Thesis submitted in fulfilment of the requirements for the degree Doctor of Technology: Biomedical Technology in the Faculty of Health and Wellness Sciences at the Cape Peninsula University of Technology 2014 / According to the World Health Organization, cancer is the leading cause of death in the developed world, while it is the second leading cause of death in the developing world. In particular, liver cancer is the fifth most commonly diagnosed cancer in men, however, it is the second most frequent cause of death, responsible for an estimated 700,000 deaths annually. General limited access to health services, including treatment and the overall management of cancer in developing countries often contribute to the increased mortality rates when compared to developed countries. For centuries, medicinal plants have been used to prevent, and to a certain extent, treat cancer as a readily available and affordable alternative. In many instances, the curative or preventative claims still remain anecdotal. However, increasing evidence suggest that polyphenolic components of plants possess antioxidant activities, which are credited with curative/beneficial properties of medicinal plants. The curative properties could either be related to the primary compounds present in the plant itself, or the bio-activation products of plant components affecting hepatic drug metabolising and antioxidant enzymes systems related to carcinogen metabolism and maintaining oxidative homeostasis, respectively. Similarly, chronic consumption of medicinal plants could also result in hepatotoxicity, either caused by the primary plant components or bio-activation products. Due to these observations it is paramount to understand the mechanisms involved in the metabolism of plant components to critically assess beneficial versus potential harmful properties associated with chronic consumption. The focus of the current study was aimed at elucidating the bio-activity of four multipurpose indigenous plants to Southern Africa, i.e. Adansonia digitata, Agathosma betulina, Siphonochilus aethiopicus and Myrothamnus flabellifolius. Traditionally, A. digitata has been used as an immunostimulant, anti-inflammatory and analgesic agent, while also as an antipyretic agent in the treatment of diarrhoea and dysentery. Similarly, traditional medicinal uses of A. betulina include treatment cholera, haematuria, calculus, kidney diseases, as well as infections of the bladder, urethra, and prostate among others. S. aethiopicus was traditionally employed to treat infections associated with pains and fevers, whereas M. flabellifolius served as treatment of conditions ranging from respiratory ailments, backache, kidney problems, haemorrhoids, chest pain, and asthma. In the first part of this study, the polyphenolic contents and antioxidant capacities of the four plants were characterised. The emphasis was placed on using different solvents, namely water, ethanol and acetone for the extraction of the plant material and different methodologies to assess the antioxidant contents and -capacities of the various extracts as both these factors can influence the outcome. When considering the antioxidant contents, total polyphenols, flavanols, and flavonols of the different solvent extracts prepared from the four plants were determined, whereas three different assays were used for the antioxidant capacities, i.e. oxygen radical absorbance capacity (ORAC), trolox equivalent antioxidant capacity (TEAC) and ferric-reducing antioxidant power (FRAP) assays. The A. digitata acetone extract had the highest (7.121 mg gallic acid equivalent (GAE)/milligram (mg) soluble solids), whereas the water extract of the same plant had the lowest total phenolic content (0.008 mg GAE/mg soluble solids). In general, the acetone extracts demonstrated the highest total polyphenol, flavanol, and flavonol contents, followed by the ethanol extracts, with the water extracts having the lowest contents. M. flabellifolius was the only distinct deviation from this rule, where the water extract demonstrated the highest total polyphenol content. Considering antioxidant capacities, the acetone extracts provided the highest antioxidant capacities for all plants when assessed using the TEAC (8.56-32.68 milimole (mmole) trolox equivalent (TE)/mg soluble solids) and FRAP (5.69-37.39 mmole ascorbic acid equivalent/mg soluble solids) antioxidant assays, with the exception of M. flabellifolius where the water extract demonstrated the highest activity (22.73 mmole ascorbic acid equivalent/mg soluble solids). Antioxidant capacity determinations with TEAC and FRAP assays followed similar patterns, which were different from capacities determined by the ORAC (0.46-533.54 mmoleTE/mg of soluble solids) assay. Corroborating the antioxidant content findings, the acetone extracts also demonstrated the highest antioxidant capacities (140.41-533.54 mmoleTE/mg of soluble solids), followed by ethanol (94.62-151.29 mmoleTE/mg of soluble solids) and water (0.46-134.02 mmoleTE/mg of soluble solids). Only M. flabellifolius (TEAC and FRAP) and S. aethiopicus (FRAP) deviated from this trend. Correlations between the polyphenolic contents and antioxidant capacities indicated that acetone and ethanol were more effective in extracting polyphenolic compounds than water, while also providing extracts with superior antioxidant activities. Furthermore, ORAC assay was the antioxidant capacity determining assay of choice for the aqueous plant extracts, whereas the TEAC and FRAP assays were more suitable when determining the antioxidant capacities of the acetone and ethanol plant extracts. These results confirm the notion that no single assay can comprehensively determine the antioxidant activities of plant extracts and that a battery of assays should be used, as the various antioxidant capacity determination techniques use different substrates with different targets for measurement. The second part of this study comprised an in vivo experimental animal model to assess the potential toxicity, antioxidant status and modulation of the hepatic phase 2 drug metabolising enzymes following chronic consumption of the various plant extracts in male Fisher rats. Rats consumed aqueous extracts of the various plants (2% and 5% (w/v)) as the sole source of drinking fluid for 90 days, and the serum chemical pathology parameters for monitoring liver and kidney function conducted. These included alkaline phosphatase (ALP), aspartate transaminase (AST), alanine transaminase (ALT), total iron (Fe), and creatinine (CREA). Parameters for blood and hepatic redox status included total polyphenols, ORAC, reduced glutathione (GSH), oxidised glutathione (GSSG), their ratio (GSH:GSSG), conjugated dienes (CD) and thiobarbituric acid reactive substances (TBARS). Assessment of the phase 2 hepatic xenobiotic metabolising enzymes included glutathione S-transferase (GST)  and activity in the cytosolic fraction and, UDP-glucuronosyltransferase (UDP-GT) activity in liver microsomes. When considering the liver and kidney function none of the plant extracts induced any significant toxicity, while 2% A. digitata significantly increased serum Fe. When considering the redox status, the whole blood and liver samples yielded similar results, with significant decreases in oxidised glutathione (GSSG) in rats consuming the 2% M. flabellifolius (82.76 mole/L) and 5% A. digitata (90.42 mole/L) with a resultant significant increase in the glutathione redox status (GSH:GSSG ratio of 5.69 and 5.64, respectively) when compared to rats consuming water (4.77). The GSH:GSSG ratio was also significantly increased by consumption of 2% A. betulina (8.45) and 5% S. aethiopicus (5.99). The consumption of all plant extracts, except 5% A. betulina and M. flabellifolius, significantly increased lipid peroxidation in the plasma CDs assay. These results indicated an increased antioxidant capacity in the liver with/without an associated reduced cellular oxidative stress status, which could be interpreted as a reduced susceptibility to oxidative damage. When considering the phase 2 hepatic enzymes, none of the plant extracts caused any significant changes in GST, GST or UDP-GT activities. The third part investigated the chemoprotective properties against cancer promotion in the liver utilising diethylnitrosamine (DEN) as cancer initiator and maize culture material of Fusarium verticillioides, containing the fumonisin B mycotoxins, as promoters in male Fischer rats. The rats consumed 2% (w/v) aqueous extracts of A. digitata, A. betulina, and S. aethiopicus over 28 days after cancer initiation and liver sections subjected to glutathione-S-transferase placental form positive GSTP+ staining and pre-cancerous liver foci categorised according to size. In addition, blood and liver analyses were done as described in the chronic feeding study above. Consumption of the A. digitata and, to a certain extent, S. aethiopicus extracts, altered the oxidative stress status in the liver as indicated by the increased lipid peroxidation, as determined by significantly increased liver CDs and the decreased GSH:GSSG ratio in the blood. This can be related to a subchronic toxicity due to the high total polyphenol intake as mentioned above. These underlying sub chronic toxic effects of A. digitata and S. aethiopicus are likely to be responsible for the observed inhibitory effect on the proliferation of GSTP+ minifoci in the liver. Hepatic phase 2 metabolising enzyme activities were not significantly altered by A. digitata and S. aethiopicus consumption, while GST activity was significantly increased by A. betulina treatment. Based on the findings of the current study, aqueous extracts of A. digitata, A. betulina, and S. aethiopicus may serve as hepatoprotectors with a potential to modulate liver carcinogenesis, specifically cancer promotion. To our knowledge, no other studies have attempted to describe the possible chemoprevention mechanisms of these indigenous medicinal plants. Assessments of phase 1 hepatic enzymes and other antioxidant enzymes are suggested for future studies to further describe biochemical and molecular mechanisms associated with consumption of these extracts. Additionally, identifying main compounds present in the plant extracts could culminate in development of drugs and novel nutraceuticals. It is also recommended that increasing concentrations of the plant extracts and/or the ethanol extracts to be used in future studies to better describe dose-responses of the different plants in liver carcinogenesis.

Magister Philosophiae - MPhil / Increasing energy needs globally have recently led to an interest in effectively bringing energy services in the trading system. Energy services were part of the Uruguay Round of negotiations, whose main achievement was the General Agreement on Trade in Services (GATS). The objective of the GATS is to achieve progressive liberalisation and reduction or elimination of trade barriers of all services sectors, including energy services. The GATS has made commendable progress in liberalising many service sectors, however it has not made meaningful progress with energy services. Furthermore though the SADC region engages in energy services trade through bilateral and regional agreements, a variety of barriers inhibit major successes from being achieved. Effective energy services trade and liberalisation has therefore proved problematic both at the multilateral, regional and bilateral level. This study, seeks to investigate why energy services liberalisation and trade at the multilateral, regional and bilateral level is problematic, with a particular focus on the SADC region. / South Africa

Magister Philosophiae - MPhil / Increasing energy needs globally have recently led to an interest in effectively bringing energy services in the trading system. Energy services were part of the Uruguay Round of negotiations, whose main achievement was the General Agreement on Trade in Services(GATS). The objective of the GATS is to achieve progressive liberalisation and reduction or elimination of trade barriers of all services sectors, including energy services. The GATS has made commendable progress in liberalising many service sectors, however it has not made meaningful progress with energy services. Furthermore though the SADC region engages in energy services trade through bilateral and regional agreements, a variety of barriers inhibit major successes from being achieved. Effective energy services trade and liberalisation has therefore proved problematic both at the multilateral, regional and bilateral level. This study,seeks to investigate why energy services liberalisation and trade at the multilateral, regional and bilateral level is problematic, with a particular focus on the SADC region.

Magister Legum - LLM / Trade has been widely accepted as an important tool in spearheading economic growth and development. In many different parts of the world, countries have alleviated poverty and economically prospered through effective trade. Despite the efforts to dismantle trade restrictions and create a common market, the problem remains that of African disintegrated markets which then lead to poor intra-African trade. There is therefore the need to use intra-African trade as an instrument that effectively serve in the attainment of rapid and sustainable social and economic development. The aim of this study is to therefore determine whether the solution to increasing intra-African trade can be found in the Tripartite FTA.

Magister Legum - LLM / Money laundering is a global problem that has adverse effects on both the developed and developing countries. If unchecked, it accelerates crime and criminal activities, affects the economy, undermines the integrity of financial markets, undermines the legitimate private sector, causes loss of revenue, poses security threats to privatisation efforts and brings about reputational risks as well as social costs.1 Given the transnational and cross-border nature of money laundering, the fight against it is global. This is why in 1989 the G72 countries decided to set up the FATF3 as a global standard-setting body for Anti-Money Laundering (AML) and combating of terrorist financing (CFT). The FATF has since developed standards for countries across the globe to adopt so as to facilitate this global fight. The standards are in the form of recommendations, and so far there are Forty Recommendations on money laundering (hereafter referred so as the Recommendations), Eight Special Recommendations on CFT, and a Ninth Special Recommendation on cash-couriers. In order to enhance its work and the adoption of its Recommendations, the FATF has also facilitated the establishment of FATF- styled regional bodies (hereinafter referred to as FSRBs) across the world. One such group is ESAAMLG, which was established in 1999. Its mandate is to coordinate and guide its member countries in the implementation of the Recommendations and guidelines. Currently, it has 15 member countries.8 Over the first ten years of its existence, among other things, ESAAMLG has through its members, achieved the following in its mandate: all members except Uganda have enacted AML legislation and some have set up structures that are essential for the implementation of the Recommendations Despite the above-mentioned achievements, the overall implementation of the Recommendations has been generally slow and low. Most of the member countries have not yet enforced their enacted AML legislation as evidenced by low rate of money laundering prosecutions in the region. Some have not yet established financial intelligence Units (FIUs) nor ratified or domesticated important AML related international legal instruments, let alone train personnel adequately. The international instruments comprise the 2000 United Nations Convention against Transnational Organised Crime (Palermo Convention) and the1988 United Nations Convention against Illicit Traffic in Narcotic Drugs and Psychotropic Substances (Vienna Convention). There are also inordinate delays in the passing of AML legislation as well as the amendment of other domestic legislation, which is necessary in order to harmonise such laws with the AML standards. This gives rise to unevenness, disconnectedness and time variability in the implementation of the Recommendations among the member countries. The main question this paper seeks to answer is this: Are there obstacles to the implementation of the Recommendations in Eastern and Southern Africa?

Magister Legum - LLM / This mini thesis examines the repercussions of the establishment of institutions under the Tripartite Free Trade Area (TFTA) Agreement on countries that have multiple regional economic community (REC) memberships. The study notes that even though the TFTA initiative is being touted as a major milestone towards the ultimate establishment of the African economic Community (AEC) and that it will help solve problems associated with multiple REC memberships, the initiative may come at a cost to countries, especially those that have maintained multiple REC memberships. The study observes that the institutions that have been established under the TFTA are a mirror reflection of the already existing institutions in the three existing regional blocks forming the TFTA, thus the Common Market for Eastern and Southern Africa (COMESA), East African Community (EAC) and the Southern Africa Development Community (SADC). The mini thesis argues that the creation of new TFTA institutions, in addition to the similar existing regional institutions, will translate into more financial costs; increased human resource cost for government officers; increase in non-financial treaty related obligations; and a high possibility of reaping fewer trade related gains, for countries that have multiple REC memberships. The mini thesis further demonstrates how the lack of clarity and hierarchy in the relationship between the TFTA institutions and the RECs’ institutions may negatively impact on countries that have multiple REC memberships. The mini thesis concludes by offering recommendations on how these challenges or costs on countries with multiple REC memberships can be addressed or ameliorated. / Government of Malawi

This paper critically analyses the use of anti-dumping regulation in the Southern African Customs Union (SACU) with specific reference to Botswana. Dumping takes place where products are introduced into the commerce of another country at less than its normal value, if the export price of the product exported from one country to another is less than the comparable price, in the ordinary course of trade, for the like product when destined for consumption in the exporting country. Anti-dumping duties are an internationally-recognised exception to three core WTO principles namely, bound tariff commitments, most-favoured-nation MFN) and national treatment. The use of anti-dumping in SACU has always been in accordance with existing WTO rules. The new SACU Agreement has important implications for the anti-dumping regime within the customs union. It changed the way in which tariff decisions, including anti-dumping tariffs, are made and it also requires member states to develop legislation on contingency trade remedies such as anti-dumping for the region and to establish national bodies to administer these remedies within different countries. The SACU council has given mandate to the International Trade Administration Commission of South Africa (ITAC) to undertake all trade remedies investigations and imposition of the necessary duties. From the inception of ITAC till now ninety-five percent (95%) of all anti-dumping applications filed at ITAC alleging dumping are instituted by South Africa industries. Thus, only five percent (5%) of all anti-dumping applications are from Botswana, Lesotho, Namibia and Swaziland (BLNS countries). The findings in this paper reveal that Botswana has never filed an application seeking protection of any of its industries. In terms of Article 14 of the SACU Agreement Botswana is in the process of establishing its national body like ITAC. The argument is that, since Botswana has never filed for any trade remedy how effective will this national body going to be? Therefore, critically analyzing the use of anti-dumping regulation is very crucial for Botswana as a SACU member. Such analysis will help assess the effectiveness of SACU institutions such as the tariff board and the Botswana national body to be established. / Dissertation (LLM)--University of Pretoria, 2010. / Centre for Human Rights / unrestricted

The focus of this study is the Southern African Development Community (SADC) preventive diplomacy interventions in Lesotho in 1994, 1998 and 2007. The core aim of the study was to evaluate the efficacy of the SADC security mechanism (the Organ on Politics, Defence and Security (OPDS) in conflict prevention, management and resolution on the basis of the Lesotho experience. Data for this qualitative case study was collected through interviews and document analysis. The twenty four participants for the study were drawn from the SADC OPDS unit, Lesotho political parties, Civil Society Organisations (CSOs), Academics from the University of Botswana (UB) and the National University of Lesotho (NUL), retired Botswana Defence officers who participated in the Lesotho missions and office of the post-2007election dispute dialogue facilitator in Lesotho. Documents on the SADC Treaties, Protocols, Communiqués and interventions in other set ups were used to highlight its operational policies, mandate, structures, successes and challenges. Lesotho was chosen as a case study because SADC employed both non-coercive (SADC Troika and Eminent Person mediation, 1994 and 2007 respectively) and coercive measures (the 1998 military intervention). The findings of the study revealed that SADC as a regional body had its own successes and challenges. Different perceptions on the SADC interventions in Lesotho emerged mainly between the participants from the ruling party and the opposition parties. While the former commended SADC for successfully mitigating the calamitous effects of 1994, 1998 and 2007 post-electoral violence, the opposition parties viewed the regional organisations as engaged in illegal interference in the domestic affairs of the country to defend the incumbent governing party. It also emerged from the study that the SADC security mechanism has numerous structural and operational flaws. There were several unanswered questions revolving around the legality and mandate of some of the missions. For instance, no concrete evidence emerged as to whether the 1998 military intervention was authorised by the SADC. The study also revealed that SADC has learnt valuable lessons from the Lesotho missions. Some of the reforms which the SADC has introduced in the OPDS such as the establishment of the SADC Stand by Force, Early Warning structures, the Mediation Unit, and a panel of expert mediators emanated mainly from the Lesotho experiences. The study recommends that SADC needs to harmonise the efforts of its OPDS structures such as the Mediation Unit; the Troika; the Inter-State Defence and Security Committee (ISDSC); the Inter-State Politics and Diplomacy Committee (ISPDC) and the Summit of Heads of States and Governments for rapid, coherent and well coordinated interventions in future regional preventive missions. It is also recommended that SADC should focus on identifying and mitigating underlying causal factors such as underdevelopment; poverty; deprivation of freedoms, marginalisation and other forms of social stratifications and oppression in its preventive diplomacy missions if durable peace is to be achieved in Lesotho and any other future cases.

In order to assess the question as to whether the Southern African Development Community in principle attracts foreign direct investment, literature and data have been cited that compared the economic profiles and relative privatisation programs of member states of the Southern African Development Community with relative high inflows of foreign direct investment with the economic profiles and relative privatisation programs of member states with relative low inflows of foreign direct investment. Qualitative research has been conducted, and the author is of opinion that generally the member states with relative high foreign direct investment inflows had economic profiles and relative privatisation programs that encouraged foreign direct investment, and member states with relative low foreign direct investment inflows had economic profiles and relative privatisation programs that deterred foreign direct investment inflows. From the above the author has inferred that the Southern African Development Community in principle attracted foreign direct investment, but not to its full potential. The reason may be that although the member states’ economic profiles are favourable for foreign direct investment, their economic growths tend to fluctuate, and the future economic profiles are therefore unpredictable. With the above in mind this mini-treatise aims to address the question as to what steps the governments of the Southern African Development Community’s member states could follow in order to attract foreign direct investment. iii The author is of opinion that the member states should endeavour to maintain an annual average economic growth rate, and that the governments continue to engage into privatisation programs in order to encourage foreign direct investment.

Infectious diseases are the world’s leading cause of premature deaths in humans and animals. The resistance to antibiotics and the emergence of new infectious diseases has increased the need for additional effective antimicrobial products. Despite numerous publications investigating antimicrobial activity of plant extracts it appears that no effective single product antimicrobial has yet been developed from plants. In many cases, however crude plant extracts have excellent activity and may provide useful products. Plants are frequently selected based on traditional use. Traditional healers usually use aqueous extracts of plants which in our experience generally have very low activities and it may be one of the reasons why no new products were developed from plants. Another approach to select plants for research is to use the taxonomic approach based on the premises that: (1) there is a correlation between active chemical compounds and antimicrobial activity; and (2) species in a family or order may have similar activities if the chemical precursors are inherited from a common ancestor. Future screening programmes could then concentrate on close relatives of species within these promising families and orders. The main aim of this study was to randomly screen leaf extracts of several hundred southern African tree species against important microbial pathogens to determine which taxa have the highest activity and may yield useful products to treat infections in human and animal health markets. A wide selection of plant species improved the possibility of finding promising extracts and has the advantage that active compounds may be discovered from plants that are not used traditionally. To ensure sustainable use only leaves of trees were examined. A spin off of this study would also indicate the susceptibility of different organisms, correlate the antimicrobial activities of the different organisms and determine what minimum inhibitory concentration (MIC) represents a good activity based on investigating many extracts against many microbes. The antimicrobial activity was determined by using a sensitive serial dilution microplate method. Acetone extracts were tested against two Gram-positive bacteria, two Gram-negative bacteria and two fungi, i.e. Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, Candida albicans and Cryptococcus neoformans. Small and mostly insignificant differences were found between the susceptibility of the microbial pathogens to the extracts. E. faecalis was the most sensitive bacterium and C. neoformans the most sensitive fungal organism. The strongest correlations in activities among the pathogens were between C. albicans and C. neoformans, and among the pathogen classes between Gram-positive and Gram-negative bacteria. The tree extracts analysed in the present study had a wide range of activities against the different pathogens. Twenty six per cent of the extracts inhibited the pathogens at MIC levels of 0.16 mg/ml. This clearly shows that 0.16 mg/ml is not low enough to discriminate between promising species. Some of the extracts inhibited the growth of more than one pathogen while other extracts had selective activities and could be the most promising to follow up. The study identified families and orders with either statistically significantly higher or lower antimicrobial activities. Among the large families, Combretaceae and Fabaceae had high mean activities against all test pathogens. The families Anacardiaceae and Moraceae had high activities against both Gram-positive and Gram-negative bacteria whereas the families Proteaceae and Meliaceae had higher antifungal activities. Among the large orders, Fabales had relatively high activities against all the pathogen classes. Considering that plants in related taxa often contain similar compounds and therefore similar activities, future studies could analyse more representative species in the promising taxa. Many tree species, genera, families and orders, including well-known and lesser known medicinal taxa in southern Africa, were identified with promising activities. To evaluate the potential use of these results, additional cytotoxicity, phytochemical and pharmacological studies should be carried out. The study, although still exploratory, underlined the potential of southern African tree extracts as sources of antimicrobial products. Application of these results within the Phytomedicine Programme has led to patents and products that were as good as commercial products in animal and field trials. We hope that our results will provided a starting point for discovering new products with useful activities. / Thesis (PhD)--University of Pretoria, 2014. / gm2014 / Paraclinical Sciences / Unrestricted