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The effect of butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), tertiary-butyl hydroquinone (TBHQ) and 8- hydroxyquinoline (8-0HQ) upon cell growth and spore germination of Clostridium botulinum type 62A was studied using a complex liquid medium. The inhibition of spore germination was further investigated using a defined liquid medium containing L-alanine, L-lactic acid and NaHC0₃. Cell growth was monitored by optical density (600 nm) increase and germination was monitored either by optical density (600 nm) decrease or by increase in percent phase dark spores viewed under phase contrast microscopy. Strict anaerobic conditions were maintained throughout the study. As the concentration of each chemical was increased the rate of germination or growth decreased. This occurred for each pH tested. GHA, BHT, TBHQ and 8-0HQ were more effective inhibitors of growth and germination as the pH was lowered from 7.2 to 5.7. At pH 7.2 50 µg/ml BHA, 200 µg/ml TBHQ and 200 µg/ml 8-0HQ were the minimum concentrations necessary for 90% inhibition of germination. BHT (400 µg/ml) inhibited germination by 65% at pH 7.2. Minimum concentrations at which growth at. pH 7.2 was inhibited by 95% were 60 µg/ml BHA, 20 µg/ml BHT, 250 µg/ml TBHQ and 20 µg/ml 8-0HQ. Inhibition of growth and germination by each chemical was found to be reversible with the exception of the inhibition of growth by BHT. / M.S.

Thesis submitted in fulfilment of the requirements for the Degree of Masters of Technology: Organic Chemistry, Durban University of Technology, 2011. / One of the greatest medical challenges facing mankind is the Human Immunodeficiency Virus/Acquired Immunodeficiency Syndrome (HIV/AIDS) which has now become a major epidemic with more than 40 million people infected worldwide. Of equal concern is its implication in high mortality and the onset of a number of opportunist mycobacterial infections, principally tuberculosis. In spite of the discovery of some relatively effective antiretroviral (ARV) drugs such as Azido Thymidine (AZT), Nevirapine (NVP) and Efavirenz (EFV), its’ application as either a single or combinational form causes side effects by harming the bone marrow. Drug resistance is a key cause of failure for treatment of HIV infection. Hence greater interdisciplinary efforts, involving both natural and social sciences, are needed urgently to combat this HIV/AIDS pandemic. Heterocyclic nitrogen based compounds, obtained from either natural sources or synthesis are adequately documented to have increased biological activity against several diseases. Recently a study of drugs containing the naphthyridine scaffold has acquired increasing attention because of its potential against HIV/AIDS. Generally, naphthyridines demonstrate good potency in both the enzyme and cellular systems and this prompted our interest in the synthesis of naphthyridine derivatives from simple and readily available starting compounds. Furthermore we wanted to form an intermediate quinolone moiety since it has good biological potential. In this study we report the synthesis of three naphthyridine derivatives, i.e. 6-phenyl-dibenzo [b, h] [1, 6] naphthyridine, 4-methyl-6-phenyl-dibenzo [b, h] [1, 6] naphthyridine and 2- methyl-6-phenyl-dibenzo [b, h] [1, 6] naphthyridine from easily available chemicals such as aniline, ortho-toludine, para-toluidine and ethyl benzoylacetate via a five step reaction scheme using either conventional reflux, microwave irradiation or both methodologies. It was found that microwave irradiation was several folds faster than conventional reflux methodology and the yield of the product was higher. The first step of the reaction scheme is a simple condensation reaction: three acrylate derivatives, viz. ethyl-3-aniline-3-phenyl acrylate, ethyl-3-phenyl-3-(ortho-tolylamino) acrylate and ethyl-3-phenyl-3-(para-tolylamino) acrylate were synthesized by refluxing ethyl benzoylacetate in an acidified ethanolic solution with aniline, ortho-toluidine and paratoluidine respectively for three hours; the yields were 95, 87.5 and 80 % respectively. Page v In the second step, thermal cyclisation was achieved for the synthesis of three quinoline derivatives, viz. 2-phenylquinoline-4(1H)-one, 8-methyl-2-phenylquinoline-4(1H)-one and 6- methyl-2-phenylquinoline-4(1H)-one from their respective acrylates under microwave irradiation for 5 minutes at 180 °C and 250 watts; the yields were 92, 84 and 80 % respectively. In the third step of the reaction, synthesis of 4-chloro-2-phenylquinoline, 4- chloro-8-methyl- 2-phenylquinoline and 4- chloro-6-methyl-2-phenylquinoline was achieved from a mixture of POCl3 and their respective quinolines via microwave irradiation for 3 minutes at 75 °C and 150 watts and via conventional reflux for 5 hours. It was found that under microwave irradiation, the reaction occurred nearly 100 fold faster but the % yield of the product was marginally higher. The fourth step of the reaction resulted in the formation of three schiff’s base, viz. 4-(Nphenyl)- 2-phenyl-4-aminoquinoline, 8-methyl-4-(N-phenyl)-2-phenyl-4-aminoquinoline and 6-methyl-4-(N-phenyl)-2-phenyl-4-aminoquinoline from their respective quinolines via microwave irradiation for 20 minutes at 180 °C and 180 watts and via conventional reflux for 2 hours. It was found that under microwave irradiation, the reaction occurred nearly 6 fold faster and the % yield of the product was over 10 % higher. The final step of the reaction was achieved by a Vilsmeir Haack reaction and in situ base catalyzed thermal cyclisation: 6-phenyl-dibenzo [b, h] [1, 6] naphthyridine, 4-methyl-6- phenyl-dibenzo [b, h] [1, 6] naphthyridine and 2-methyl-6-phenyl-dibenzo [b, h] [1, 6] naphthyridine were synthesized from their respective schiffs base via microwave irradiation for 20 minutes at 75 °C at 120 watts and via conventional reflux for 21 hours. It was found that under microwave irradiation, the reaction occurred over 60 fold faster and the % yield of the product was over 20 % higher.The outline for the five step synthesis of the three naphthyridines is presented graphically below: Page vi Key: (a) R1= H; R2=H (b) R1 = H; R2 = CH3 (c) R1 = CH3; R2 =H Reaction Conditions: 1) conc.HCl, EtOH, 3hrs, 50 °C; 2) conc. HCl, hand stirring 10 min; 3) 180 °C, MWI, 250 watts, 5 min; 4) POCl₃, MWI, 75 °C, 150 watts, 2 min; 5) POCl₃, 100 oC, 5 hrs; 6) aniline, t-BuOH, MWI, 180 °C, 180 watts, 20 min; 7) aniline, t-BuOH, 80 °C, 3 hrs; 8) DMF, POCl₃, MWI, 75 °C,120 watts 20 minute; 9) DMF, POCl3, 100 oC, 21 hrs.

Submitted in fulfillment of the requirements of the degree of Master in Technology, Department of Biomedical Technology and Clinical Technology, Durban University of Technology, Durban, South Africa, 2016. / Introduction In Africa, use of phytotherapy for treatment of diabetes mellitus is a common form of practice. Considering the increasing burden of non-communicable diseases in South Africa efforts are directed at simple, cost effective, non-hazardous and efficient methods to treat cancer, cardiovascular diseases and diabetes. The role of phytonanotherapy is an attractive proposition for advancing new therapies. Metal nanoparticles are a possible means for delivery of such therapies. However, this requires investigation on interactions, mechanisms and therapeutic efficacy upon co-administering ethnobotanicals with metal nanoparticles and existing drug therapy in human beings. Aim The primary aim of the study was to test the in vitro antidiabetic and antibacterial activity of Ocimum sanctum (leaf extracts and flower extracts), Ocimum basilicum (leaf extracts and flower extracts), and a combination of the leaf extracts of both, and to observe whether any antidiabetic and antibacterial activity was enhanced in due to phyto-synthesised bimetallic gold-silver (Au-Ag) nanoparticles and silver nanoparticles. Methods Aqueous and ethanol extracts of O. sanctum and O. basilicum leaf and flowers alone and combined (leaf + flower) were prepared using hot vs cold water extraction techniques and 60% and 70% ethanol as polar solvents. A simple, rapid, cost effective and reproducible green chemistry method synthesised alloyed bimetallic (Au-Ag) nanoparticles using O. basilicum leaf and flower aqueous extracts and prepared silver nanoparticles (AgNps) using O. basilicum and O. sanctum leaf aqueous extracts singly and in combination (O. sanctum + O. basilicum). The size, shape and elemental analysis of the nanoparticles was carried out using UV-Visible spectroscopy, transmission electron microscopy (TEM), scanning electron microscopy coupled with energy-dispersive X-ray (SEM-EDX), dynamic light scattering (DLS) and zeta potential. Fourier transform infrared spectroscopy (FT-IR) supported by gas chromatography mass spectroscopy (GC-MS) identified the bio-capping agents. Antidiabetic carbohydrate metabolising enzymes, α-amylase (porcine) and Bacillus stearothermophilus α-glucosidase as models tested the in vitro inhibitory potential of the aqueous and ethanol plant extracts and the phyto-synthesised (Au-Ag) bimetallic and AgNps. In addition, the study investigated the antibacterial potential for the aqueous plant preparations and their respective phyto-synthesised bimetallic and AgNps against the bacterial species Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Salmonella species and Pseudomonas aeruginosa compared to gentamycin and vancomycin. Results Bimetallic nanoparticles (synthesised from leaf and flower aqueous extracts) displayed inhibitory activity that showed uncompetitive inhibition (leaf extract), and non-competitive inhibition (flower extract) of α-amylase and competitive (leaf extract) and uncompetitive inhibition (flower extract) of α-glucosidase. Bimetallic nanoparticles were higher in inhibitory activity than acarbose and the crude O. basilicum ethanol and aqueous leaf and flower extracts. In the antibacterial analysis, bimetallic nanoparticles derived from O. basilicum leaf showed inhibition against Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Pseudomonas aeruginosa and were greater in activity compared to the crude aqueous leaf extract from O. basilicum. The in vitro inhibitory effect of AgNps derived from O. sanctum and AgNps derived from O. basilicum on both enzymes was higher in activity than acarbose and their respective crude extracts. However, in combination (O. sanctum + O. basilicum), the derived AgNps appeared to be a less potent inhibitor of α-amylase and α-glucosidase enzyme and was lower than acarbose. AgNps synthesised from the combination of O. sanctum and O. basilicum showed the highest percentage inhibition against Bacillus stearothermophilus α-glucosidase, and AgNps derived from O. sanctum and AgNps derived from O. basilicum displayed competitive type of inhibition. In the antibacterial analysis, AgNps derived from the various extracts showed zones of inhibition against the Gram negative and Gram positive bacterial test strains. However, AgNps synthesised from the O. sanctum leaf extract showed higher inhibition against Escherichia coli than the positive control gentamycin and higher inhibition against Staphylococcus aureus compared to vancomycin. In addition, AgNps from O. sanctum leaf extract displayed inhibition against Bacillus subtilis, Pseudomonas aeruginosa and Salmonella species, thus representing the highest antibacterial potential. Conclusion The results demonstrate the possibility of synthesis of stable silver and bimetallic nanoparticles of Ocimum sp. The synthesised silver nanoparticles and first time synthesis of bimetallic (Au-Ag) nanoparticles displayed enhanced antihyperglycaemic properties compared to their respective crude extracts and, therefore, show promising effects in lowering postprandial hyperglycaemia in diabetic patients with dual potential for antibacterial treatment. However, the antidiabetic and antibacterial effect will need to be further affirmed in a clinical context. Medicinal plants with therapeutic value may create a new platform for further research to explore the potential for herbal medicine and nanoscience as effective biomedical and industrial applications, and for improving existing drug delivery systems in diabetic patients. Investigations into the cytotoxicity of these extracts and phytosynthesised nanoparticles is recommended. / M

Mini-dissertation submitted in partial compliance with the requirements of the Master's Degree in Technology: Homoeopathy, Durban Institute of Technology, 2002. / The purpose of this study was to determine the effect that Tulbagia violacea ethanolic herbal tincture, and Tulbagia violacea IX and 6X homoeopathic potencies, had 011 the in vitro growth inhibition of Escherichia coli, Klebsiella pneumoniae, Staphylococcus ourens. Pseudomonas aeruginosa and Bacil/us cereus respectively, as compared to a ethanol negative control. The final results were expressed as a ratio to the values obtained from gentamyein and vancomycin. Measurement was by means of the discdiffusion assay. For this study fifteen Mueller-Hinton agar plates were prepared and inoculated with each test bacteria in turn. Filter paper discs were individually inoculated with the sample substances and the control using a micropipette, before being allowed to air dry, One disc each of the Tulbagia violacea herbal tincture, 1X potency, 6X potency, ethanol control, as well as a gentamyein and vancomycin disc were placed equidistantly apart on each plate, The gentamyein and vancomycin discs were included in the experiment with the sole purpose of accounting for plate-to-plate variations in the pharmacological sensitivity of the same species of bacteria, The plates were incubated at 37\xB0C, and the zones of inhibitions measured with a pair of Vernier callipers at ] 8 hour, 24 hour and 36 hour intervals. Il Data entry and analysis was done using the SPSS\xAE statistical package, The Friedman test was used for intra-group comparison of each test or control substance at 18 hours, 24 hours and 36 hours. The Mann-Whitney U test was used to compare the mean inhibition zones produced by the test and control substance after 18 hours, 24 hours and 36 hours of incubation. The tests were performed at a=O.05 (5%) level of significance, The results obtained were that the Tulbagia violacea herbal tincture, and IX and 6X nomoeopathic potencies did not produce a statistically significant inhibitive effect on / M

Diseases caused by pathogenic bacteria continue to be major health concerns. For example, it is estimated that in the year 2000 typhoid fever caused over 21,000,000 illnesses and ~200,000 deaths (Crump et al., 2004). The disease is caused by S. typhi, a closely-related serotype of S. typhiumurium, the salmonella strain in which BipA was first identified. The CDC estimated that in 2013, multidrug resistant bacteria caused over 2 million infections in the United States, ending in more than 23,000 deaths (CDC, 2013). This number is set to rise as more bacteria become resilient to the collection of conventional antibiotics. The increasing number of multidrug resistant bacterial strains necessitates the development of new antimicrobial drugs. BipA is an attractive target for drug research. As mentioned in Section 2.5.2, BipA is ubiquitous in eubacteria and lower eukaryotes such as protozoa, but is absent from higher-order eukaryotes such as humans. Because the protein is essential for bacterial survival, BipA presents a major vulnerability of pathogenic bacteria. A drug targeting the protein itself or its interactions to the ribosome will disable only the bacteria, but have no effect on the eukaryotic host. A comprehensive model of BipA bound to the 70S ribosome will provide unparalleled insight into BipA's binding site and its mechanism. Toward this goal, cryo-EM techniques were employed to visualize the binding site of BipA on the 70S ribosome, characterize its interactions with the ribosome, and elucidate its mechanism on the ribosome. An X-ray structure of isolated BipA-GMPPNP was elucidated, by collaborators, and used for further molecular modeling of the protein to reveal possible atomic interactions between BipA and 70S ribosome. Additional biochemical studies were performed to fully characterize the specific ribosomal complex that optimizes binding of the factor. Together, the cryo-EM reconstruction, the BipA X-ray structure, the subsequent molecular modeling, and the additional biochemical studies provide a comprehensive model for BipA binding. Over the last years, the introduction of new automated algorithms for particle selection (AutoPicker) and classification (RELION) for the cryo-EM technique has revolutionized the workflow of the entire imaging and reconstruction process. The BipA dataset was primed to be used as a test bed for these algorithms and classification technique, respectively. Using old and new techniques to process the dataset allows a discussion of how the single particle reconstruction process can be vastly improved, with greater automation and efficiency.

Thesis (M.Sc. (Biochemistry)) --University of Limpopo, 2013 / This study was carried out to investigate antioxidant and antibacterial properties of 9 indigenous medicinal plants, viz., Euclea undulata (mogweregwere), Momordica balsamia (mogapu badimo), sefapa badimo, Senecio asperulus (makgonatšohle), Stiburus alopecuroides (mošalašuping), serolana, Euphorbia inaequilatera (kgama-maswana), mokgagapitsi and Clerodendrum glabrum (mohlokohloko) and to further isolate compounds that relate to these properties. Four extracting solvents with varying polarities viz. n-hexane, dichloromethane, acetone and methanol were used to extract the bioactive compounds from the ground powdered plant materials. The TLC plates, developed in three solvent systems viz., benzene, ethanol and ammonia (BEA, 18:10:0.2, v/v/v); ethyl acetate, methanol and water (EMW, 10:1.35:1, v/v/v) and chloroform, ethyl acetate and formic acid (CEF, 10:8:2, v/v/v), were visualised using DPPH, vanillin-sulphuric acid, visible light at 366 nm, UV light at 254 nm and bioautography for the presence of potential antioxidant and antibacterial compounds. The results of the screening process showed that only four plants possessed antioxidant compound(s) while six plants had antibacterial activity against Staphylococcus aureus. Euclea undulata “MKK” was observed to possess both antibacterial and antioxidant active compounds. Two antioxidant active compounds were isolated from two plants, viz., Euphorbia inaequilatera and Dicerocaryum senecioides. Solvent-solvent extraction, column chromatography and preparative TLC were used to further isolate and characterise target compounds. The antioxidant active compounds were found to separate well under EMW, an indication that the compounds are polar and intermediate-polar. The NMR spectra of the compound isolated from the D. senecioides revealed that the compound is a stilbenoid. For the first time, we report that the anti-inflammatory, antioxidant and antiproliferation properties of the D. senecioides reported by other studies performed in this laboratory could be due to this isolated stilbenoid compound. However, further studies are still necessary to confirm this assertion.

Microbial contamination of contact lenses is a significant risk factor leading to adverse responses. Adhesion of microorganisms to a contact lens is the first step in a series of events that leads to contact lens-related infections or inflammation. Recently, some of the non-steroidal anti-inflammatory drugs (NSAIDs) have been shown to have the ability to interfere with microbial biofilm formation. In this project, antibacterial properties of commonly used NSAIDs (salicylic acid, sodium diclofenac and ketorolac) were assessed and characterised using biological assays and molecular biological techniques. Salicylic acid, ketorolac and diclofenac reduced adhesion of a range of bacterial species isolated from corneal infection and inflammatory events to contact lenses in a dose-dependent manner. Salicylic acid also decreased the adhesion of Pseudomonas aeruginosa and Staphylococcus epidermidis to human corneal epithelial cells in a dose-dependent manner. Results further demonstrated that NSAIDs had a significant impact on the production of virulence factors such as Type IV pili mediated (twitching) motility, flagella mediated swimming, elastase, protease IV and alkaline protease and affected the production of acylated homoserine lactones of P. aeruginosa. Salicylic acid and ketorolac affect the expression of P. aeruginosa outer membrane proteins. In the presence of the salicylic acid and ketorolac more than 85% of all detectable outer membrane proteins changed and most were down-regulated. Moreover, in the presence of salicylic acid at least five gene products, including Na+ - translocating NADH (Nrq1), choline dehydrogenase (CHDH), a hypothetical protein of unknown function, a gene product with no similarity to any known sequence in the database and a sequence similar to 23S rRNA of P. aeruginosa, were down-regulated. The results of this study clearly demonstrated that NSAIDs have a significant impact on virulence factors and the expression of acylated homoserine lactones by P. aeruginosa. This thesis has illustrated the potential of NSAIDs for preventing bacterial contamination of contact lenses by ocular pathogens and highlights the potential for NSAIDs as antibacterial agents. Therefore, this class of compound should be investigated further for their therapeutic efficacy in vivo.

The disposition of four therapeutically important antimicrobial agents was studied in llamas following intravenous bolus administration. Six llamas were each given ampicillin, tobramycin, trimethoprim and enrofloxacin at a dose of 12 mg/kg, 1 mg/kg, 3 mg/kg and 5 mg/kg of body weight with a wash out period of 3 days between each treatment. Plasma concentrations of these antimicrobial agents over 12 hours following IV bolus dosing were determined by reverse phase HPLC. Dispositions of these four antimicrobial agents were described by two compartment open model with elimination from the central compartment, and also by non-compartmental methods. From compartmental analysis, the elimination rate constant, half-life, and apparent volume of distribution in the central compartment were determined. Statistical moment theory was used to determine non-compartmental pharmacokinetic parameters of mean residence time, clearance, and volume of distribution at steady-state. Based on the disposition parameters determined, a dose and dosing interval for each of the four antimicrobial agents was suggested for llamas. Steady state peak and trough plasma levels were also predicted for the drugs in this study for llamas. / Graduation date: 1994

Thesis (M.S.)--Oregon State University, 2007. / Printout. Includes bibliographical references (leaves 35-40). Also available on the World Wide Web.