Diabetes mellitus (DM) represents a series of metabolic conditions associated with hyperglycemia and caused by defects in insulin secretion, and/ insulin action. Exposure to chronic hyperglycemia may result in microvascular complications in the retina, kidney or peripheral nerves. According to the World Health Organization (WHO) global burden of disease, more than 176 million people are diabetic with about two thirds of these living in developing countries. With a long course and serious complications that often result in high incidences of mobility and mortality rate, the treatment of diabetes is often costly. The management of this disease is not without side effects and this is a challenge to the medical system. This has led the researches to seek new antidiabetic agents from plants. Acetone extract of 8 plants namely Terminalia sericea Burch. Ex DC, Euclea natalensis A.DC, Warbugia salutaris Bertol.f.) Chiov., Artemisia afra Jacq.ex Willd., Aloe ferox Mill, Sclerocarya birrea (A.Richi.) Hochst. subsp. caffra , Spirostachys Africana Sond and Psidium guajava L were evaluated for antidiabetic and antioxidant properties. In addition extracts were tested for cytotoxicity. Different parts of all these plants are traditionally used in South Africa for diabetes treatment. Plants were selected based on ethnobotanical information and phytochemical constituents. For determining inhibitory activity against each enzyme (α-glucosidase and α- amylase), all extracts were tested at concentration that ranged from 2x10-5 to 0.2mg/ml for α-glucosidase and 0.025 to 1.25mg/ml for α-amylase and fifty percent inhibition or higher was taken as significant (p<0.05). The extracts of A. ferox and S. africana showed no inhibition against α-glucosidase at the highest concentration tested (0.2mg/ml) whereas A. afra showed weak inhibition (47.15%). T. sericea showed to be a potent inhibitor of α-glucosidase exhibiting 97.44 % inhibition of the enzyme (p<0.05). W. salutaris, S birrea and E. natalensis also showed good activity on α-glucosidase as they demonstrated 71.84; 97.44 and 92.60 % inhibition respectively (p<0.05). Other plant extracts such as A. ferox and S. africana did not exhibit any activity on α-glucosidase. T. sericea and S. birrea showed the best inhibitory activity on α-amylase enzyme, exhibiting 91.91 and 94.94 % inhibition respectively at 1.25mg/ml. A. afra, E. natalensis, P. guajava and W. salutaris also showed good inhibitory activity on -amylase enzyme at 1.25mg/ml which was the highest concentration tested (p<0.05). Low levels of plasma antioxidants is a risk factor associated with diabetes therefore, it has been suggested that plant-based medicines that contain antioxidant properties add an advantage in curbing complications that arise during DM aetiology. The antioxidant activity of plant extracts was carried out using 2, 2-Diphenyl-1-Picrylhydrazyl (DPPH) assay. Six plant extracts which showed good α-glucosidase and α-amylase inhibitory activity were evaluated for antioxidant activity. The radical scavenging activity was measured in terms of the amount of antioxidants necessary to decrease the initial DPPH absorbance (EC50). The EC50 is the amount of antioxidants necessary to decrease initial DPPH absorbance by 50%. All 6 tested plant extracts showed good activity. W. salutaris and T. sericea demonstrated the highest activity exhibiting EC50 values of 5.08 and 5.56βg/ml respectively as compared to ascorbic acid/Vitamin C (EC50=2.52μg/ml), a well- known potent antioxidant. This was followed by P. guajava (EC50=6.97μg/ml); E. natalensis (EC50=8.46μg/ml) and S. birrea (EC50=9.41μg/ml). A. ferox showed EC50 value of 48.53μg/ml. It has been suggested that plant extracts and compounds must undergo toxicity test for safety before drug discovery is taken into consideration. Due to the large number of plants screened in this study and limited resources in our laboratory, only the acetone extract of T. sericea (which demonstrated good α-glucosidase and -amylase inhibitory activities) was tested for cytotoxicity. Acetone extract of T. sericea demonstrated moderate toxicity against primary vervet monkey kidney cells (VK) cells exhibiting IC50 values of 20.94 μg/ml when tested at 400μg/ml. Consequently, the acetone extract of T. sericea was selected for the isolation and identification of bioactive compounds. A bio-assay guided fractionation of the acetone extract of T. sericea led to the isolation of 4 pure compounds namely β-sitosterol, β-sitosterol-3-acetate, lupeol and 3-onestigmasterol and two sets of mixtures of isomers (epicatechin-catechin; MI1 and epigallocatechingallocatechin; MI2). Antidiabetic, antioxidant and cytotoxicity activities of isolated compounds were evaluated. μ–Sitosterol and lupeol showed best inhibitory activity on α-glucosidase exhibiting 50% inhibitory concentration (IC50) value of 54.50 μM and 66.48 μM respectively (p<0.05). This was followed by the MI2; epigallocatechin-gallocatechin (IC50=119.34 μM); β-sitosterol-3-acetate (IC50=129.34 μM); 3-one-stigmasterol (IC50=164.87 μM) and the MI1; epicatechin-catechin (IC50=255.76 μM). During the evaluation of purified compound’s inhibitory activity on α-amylase, compounds of interest were lupeol and β-sitosterol which exhibited IC50 values of 140.72 μM and 216.02 μM respectively as compared to the positive drug-control acarbose (IC50=65.25 μM). Epicatechincatechin and epigallocatechin-gallocatechin also demonstrated α-amylase inhibitory properties and the IC50 values were found to be lower than 100μg/ml. Epigallocatechin-gallocatechin, epicatechin-catechin and lupeol showed good free radical scavenging activity as they inhibited DPPH by 98.19; 96.98 and 70.90 % at 100ìg/ml respectively (p<0.05). The DPPH scavenging activity was very low in case of 3-one-stigmasterol (21.5% inhibition), whilst β-sitosterol and its derivative β-sitosterol-3-acetate did not show any activity. During cytotoxicity evaluation of pure compounds against monkey kidney cells, all the compounds except β-sitosterol did not inhibit the growth of these cells lines at the highest concentration tested (200μg/ml). β-Sitosterol showed moderate toxicity exhibiting IC50 values of 197.72 μM. β- Sitosterol-3-acetate, epicatechin-catechin, lupeol and epigallocatechin-gallocatechin were found to be non-toxic to Vero cells as 100% cell viability was observed when Vero cells were exposed to these samples at 200μg/ml. The compounds isolated and the extract of T. sericea demonstrated significant antidiabetic and antioxidant properties as compared to well known drugs acarbose (a known -glucosidase and α- amylase inhibitor) and Vitamin C (a well known antioxidant). This study is the first to report α- glucosidase, α-amylase and antioxidant properties of epicatechin-catechin, epigallocatechingallocatechin, β-sitosterol-3-acetate and stigma-4-ene-3-one isolated from T. sericea. In addition, epicatechin-catechin, epigallocatechin-gallocatechin, β-sitosterol-3-acetate and stigma-4-ene-3-one are isolated from T. sericea for the first time. Overall all results scientifically validated the traditional use of the bark of T. sericea for diabetes in South Africa. / Dissertation (MSc)--University of Pretoria, 2011. / Plant Science / unrestricted

Among Nature’s creation, terpenoids are more versatile and exciting natural products. In a remarkable display of synthetic ingenuity and creativity, nature has endowed terpenes with a bewildering array of carbocyclic frameworks with unusual assemblage of rings and functionalities. This phenomenal structural diversity of terpenes make them ideal targets for developing and testing new synthetic strategies for efficient articulation of carbocyclic frameworks. The thesis entitled “Total synthesis of sesquiterpenes acorenols, chamigrenes, and laurokamurene B; and Enantiospecific synthesis of ABC-ring system of A-nor and abeo pentacyclic triterpenes” describes the studies directed towards the total synthesis of the sesquiterpenes mentioned in the title and exploratory studies towards triterpenoids. In each chapter of the thesis, the compounds are sequentially numbered (bold) and references are marked sequentially as superscripts and listed at the end of the chapter. All the spectra included in the thesis were obtained by xeroxing the original NMR spectra. The sesquiterpenes acorenols, containing an interesting spiro[4.5]decane carbon framework, was first isolated in 1970 by the research group of Tomita from the wood of Juniperus rigida. Recently, in 2003, Braun and coworkers reported the isolation of epi α- and epi β-acorenols along with α- and β-acorenols from the sandal wood oil Santalum spicatum. Total synthesis of all the four acorenols has been described in the first part of the first chapter of the thesis. Initially, a model study has been carried out for the spirocyclopentannulation of cyclohexanone employing a combination of Ireland ester Claisen rearrangement and ring closing metathesis reaction to furnish methyl 4-methylspiro[4.5]dec-3-en-1-carboxylate. The same methodology has been extended for the total synthesis of all the four acorenols starting from cyclohexane-1,4-dione via cis and trans isomers of methyl 4-methyl-8-methylene-spiro[4.5]dec-3-ene-1-carboxylate. Total synthesis of β-chamigrene, γ-chamigrene and laurencenone C, containing spiro[5.5]undecane carbon framework, has been described in the second part of the first chapter. As a model study, cyclohexanone has been transformed into 1,5,5-trimethylspiro-[5.5]undec-4-en-3-one employing a combination of Ireland ester Claisen rearrangement and intramolecular type-II carbonyl ene reactions. The methodology has been extended to chamigrenes starting from cyclohexane-1,4-dione via methyl 2-(1-isopropenyl-4-oxocyclo-hexyl)-2-methylpropanoate and 5,5-dimethyl-1,9-ismethylenespiro[5.5]undecan-3-ol. The marine sesquiterpenes laurokamurenes were first isolated in 2006 by Mao and Guo from Laurencia okamurai Yamada. First total synthesis of (±)-laurokamurene B has been described in the first part of the second chapter. To begin with Ireland ester Claisen rearrangement of but-2-enyl 2-methylpropionate furnished methyl 2,2,3-trimethylpent-4-enoate, which was then transformed into 4,5,5-trimethyl-3-(4-methylphenyl)hepta-1,6-dien-3-ol. RCM reaction followed by reductive deoxygeneation transformed 4,5,5-trimethyl-3-(4-methylphenyl)hepta-1,6-dien-3-ol into (±)-laurokamurene B. Subsequently, an enantioselective total synthesis of (+)-laurokamurene B has been accomplished. Stereoselective hydrogenation of methyl campholenoate furnished methyl 2-[(1S,3S)-2,2,3-trimethyl-cyclopent-1-yl]acetate, which was then transformed into (+)-laurokamurene B via degradation of the two carbon side chain and introduction of the aryl moiety, which established the absolute configuration of laurokamurenes. The third chapter deals with the enantiospecific generation of ABC-ring system of A-nor and abeo 4(3 → 2) tetra and pentacyclic triterpenes. To begin with (R)-carvone was identified as B-ring of ABC-ring system of A-nor and abeo tetra and pentacyclic triterpenes, as the absolute configuration at the C-5 position of the targets correlate to the stereo centre of carvone, and isopropenyl group can serve as the C-4 carbon of the targets along with the two gem-dimethyl groups. A lithium liquid ammonia mediated cyclisation of δ,ε-unsaturated esters was employed for the construction of the A ring and an RCM reaction was opted for the construction of the C ring. (R)-Carvone has been converted into 2-(1-ethoxyethoxy)-1,3,7,7-tetramethylbicyclo[4.3.0]non-3-en-8-ol via lithium liquid ammonia mediated cyclisation of methyl 2-(1-ethoxyethoxy)-6-isopropenyl-1,3-dimethylcyclohex-3-enyl]acetate, which was then transformed into 4-methoxymethoxy-2,5,5,9-tetramethyltricyclo[7.4.0.02,6]tridec-11-en-8-one via the RCM reaction of 3,4-bisallyl-8-methoxymethoxy-4,6,9,9-tetramethylbicyclo-[4.3.0]nonan-3-one. The strategy has been further extended to the synthesis of 4-methylene-2,5,5,9-tetramethyltricyclo[7.4.0.02,6]tridec-11-en-8-one, which contains the ABC ring system of abeo 4(3→2) tetra and pentacyclic triterpenes.