograms. Data were analysed using t-test, correlations, chi-square and ANOVA tests at a 0.05 level of significance. The PIL was successful in improving patient knowledge of the disease, TB medicine-taking, side effects, drug-resistant TB and HIV and TB co-infection. At baseline, there was no significant difference in the overall mean percentage knowledge score between the control and experimental groups (p=0.074). At follow-up, the percentage knowledge score for the experimental group increased significantly from 59.0% to 84.6% (p<0.001) and showed a significantly higher score than the control group (p<0.001), displaying evidence of the impact of the PIL as a counselling tool on patient knowledge. The PIL generated a highly positive response in the experimental group who indicated that they had referred to the leaflet over the last month and that it had played an important role in improving their TB medicine-related knowledge. This was reflected in the experimental group knowledge score of greater than 80% for almost three quarters of the patients whereas only 14% in the control group achieved this score. Patients appreciated the inclusion of pictograms and strongly felt that they helped them to recall and understand the textual PIL content. The study found that patients want side effect information and, interestingly, did not perceive the presentation of side effects in pictorial form to constitute a risk factor for nonadherence. Use of the illustrated PIL (experimental group) resulted in a significant improvement in patient self-efficacy (p=0.002), but showed no effect on self-reported adherence (p=0.563). Neither self-efficacy nor adherence was influenced by gender, age or education. An education effect on knowledge was only observed in the control group at baseline. The newly developed MLT was shown to be a valid and reliable tool and a moderate, positive and significant correlation was noted between the MLT score and baseline TB medicine-related knowledge in both the control and experimental groups. As there is a paucity of studies investigating the influence of take-home written leaflets on TB medicine knowledge and on patient behaviour, this study represents a significant knowledge contribution. It is the first study to report the development and evaluation of a patient-centred PIL to address the dearth of available TB medicine information. The use of targeted user-friendly, illustrated information leaflets can be a valuable counselling aid to improve patient knowledge and self-efficacy, particularly among patients with limited literacy. However, careful consideration of the design and content, with input from the endusers at all stages of the process, will optimise its effectiveness. The proposed framework for the development and implementation of patient-centred health and medicines information in a developing country context presented in this thesis could be used as a theoretical basis for informing the development of effective information materials targeting other disease states. Local patients taking TB medicines identified nurses, WMI and media as their current sources of information but they expressed a strong desire to know more about their treatment. Targeted public health interventions that focus on medicine-taking information and behaviours and encourage patients to adopt a more active, questioning role in health consultations could improve health literacy and empower patients in their medicine-taking practices. , Thesis (PhD) -- Faculty of Pharmacy, Pharmacy, 2015
- Full Text:
- Date Issued: 2015
- Authors: Patel, Sonal
- Date: 2015
- Subjects: Tuberculosis Patients , Health literacy , Patient education , Communication in medicine , Picture-writing
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/194071 , vital:45420 , DOI 10.21504/10962/194071
- Description: South Africa carries a significant TB burden as evidenced in the 2013 statistics which report 450 000 new active TB cases and 890 000 TB-related mortalities. For successful treatment outcomes, 90% adherence is necessary, but many patients prematurely discontinue treatment due to poor knowledge and understanding of their complex TB medicines. Patient education is pivotal in improving knowledge, health literacy and behavioural outcomes such as health information seeking, self-efficacy and adherence. In the under-resourced South African healthcare system, time and capacity to adequately counsel patients are limited. The value of written medicine information (WMI) to supplement the verbal information provided by healthcare professionals (HCPs) has been widely investigated but minimal South African research is available. Current WMI distributed in South Africa is mainly generated by pharmaceutical manufacturers and is complex, incomprehensible and undesirable to patients. TB-related WMI focuses mainly on the disease, with little information relating to TB medicines and their use. The overall aim of this project was to improve patient knowledge about their TB medicines through the use of a simple illustrated patient information leaflet (PIL). Objectives to achieve this aim included: investigation of the medicine information seeking behaviour (MISB) of long term patients attending public health sector facilities; the development and validation of a medicine literacy test (MLT) to identify patients with limited health literacy requiring additional support and counselling; the development and evaluation of a patient-centred illustrated PIL for first-line TB treatment; the assessment of self-efficacy and adherence using modified versions of the HIV Treatment Adherence Self-Efficacy Scale (HIV-ASES) and Morisky 8-item Medicine Adherence Scale (MMAS-8), respectively, and the investigation of the impact of the PIL on patient knowledge and these health-related behaviours. Six focus group discussions (FGDs) conducted in 34 isiXhosa-speaking patients with limited formal education taking long-term treatment explored themes related to information needs, information-seeking practices and awareness of and ability to utilize information sources. Codes were analysed and potential themes and subthemes were identified and refined. The findings of this study reflected a passive, disempowered patient due to both patient-related and systemic healthcare factors. Poor awareness of information sources, lack of health-related knowledge, stigma and lack of awareness of the importance of appropriate medicine-related knowledge contributed to a lack of information-seeking practice. Patients neither asked questions nor were encouraged to do so. All expressed an unmet need for information and a desire for receiving relevant, appropriate, written medicine-related information. Feedback from this phase of the study was used to inform the development of the targeted patientcentred PIL. A double-sided A4 PIL containing information about TB medicines was designed giving careful consideration to content, format and layout features. Twenty five pictograms were designed through a rigorous, iterative design process and were included in the PIL that was evaluated in a randomised control trial (RCT) conducted amongst 120 TB patients attending a high burden TB clinic in South Africa. Interviews were conducted in either isiXhosa or Afrikaans via a trained interpreter. Patients were randomly allocated to either a control (standard care) or an experimental group (standard care plus brief counselling using the PIL). Two interviews were conducted using a prepared questionnaire; one at baseline followed by a 4-week follow-up. Baseline data included demographics, medicine literacy test, health information sources, knowledge of TB medicines, self-reported adherence and self-efficacy. Data collected at the 4-week follow-up interview included TB knowledge, self-reported adherence, self-efficacy, opinion of TB medicine information and interpretation of pictograms. Data were analysed using t-test, correlations, chi-square and ANOVA tests at a 0.05 level of significance. The PIL was successful in improving patient knowledge of the disease, TB medicine-taking, side effects, drug-resistant TB and HIV and TB co-infection. At baseline, there was no significant difference in the overall mean percentage knowledge score between the control and experimental groups (p=0.074). At follow-up, the percentage knowledge score for the experimental group increased significantly from 59.0% to 84.6% (p<0.001) and showed a significantly higher score than the control group (p<0.001), displaying evidence of the impact of the PIL as a counselling tool on patient knowledge. The PIL generated a highly positive response in the experimental group who indicated that they had referred to the leaflet over the last month and that it had played an important role in improving their TB medicine-related knowledge. This was reflected in the experimental group knowledge score of greater than 80% for almost three quarters of the patients whereas only 14% in the control group achieved this score. Patients appreciated the inclusion of pictograms and strongly felt that they helped them to recall and understand the textual PIL content. The study found that patients want side effect information and, interestingly, did not perceive the presentation of side effects in pictorial form to constitute a risk factor for nonadherence. Use of the illustrated PIL (experimental group) resulted in a significant improvement in patient self-efficacy (p=0.002), but showed no effect on self-reported adherence (p=0.563). Neither self-efficacy nor adherence was influenced by gender, age or education. An education effect on knowledge was only observed in the control group at baseline. The newly developed MLT was shown to be a valid and reliable tool and a moderate, positive and significant correlation was noted between the MLT score and baseline TB medicine-related knowledge in both the control and experimental groups. As there is a paucity of studies investigating the influence of take-home written leaflets on TB medicine knowledge and on patient behaviour, this study represents a significant knowledge contribution. It is the first study to report the development and evaluation of a patient-centred PIL to address the dearth of available TB medicine information. The use of targeted user-friendly, illustrated information leaflets can be a valuable counselling aid to improve patient knowledge and self-efficacy, particularly among patients with limited literacy. However, careful consideration of the design and content, with input from the endusers at all stages of the process, will optimise its effectiveness. The proposed framework for the development and implementation of patient-centred health and medicines information in a developing country context presented in this thesis could be used as a theoretical basis for informing the development of effective information materials targeting other disease states. Local patients taking TB medicines identified nurses, WMI and media as their current sources of information but they expressed a strong desire to know more about their treatment. Targeted public health interventions that focus on medicine-taking information and behaviours and encourage patients to adopt a more active, questioning role in health consultations could improve health literacy and empower patients in their medicine-taking practices. , Thesis (PhD) -- Faculty of Pharmacy, Pharmacy, 2015
- Full Text:
- Date Issued: 2015
Behaviour and trophic ecology of oceanic triggerfish (Canthidermis maculata) and rainbow runner (Elagatis bipinnulata) associated with floating objects in the open ocean
- Forget, Fabien Rocky Gilbert
- Authors: Forget, Fabien Rocky Gilbert
- Date: 2016
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193531 , vital:45340
- Description: The oceanic triggerfish (Canthidermis maculata) and rainbow runner (Elagatis bipinnulata) are epipelagic species that commonly associate with floating objects throughout the world’s tropical oceans. Besides tunas, these two species often form the bulk of the aggregation and are major non-target species (bycatch) incidentally captured at fish aggregation devices (FADs) by tropical tuna purse seiners. Despite being abundant, little is known about their basic biology, ecology and associative behavior with floating objects. This thesis aims to address knowledge gaps in their associative behavior and ecology at floating objects and attempts to explore potential bycatch mitigation methods. Acoustic telemetry was used to characterise several aspects of the two species’ behavior at floating objects. Fish were captured and tagged at drifting FADs in the core of the western Indian Ocean tuna fishing grounds. Prolonged remote observations were obtained through satellite linked receivers which provided fine-scale data on the behavior of oceanic triggerfish (n= 46 919 cumulated observation days) and rainbow runner (n= 24 538 cumulated observation days). The two species remained associated with the same floating object for periods extending up to several months. The mean residence time estimated using a survival analysis was 65 days for oceanic triggerfish and 94 days for rainbow runner. A distinct diel pattern in association was observed; the two species were closely associated to the FAD during the night and increased their home range during the day while performing short excursions (oceanic triggerfish mean: 2.0 hours ±1.6 SD; rainbow runner mean: 2.4 hours ±2.2 SD) away from the FAD. Similarly, distinct patterns in the vertical movements and locomotory activity (measured using accelerometer tags) were observed as the two species increased their vertical movements and activity levels during the day. Generally, their vertical movements were largely restricted to the mixed layer (< 60 m). At night, the two species remained close to the surface (0-5 m) where they appear to be in a resting state. An interesting behavioural switch in activity levels and vertical distribution was observed for oceanic triggerfish which coincided with a change in the environment as the FAD drifted close to a cold-core cyclonic eddy. Ambient light appears to be the key stimulus triggering the observed diel behavioural patterns. species-specific vulnerability for target species, skipjack (Katsuwonus pelamis), yellowfin (Thunnus albacares), bigeye tuna (Thunnus obesus) and non-target species, including silky shark (Carcharhinus falciformis), oceanic triggerfish, and rainbow runner, to the purse seine fishing gear were investigated. Hourly presence rates at FADs over 24 hours were examined to assess whether the capture rate of non-target species could be reduced at specific times during the day. Silky sharks displayed a similar associative pattern to that of targeted tunas. Moreover, the vulnerability of both target and non-target species were close to the maximum during the period of behavioural transitions at sunrise when the majority of the fishing sets are made. As such, a change in set time in the western Indian Ocean does not appear to be a feasible method to reduce capture rates of non-target species. Stomach content and stable isotope analysis were conducted on oceanic triggerfish (stomach n = 152, isotope n = 65) and rainbow runner (stomach n = 156, isotope n = 71) incidentally captured individuals at FADs by purse seiners. Typical FAD-associated prey items represented a minor component of their diet as they predominantly foraged on zooplankton and micronekton in the water column, suggesting that oceanic triggerfish and rainbow runner do not associate to floating objects for direct trophic advantages. Overlap in their dietary and isotopic niches indicates an intermediate level of competition. Overall, the spatial and trophic ecology of both species are remarkably similar and prolonged associations with FADs indicates that floating objects play a significant ecological role in the pelagic environment for the two species. , Thesis (PhD) -- Faculty of Science, Ichthyology and Fisheries Science, 2016
- Full Text:
- Date Issued: 2016
- Authors: Forget, Fabien Rocky Gilbert
- Date: 2016
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/193531 , vital:45340
- Description: The oceanic triggerfish (Canthidermis maculata) and rainbow runner (Elagatis bipinnulata) are epipelagic species that commonly associate with floating objects throughout the world’s tropical oceans. Besides tunas, these two species often form the bulk of the aggregation and are major non-target species (bycatch) incidentally captured at fish aggregation devices (FADs) by tropical tuna purse seiners. Despite being abundant, little is known about their basic biology, ecology and associative behavior with floating objects. This thesis aims to address knowledge gaps in their associative behavior and ecology at floating objects and attempts to explore potential bycatch mitigation methods. Acoustic telemetry was used to characterise several aspects of the two species’ behavior at floating objects. Fish were captured and tagged at drifting FADs in the core of the western Indian Ocean tuna fishing grounds. Prolonged remote observations were obtained through satellite linked receivers which provided fine-scale data on the behavior of oceanic triggerfish (n= 46 919 cumulated observation days) and rainbow runner (n= 24 538 cumulated observation days). The two species remained associated with the same floating object for periods extending up to several months. The mean residence time estimated using a survival analysis was 65 days for oceanic triggerfish and 94 days for rainbow runner. A distinct diel pattern in association was observed; the two species were closely associated to the FAD during the night and increased their home range during the day while performing short excursions (oceanic triggerfish mean: 2.0 hours ±1.6 SD; rainbow runner mean: 2.4 hours ±2.2 SD) away from the FAD. Similarly, distinct patterns in the vertical movements and locomotory activity (measured using accelerometer tags) were observed as the two species increased their vertical movements and activity levels during the day. Generally, their vertical movements were largely restricted to the mixed layer (< 60 m). At night, the two species remained close to the surface (0-5 m) where they appear to be in a resting state. An interesting behavioural switch in activity levels and vertical distribution was observed for oceanic triggerfish which coincided with a change in the environment as the FAD drifted close to a cold-core cyclonic eddy. Ambient light appears to be the key stimulus triggering the observed diel behavioural patterns. species-specific vulnerability for target species, skipjack (Katsuwonus pelamis), yellowfin (Thunnus albacares), bigeye tuna (Thunnus obesus) and non-target species, including silky shark (Carcharhinus falciformis), oceanic triggerfish, and rainbow runner, to the purse seine fishing gear were investigated. Hourly presence rates at FADs over 24 hours were examined to assess whether the capture rate of non-target species could be reduced at specific times during the day. Silky sharks displayed a similar associative pattern to that of targeted tunas. Moreover, the vulnerability of both target and non-target species were close to the maximum during the period of behavioural transitions at sunrise when the majority of the fishing sets are made. As such, a change in set time in the western Indian Ocean does not appear to be a feasible method to reduce capture rates of non-target species. Stomach content and stable isotope analysis were conducted on oceanic triggerfish (stomach n = 152, isotope n = 65) and rainbow runner (stomach n = 156, isotope n = 71) incidentally captured individuals at FADs by purse seiners. Typical FAD-associated prey items represented a minor component of their diet as they predominantly foraged on zooplankton and micronekton in the water column, suggesting that oceanic triggerfish and rainbow runner do not associate to floating objects for direct trophic advantages. Overlap in their dietary and isotopic niches indicates an intermediate level of competition. Overall, the spatial and trophic ecology of both species are remarkably similar and prolonged associations with FADs indicates that floating objects play a significant ecological role in the pelagic environment for the two species. , Thesis (PhD) -- Faculty of Science, Ichthyology and Fisheries Science, 2016
- Full Text:
- Date Issued: 2016
Soil microbial properties and apple tree performance under conventional and organic management
- Authors: Meyer, André Harold
- Date: 2016
- Subjects: Soil management South Africa Western Cape , Agricultural chemicals Environmental aspects , Soil microbiology South Africa Western Cape , Vesicular-arbuscular mycorrhizas , Enzymes Biotechnology , Apples Organic farming South Africa Western Cape
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64545 , vital:28557
- Description: Conventional (CON) soil management that permits the use of agrochemicals is currently the most common form of management in Western Cape deciduous fruit orchards. There is increasing pressure to minimise or eliminate synthetic agrochemical usage due to its potentially harmful effect on the environment, particularly to non-target soil microorganisms, and to the functions and processes they perform or mediate. In apple orchards, organic (ORG) practices exclude the use of synthetic pesticides and herbicides making use instead of organic fertilisers and naturally derived products as defined by organic certification programs. ORG practices aim to improve nutrient availability, yield, and long-term orchard sustainability relative to CON orchard management practices. If ORG and CON orchard floor management practices affect orchard ecosystems differently, such differences should be measurable in terms of differences in microbiological parameters. In this thesis it is hypothesised that ORG practices would induce positive soil microbiological responses in Western Cape apple orchards relative to CON practices, and by inference general soil health and apple tree performance. To test this hypothesis a polyphasic approach was adopted. This involved measurement of soil microbial activities and functional diversities, by enzyme activity (using colorimetric assays) and carbon-substrate utilisation (using the BIOLOG™ system), respectively. With reference to the enzyme analyses, the performance of a literature-validated, enzyme-based soil health index was also tested. The analyses were supported by coarse-level comparisons of the magnitude of bacteria, fungi, actinobacteria and total heterotroph populations using traditional culturing techniques (dilution plating on growth media). The extent to which the microbial status differed between the applied ORG and CON treatments was thought likely to reflect such treatment-induced variables as soil nutrient status, apple tree nutritional response, tree growth and yield, all of which were determined. Because the root systems of deciduous fruit trees commonly extend to depths >60 cm in well-prepared soils, microbial enzyme activities in the soil depth intervals corresponding to the lower rootzone, were also investigated. This research was carried out in a randomized field trial. Finally, to gain a broader understanding of the effects of contrasting soil management systems on soil microbiology under a greater variety of environmental conditions, arbuscular mycorrhizal (AM) fungal dynamics were explored in a survey of commercial apple orchards. These orchards were selected to span the range of environmental conditions that occur in the apple production areas of the Western Cape. Orchard soils under ORG management promoted richer microbial ecosystems, and appeared to be better able to sustain community metabolic diversity and, by inference, the functions mediated by soil microbial communities, than those under CON management. This implies that ORG approaches possibly afford a better option to sustain critical ecosystem functions than CON management. This possibly explains why use of straw mulches and compost in accordance with ORG practices, compared with CON practices, promoted β-glucosidase, acid phosphatase and urease activities rather than affecting the abundance of the micro-organisms that produce these enzymes. Enzyme activities in the 0–30 cm soil intervals were also more effectively promoted by ORG than CON practices, although no differences were observed at lower depth intervals. ORG practices promoted functional AM associations more effectively than CON practices, but the abundance of glomalin, a beneficial by-product of AM fungi, was unaffected. The greater enzyme activities and higher root colonisation levels in the ORG treatments probably contributed to improved nutritional effects that caused greater vegetative growth, but lower yields, in the ORG treatments. Yield suppression was conceivably due to excessive vegetative growth induced by oversupply of compost and the mineral nutrients contained therein. The survey of Western Cape apple orchards suggested that neither glomalin nor root colonisation bore any specific relationship to production area, cultivation practice, scion x rootstock combination, or, in the case of root colonisation, with any chemical parameters. However, the effect of season on glomalin was conclusively shown, being higher in summer than in spring, as was the lack of any effect of year on glomalin and root colonisation. Collectively, these results showed that ORG soil management promote soil microbiology, soil nutrient status, and apple tree performance compared to CON management. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2016
- Full Text:
- Date Issued: 2016
- Authors: Meyer, André Harold
- Date: 2016
- Subjects: Soil management South Africa Western Cape , Agricultural chemicals Environmental aspects , Soil microbiology South Africa Western Cape , Vesicular-arbuscular mycorrhizas , Enzymes Biotechnology , Apples Organic farming South Africa Western Cape
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64545 , vital:28557
- Description: Conventional (CON) soil management that permits the use of agrochemicals is currently the most common form of management in Western Cape deciduous fruit orchards. There is increasing pressure to minimise or eliminate synthetic agrochemical usage due to its potentially harmful effect on the environment, particularly to non-target soil microorganisms, and to the functions and processes they perform or mediate. In apple orchards, organic (ORG) practices exclude the use of synthetic pesticides and herbicides making use instead of organic fertilisers and naturally derived products as defined by organic certification programs. ORG practices aim to improve nutrient availability, yield, and long-term orchard sustainability relative to CON orchard management practices. If ORG and CON orchard floor management practices affect orchard ecosystems differently, such differences should be measurable in terms of differences in microbiological parameters. In this thesis it is hypothesised that ORG practices would induce positive soil microbiological responses in Western Cape apple orchards relative to CON practices, and by inference general soil health and apple tree performance. To test this hypothesis a polyphasic approach was adopted. This involved measurement of soil microbial activities and functional diversities, by enzyme activity (using colorimetric assays) and carbon-substrate utilisation (using the BIOLOG™ system), respectively. With reference to the enzyme analyses, the performance of a literature-validated, enzyme-based soil health index was also tested. The analyses were supported by coarse-level comparisons of the magnitude of bacteria, fungi, actinobacteria and total heterotroph populations using traditional culturing techniques (dilution plating on growth media). The extent to which the microbial status differed between the applied ORG and CON treatments was thought likely to reflect such treatment-induced variables as soil nutrient status, apple tree nutritional response, tree growth and yield, all of which were determined. Because the root systems of deciduous fruit trees commonly extend to depths >60 cm in well-prepared soils, microbial enzyme activities in the soil depth intervals corresponding to the lower rootzone, were also investigated. This research was carried out in a randomized field trial. Finally, to gain a broader understanding of the effects of contrasting soil management systems on soil microbiology under a greater variety of environmental conditions, arbuscular mycorrhizal (AM) fungal dynamics were explored in a survey of commercial apple orchards. These orchards were selected to span the range of environmental conditions that occur in the apple production areas of the Western Cape. Orchard soils under ORG management promoted richer microbial ecosystems, and appeared to be better able to sustain community metabolic diversity and, by inference, the functions mediated by soil microbial communities, than those under CON management. This implies that ORG approaches possibly afford a better option to sustain critical ecosystem functions than CON management. This possibly explains why use of straw mulches and compost in accordance with ORG practices, compared with CON practices, promoted β-glucosidase, acid phosphatase and urease activities rather than affecting the abundance of the micro-organisms that produce these enzymes. Enzyme activities in the 0–30 cm soil intervals were also more effectively promoted by ORG than CON practices, although no differences were observed at lower depth intervals. ORG practices promoted functional AM associations more effectively than CON practices, but the abundance of glomalin, a beneficial by-product of AM fungi, was unaffected. The greater enzyme activities and higher root colonisation levels in the ORG treatments probably contributed to improved nutritional effects that caused greater vegetative growth, but lower yields, in the ORG treatments. Yield suppression was conceivably due to excessive vegetative growth induced by oversupply of compost and the mineral nutrients contained therein. The survey of Western Cape apple orchards suggested that neither glomalin nor root colonisation bore any specific relationship to production area, cultivation practice, scion x rootstock combination, or, in the case of root colonisation, with any chemical parameters. However, the effect of season on glomalin was conclusively shown, being higher in summer than in spring, as was the lack of any effect of year on glomalin and root colonisation. Collectively, these results showed that ORG soil management promote soil microbiology, soil nutrient status, and apple tree performance compared to CON management. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2016
- Full Text:
- Date Issued: 2016
A systems thinking approach to e-government strategy formulation for water service delivery in South African local municipalities
- Authors: Osah, Umeoniso Joshua
- Date: 2017
- Subjects: Internet in public administration South Africa , Electronic government information South Africa , Communication in public administration South Africa , Municipal services South Africa , Municipal services Information technology , System theory
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64471 , vital:28547 , DOI 10.21504/10962/64471
- Description: E-Government deployments by stakeholders within the South African water service sector, provide certain benefits for the sector. While deemed beneficial and of considerable value, e-Government deployments and implementations in the water sector of local governments of South Africa have not always been successful. One important reason for e-Government failures, among several others, is the lack of coherent strategies, informed by key representing stakeholder views, to guide implementation and deployment of e-Government tools. Without strategies, it is highly likely that ICT integration will be conducted haphazardly. As a point of note, more than the deliverable (strategy document) that represents the output of the strategy development process, it serves more purpose to understand the process that results in the strategy. Importantly, understanding the process helps to account for the formed relationships between the various stakeholders that need to buy into the strategy. The research study develops an e-Government strategy formulation framework based on a systems thinking approach, intended to support the strategy formulation process of e-Government strategies – to underpin the effective integration, deployment and sustained use of ICT solutions for water service delivery at the local government level. A systems thinking approach is considered due to its emphasis on the strategy being informed by a holistic assessment. Where there is some knowledge about the processes by which a strategy is formulated – over time ideas may be derived on the types of processes that may produce efficient e-Government strategies. The research is conducted using the Design Science research paradigm. The Design Science paradigm is comprised of two processes – build and evaluate (Hevner et al., 2004). The build process, as related to this research concentrates on the progression through which the theoretical e-Government strategy formulation framework is derived. Weick’s (1989) theorizing approach is ascribed, supporting the design of the theoretical framework. In applying Weick’s theorizing approach, firstly, the lack of knowledge on how e-Government strategies should be formulated in South African local municipalities – is explicated in an intelligible manner. Once the problem is properly articulated, a trial and error selection process is undertaken of existing approaches on strategy formulation – thought to possess the potential to contribute to the development of an e-Government strategy formulation framework, suited to local governments in South Africa. With good reason, 10 (ten) strategy approaches are selected from, e-Government programmes in developed countries, Non-Governmental organizational strategy approaches, and business related strategy formulation approaches. Lastly, as all possible approaches that may contribute to the framework development process cannot be selected, criteria is specified to limit the number of possible selections. Furthermore, in deriving the framework, foundations for systematically dealing with unstructured problems, such as, strategy formulation are consulted. This foundation along with the research goals, informs the development of a template used to comparatively analyse the 10 selected approaches on strategy formulation. This analysis aids in revealing the components of an e-Government strategy formulation process. With the developed framework, the evaluation process of the design science research commences, seeking to determine the utility of the framework (suitability and shortcomings). The framework is applied to the procedural formulation of a strategy for a tentative e-Government project called MobiSAM, which aims to enhance citizen engagement with local government through the use of mobile phones. The strategy formulation application process in the project environment and local government reveals lessons that inform revisions to the framework. The e-Government strategy formulation framework, therefore represents a fundamental tool for e-Government strategy development in local municipalities, and may be customized to fit the requirements of varying local municipalities. , Thesis (PhD) -- Faculty of Commerce, Information Systems, 2017
- Full Text:
- Date Issued: 2017
- Authors: Osah, Umeoniso Joshua
- Date: 2017
- Subjects: Internet in public administration South Africa , Electronic government information South Africa , Communication in public administration South Africa , Municipal services South Africa , Municipal services Information technology , System theory
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64471 , vital:28547 , DOI 10.21504/10962/64471
- Description: E-Government deployments by stakeholders within the South African water service sector, provide certain benefits for the sector. While deemed beneficial and of considerable value, e-Government deployments and implementations in the water sector of local governments of South Africa have not always been successful. One important reason for e-Government failures, among several others, is the lack of coherent strategies, informed by key representing stakeholder views, to guide implementation and deployment of e-Government tools. Without strategies, it is highly likely that ICT integration will be conducted haphazardly. As a point of note, more than the deliverable (strategy document) that represents the output of the strategy development process, it serves more purpose to understand the process that results in the strategy. Importantly, understanding the process helps to account for the formed relationships between the various stakeholders that need to buy into the strategy. The research study develops an e-Government strategy formulation framework based on a systems thinking approach, intended to support the strategy formulation process of e-Government strategies – to underpin the effective integration, deployment and sustained use of ICT solutions for water service delivery at the local government level. A systems thinking approach is considered due to its emphasis on the strategy being informed by a holistic assessment. Where there is some knowledge about the processes by which a strategy is formulated – over time ideas may be derived on the types of processes that may produce efficient e-Government strategies. The research is conducted using the Design Science research paradigm. The Design Science paradigm is comprised of two processes – build and evaluate (Hevner et al., 2004). The build process, as related to this research concentrates on the progression through which the theoretical e-Government strategy formulation framework is derived. Weick’s (1989) theorizing approach is ascribed, supporting the design of the theoretical framework. In applying Weick’s theorizing approach, firstly, the lack of knowledge on how e-Government strategies should be formulated in South African local municipalities – is explicated in an intelligible manner. Once the problem is properly articulated, a trial and error selection process is undertaken of existing approaches on strategy formulation – thought to possess the potential to contribute to the development of an e-Government strategy formulation framework, suited to local governments in South Africa. With good reason, 10 (ten) strategy approaches are selected from, e-Government programmes in developed countries, Non-Governmental organizational strategy approaches, and business related strategy formulation approaches. Lastly, as all possible approaches that may contribute to the framework development process cannot be selected, criteria is specified to limit the number of possible selections. Furthermore, in deriving the framework, foundations for systematically dealing with unstructured problems, such as, strategy formulation are consulted. This foundation along with the research goals, informs the development of a template used to comparatively analyse the 10 selected approaches on strategy formulation. This analysis aids in revealing the components of an e-Government strategy formulation process. With the developed framework, the evaluation process of the design science research commences, seeking to determine the utility of the framework (suitability and shortcomings). The framework is applied to the procedural formulation of a strategy for a tentative e-Government project called MobiSAM, which aims to enhance citizen engagement with local government through the use of mobile phones. The strategy formulation application process in the project environment and local government reveals lessons that inform revisions to the framework. The e-Government strategy formulation framework, therefore represents a fundamental tool for e-Government strategy development in local municipalities, and may be customized to fit the requirements of varying local municipalities. , Thesis (PhD) -- Faculty of Commerce, Information Systems, 2017
- Full Text:
- Date Issued: 2017
Assessing the potential role of microorganisms in the production of seedlings for the restoration of Albany Thicket
- Authors: Mpama, Nelisa
- Date: 2017
- Subjects: Vesicular-arbuscular mycorrhizas , Rhizobacteria , Restoration ecology South Africa Albany , Microorganisms
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64555 , vital:28558
- Description: The role of microorganisms in restoration of the Albany Thicket has not been well documented, although the benefits to plants of these various interactions has been well documented. Microorganisms are chief ecological engineers and assist in resolving environmental problems and act to restore degraded ecosystem function by forming mutual relationships with the roots of the plants. The aim of this study was to assess the potential of microorganisms for the improved biomass production of selected woody and succulent seedlings used in mesic thicket restoration. Three tree species were selected for propagation in this study namely; Mystroxylon aethiopicum Scutia myrtina and Aloe ferox. Soil samples were collected from a degraded and intact thicket site from Bathurst, South Africa. Soils were evaluated for number of arbuscular mycorrhizal (AM) spores, mycorrhizal infectivity potential and nutrient availability both before and after seedling propagation. Pasteurized soil from the degraded site was used in a pot trial. Ten replicates seedling for plant species were planted and subjected to four treatments which included inoculation with AM fungi and the rhizobacterium, Enterobacter sp., alone and in combination; the fourth treatment was an un-inouclated control. Plant growth parameters were recorded at regular intervals where appropriate and seedlings were harvested after 24 weeks for biomass measurements and AM colonisation assessments. Although generally low (< 1 spore per gram) the density of AM fungal spores was significantly higher in soils from the intact site when compared with soils from the degraded site. The mycorrhizal potential of the soils was however not significantly different. Mystroxylon aethiopicum seedling shoot height, canopy diameter and shoot biomass showed a significant increase when inoculated with AM fungi while S. myrtina seedlings showed increased shoot height when inoculated with both AM fungi and Enterobacter sp. Aloe ferox seedlings did not respond to microbial inoculation. The concentration of soil P and Na increased in treatments with Enterobacter sp. alone and in combination with AM fungi. Mystroxylon aethiopicum and S. myrtina seedlings showed a dependency on microbial inoculants indicating the importance of inoculation in the nursery before planting out into the field. Overall AM fungal inoculants applied to seedlings can be used to compensate for nutrient deficiency in soils. Although the Enterobacter isolate used was known to have various plant growth promoting capabilities. It is recommended that other rhizobacterial isolates be investigated. , Thesis (MSc) -- Faculty of Science, Environmental Science, 2017
- Full Text:
- Date Issued: 2017
- Authors: Mpama, Nelisa
- Date: 2017
- Subjects: Vesicular-arbuscular mycorrhizas , Rhizobacteria , Restoration ecology South Africa Albany , Microorganisms
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64555 , vital:28558
- Description: The role of microorganisms in restoration of the Albany Thicket has not been well documented, although the benefits to plants of these various interactions has been well documented. Microorganisms are chief ecological engineers and assist in resolving environmental problems and act to restore degraded ecosystem function by forming mutual relationships with the roots of the plants. The aim of this study was to assess the potential of microorganisms for the improved biomass production of selected woody and succulent seedlings used in mesic thicket restoration. Three tree species were selected for propagation in this study namely; Mystroxylon aethiopicum Scutia myrtina and Aloe ferox. Soil samples were collected from a degraded and intact thicket site from Bathurst, South Africa. Soils were evaluated for number of arbuscular mycorrhizal (AM) spores, mycorrhizal infectivity potential and nutrient availability both before and after seedling propagation. Pasteurized soil from the degraded site was used in a pot trial. Ten replicates seedling for plant species were planted and subjected to four treatments which included inoculation with AM fungi and the rhizobacterium, Enterobacter sp., alone and in combination; the fourth treatment was an un-inouclated control. Plant growth parameters were recorded at regular intervals where appropriate and seedlings were harvested after 24 weeks for biomass measurements and AM colonisation assessments. Although generally low (< 1 spore per gram) the density of AM fungal spores was significantly higher in soils from the intact site when compared with soils from the degraded site. The mycorrhizal potential of the soils was however not significantly different. Mystroxylon aethiopicum seedling shoot height, canopy diameter and shoot biomass showed a significant increase when inoculated with AM fungi while S. myrtina seedlings showed increased shoot height when inoculated with both AM fungi and Enterobacter sp. Aloe ferox seedlings did not respond to microbial inoculation. The concentration of soil P and Na increased in treatments with Enterobacter sp. alone and in combination with AM fungi. Mystroxylon aethiopicum and S. myrtina seedlings showed a dependency on microbial inoculants indicating the importance of inoculation in the nursery before planting out into the field. Overall AM fungal inoculants applied to seedlings can be used to compensate for nutrient deficiency in soils. Although the Enterobacter isolate used was known to have various plant growth promoting capabilities. It is recommended that other rhizobacterial isolates be investigated. , Thesis (MSc) -- Faculty of Science, Environmental Science, 2017
- Full Text:
- Date Issued: 2017
Comparative fish ecology in three periodically connected rivers in the upper Zambezi and Okavango ecoregions
- Authors: Taylor, Geraldine Claire
- Date: 2017
- Subjects: Fishes Ecology Zambia Zambezi District , Floodplain ecology Zambia Zambezi District , Stable isotopes , Fishes Food Zambia Zambezi District , Fishes Mortality Zambia Zambezi District , Fish populations Zambia Zambezi District , Fishes Growth
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/65055 , vital:28660 , DOI https://doi.org/10.21504/10962/65055
- Description: The Upper Zambezi, Kavango and Kwando rivers are three periodically interlinked floodplain rivers which share the same Upper Zambezian floodplain ichthyofauna. The aim of this thesis was to compare the biology and ecology of the fish communities in these three rivers. The objective was to test the hypothesis that fish community composition and assemblage structure, fish diets, food web structure and trophic dynamics, fish growth rates and total mortality are influenced by the differing flood magnitudes of the three rivers, in support of the flood pulse concept. To understand the abiotic characteristics of each river, water temperature, flood regime, total dissolved inorganic nitrogen concentrations and water quality parameters were measured and compared between rivers. Water temperatures varied seasonally, and seven day moving averages peaked above 30 °C in January, and fell to between 16 and 19 °C in June. The Zambezi River had the largest flood (6.14 m), followed by the Kavango River (3.80 m), while the Kwando River had the smallest flood (0.65 m). Total dissolved inorganic nitrogen concentrations were low in the Kavango and Zambezi Rivers (0.2 - 0.6 mg/l), and slightly higher in the Kwando River (<1 mg/l). Conductivity, total dissolved solids and total dissolved inorganic nitrogen concentrations decreased with the flood (dilution effect). Using biomass catch per unit effort data from experimental gillnets, fish community composition and assemblage structure was described, and differed between rivers in all hydrological seasons. In the Zambezi and Kavango rivers, fish assemblages varied with hydrological season as a result of the homogenising influence of the flood pulse, while in the Kwando River fish assemblages did not differ seasonally as flood pulses were small and often irregular. Differences in community composition were attributed to the abundance of Hydrocynus vittatus, a large bodied open water predator, in the Zambezi and Kavango rivers, and its relative absence in the Kwando River. Based on the results of the community composition, six focus species were chosen that were abundant and representative of the various feeding modes and life history strategies of the fish community. These were the striped robber Brycinus lateralis, sharptooth catfish Clarias gariepinus, blunttooth catfish Clarias ngamensis, African pike Hepsetus cuvieri, silver catfish Schilbe intermedius and purpleface largemouth Serranochromis macrocephalus. Stomach contents analysis was then used to compare the feeding ecology of the six example species between rivers. Clarias gariepinus, C. ngamensis and S. intermedius were piscivorous in the Zambezi and Kavango rivers, and preyed upon more invertebrates in the Kwando River, while Hepsetus cuvieri and S. macrocephalus were piscivorous in all three rivers. Differences in diets were attributed to seasonal prey abundance, with prey fishes abundant during falling and low water when the Zambezi and Kavango rivers were sampled, while invertebrates were abundant during rising and high water when the Kwando River was sampled. Prey mastication by B. lateralis made prey identification difficult. For other predators, the usefulness of stomach contents analysis for dietary descriptions was restricted by the high proportion of empty stomachs. As a result, whole ecosystem stable isotope analysis was used to gain a holistic understanding of the food web structure and fish feeding ecology of the three rivers. The Zambezi and Kavango river food webs were supported by C enriched resources such as C4 and C3 riparian vegetation from the floodplain, while the Kwando River food web was based on C depleted resources such as filamentous algae and aquatic macrophytes. The Zambezi River food web had a restricted nitrogen range, with reduced food chain length and the predators in this river did not occupy such elevated trophic positions compared to in the Kavango and Kwando river food webs. This was attributed to the overfishing of the primary and tertiary consumers in the Zambezi River, a phenomenon known to reduce food chain length. Focussing on predator communities, in the Zambezi and Kavango rivers, H. vittatus isotopic niche width was large and overlapped significantly with most other predators, while in the Kwando River predator niches were more distinct. This supported previously proposed hypotheses by describing H. vittatus as a dominant predator which excludes all other fishes by predation or competition. Despite the dominance of H. vittatus, C. gariepinus occupied the position of top predator in all three rivers, and information on the habitat use, feeding habits and trophic niches of the serranochromine cichlids added understanding of their ecology. Lastly, age was determined using sectioned sagittal otoliths for C. gariepinus, C. ngamensis, S. intermedius and S. macrocephalus and using whole asteriscus otoliths for B. lateralis and H. cuvieri, and growth was modelled using the von Bertalanffy growth equation. Growth performance was high in the Zambezi and Kavango rivers, and lower in the Kwando River, most likely in response to the varying flood magnitudes. Total mortality rates, estimated using Hoenig’s maximum-age based equation, were high in the Zambezi River as a result of the high fishing pressure on this river. Overall floodplain fish ecology in the Zambezi, Kavango and Kwando rivers was influenced by the flood pulse, as was predicted by the flood pulse concept. Periodic and equilibrium life history strategists were found to adapt either to the pulsing environments of the Zambezi and Kavango rivers, or to the more stable environment of the Kwando River, and large bodied, long lived periodic strategists such as C. gariepinus tended to be highly plastic and able to thrive in most conditions. Data also suggested that Zambezi River food web structure and fish mortality rates have been impacted by overfishing, for which more information is needed to conserve and manage this system. , Thesis (PhD) -- Faculty of Science, Ichthyology and Fisheries Science, 2018
- Full Text:
- Date Issued: 2017
- Authors: Taylor, Geraldine Claire
- Date: 2017
- Subjects: Fishes Ecology Zambia Zambezi District , Floodplain ecology Zambia Zambezi District , Stable isotopes , Fishes Food Zambia Zambezi District , Fishes Mortality Zambia Zambezi District , Fish populations Zambia Zambezi District , Fishes Growth
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/65055 , vital:28660 , DOI https://doi.org/10.21504/10962/65055
- Description: The Upper Zambezi, Kavango and Kwando rivers are three periodically interlinked floodplain rivers which share the same Upper Zambezian floodplain ichthyofauna. The aim of this thesis was to compare the biology and ecology of the fish communities in these three rivers. The objective was to test the hypothesis that fish community composition and assemblage structure, fish diets, food web structure and trophic dynamics, fish growth rates and total mortality are influenced by the differing flood magnitudes of the three rivers, in support of the flood pulse concept. To understand the abiotic characteristics of each river, water temperature, flood regime, total dissolved inorganic nitrogen concentrations and water quality parameters were measured and compared between rivers. Water temperatures varied seasonally, and seven day moving averages peaked above 30 °C in January, and fell to between 16 and 19 °C in June. The Zambezi River had the largest flood (6.14 m), followed by the Kavango River (3.80 m), while the Kwando River had the smallest flood (0.65 m). Total dissolved inorganic nitrogen concentrations were low in the Kavango and Zambezi Rivers (0.2 - 0.6 mg/l), and slightly higher in the Kwando River (<1 mg/l). Conductivity, total dissolved solids and total dissolved inorganic nitrogen concentrations decreased with the flood (dilution effect). Using biomass catch per unit effort data from experimental gillnets, fish community composition and assemblage structure was described, and differed between rivers in all hydrological seasons. In the Zambezi and Kavango rivers, fish assemblages varied with hydrological season as a result of the homogenising influence of the flood pulse, while in the Kwando River fish assemblages did not differ seasonally as flood pulses were small and often irregular. Differences in community composition were attributed to the abundance of Hydrocynus vittatus, a large bodied open water predator, in the Zambezi and Kavango rivers, and its relative absence in the Kwando River. Based on the results of the community composition, six focus species were chosen that were abundant and representative of the various feeding modes and life history strategies of the fish community. These were the striped robber Brycinus lateralis, sharptooth catfish Clarias gariepinus, blunttooth catfish Clarias ngamensis, African pike Hepsetus cuvieri, silver catfish Schilbe intermedius and purpleface largemouth Serranochromis macrocephalus. Stomach contents analysis was then used to compare the feeding ecology of the six example species between rivers. Clarias gariepinus, C. ngamensis and S. intermedius were piscivorous in the Zambezi and Kavango rivers, and preyed upon more invertebrates in the Kwando River, while Hepsetus cuvieri and S. macrocephalus were piscivorous in all three rivers. Differences in diets were attributed to seasonal prey abundance, with prey fishes abundant during falling and low water when the Zambezi and Kavango rivers were sampled, while invertebrates were abundant during rising and high water when the Kwando River was sampled. Prey mastication by B. lateralis made prey identification difficult. For other predators, the usefulness of stomach contents analysis for dietary descriptions was restricted by the high proportion of empty stomachs. As a result, whole ecosystem stable isotope analysis was used to gain a holistic understanding of the food web structure and fish feeding ecology of the three rivers. The Zambezi and Kavango river food webs were supported by C enriched resources such as C4 and C3 riparian vegetation from the floodplain, while the Kwando River food web was based on C depleted resources such as filamentous algae and aquatic macrophytes. The Zambezi River food web had a restricted nitrogen range, with reduced food chain length and the predators in this river did not occupy such elevated trophic positions compared to in the Kavango and Kwando river food webs. This was attributed to the overfishing of the primary and tertiary consumers in the Zambezi River, a phenomenon known to reduce food chain length. Focussing on predator communities, in the Zambezi and Kavango rivers, H. vittatus isotopic niche width was large and overlapped significantly with most other predators, while in the Kwando River predator niches were more distinct. This supported previously proposed hypotheses by describing H. vittatus as a dominant predator which excludes all other fishes by predation or competition. Despite the dominance of H. vittatus, C. gariepinus occupied the position of top predator in all three rivers, and information on the habitat use, feeding habits and trophic niches of the serranochromine cichlids added understanding of their ecology. Lastly, age was determined using sectioned sagittal otoliths for C. gariepinus, C. ngamensis, S. intermedius and S. macrocephalus and using whole asteriscus otoliths for B. lateralis and H. cuvieri, and growth was modelled using the von Bertalanffy growth equation. Growth performance was high in the Zambezi and Kavango rivers, and lower in the Kwando River, most likely in response to the varying flood magnitudes. Total mortality rates, estimated using Hoenig’s maximum-age based equation, were high in the Zambezi River as a result of the high fishing pressure on this river. Overall floodplain fish ecology in the Zambezi, Kavango and Kwando rivers was influenced by the flood pulse, as was predicted by the flood pulse concept. Periodic and equilibrium life history strategists were found to adapt either to the pulsing environments of the Zambezi and Kavango rivers, or to the more stable environment of the Kwando River, and large bodied, long lived periodic strategists such as C. gariepinus tended to be highly plastic and able to thrive in most conditions. Data also suggested that Zambezi River food web structure and fish mortality rates have been impacted by overfishing, for which more information is needed to conserve and manage this system. , Thesis (PhD) -- Faculty of Science, Ichthyology and Fisheries Science, 2018
- Full Text:
- Date Issued: 2017
Contributions to the study of nonholonomic Riemannian manifolds
- Authors: Barrett, Dennis Ian
- Date: 2017
- Subjects: Riemannian manifolds , Curvature , Lie groups , Geometry, Riemannian , Tensor fields
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/7554 , vital:21272 , DOI https://doi.org/10.21504/10962/7554
- Description: In this thesis we consider nonholonomic Riemannian manifolds, and in particular, left- invariant nonholonomic Riemannian structures on Lie groups. These structures are closely related to mechanical systems with (positive definite) quadratic Lagrangians and nonholo- nomic constraints linear in velocities. In the first chapter, we review basic concepts of non- holonomic Riemannian geometry, including the left-invariant structures. We also examine the class of left-invariant structures with so-called Cartan-Schouten connections. The second chapter investigates the curvature of nonholonomic Riemannian manifolds and the Schouten and Wagner curvature tensors. The Schouten tensor is canonically associated to every non- holonomic Riemannian structure (in particular, we use it to define isometric invariants for structures on three-dimensional manifolds). By contrast, the Wagner tensor is not generally intrinsic, but can be used to characterise flat structures (i.e., those whose associated parallel transport is path-independent). The third chapter considers equivalence of nonholonomic Rie- mannian manifolds, particularly up to nonholonomic isometry. We also introduce the notion of a nonholonomic Riemannian submanifold, and investigate the conditions under which such a submanifold inherits its geometry from the enveloping space. The latter problem involves the concept of a geodesically invariant distribution, and we show it is also related to the curvature. In the last chapter we specialise to three-dimensional nonholonomic Riemannian manifolds. We consider the equivalence of such structures up to nonholonomic isometry and rescaling, and classify the left-invariant structures on the (three-dimensional) simply connected Lie groups. We also characterise the flat structures in three dimensions, and then classify the flat structures on the simply connected Lie groups. Lastly, we consider three typical examples of (left-invariant) nonholonomic Riemannian structures on three-dimensional Lie groups, two of which arise from problems in classical mechanics (viz., the Chaplygin problem and the Suslov problem). , Thesis (PhD) -- Faculty of Science, Mathematics (Pure and Applied), 2017
- Full Text:
- Date Issued: 2017
- Authors: Barrett, Dennis Ian
- Date: 2017
- Subjects: Riemannian manifolds , Curvature , Lie groups , Geometry, Riemannian , Tensor fields
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/7554 , vital:21272 , DOI https://doi.org/10.21504/10962/7554
- Description: In this thesis we consider nonholonomic Riemannian manifolds, and in particular, left- invariant nonholonomic Riemannian structures on Lie groups. These structures are closely related to mechanical systems with (positive definite) quadratic Lagrangians and nonholo- nomic constraints linear in velocities. In the first chapter, we review basic concepts of non- holonomic Riemannian geometry, including the left-invariant structures. We also examine the class of left-invariant structures with so-called Cartan-Schouten connections. The second chapter investigates the curvature of nonholonomic Riemannian manifolds and the Schouten and Wagner curvature tensors. The Schouten tensor is canonically associated to every non- holonomic Riemannian structure (in particular, we use it to define isometric invariants for structures on three-dimensional manifolds). By contrast, the Wagner tensor is not generally intrinsic, but can be used to characterise flat structures (i.e., those whose associated parallel transport is path-independent). The third chapter considers equivalence of nonholonomic Rie- mannian manifolds, particularly up to nonholonomic isometry. We also introduce the notion of a nonholonomic Riemannian submanifold, and investigate the conditions under which such a submanifold inherits its geometry from the enveloping space. The latter problem involves the concept of a geodesically invariant distribution, and we show it is also related to the curvature. In the last chapter we specialise to three-dimensional nonholonomic Riemannian manifolds. We consider the equivalence of such structures up to nonholonomic isometry and rescaling, and classify the left-invariant structures on the (three-dimensional) simply connected Lie groups. We also characterise the flat structures in three dimensions, and then classify the flat structures on the simply connected Lie groups. Lastly, we consider three typical examples of (left-invariant) nonholonomic Riemannian structures on three-dimensional Lie groups, two of which arise from problems in classical mechanics (viz., the Chaplygin problem and the Suslov problem). , Thesis (PhD) -- Faculty of Science, Mathematics (Pure and Applied), 2017
- Full Text:
- Date Issued: 2017
Outcome and impact assessment in the comprehensive evaluation of rural ICT projects in developing countries
- Mtkoko, Hafeni Tulimewawa Wilhelmina Lyatenda
- Authors: Mtkoko, Hafeni Tulimewawa Wilhelmina Lyatenda
- Date: 2017
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64483 , vital:28548 , DOI 10.21504/10962/64483
- Description: Despite existing literature that indicates that Information and communication technologies (ICTs) act as tools for social change and development, there is still limited empirical evidence that demonstrates this. An outcome and impact assessment based on a comprehensive (holistic) evaluation is deemed appropriate at a time when many ICT4D programmes fail to effectively demonstrate their impact towards rural development. A comprehensive evaluation is one that incorporates the evaluation of the need, design, implementation, outcome and impact, efficiency, and scalability of a programme. These stages make up the different domains of an evaluation lifecycle. The following study aims to develop an outcome and impact assessment framework for ICT4D programmes. It forms part of continuous research associated with the development of a rural ICT Comprehensive Evaluation framework. A theoretical approach, using Design Science and Weick’s theorizing process, was applied to investigate the development of a framework (design artefact) for outcome and impact assessment of ICT4D programmes. The theorizing process analysed existing outcome and impact assessment frameworks from social programmes, information systems/technology programmes, and ICT4D programmes. The output of the theorizing process proposes five critical themes of outcome and impact assessment of rural ICT4D programmes that should be assessed. These themes include: Strategic Value, Most Significant Change, Empowerment, Livelihoods, and Sustainability. To assess its utility, the framework was implemented in the Siyakhula Living Lab and Information and Communication Technology for Rural Education (ICT4RED) projects in South Africa. Through the application of the framework in real life ICT4D contexts, the lessons learned contributed to its revision and enhancement. The proposed framework aims to guide evaluators through the assessment of outcomes and impacts in ICT4D programmes. It provides a foundation and justification for the selected outcome and impact assessment themes that contribute to a comprehensive evaluation. An outcome and impact assessment that is informed by: baseline; needs assessment; programme theory assessment; and process assessment data, provides ICT4D evaluators and project stakeholders with meaningful outcome and impact feedback. Having such an approach to outcome and impact assessment ensures that the evaluation process is seen more holistically as part of the ICT4D project as a whole. , Thesis (PhD) -- Faculty of Commerce, Information Systems, 2017
- Full Text:
- Date Issued: 2017
- Authors: Mtkoko, Hafeni Tulimewawa Wilhelmina Lyatenda
- Date: 2017
- Subjects: Uncatalogued
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64483 , vital:28548 , DOI 10.21504/10962/64483
- Description: Despite existing literature that indicates that Information and communication technologies (ICTs) act as tools for social change and development, there is still limited empirical evidence that demonstrates this. An outcome and impact assessment based on a comprehensive (holistic) evaluation is deemed appropriate at a time when many ICT4D programmes fail to effectively demonstrate their impact towards rural development. A comprehensive evaluation is one that incorporates the evaluation of the need, design, implementation, outcome and impact, efficiency, and scalability of a programme. These stages make up the different domains of an evaluation lifecycle. The following study aims to develop an outcome and impact assessment framework for ICT4D programmes. It forms part of continuous research associated with the development of a rural ICT Comprehensive Evaluation framework. A theoretical approach, using Design Science and Weick’s theorizing process, was applied to investigate the development of a framework (design artefact) for outcome and impact assessment of ICT4D programmes. The theorizing process analysed existing outcome and impact assessment frameworks from social programmes, information systems/technology programmes, and ICT4D programmes. The output of the theorizing process proposes five critical themes of outcome and impact assessment of rural ICT4D programmes that should be assessed. These themes include: Strategic Value, Most Significant Change, Empowerment, Livelihoods, and Sustainability. To assess its utility, the framework was implemented in the Siyakhula Living Lab and Information and Communication Technology for Rural Education (ICT4RED) projects in South Africa. Through the application of the framework in real life ICT4D contexts, the lessons learned contributed to its revision and enhancement. The proposed framework aims to guide evaluators through the assessment of outcomes and impacts in ICT4D programmes. It provides a foundation and justification for the selected outcome and impact assessment themes that contribute to a comprehensive evaluation. An outcome and impact assessment that is informed by: baseline; needs assessment; programme theory assessment; and process assessment data, provides ICT4D evaluators and project stakeholders with meaningful outcome and impact feedback. Having such an approach to outcome and impact assessment ensures that the evaluation process is seen more holistically as part of the ICT4D project as a whole. , Thesis (PhD) -- Faculty of Commerce, Information Systems, 2017
- Full Text:
- Date Issued: 2017
The Role of HOP in Emerin-Mediated Nuclear Structure
- Authors: Kituyi, Sarah Naulikha
- Date: 2017
- Subjects: Heat shock proteins , Nuclear structure , Nuclear membranes , Cancer Treatment , Molecular chaperones , Cytoskeleton , Cytoplasm
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/59230 , vital:27485 , DOI 10.21504/10962/59230
- Description: A vital component of the integral nuclear membrane is emerin, a Lamin Emerin and Man1 (LEM) domain protein whose concentration determines the levels of partner proteins that together constitute the structure of the nuclear envelope. Deficiencies in any of these proteins causes the failure of the structure and assembly and disassembly of the nuclear envelope, which disrupts chromosome segregation and nuclear compartmentalization that are both associated with disease. Emerin also localizes in the cytoplasm where it is implicated in the structure of the cytoskeleton via interaction with tubulin and actin and thus its deficiency may equally contribute to the collapse of the cytoskeleton. The Hsp70-Hsp90 organising protein (Hop) functions as a cochaperone for entry of client proteins into the Hsp90 folding cycle. Hop is upregulated in cancer and regulates a number of cell biology processes via interactions with proteins independently of Hsp90. In a previous study using global whole cell mass spectrometry, emerin was shown to be the most significantly down regulated protein in Hop depleted cell lysates. In this current study, it was postulated that emerin interacts with Hop, and this interaction regulates the stability, and level of emerin in the nucleus which impacts on the structure of the nuclear envelope. We used HEK293T cell lines stably expressing shRNA against Hop, emerin and a non-targeting control alongside the over expression of Hop in HEK293 cells to determine the effect of Hop levels on emerin expression and vice versa via Western blotting. The effect of Hop on the localization of emerin was assessed via subcellullar fractionation and confocal microscopy, while the impact on the structure of the nucleus was determined by transmission electron microscopy (TEM). We established that the depletion of Hop using shRNA and the over expression of Hop both result in the proteasomal and lysosomal degradation of emerin. Co-immunoprecipitation assays confirmed that Hop and emerin are in a common complex, which was not dependent on the presence of Hsp90. Loss of Hop or emerin led to a deformation of nuclear structure and a statistically significant decrease in nuclear size compared to control cells and was associated with an increase in the levels of nuclear protein, lamin A-C. Loss of emerin and Hop resulted in increased long term cell survival, but only after restriction of the nucleus when the cells had migrated across a transwell membrane. Taken together, the results obtained suggest that Hop acts as a scaffold for the stabilization of emerin and that the effects of Hop depletion on the structure of the nucleus and long term survival are mediated via the depletion of emerin. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2017
- Full Text:
- Date Issued: 2017
- Authors: Kituyi, Sarah Naulikha
- Date: 2017
- Subjects: Heat shock proteins , Nuclear structure , Nuclear membranes , Cancer Treatment , Molecular chaperones , Cytoskeleton , Cytoplasm
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/59230 , vital:27485 , DOI 10.21504/10962/59230
- Description: A vital component of the integral nuclear membrane is emerin, a Lamin Emerin and Man1 (LEM) domain protein whose concentration determines the levels of partner proteins that together constitute the structure of the nuclear envelope. Deficiencies in any of these proteins causes the failure of the structure and assembly and disassembly of the nuclear envelope, which disrupts chromosome segregation and nuclear compartmentalization that are both associated with disease. Emerin also localizes in the cytoplasm where it is implicated in the structure of the cytoskeleton via interaction with tubulin and actin and thus its deficiency may equally contribute to the collapse of the cytoskeleton. The Hsp70-Hsp90 organising protein (Hop) functions as a cochaperone for entry of client proteins into the Hsp90 folding cycle. Hop is upregulated in cancer and regulates a number of cell biology processes via interactions with proteins independently of Hsp90. In a previous study using global whole cell mass spectrometry, emerin was shown to be the most significantly down regulated protein in Hop depleted cell lysates. In this current study, it was postulated that emerin interacts with Hop, and this interaction regulates the stability, and level of emerin in the nucleus which impacts on the structure of the nuclear envelope. We used HEK293T cell lines stably expressing shRNA against Hop, emerin and a non-targeting control alongside the over expression of Hop in HEK293 cells to determine the effect of Hop levels on emerin expression and vice versa via Western blotting. The effect of Hop on the localization of emerin was assessed via subcellullar fractionation and confocal microscopy, while the impact on the structure of the nucleus was determined by transmission electron microscopy (TEM). We established that the depletion of Hop using shRNA and the over expression of Hop both result in the proteasomal and lysosomal degradation of emerin. Co-immunoprecipitation assays confirmed that Hop and emerin are in a common complex, which was not dependent on the presence of Hsp90. Loss of Hop or emerin led to a deformation of nuclear structure and a statistically significant decrease in nuclear size compared to control cells and was associated with an increase in the levels of nuclear protein, lamin A-C. Loss of emerin and Hop resulted in increased long term cell survival, but only after restriction of the nucleus when the cells had migrated across a transwell membrane. Taken together, the results obtained suggest that Hop acts as a scaffold for the stabilization of emerin and that the effects of Hop depletion on the structure of the nucleus and long term survival are mediated via the depletion of emerin. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2017
- Full Text:
- Date Issued: 2017
Synthesis and biolgical screening of potential plasmodium falciparum DXR inhibitors
- Authors: Adeyemi, Christiana Modupe
- Date: 2017-04
- Subjects: Plasmodium falciparum , Enzyme inhibitors , Malaria , Antimalarials , Drug development , Malaria -- Chemotherapy , Isopentenoids -- Synthesis , Fosmidomycin , 1-Deoxy-D-xylulose 5-phosphate
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61790 , vital:28060
- Description: The non-mevalonate isoprenoid pathway, also known as the 1-deoxy-D-xylulose-5- phosphate DXP pathway, is absent in humans, but present in the anopheles mosquito responsible for the transmission of malaria. DXP reductoisomerase - a key enzyme in the DXP pathway in Plasmodium falciparum (PfDXR) has been identified as a target for the design of novel anti-malarial drugs. Fosmidomycin and its acetyl analogue (FR900098) are known to be inhibitors of PfDXR and, in this study, synthetic variations of the fosmidomycin scaffold have led to four series of novel analogues. Particular attention has been centred on the introduction of various substituted benzyl groups in each of these series in order to occupy a recently discovered vacant pocket in the PfDXR active-site and thus enhance ligand-enzyme binding. In the process 160 ligands and precursors have been prepared, no less than 119 of them novel. Fistly, a series of C-benzylated phosphonate esters and phosphonic acids were synthesised, in which the fosmidomycin hydroxamate Mg2+- coordinating moiety was replaced by an amide funtionality and the number of methylene groups in the “hydrophobic patch” between the phosphonate and the hydroxamate moiety was decreased from two to one. Several approaches were explored for this series, the most successful involving reaction of 3- substituted anilines with a-bromo propanoic acid in the presence of the coupling agent 1,1'- carbonyldiimidazole (CDI), followed by Michaelis-Arbuzov phosphonation using triethyl phosphite. Reaction of the resulting chiral phosphonate esters with bromotrimethylsilane gave the corresponding phosphonic acids in good yields. In order to obviate chirality issues, a second series of potential “reverse” fosmidomycin analogues was synthesised by replacing the methylene group adjacent to the the phosphonate moiety with a nitrogen atom. Deprotonation, alkylation and phosphorylation of various amines gave diethyl #-benzylphosphoramidate ester intermediate. Aza-Michael addition of these intermediates, followed by hydrolysis gave the corresponding carboxylic acids which could be reacted with different hydroxylamine hydrochloride derivatives to afford the novel hydroxamic acid derivatives in good yields. Thirdly, a series of a novel #-benzylated phosphoramidate derivatives were prepared as aza- FR900098 analogues. Alkylation of different amines using bromoacetalde-hyde diethylacetal gave a series of N-benzyl-2,2-diethoxyethylamine compounds, which were then elaborated via a futher six steps to afford novel #-benzylated phosphoramidate derivatives. Finally, in order to ensure syn-orientation of the donor atoms in the Mg - coordinating group and, at the same time, introduce conformational constraints in the ligand, the hydrophobic patch and the hydroxamate moiety were replaced by cyclic systems. Several approaches towards the synthesis of such conformationally constrained phosphoramidate analogues from maleic anhydride led to the unexpected isolation of an unprecedented acyclic furfuryl compound, and 1H NMR and DFT-level theoretical studies have been initiated to explore the reaction sequence. A series of #-benzylated phosphoramidate derivatives containing dihydroxy aromatic rings (as the conformationally constrained groups) to replace the hydroxamate moiety, were successfully obtained in six steps from the starting material, 3,4-dihydroxylbenzaldehyde. While in vitro assays have been conducted on all of the synthesised compounds, and some of the ligands show promising anti-malarial inhibitory activity - most especially the conformationally constrained cyclic #-benzylated phosphoramidate series. Interestingly, a number of these compounds has also shown activity against T.brucei - the causative agent of sleeping sickness. In silico docking studies of selected compounds has revealed the capacity of some of the ligands to bind effectively in the PfDXR active-site with the newly introduced benzyl group occupying the adjacent vacant pocket. The physico-chemical properties of these ligands were also explored in order to predict the oral-bioavailability. Most of the ligands obeyed the Lipinski rule of 5, while QSAR methods have been used in an attempt to correlate structural variations and calculated molecular properties with the bioassay data. , Thesis (PhD) -- Faculty of Science, Chemistry, 2017
- Full Text:
- Date Issued: 2017-04
- Authors: Adeyemi, Christiana Modupe
- Date: 2017-04
- Subjects: Plasmodium falciparum , Enzyme inhibitors , Malaria , Antimalarials , Drug development , Malaria -- Chemotherapy , Isopentenoids -- Synthesis , Fosmidomycin , 1-Deoxy-D-xylulose 5-phosphate
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61790 , vital:28060
- Description: The non-mevalonate isoprenoid pathway, also known as the 1-deoxy-D-xylulose-5- phosphate DXP pathway, is absent in humans, but present in the anopheles mosquito responsible for the transmission of malaria. DXP reductoisomerase - a key enzyme in the DXP pathway in Plasmodium falciparum (PfDXR) has been identified as a target for the design of novel anti-malarial drugs. Fosmidomycin and its acetyl analogue (FR900098) are known to be inhibitors of PfDXR and, in this study, synthetic variations of the fosmidomycin scaffold have led to four series of novel analogues. Particular attention has been centred on the introduction of various substituted benzyl groups in each of these series in order to occupy a recently discovered vacant pocket in the PfDXR active-site and thus enhance ligand-enzyme binding. In the process 160 ligands and precursors have been prepared, no less than 119 of them novel. Fistly, a series of C-benzylated phosphonate esters and phosphonic acids were synthesised, in which the fosmidomycin hydroxamate Mg2+- coordinating moiety was replaced by an amide funtionality and the number of methylene groups in the “hydrophobic patch” between the phosphonate and the hydroxamate moiety was decreased from two to one. Several approaches were explored for this series, the most successful involving reaction of 3- substituted anilines with a-bromo propanoic acid in the presence of the coupling agent 1,1'- carbonyldiimidazole (CDI), followed by Michaelis-Arbuzov phosphonation using triethyl phosphite. Reaction of the resulting chiral phosphonate esters with bromotrimethylsilane gave the corresponding phosphonic acids in good yields. In order to obviate chirality issues, a second series of potential “reverse” fosmidomycin analogues was synthesised by replacing the methylene group adjacent to the the phosphonate moiety with a nitrogen atom. Deprotonation, alkylation and phosphorylation of various amines gave diethyl #-benzylphosphoramidate ester intermediate. Aza-Michael addition of these intermediates, followed by hydrolysis gave the corresponding carboxylic acids which could be reacted with different hydroxylamine hydrochloride derivatives to afford the novel hydroxamic acid derivatives in good yields. Thirdly, a series of a novel #-benzylated phosphoramidate derivatives were prepared as aza- FR900098 analogues. Alkylation of different amines using bromoacetalde-hyde diethylacetal gave a series of N-benzyl-2,2-diethoxyethylamine compounds, which were then elaborated via a futher six steps to afford novel #-benzylated phosphoramidate derivatives. Finally, in order to ensure syn-orientation of the donor atoms in the Mg - coordinating group and, at the same time, introduce conformational constraints in the ligand, the hydrophobic patch and the hydroxamate moiety were replaced by cyclic systems. Several approaches towards the synthesis of such conformationally constrained phosphoramidate analogues from maleic anhydride led to the unexpected isolation of an unprecedented acyclic furfuryl compound, and 1H NMR and DFT-level theoretical studies have been initiated to explore the reaction sequence. A series of #-benzylated phosphoramidate derivatives containing dihydroxy aromatic rings (as the conformationally constrained groups) to replace the hydroxamate moiety, were successfully obtained in six steps from the starting material, 3,4-dihydroxylbenzaldehyde. While in vitro assays have been conducted on all of the synthesised compounds, and some of the ligands show promising anti-malarial inhibitory activity - most especially the conformationally constrained cyclic #-benzylated phosphoramidate series. Interestingly, a number of these compounds has also shown activity against T.brucei - the causative agent of sleeping sickness. In silico docking studies of selected compounds has revealed the capacity of some of the ligands to bind effectively in the PfDXR active-site with the newly introduced benzyl group occupying the adjacent vacant pocket. The physico-chemical properties of these ligands were also explored in order to predict the oral-bioavailability. Most of the ligands obeyed the Lipinski rule of 5, while QSAR methods have been used in an attempt to correlate structural variations and calculated molecular properties with the bioassay data. , Thesis (PhD) -- Faculty of Science, Chemistry, 2017
- Full Text:
- Date Issued: 2017-04
Bioprospecting for amylases, cellulases and xylanases from ericoid associated fungi, their production and characterisation for the bio-economy
- Authors: Adeoyo, Olusegun Richard
- Date: 2018
- Subjects: Mycorrhizal fungi , Hydrolases , Ericaceae South Africa , Ericaceae Molecular aspects
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64327 , vital:28533
- Description: South Africa is one of the most productive areas for ericaceous plants with about 850 identified species in the Cape Floral Region. The Albany Centre of Endemism where all fungi used in this study were isolated from, falls within this region. Ericaceous plants interact with some fungi via an association called the ericoid mycorrhizal (ERM) association. All fungi used in this study were isolated from roots of six ericaceous plants; Erica cerinthoides, Erica demissa, Erica chamissonis, Erica glumiflora, Erica caffra and Erica nemorosa. Fungal enzymes are known to play a significant role in the food, brewing, detergent, pharmaceutical and biofuel industries. The enzyme industry is among the major sectors of the world, and additional novel sources are being explored from time to time. This study focussed on amylases (amyloglucosidase, AMG), cellulases (endoglucanase) and xylanases (endo-1,4-P-xylanase) production from ERM fungal isolates. Out of the fifty-one (51), fungal isolates screened, ChemRU330 (Leohumicola sp.), EdRU083 and EdRU002 were among the fungi that had the highest activities of all the enzymes. They were tested for the ability to produce amylases and cellulases under different pH and nutritional conditions that included: carbon sources, nitrogen sources and metal ions, at an optimum temperature of 28°C in a modified Melin-Norkrans (MMN) liquid medium. Cellulase specific activity of 3.99, 2.18 and 4.31 (U/mg protein) for isolates EdRU083, EdRU002 and ChemRU330, respectively, was produced at an optimal pH of 5.0. For amylase, ChemRU330 had the highest specific activity of 1.11 U/mg protein while EdRU083 and EdRU02 had a specific activity of 0.80 and 0.92 U/mg protein, respectively, at the same pH with corresponding biomass yield of 113, 125 and 97 mg/50 ml, respectively. Increased enzyme activities and improved mycelial biomass production were obtained in the presence of supplements such as potassium, sodium, glucose, maltose, cellobiose, tryptone and peptone, while NaFe-EDTA and cobalt inhibited enzyme activity. ChemRU330 was selected to determine the consistency and amount of amylase, cellulase and xylanase formed after several in vitro subculturing events. AMG and endo-1,4-P-xylanase were found to have the most consistent production throughout the study period. The AMG was stable at 45oC (pH 5.0), retaining approximately 65% activity over a period of 24 h. The molecular mass of AMG and endo-1,4-P-xylanase were estimated to be 101 kDa and 72 kDa, respectively. The Km and kcat were 0.38 mg/ml and 70 s-1, respectively, using soluble starch (AMG). For endo-1,4-P-xylanase, the Km and Vmax were 0.93 mg/ml and 8.54 U/ml, respectively, using beechwood xylan (endo-1,4-P-xylanase) as substrate. Additionally, crude extracts of five root endophytes with unique morphological characteristics were screened for antibacterial properties and was followed by determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). L. incrustata (ChemRU330) and Chaetomium sp. extracts exhibited varying degrees of inhibition against two Gram-positive and Gram-negative bacteria. The crude extract of L. incrustata was the most effective which was found to inhibit Staphylococcus aureus (MIC: 1 mg/ml), Bacillus subtilis (MIC: 2 mg/ml) and Proteus vulgaris (MIC: 16 mg/ml). The L. incrustata displayed potential for antibacterial production and could be considered as an additional source of new antimicrobial agents in drug and food preservation. Also, the three isolates used for enzyme production were identified to genus and species levels, i.e., Leohumicola incrustata (ChemRU330), Leohumicola sp. (EdRU083) and Oidiodendron sp. (EdRU002) using both ITS and Cox1 DNA regions. The molecular analysis results indicated that these ERM mycorrhizal fungi were similar to those successfully described by some researchers in South Africa and Australia. Therefore, this study opens new opportunities for exploring ERM fungal biomolecules for the bio-economy. The promising physicochemical properties, starch and xylan hydrolysis end- products, and being non-pathogenic make AMG and endo-1,4-P-xylanase potential candidates for future applications as additives in the food industry for the production of glucose, glucose syrups, high-fructose corn syrups, and as well as the production of bioethanol. Finally, the findings of this study revealed that it is possible to produce hydrolytic enzymes from ERM fungi in vitro using chemically defined media. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018
- Authors: Adeoyo, Olusegun Richard
- Date: 2018
- Subjects: Mycorrhizal fungi , Hydrolases , Ericaceae South Africa , Ericaceae Molecular aspects
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/64327 , vital:28533
- Description: South Africa is one of the most productive areas for ericaceous plants with about 850 identified species in the Cape Floral Region. The Albany Centre of Endemism where all fungi used in this study were isolated from, falls within this region. Ericaceous plants interact with some fungi via an association called the ericoid mycorrhizal (ERM) association. All fungi used in this study were isolated from roots of six ericaceous plants; Erica cerinthoides, Erica demissa, Erica chamissonis, Erica glumiflora, Erica caffra and Erica nemorosa. Fungal enzymes are known to play a significant role in the food, brewing, detergent, pharmaceutical and biofuel industries. The enzyme industry is among the major sectors of the world, and additional novel sources are being explored from time to time. This study focussed on amylases (amyloglucosidase, AMG), cellulases (endoglucanase) and xylanases (endo-1,4-P-xylanase) production from ERM fungal isolates. Out of the fifty-one (51), fungal isolates screened, ChemRU330 (Leohumicola sp.), EdRU083 and EdRU002 were among the fungi that had the highest activities of all the enzymes. They were tested for the ability to produce amylases and cellulases under different pH and nutritional conditions that included: carbon sources, nitrogen sources and metal ions, at an optimum temperature of 28°C in a modified Melin-Norkrans (MMN) liquid medium. Cellulase specific activity of 3.99, 2.18 and 4.31 (U/mg protein) for isolates EdRU083, EdRU002 and ChemRU330, respectively, was produced at an optimal pH of 5.0. For amylase, ChemRU330 had the highest specific activity of 1.11 U/mg protein while EdRU083 and EdRU02 had a specific activity of 0.80 and 0.92 U/mg protein, respectively, at the same pH with corresponding biomass yield of 113, 125 and 97 mg/50 ml, respectively. Increased enzyme activities and improved mycelial biomass production were obtained in the presence of supplements such as potassium, sodium, glucose, maltose, cellobiose, tryptone and peptone, while NaFe-EDTA and cobalt inhibited enzyme activity. ChemRU330 was selected to determine the consistency and amount of amylase, cellulase and xylanase formed after several in vitro subculturing events. AMG and endo-1,4-P-xylanase were found to have the most consistent production throughout the study period. The AMG was stable at 45oC (pH 5.0), retaining approximately 65% activity over a period of 24 h. The molecular mass of AMG and endo-1,4-P-xylanase were estimated to be 101 kDa and 72 kDa, respectively. The Km and kcat were 0.38 mg/ml and 70 s-1, respectively, using soluble starch (AMG). For endo-1,4-P-xylanase, the Km and Vmax were 0.93 mg/ml and 8.54 U/ml, respectively, using beechwood xylan (endo-1,4-P-xylanase) as substrate. Additionally, crude extracts of five root endophytes with unique morphological characteristics were screened for antibacterial properties and was followed by determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). L. incrustata (ChemRU330) and Chaetomium sp. extracts exhibited varying degrees of inhibition against two Gram-positive and Gram-negative bacteria. The crude extract of L. incrustata was the most effective which was found to inhibit Staphylococcus aureus (MIC: 1 mg/ml), Bacillus subtilis (MIC: 2 mg/ml) and Proteus vulgaris (MIC: 16 mg/ml). The L. incrustata displayed potential for antibacterial production and could be considered as an additional source of new antimicrobial agents in drug and food preservation. Also, the three isolates used for enzyme production were identified to genus and species levels, i.e., Leohumicola incrustata (ChemRU330), Leohumicola sp. (EdRU083) and Oidiodendron sp. (EdRU002) using both ITS and Cox1 DNA regions. The molecular analysis results indicated that these ERM mycorrhizal fungi were similar to those successfully described by some researchers in South Africa and Australia. Therefore, this study opens new opportunities for exploring ERM fungal biomolecules for the bio-economy. The promising physicochemical properties, starch and xylan hydrolysis end- products, and being non-pathogenic make AMG and endo-1,4-P-xylanase potential candidates for future applications as additives in the food industry for the production of glucose, glucose syrups, high-fructose corn syrups, and as well as the production of bioethanol. Finally, the findings of this study revealed that it is possible to produce hydrolytic enzymes from ERM fungi in vitro using chemically defined media. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018
Exploring the potential of imines as antiprotozoan agents with focus on t. Brucei and p. Falciparum
- Authors: Oluwafemi, Kola Augustus
- Date: 2018
- Subjects: Protozoa , Parasites , Imines , Nuclear magnetic resonance , HeLa cells , Plasmodium falciparum , Trypanosoma brucei , Isomerism
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/62235 , vital:28145 , DOI 10.21504/10962/62235
- Description: This work focuses on the design, synthesis and evaluation of imine-containing heterocyclic and acyclic compounds with special focus on their bioactivity against parasitic protozoans (P. falciparum and T. brucei) - given the context of drug resistance in the treatment of malaria and Human African sleeping sickness and the fact that several bioactive organic compounds have been reported to possess the imino group. Starting from 2-aminopyridine, novel #-alkylated-5-bromo-7-azabenzimidazoles and substituted 5-bromo-1-(carbamoylmethy)-7-azabenzimidazole derivatives were prepared, and their bioactivity against parasitic protozoans was assessed. NMR spectra of the substituted 5- bromo-1-(carbamoylmethy)-7-azabenzimidazole derivatives exhibited rotational isomerism, and a dynamic NMR study was used in the estimation of the rate constants and the free- energies of activation for rotation. The free-energy differences between the two rotamers were determined and the more stable conformations were predicted. Novel 2-phenyl-7-azabenzimidazoles were also synthesised from 2-aminopyridine. A convenient method for the regioselective formylation of 2,3-diaminopyridines into 2-amino- 7-(benzylimino)pyridine analogues of 2-phenyl-7-azabenzimidazole was developed, and some of the resulting imino derivatives were hydrogenated to verify the importance of the imino moiety for bioactivity. The 2-phenyl-7-azabenzimidazoles and the 2-amino-7- (benzylimino)pyridine analogues were screened for their anti-protozoal activity and their cytotoxicity level was determined against the HeLa cell line. In order to validate the importance of the pyridine moiety, novel #-(phenyl)-2- hydroxybenzylimines, #-(benzyl)-2-hydroxybenzylimines and (±)-trans-1,2-bis[2- hydroxybenzylimino]cyclohexanes were also synthesized and screened for activity against the parasitic protozoans and for cytotoxicity against the HeLa cell line. The biological assay results indicated that these compounds are not significantly cytotoxic and a good number of them show potential as lead compounds for the development of new malaria and trypanosomiasis drugs. , Thesis (PhD) -- Faculty of Science, Chemistry, 2018
- Full Text:
- Date Issued: 2018
- Authors: Oluwafemi, Kola Augustus
- Date: 2018
- Subjects: Protozoa , Parasites , Imines , Nuclear magnetic resonance , HeLa cells , Plasmodium falciparum , Trypanosoma brucei , Isomerism
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/62235 , vital:28145 , DOI 10.21504/10962/62235
- Description: This work focuses on the design, synthesis and evaluation of imine-containing heterocyclic and acyclic compounds with special focus on their bioactivity against parasitic protozoans (P. falciparum and T. brucei) - given the context of drug resistance in the treatment of malaria and Human African sleeping sickness and the fact that several bioactive organic compounds have been reported to possess the imino group. Starting from 2-aminopyridine, novel #-alkylated-5-bromo-7-azabenzimidazoles and substituted 5-bromo-1-(carbamoylmethy)-7-azabenzimidazole derivatives were prepared, and their bioactivity against parasitic protozoans was assessed. NMR spectra of the substituted 5- bromo-1-(carbamoylmethy)-7-azabenzimidazole derivatives exhibited rotational isomerism, and a dynamic NMR study was used in the estimation of the rate constants and the free- energies of activation for rotation. The free-energy differences between the two rotamers were determined and the more stable conformations were predicted. Novel 2-phenyl-7-azabenzimidazoles were also synthesised from 2-aminopyridine. A convenient method for the regioselective formylation of 2,3-diaminopyridines into 2-amino- 7-(benzylimino)pyridine analogues of 2-phenyl-7-azabenzimidazole was developed, and some of the resulting imino derivatives were hydrogenated to verify the importance of the imino moiety for bioactivity. The 2-phenyl-7-azabenzimidazoles and the 2-amino-7- (benzylimino)pyridine analogues were screened for their anti-protozoal activity and their cytotoxicity level was determined against the HeLa cell line. In order to validate the importance of the pyridine moiety, novel #-(phenyl)-2- hydroxybenzylimines, #-(benzyl)-2-hydroxybenzylimines and (±)-trans-1,2-bis[2- hydroxybenzylimino]cyclohexanes were also synthesized and screened for activity against the parasitic protozoans and for cytotoxicity against the HeLa cell line. The biological assay results indicated that these compounds are not significantly cytotoxic and a good number of them show potential as lead compounds for the development of new malaria and trypanosomiasis drugs. , Thesis (PhD) -- Faculty of Science, Chemistry, 2018
- Full Text:
- Date Issued: 2018
Formulation of an enzyme cocktail, HoloMix, using cellulolytic and xylanolytic enzyme core-sets for effective degradation of various pre-treated hardwoods
- Authors: Malgas, Samkelo
- Date: 2018
- Subjects: Biomass , Cellulase , Hardwoods , Xylanases
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/62827 , vital:28297 , DOI https://doi.org/10.21504/10962/62827
- Description: Currently, there is a growing interest in utilising hardwoods as feedstocks for bioethanol production due to the vast advantages they have over other feedstocks for fermentable sugar production. In this study, two selected hardwoods, Acacia and Populus spp., were subjected to two pre-treatment processes (Sodium chlorite delignification and Steam explosion) and compared with respect to how these pre-treatments affect their enzymatic saccharification. Hardwoods were selected for this study, because hardwoods are easier to delignify when compared to softwoods, and therefore their polysaccharides are more easily accessible by enzymes for the purpose of producing fermentable sugars. Currently available commercial enzyme mixtures have been developed for optimal hydrolysis of acid-pre-treated corn stover and are therefore not optimal for saccharification of pre-treated hardwoods. In this work, we attempted the empirical design of a hardwood specific enzyme cocktail, HoloMix. Firstly, a cellulolytic core-set, CelMix (in a ratio of Egl 68%: Cel7A 17%: Cel6A 6%: Bgl1 9%), for the optimal release of glucose, and a xylanolytic core-set, XynMix (in a ratio of Xyn2A 60%: XT6 20%: AguA 11%: SXA 9%), for the optimal release of xylose, were formulated using an empirical enzyme ratio approach after biochemically characterising these enzymes. As it is well ̶ known that biomass pre-treatment may result in the generation of compounds that hamper enzymatic hydrolysis and microbial fermentation, the effects of these compounds on CelMix and XynMix were evaluated. Using the optimised CelMix and XynMix cocktails, a HoloMix cocktail was established for optimal reducing sugar, glucose and xylose release from the various pre-treated hardwoods. For delignified biomass, the optimized HoloMix consisted of CelMix to XynMix at 75% to 25% protein loading, while for the untreated and steam exploded biomass the HoloMix consisted of CelMix to XynMix at 93.75% to 6.25% protein loading. Sugar release by the HoloMix at a loading of 27.5 mg protein/g of biomass (or 55 mg protein/g of glucan) after 24 h gave 70-100% sugar yield. Treatment of the hardwoods with a laccase from Agaricus bisporus, especially wood biomass with a higher proportion of lignin, significantly improved saccharification by the formulated HoloMix enzyme cocktails. This study provided insights into the enzymatic hydrolysis of various pre-treated hardwood substrates and assessed whether the same lignocellulolytic cocktail can be used to efficiently hydrolyse different hardwood species. The present study also demonstrated that the hydrolysis efficiency of the optimised HoloMix was comparable to (if not better) than commercial enzyme preparations during hardwood biomass saccharification. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018
- Authors: Malgas, Samkelo
- Date: 2018
- Subjects: Biomass , Cellulase , Hardwoods , Xylanases
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/62827 , vital:28297 , DOI https://doi.org/10.21504/10962/62827
- Description: Currently, there is a growing interest in utilising hardwoods as feedstocks for bioethanol production due to the vast advantages they have over other feedstocks for fermentable sugar production. In this study, two selected hardwoods, Acacia and Populus spp., were subjected to two pre-treatment processes (Sodium chlorite delignification and Steam explosion) and compared with respect to how these pre-treatments affect their enzymatic saccharification. Hardwoods were selected for this study, because hardwoods are easier to delignify when compared to softwoods, and therefore their polysaccharides are more easily accessible by enzymes for the purpose of producing fermentable sugars. Currently available commercial enzyme mixtures have been developed for optimal hydrolysis of acid-pre-treated corn stover and are therefore not optimal for saccharification of pre-treated hardwoods. In this work, we attempted the empirical design of a hardwood specific enzyme cocktail, HoloMix. Firstly, a cellulolytic core-set, CelMix (in a ratio of Egl 68%: Cel7A 17%: Cel6A 6%: Bgl1 9%), for the optimal release of glucose, and a xylanolytic core-set, XynMix (in a ratio of Xyn2A 60%: XT6 20%: AguA 11%: SXA 9%), for the optimal release of xylose, were formulated using an empirical enzyme ratio approach after biochemically characterising these enzymes. As it is well ̶ known that biomass pre-treatment may result in the generation of compounds that hamper enzymatic hydrolysis and microbial fermentation, the effects of these compounds on CelMix and XynMix were evaluated. Using the optimised CelMix and XynMix cocktails, a HoloMix cocktail was established for optimal reducing sugar, glucose and xylose release from the various pre-treated hardwoods. For delignified biomass, the optimized HoloMix consisted of CelMix to XynMix at 75% to 25% protein loading, while for the untreated and steam exploded biomass the HoloMix consisted of CelMix to XynMix at 93.75% to 6.25% protein loading. Sugar release by the HoloMix at a loading of 27.5 mg protein/g of biomass (or 55 mg protein/g of glucan) after 24 h gave 70-100% sugar yield. Treatment of the hardwoods with a laccase from Agaricus bisporus, especially wood biomass with a higher proportion of lignin, significantly improved saccharification by the formulated HoloMix enzyme cocktails. This study provided insights into the enzymatic hydrolysis of various pre-treated hardwood substrates and assessed whether the same lignocellulolytic cocktail can be used to efficiently hydrolyse different hardwood species. The present study also demonstrated that the hydrolysis efficiency of the optimised HoloMix was comparable to (if not better) than commercial enzyme preparations during hardwood biomass saccharification. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018
Interaction of catechol O-methyltransferase with gold and silver nanoparticles
- Authors: Usman, Aminu
- Date: 2018
- Subjects: Parkinson's disease , Methyltransferases , Catechol , Nanoparticles
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61818 , vital:28063 , DOI https://doi.org/10.21504/10962/61818
- Description: Catechol O-methyltransferase (S-adenosyl-Z-methionine: catechol O-methyltransferase; COMT; EC 2.1.1.6) is a ubiquitous enzyme that catalyses the transfer of a methyl group from the cofactor, S-adenosyl-Z-methionine (SAM) to a hydroxyl group of endogenous and exogenous catechol-containing moieties. The physiological role of this enzyme is the methylation and thereby inactivation of the catechol-containing bio-active and bio-toxic compounds, including catechol-neurotransmitters, catechol-estrogens and catechol-containing drugs. Activity of this enzyme is implicated in the treatment of Parkinson’s disease and is associated with other diseases including breast cancer and an array neuropsychological disorders, such as schizophrenia. This thesis explores the use of gold and silver nanoparticles (NPs) (AuNPs and AgNPs) to inhibit the catalytic activity of mammalian COMT. Because of its accessibility and availability, we initially investigated bovine soluble COMT (BSCOMT) from liver tissue. Bioinformatic analyses and structural modeling revealed high (>90%) sequence similarity between BSCOMT and human soluble COMT (HSCOMT). BSCOMT was partially purified to 7.78 fold, 1.65% yield and had a specific activity of 0.052 U/mg. It had pH and temperature optima of 8.5 and 40oC, respectively. The Km, Vmax, Kcat and Kcat/Km towards esculetin methylation were respectively 1.475±0.130 pM, 0.0353±0.001 pmol/ml/min, 1.748 x 10-2±5.0x10-4 min-1 and 1.18x10-2 M-1. min-1. HSCOMT was expressed in Escherichia coli BL21(DE3) which showed optimal activity for esculetin methylation at pH and temperature of 7.0 and 30°C, respectively. It was purified to 5.62 fold, 22.6% yield with a specific activity of 3.85 U/mg. HSCOMT kinetic plots, upon incubation of the reaction mixture at 30°C for 5 min before addition of SAM was hyperbolic with Km, Vmax, Kcat and Kcat/Km values of 1.79 pM, 0.412 pmol/ml/min, 2.08 min-1 and 1.165 M-1. min-1, respectively. AuNPs and AgNPs showed a concentration dependent inhibition of HSCOMT activity upon increasing the 5 min incubation time to 1 h. Interestingly, HSCOMT kinetics, with 1 h incubation at 30°C, showed a sigmoidal curve, as well as increased activity. Incubation of the reaction mixture in the presence of 60 pM AuNPs and/or AgNPs for 1 hreversed the observed sigmoidal to a hyperbolic curve, with kinetic parameters comparable to those of 5 min incubation. SDS-PAGE analyses of HSCOMT after the kinetic experiments showed the enzyme incubated for 5 min as a monomer, while that which was incubated for 1 h migrated substantially as dimer. However, the HSCOMT incubated for 1 h in the presence of 60 pM AuNPs and/or AgNPs migrated as a monomer. This indicated that the extension of the incubation period allowed the dimerization of HSCOMT, which exhibited sigmoidal kinetics and higher activity. The presence of NPs impeded the HSCOMT dimerization which decreased the activity. Varying the concentration of SAM suggested that SAM had an allosteric modulatory effect on HSCOMT. Absorption spectroscopy indicated adsorption of HSCOMT on the gold and silver NP surfaces and the formation of NPs-HSCOMT corona. Fluorescence spectroscopy showed that the interaction of HSCOMT with both gold and silver NPs was governed by a static quenching mechanism, implying the formation of a non-fluorescent fluorophore-NP complex at the ground state. Further fluorometric analyses indicated that both gold and silver NPs had contact with Trp143; that the interactions were spontaneous and were driven by electrostatic interactions. Fourier transform infrared spectroscopic studies showed the adsorption of HSCOMT of the NPs surfaces to cause relaxation of the enzyme’s B-sheet structures. Molecular docking studies indicated involvement of largely hydrophilic amino acids, with the interacting distances of less than 3.5A. These findings signify the potential of nanotechnology in the control of COMT catalytic activity for the management of the COMT-related disorders. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018
- Authors: Usman, Aminu
- Date: 2018
- Subjects: Parkinson's disease , Methyltransferases , Catechol , Nanoparticles
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61818 , vital:28063 , DOI https://doi.org/10.21504/10962/61818
- Description: Catechol O-methyltransferase (S-adenosyl-Z-methionine: catechol O-methyltransferase; COMT; EC 2.1.1.6) is a ubiquitous enzyme that catalyses the transfer of a methyl group from the cofactor, S-adenosyl-Z-methionine (SAM) to a hydroxyl group of endogenous and exogenous catechol-containing moieties. The physiological role of this enzyme is the methylation and thereby inactivation of the catechol-containing bio-active and bio-toxic compounds, including catechol-neurotransmitters, catechol-estrogens and catechol-containing drugs. Activity of this enzyme is implicated in the treatment of Parkinson’s disease and is associated with other diseases including breast cancer and an array neuropsychological disorders, such as schizophrenia. This thesis explores the use of gold and silver nanoparticles (NPs) (AuNPs and AgNPs) to inhibit the catalytic activity of mammalian COMT. Because of its accessibility and availability, we initially investigated bovine soluble COMT (BSCOMT) from liver tissue. Bioinformatic analyses and structural modeling revealed high (>90%) sequence similarity between BSCOMT and human soluble COMT (HSCOMT). BSCOMT was partially purified to 7.78 fold, 1.65% yield and had a specific activity of 0.052 U/mg. It had pH and temperature optima of 8.5 and 40oC, respectively. The Km, Vmax, Kcat and Kcat/Km towards esculetin methylation were respectively 1.475±0.130 pM, 0.0353±0.001 pmol/ml/min, 1.748 x 10-2±5.0x10-4 min-1 and 1.18x10-2 M-1. min-1. HSCOMT was expressed in Escherichia coli BL21(DE3) which showed optimal activity for esculetin methylation at pH and temperature of 7.0 and 30°C, respectively. It was purified to 5.62 fold, 22.6% yield with a specific activity of 3.85 U/mg. HSCOMT kinetic plots, upon incubation of the reaction mixture at 30°C for 5 min before addition of SAM was hyperbolic with Km, Vmax, Kcat and Kcat/Km values of 1.79 pM, 0.412 pmol/ml/min, 2.08 min-1 and 1.165 M-1. min-1, respectively. AuNPs and AgNPs showed a concentration dependent inhibition of HSCOMT activity upon increasing the 5 min incubation time to 1 h. Interestingly, HSCOMT kinetics, with 1 h incubation at 30°C, showed a sigmoidal curve, as well as increased activity. Incubation of the reaction mixture in the presence of 60 pM AuNPs and/or AgNPs for 1 hreversed the observed sigmoidal to a hyperbolic curve, with kinetic parameters comparable to those of 5 min incubation. SDS-PAGE analyses of HSCOMT after the kinetic experiments showed the enzyme incubated for 5 min as a monomer, while that which was incubated for 1 h migrated substantially as dimer. However, the HSCOMT incubated for 1 h in the presence of 60 pM AuNPs and/or AgNPs migrated as a monomer. This indicated that the extension of the incubation period allowed the dimerization of HSCOMT, which exhibited sigmoidal kinetics and higher activity. The presence of NPs impeded the HSCOMT dimerization which decreased the activity. Varying the concentration of SAM suggested that SAM had an allosteric modulatory effect on HSCOMT. Absorption spectroscopy indicated adsorption of HSCOMT on the gold and silver NP surfaces and the formation of NPs-HSCOMT corona. Fluorescence spectroscopy showed that the interaction of HSCOMT with both gold and silver NPs was governed by a static quenching mechanism, implying the formation of a non-fluorescent fluorophore-NP complex at the ground state. Further fluorometric analyses indicated that both gold and silver NPs had contact with Trp143; that the interactions were spontaneous and were driven by electrostatic interactions. Fourier transform infrared spectroscopic studies showed the adsorption of HSCOMT of the NPs surfaces to cause relaxation of the enzyme’s B-sheet structures. Molecular docking studies indicated involvement of largely hydrophilic amino acids, with the interacting distances of less than 3.5A. These findings signify the potential of nanotechnology in the control of COMT catalytic activity for the management of the COMT-related disorders. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2018
- Full Text:
- Date Issued: 2018