African traditional medicine relies largely on the abundant African flora estimated at several tens of thousands of species. These plants, like other living organisms, produce natural products which are organic molecules exhibiting a remarkable wide range of chemical diversity and a multiplicity of biological properties. Over the past 20 years, interest in drugs of plants origin has been reviving and growing steadily. Among the broad spectrum of natural products that are showing promise as possible leads to useful therapeutic agents are the terpenoids. In the present study, selected African medicinal plants were investigated for the presence of extractable and exploitable terpenoids as leads or raw materials for producing more potent bioactive compounds for pre-clinical drugs discovery programme for chemoprotective agents against cancer, HIV/AIDS, diabetes, hypertension, malaria and other chronic diseases. The plants investigated in this study included Callistemon salignus, C. viminalis, Melaleuca bracteata var. revolution gold, M. bracteata var. revolution green, M. trichostachya var. compata, Syzygium aromaticum and Tectona grandis. These plants were subjected to two separate regimes of phytochemical extraction protocols namely volatile and non-volatile-extraction protocol. The Callistemon species and Melaleuca species upon hydrodistillation afforded essential oils. The gas chromatographic and mass spectrometric (GC-MS) analysis of these essential oils reveals that 1,8-cineole was the major constituent of the Callistemon oils. Similarly, 1,8-cineole was the major constituents of the essential oil of M. trichostachya var. compata, while methyl eugenol was the predominant constituent of the oils of Melaleuca bracteata var. revolution gold, and M. bracteata var. revolution green. Antibacterial investigation of the essential oils showed that they possess strong to moderate inhibitory effect against selected bacteria. In the non-volatile extraction protocol, various parts of the plants were sequentially extracted with organic solvents to obtain crude extracts which were subjected to fractionation and purification protocols (chromatographic techniques and re-crystallization). The crude extracts from the leaves of the Callistemon and Melaleuca species gave a crystalline mixture of betulinic acid and oleanolic acid in an appreciable yield. The crude extract from the cloves of Syzygium aromaticum yielded oleanolic acid as the major extractive and maslinic acid as minor extractive. The crude extracts from Tectona grandis afforded betulinic acid in an appreciable yield. The elucidation of the structures of the pure extractives was achieved by extensive 1D and 2D nuclear magnetic resonance (NMR) spectroscopy as well as infra-red spectroscopy (FT-IR) and mass spectrometry (MS). Betulinic acid and oleanolic acid were chosen as seed molecules for making known and unknown derivatives for lead optimization study. The semi-synthesized compounds were 3- acetoxyoleanolic acid, 3-acetoxyoleanolic hydrazide, 3-acetoxyloleanolic hydrazone, 3-succinyl oleanolic acid, 3-acetoxybetulinic acid, 3-succinylbetulinic acid and maslinic acid di-acetate. / Thesis (Ph.D.)-University of KwaZulu-Natal, Westville, 2010.

No abstract available. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2003.

M.Sc. / A biosystematic study of the genus Sutherlandia (L.) R. Br., a poorly studied genus with confusing geographical variants, is presented. The species of Sutherlandia are all endemic to southern Africa. The species are very closely related and problems regarding their taxonomy are discussed. A few morphological characters that are useful in distinguishing amongst species are illustrated and discussed in detail. Morphological data are used to investigate infrageneric relationships in a phenetic analysis of 51 geographically separated populations. Sutherlandia has traditional medicinal uses, mainly as an anti-cancer treatment for internal cancers and as a general tonic. A survey of chemical compounds was done and the results are illustrated and presented in tables. The nature of this study did not allow detailed medical investigations, but the medicinal value of Sutherlandia and the compounds detected are highlighted. It is suggested that the anti-cancer activity can mainly be ascribed to the high levels of canavanine, a non-protein amino acid, in the leaves of the plant. Canavanine, an arginine analogue, is known for its antitumourigenic properties. The value of the plant as a bitter tonic is probably related to the presence of several triterpenoids, some of which may well also have other beneficial effects. Enzyme electrophoresis was done to explore genetic relationships amongst the numerous regional forms of Sutherlandia. A study of 19 populations showed that they are all closely related and that a more conservative treatment of the taxa is called for. As a result the number of taxa is reduced. A complete taxonomic revision of the genus is presented. The number of species is reduced from six to two, namely S. frutescens and S. tomentosa. The former is divided into three subspecies, namely subsp. frutescens, subsp. microphylla and subsp. speciosa. Some regional forms are described and illustrated, but these are not formally recognised as taxa. A key to the species, subspecies and regional forms is provided, and the. nomenclature, typification, description and geographical distribution for each of the taxa are given. The multidisciplinary approach of this study provided a better understanding of the morphological, chemical and genetic variation in this relatively poorly known but potentially valuable ornamental and medicinal plant.

In Africa, the importance of medicinal plants in folklore medicine and their contribution to primary healthcare is well recognized. Across the continent, local herbal mixtures still provide the only therapeutic option for about 80% of the population. The vast floral diversity and the intrinsic ethnobotanical knowledge has been the backbone of localized traditional herbal medical practices. In Africa, an estimated 5400 of the 60000 described plant taxa possess over 16300 therapeutic uses. Similarly, with a therapeutic flora comprising of approximately 650 species, herbal medical practitioners in South Africa, make use of a plethora of plants to treat different human diseases and infections. Over the years, studies have identified numerous plant species with potential against chronic metabolic diseases including type 2 diabetes mellitus (T2DM). Globally, the incidence and prevalence of T2DM have reached epidemic proportions affecting people of all ages, nationalities and ethnicity. Considered the fourth leading cause of deaths by disease, T2DM is a global health crisis with an estimated diagnosis and mortality frequency of 1 every 5 seconds and 1 every 7 seconds respectively. Though the exact pathophysiology of T2DM is not entirely understood, initial peripheral insulin resistance in adipose tissue, liver, and skeletal muscle with subsequent pancreatic β-cell dysfunction resulting from an attempt to compensate for insulin resistance is a common feature of the disease. The current approach to treating T2DM is the use of oral antidiabetic agents (OAAs), insulin, and incretin-based drugs in an attempt to achieve glycaemic control and maintain glucose homeostasis. However, conventional anti-T2DM drugs have been shown to have limited efficacies and serious adverse effects. Hence, the need for newer, more efficacious and safer anti-T2DM agents. Sutherlandia frutescens subsp. microphylla is a flowering shrub of the pea family (Fabaceae/Leguminaceae) found mainly in the Western Cape and Karoo regions of Southern Africa. Concoctions of various parts of the plant are used in the management of different ailments including T2DM. However, despite extensive biological and pharmacological studies, few analyses exist of the chemical constituents of S. frutescens and no Triple Time of Flight Liquid Chromatography with Mass Spectrometry (Triple TOF LC/MS/MS) analysis has been performed. The initial aim of this study was to investigate the phytochemical profile of hot aqueous, cold aqueous, 80% ethanolic, 100% ethanolic, 80% methanolic and 100% methanolic extracts of a single source S. frutescens plant material using colorimetric and spectrophotometric analysis. The hot aqueous extractant was found to be the best extractant for S. frutescens, yielding 1.99 g of crude extract from 16 g fresh powdered plant material. This data suggests that application of heat and water as the extractant (hot aqueous) could play a vital role in extraction of bioactive compounds from S. frutescens and also justifies the traditional use of a tea infusion of S. frutescens. Colorimetric analysis revealed the presence of flavonoids, flavonols, tannins, and phenols in all extracts with varying intensity. The organic extracts 100% methanol, 80% and 100% ethanol exhibited high color intensity (+++) for flavonoids and flavonols respectively, while all the extracts exhibited a moderate color intensity (++) for tannins and phenols. Spectrophotometric analysis of S. frutescens extracts revealed that all the organic extracts contained a significantly higher concentration (in mg/g of extract) of flavonols and tannins when compared to the aqueous extracts. All extracts contained approximately equal levels of phenols. These data confirm the presence of all four groups of bioactive phytocompounds in the S. frutescens extracts used in this study, and also confirm that different solvent extractants possess the capability to differentially extract specific groups of phytocompounds. in individual extracts. Further comparison of these compounds with online databases of anti-diabetic phytocompounds led to the preliminary identification of 10 possible anti-diabetic compounds; α-Pinene, Limonene, Sabinene, Carvone, Myricetin, Rutin, Stigmasterol, Emodin, Sarpagine and Hypoglycin B in crude and solid phase extraction (SPE) fractions of S. frutesecens. Furthermore, using two hepatic cell lines (Chang and HepG2) as an in-vtro model system, the anti-T2DM properties of crude aqueous and organic extracts of S. frutescents was investigated and compared. Both aqueous and organic extracts of S. frutescens were found to decrease gluconeogenesis, increase glucose uptake and decrease lipid accumulation (Triacylglycerol, Diacylglycerol, and Monoacylglycerol) in Chang and HepG2 hepatic cell cultures made insulin resistant (IR) following exposure to high concentration of insulin and fructose. Using real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), the aqueous and organic extracts of S. frutescens were confirmed to regulate the expression of Vesicle-associated membrane protein 3 (VAMP3), Mitogen-activated protein kinase 8 (MAPK8), and Insulin receptor substrate 1 (IRS1) in insulin resistant hepatic cells. IR-mediated downregulation of VAMP3, MAPK8, and IRS1 mRNA in IR HepG2 hepatic cell cultures was reversed in the presence of aqueous and organic extracts of S. frutescens. The hot aqueous extract displayed the highest activity in all the assays, while all the organic extracts displayed similar potency. In conclusion, this study reports that aqueous and organic extracts of S. frutescens possess numerous anti-diabetic compounds that can be further investigated for the development of new, more efficacious and less toxic anti-diabetic agents. The presence of multiple compounds in a single extract does suggest a synergistic or combinatorial therapeutic effect. These findings support the burgeoning body of in-vivo and in-vitro literature evidence on the anti-diabetic properties of S. frutescens and its use in folklore medicine.

Siphonochilus aethiopicus (Schweinf.) B.L. Burtt (Zingiberaceae), commonly known as wild ginger, is a highly sought after plant for use in traditional medicine in South Africa. Over-exploitation of this medicinal plant has resulted in regional extinction in the wild. As a result, there is great interest in the medicinal properties of S. aethiopicus, and as a plant for small scale cultivation to increase the supply for use in traditional medicine. Water, ethanol and ethyl acetate extracts were prepared from the leaves, rhizomes and roots of S. aethiopicus. These extracts were tested for in vitro anti-inflammatory activity in the cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) assays, and in the microdilution antibacterial assay. The aqueous extracts showed no significant prostaglandin synthesis inhibition in the COX-1 and COX-2 assays. The ethanol and ethyl acetate extracts of the leaves showed the highest levels of activity at a concentration of 250 µg ml¯¹ per test solution, in both the COX-1 and COX-2 assays. The ethanol and ethyl acetate extracts of the rhizomes and roots also had moderate levels of activity in the COX-1 assay. These results provide some evidence for the rational use of S. aethiopicus in traditional medicine for anti-inflammatory purposes. In the microdilution antibacterial assay, no inhibitory activity against the test bacteria was detected with the aqueous extracts. The ethanol and ethyl acetate extracts tested showed greater antibacterial activity at minimal inhibitory concentrations ranging from 0.78 to 3.13 mg ml¯¹ against the gram-positive bacteria (Bacillus subtilis, Staphylococcus aureus) than the Gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae). No distinct differences were observed between the ethanol and ethyl acetate extracts, or between the different plant parts. A serial extraction of S. aethiopicus rhizome material was conducted and the extracts were tested in the COX-1 assay and the microdilution assay as a preliminary investigation for a bulk extraction. The hexane and ethyl acetate extracts gave slightly higher COX-1 inhibition than the ethanol extract. No distinct differences were observed in the microdilution assay. A bulk ethyl acetate extract of S. aethiopicus rhizome material was prepared, yielding 6.3 g of a thin orange oil. Vacuum liquid chromatography (VLC) was used to fractionate ≈4 g of the extract. The VLC fractions were evaluated using thin layer chromatography (TLC) and a bioautographic assay, using S. aureus as a test organism. The fractions were also tested in the COX-1 assay. The bioautography revealed a number of compounds which exhibited antibacterial activity. Fraction C was purified further using preparative TLC, and 24.9 mg of a pure compound from R,0.54 (toluene:ethyl acetate 93:7) was isolated. The structure of the compound was elucidated from nuclear magnetic resonance (NMR) spectra, and mass spectroscopy of the compound was also recorded. The compound was identified as the sesquiterpenoid furanoeremophil-2-en-1-one, which is structurally identical to the recently reported compound 4aαH-3,5α,8aβ-trimethyl-4,4a,9-tetrahydro-naphtho[2,3-b]-furan-8-one. The compound showed only a very minimal bacteriostatic effect in the microdilution assay. S. aethiopicus plants were harvested before and after seasonal senescence. Ethanol extracts were prepared from fresh or dried material of the leaves, rhizomes and roots, and tested in the COX-1 assay and the microdilution assay TLC fingerprints of the various extracts were also prepared. No noteworthy changes in COX-1 inhibition, due to senescence, were observed with extracts prepared from fresh material, although there did appear to be a slight decrease in activity in the α-roots and an increase in the β-roots after senescence (fresh and dry). A decrease in the antibacterial activity of the leaves and an increase in the antibacterial activity of the α-roots was observed after senescence. These results suggest that the time of harvest may only have a minimal influence on the degree of anti-inflammatory and antibacterial activity. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 2002.

Schumanniophyton magnificum and Glypheae brevis are important medicinal plants growing wild in the West African rain forest. They are used in folkloric medicine in the treatment of epilepsy and convulsion as well as for some other diseases. The purpose of this work was to investigate the aspect of folkloric use in order to support folkloric claims and document the findings. The extracts were prepared from ground plant material by a continuous extraction method. Five hundred grams of ground plant material were continuously de-fatted with 2 L petroleum ether (60°- 80°) in a Soxhlet apparatus for about 5 h. The resulting marc was dried and the chemical constituents extracted hot in a Soxhlet apparatus for about 8 to 10 h with 2 L aqueous ethanol (70%). The efficacy of the extraction method was confirmed using standard bioassays and phytochemical analyses. The anti-convulsant activity of the crude extracts was evaluated in vivo against chemically induced convulsions using three different animal models, namely the strychnine, the picrotoxin and the pentylenetetrazole tests. The acute and delayed toxicity test results showed that in all the animal models investigated very high doses, about four times higher than the protective doses of the extracts, were required to kill 50% of the population of animal used. Phytochemical assays of the extracts indicated the presence of alkaloids only in S. magnificum root extract and glycosides in extracts from both species. The glycosides were positive to Baljet, Xanthydrol and Keller-Kiliani tests for cardiac glycosides. S. magnificum and G. brevis chemical constituents were initially isolated with a sequential fractionation method starting with a highly non-polar solvent and gradually increasing to a more polar solvent. The fractions were pooled on the basis of TLC similarity profiles when viewed under the UV light at 254 and 366 nm and were found to have two and four major UV absorbing fractions for S. magnificum and G. brevis respectively. Radio-receptor binding tests were used to assess the anti-convulsant activities of the hydro-alcoholic crude extracts, the organic and aqueous fractions of the crude extracts, partially purified components and pure components in in vitro tests against some standard GABA[A] receptor antagonists, muscimol and isoguvacine respectively. The anti-convulsant activities resided in the aqueous fractions of the hydro-alcoholic crude extracts of both plants. The purely organic fractions of G. brevis demonstrated no activity while all the fractions of the aqueous component demonstrated some degree of activity. The anti-convulsant activity of S. magnificum was found only in one fraction-Fraction 1. This Fraction was further investigated and one of the components appear to be responsible for the activity. The structure of the active constituent was 5,7dihdroxy-2 methylbenzopyran-4-one, a noreugenin. A second bioactive compound, schumanniofoside, was identified from Fraction M[5.2] from S. magnificum. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2002.

This thesis examined the in vitro and in vivo anti-oxidative and anti-diabetic activities of five African medicinal plants which are traditionally used for the treatment of diabetes mellitus viz; Ziziphus mucronata, Cassia singueana, Parkia biglobosa, Khaya senegalensis and Vitex doniana. Ethanol, ethyl acetate and aqueous crude extracts of the stem bark, root and leaf samples of each of the plants (a total of 45 crude extracts) were investigated for detailed anti-oxidative activity and the most active crude extract from each plant was selected for further fractionation with solvents of increasing polarity. Subsequently, the solvent fractions derived from these crude extracts (a total of 21 fractions) were also subjected to the anti-oxidative assays as well as α-glucosidase and α-amylase inhibitory activities assays. Results from these assays revealed that the butanol fractions from crude extracts of Z. mucronata, P. biglobosa, K. senegalensis and V. doniana and the acetone fraction from the crude extract of C. singueana were the most bioactive. Kinetic delineation of the types of enzyme inhibitions exerted by these most active fractions as well as measurement of relevant kinetic parameters (KM, Vmax and Kᵢ) were done using Lineweaver-Burke’s plot. Furthermore, the most active fractions were also subjected to GC-MS analysis and in vivo intervention trial in a type 2 diabetes (T2D) model of rats (except fraction from V. doniana). The in vivo studies revealed that all the fractions possessed potent in vivo anti-T2D activity (to varying extent) and the possible mechanisms of actions were proposed. Furthermore, most of the fractions were able to ameliorate the T2D-associated complications. Analysis of in vivo oxidative stress markers such as glutathione, thiobarbituric acid reactive substances, superoxide dismutase and catalase in the serum, liver, kidney, heart and pancreas of the animals also gave a clue in to the possible mechanism of action. Bioassay guided isolation was used to track the bioactive anti-diabetic compounds from these fractions via column chromatography. The isolated compounds were characterized by ¹H NMR, ¹³C NMR, 2D NMR (in two cases) and mass spectroscopy (in one case). From this study, 2,7-dihydroxy-4H-1-benzopyran-4-one, 3β-O-acetyl betulinic acid, lupeol and bicyclo[2.2.0]hexane-2,3,5triol were identified as the possible bioactive compounds from Z. mucronata, C. singueana, P. biglobosa, K. senegalensis solvent fractions respectively. The findings of this work are important for the relevant government agencies, pharmaceutical industries, scientific community and poor diabetic patients because it might open an avenue for the development of viable and cost effective anti-diabetic herbal products and/or novel plant-derived anti-diabetic drugs. / Ph.D. University of KwaZulu-Natal, Durban 2013.

Acquired Immunodeficiency Syndrome (AIDS) is a human disease caused by the human immunodeficiency virus type 1 (HIV-1) and it is one of the biggest social, economic and health challenges in the world. The Joint United Nations Programme on HIV/AIDS (UNAIDS) and World Health Organization (WHO) estimated that between 33.4 to 46.0 million people around the world were living with HIV/AIDS in December 2005 and the highest estimates are in the Sub-Saharan Africa (around 25 million). In more developed countries a combined antiretroviral therapy called highly active antiretroviral therapy (HAART) is used, which results in reduced progression to AIDS in most patients. Despite the beneficial effects of HAART, significant side effects are experienced by treated patients. In addition, most infected people live in countries where the treatment is very expensive or, in many cases, not available at all. These people therefore rely on medicinal plants for health care. In this study, aqueous extracts from Sutherlandia frutescens and Lobostemon trigonus were screened for potential anti-HIV activities in a series of in vitro enzymatic assays, including reverse transcriptase, HIV-1 protease and glycohydrolases. Two extracts of Sutherlandia leaves (SFL-1 and SFL-2) were prepared that inhibited HIV reverse transcriptase and a Lobostemon leaf extract (LTL) was shown to also inhibit this enzyme. All extracts were assayed at 1.25mg/ml. Tannin content was determined for all active extracts using a tannic acid assay. SFL-1 and SFL-2 were found to contain about 6 percent and 7 percent tannins, respectively, and LTL contained 31% tannins by weight. Tannins were removed using polyamide columns and three fractions were collected for each. The extracts were also fractionated with Sephadex G-25, Amberlite IR 120 and Dowex 1-X8 as size exclusion, cation exchange and anion exchange, respectively. Extracts were also fractionated by preparative thin layer chromatography where two compounds were separated from S. frutescens extract with high activity against reverse transcriptase while showing insignificant inhibition towards other enzymes tested. SFL-BFW-10 and SFL-WEF-7 inhibited reverse transcriptase by almost 100 percent and the IC50 values calculated for these compounds were 0.34 and 0.23mg/ml, respectively. Cytotoxicity of these compounds was evaluated on Chang liver cells and peripheral blood mononuclear cells (PBMCs). None of these compounds showed any significant inhibition of cell proliferation. The purity of these compounds could not be confirmed because there was insufficient material to use in the techniques required to show purity and identification. Therefore, TLC was used to determine the nature of these compounds. SFL-BFW-10 was identified as an organic acid and SFL-WEF-7 was identified as flavonoid.

Focused procedures which streamline and optimise plant prioritisation and selection in bioprospecting have the potential to save both time and resources. A variety of semiquantitative techniques were assessed for their ability to prioritise ethnomedicinal taxa in the Flora of Southern Africa (FSA) region. These techniques were subsequently expanded upon for application in plant selection for the Novel Drug Development Platform bioprospecting programme. Least squares regression analyses were used to test the hypothesis that ethnomedicinal plant use in southern Africa is strictly random, i.e. no order or family contains significantly more medicinal plants, than any other order or family. This hypothesis was falsified revealing several 'hot' plant orders. The distribution of southern African ethnomedicinal taxa was investigated, and revealed low ethnomedicinal plant usage in the Western Cape and Northern Cape. The historical settlement of Bantu tribes in the eastern regions of southern Africa was one explanation for this discrepancy. Growth forms of ethnomedicinal taxa in 'hot' orders (identified in the regression analysis) were analysed. The results indicated no clear preferences across orders, but rather a preference for particular growth forms in certain orders. With respect to distribution, endemism and Red Data List status of ethnomedicinal taxa, the Western Cape had the greatest proportion of endemics and Namibia had the highest proportion of Red Data Listed ethnomedicinal taxa. With respect to chemotaxonomy, the Asteraceae contained the highest proportion of terpenoids, the Rubiaceae the highest proportion of alkaloids and the Fabaceae the highest proportion of flavonoids. The predictive value of regression analyses was tested against an existing analysis of anti-malarials and the subsequent in vitro bioassays on Plasmodium falciparum. In particular, the ability of these analyses to identify plants with anti plasmodial IC50 values of [less than or equal to] 10 [micro]g/ml was assessed. Most species in 'hot' genera showed comparatively good antiplasmodial activities (IC50 [less than or equal to] 10 [micro]g/ml). Plant candidates were prioritised for screening anti-tuberculosis, anti-diabetes and immune-modulatory compounds, using a weighting system based on; their ethnomedicinal application, chemotaxonomic potential, frequency in ethnomedicinal trade, association with the relative disease, toxicity, Red Data status, indigenous or endemic status, and family selection in ethnomedicine (identified through regression analyses). Other taxa were short-listed due to their presence in biodiversity hotspots where few ethnomedicinal plant use records are documented, and still others were incorporated due to their taxonomic association with efficacious exotic allies. Statistical analyses of the weighting processes employed were not possible in the absence of screening results which are due only in December 2006. The legislation governing bioprospecting in South Africa is discussed and several recommendations are presented to minimise negative impacts on the industry. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2005.

Many South Africans continue to use traditional medicine in their daily lives as an alternative form of health care, also as part of their cultural heritage. Medicinal plants are proving to be an important source of novel drugs, and the knowledge provided by traditional healers is a useful tool in the search for antimicrobials. The antimicrobial activity of <M. villosus was investigated against ten bacteria and six fungi. The antibacterial assay showed that the root extract had the highest inhibition against the Gram-positive bacteria at the minimum inhibition concentration of o.1 mg/ml, as well as against the Gram-negative, E. coli, at the MIC of 1.0 mg/ml. Antifungal activity was witnessed against Cladosporium cladosporoide, C. cucumerinum&C sphaerosperum all at the minimum inhibitory concentration of 1.0 mg/ml. An attempt was made to isolate and identify the active antimicrobial compounds. A flavonol, quercetin was isolated and identified by means of UV spectral graphs, and TLC comparison of the plant extract and standard. However, a second isolated antibacterial compound could not be identified fully but it can be said that it is a triterpenoid. / Dissertation (MSc (Botany))--University of Pretoria, 2006. / Plant Science / unrestricted