<p>Nonribosomal peptides, polyketides, and fatty acids comprise a distinct subset of microbial secondary metabolites produced by similar biosynthetic methods and exhibit broad structural diversity with a high propensity for biological activity. Dedicated studies of these specific microbial small molecules have identified numerous potent actions towards human cells with many clinical translations. Interestingly, most therapeutically used nonribosomal peptides and polyketides were discovered from soil bacteria, meanwhile, bacteria that have co-evolved within a human context, the human microbiota, have barely been explored for secondary metabolites. The central goal of this thesis is to explore the secondary metabolome of human microbiota for nonribosomal peptides and polyketides, which are hypothesized to possess biological activities significant within the human host context. Candidate organisms were chosen for their established connections to human health and evidence suggestive of secondary metabolite production. Specifically, questions about gene to molecule prediction capability, metabolite production, structural diversity, and biological activity were explored from studies of the dental caries linked Streptococcus mutans UA159, from the probiotic Lactobacillus plantarum WCFS1, and the Crohn’s disease associated Faecalibacterium prausnitzii.</p> / Master of Science (MSc)

Cyclotides are small cyclic plant proteins, and this thesis addresses their cytotoxic structure-activity properties and their mode of action on human cancer cell lines. Cyclotides were isolated from Viola odorata and Viola tricolor; three novel cyclotide sequences and two known sequences, but of new origin, were identified using mass spectrometry, amino acid analysis, and Edman degradation. The cyclotide structure includes three disulphide bonds in a knotted arrangement, which forces hydrophobic amino acid residues to be exposed on the surface of the molecule; 3-D homology models of cyclotides have revealed an amphipathic surface and charged residues located at similar positions in the molecules. The charged amino acid residues were shown to play a key role in the cytotoxicity of the cyclotide cycloviolacinO2 on a human lymphoma cell line. Methylation of Glu caused a dramatic change in cytotoxicity, lowering the potency 48 times, whereas concealing the charge of Arg with 1,2-cyclohexanedione caused virtually no change in potency. Acetylation of the two Lys caused a 3-fold reduction in potency, and masking all positive charges caused a 7-fold reduction. Additionally, disturbing the amphipathic structure by reducing and alkylating the disulphide bonds abolished the cytotoxicity. The time dependency of cytotoxicity and cell gross morphology after cyclotide exposure were investigated on the lymphoma cell line. Cells exposed to 4 µM of cycloviolacinO2 showed necrotic characteristics, such as membrane disintegration, within 5 min; a membrane disruptive effect of cycloviolacinO2 was also observed in a functional assay based on liposomes at a peptide-to-lipid molar ratio of 6.5. The anti-tumour properties of cycloviolacinO2 were evaluated on three human cancer cell lines using the hollow fibre assay in vitro and in vivo. The cyclotide exhibited potent anti-tumour activity in the micro-molar concentration range on all cell lines in vitro, but no effect on tumour growth could be established in vivo.

Gilvocarcin V (GV) belongs to the angucycline class of antibiotics that possesses remarkable anticancer and antibacterial activities with low toxicity. Gilvocarcin exhibits its light induced anticancer activity by mediating crosslinking between DNA and histone H3. When photo-activated by near-UV light, the C8 vinyl group forms a [2+2] cycloadduct with thymine residues of double stranded DNA. D-fucofuranose is considered essential for histone H3 interactions. However, the poor water solubility has rendered it difficult to develop gilvocarcin as a drug. We aim to design novel gilvocarcin analogues with improved pharmaceutical properties through chemo-enzymatic synthesis and mutasynthesis. Previous studies have characterized many biosynthetic genes encoding the gilvocarcin biosynthetic skeleton. Despite these previous findings the exact functions of many other key genes are yet to be fully understood. Prior gene inactivation and cross-feeding experiments have revealed that the first isolable tetracyclic aromatic product undergoes a series of steps involving C–C bond cleavage followed by two O-methylations, a penultimate C-glycosylation and final lactone formation in order to fully develop the gilvocarcin structure. To provide a deeper understanding of these complex biochemical transformations, three specific aims were devised: 1) synthesis of the proposed intermediate and in vitro enzyme reactions revealed GilMT and GilM’s roles in gilvocaric biosynthesis; 2) utilizing in vitro studies the enzyme responsible for the C–C bond cleavage and its substrate were determined; 3) a small series of structural analogues of the intermediate from the gilvocarcin pathway was generated via chemical synthesis and fed to the mixture of the enzymes, GilMT and GilM. These reaction mixtures were then analyzed to establish the diversity of substrates tolerated by the enzymes.

<p>In South Africa, the crude aqueous extract from Hypoxis hemerocallidea is used by AIDS patients to treat opportunistic infections, such as tuberculosis. The rapid emergence of multidrug-resistant tuberculosis, and extreme drug resistant tuberculosis, in recent years, is a major threat to human health. The treatment of TB, nosocomial bacterial infections, and fungal infections is now a clinical challenge, especially in the immuno-compromised individual. There is a dire need for novel antibiotic alternatives with phytotherapies and plant-derived compounds as potentially promising alternatives. The main objective of this study was to investigate the antimycobacterial activity of Hypoxis hemerocallidea, a South African medicinal plant, using Mycobacterium smegmatis.</p>

The solvent-free (Cp*IrCl2)2 catalyzed N-alkylation of urea with benzyl alcohol has been studied. A variety of reaction conditions were studied and optimized to produce a high yield (82%) of N,N-dibenzylurea. A series of substituted benzyl alcohols were examined at the optimal reaction conditions. However, the preparation of substituted benzyl urea derivatives using conditions optimized for benzyl alcohol gave poor yields or intractable mixtures.

Os extratos de espécies de Passiflora (Passifloraceae), comumente empregados no tratamento de diversas doenças em regiões tropicais e subtropicais, têm se mostrado fonte em potencial de medicamentos. O uso popular de espécies vegetais pertencentes a esse gênero decorrente de suas propriedades sedativas consta a partir do século 17 na Europa. Após cerca de 200 anos, foram documentados os estudos farmacológicos iniciais com P. incarnata. No Brasil, uma espécie morfologicamente assemelhada a essa - P. edulis Sims. - tem encontrado uso como sedativo, ansiolítico, diurético e no tratamento de distúrbios gastrintestinais, embora esta espécie seja explorada comercialmente na produção de sucos concentrados. Pesquisadores têm avaliado a atividade de extratos de folhas de P. edulis no sistema nervoso central. Considerando o uso popular e a falta de trabalhos científicos avaliando a ação no sistema digestório, o extrato liofilizado preparado com as folhas e com os caules dessa espécie foi ensaiado em ratos, empregando-se solução de etanol acidificado como indutor de úlceras. A triagem fitoquímica e a avaliação da atividade antioxidante (DPPH, ORAC) foram realizadas para verificar o potencial uso dos metabólitos encontrados no extrato. O método espectrofotométrico foi empregado na análise do teor flavonoídico. As técnicas de Cromatografia Líquida de Alta eficiência (CLAE) e espectrofotometria UV/visível foram utilizadas para analisar as frações f1avonoídicas. A caracterização morfoanatômica das folhas e dos caules da espécie foi efetuada como auxiliar no controle de qualidade da droga vegetal. No modelo ensaiado, o extrato de P. edulis não mostrou atividade antiúlcera promissora. Nessa fase não se pode descartar seu uso no tratamento de úlceras gástricas. O extrato liofilizado apresentou teor flavonoídico de 1,2% e atividade antioxidante, demonstrada com os métodos empregando DPPH (EC50 de 160 &#181;g/mL) e ORAC (775,35 &#177; 36,12 &#181;mol eq Trolox/g de extrato). Na análise preliminar das frações flavonoídica foi evidenciada a predominância de flavonas. Caracteres morfoanatômicos importantes na análise da droga vegetal foram documentados. / The extracts of Passiflora spp. (Passifloraceae), commonly used in the treatment of several diseases from tropical and subtropical regions, have been proving to be a potential source for the preparation of medicines. Because of theirs sedating properties, popular use of vegetal species belonging to this genus has been found since the 17th century in Europe. Then, after 200 years, early pharmacologic studies with P. incarnata were documented. In Brazil, one species morphologically similar to this one - P. edulis Sims. - has been used as sedative, anxiolytic, diuretic and for the treatment of gastrointestinal diseases, although this species has been commercially explored in the production of concentrated juices. Researchers have been evaluating the effects of the extract of P. edulis leaves in the central nervous system. Considering its popular use and the lack of scientific research evaluating its effect in the digestive tract, the freeze-dried extract prepared with its leaves and stems was tested in rats. Acute gastric lesions were induced by acidified solution of ethanol. The phytochemical screening and the antioxidant effects evaluation (DPPH, ORAC) have been carried out in order to evaluate the potential use of metabolites found in the extract. The spectrophotometric method was applied in the analyses of flavonoids content. High Performance Liquid Chromatography (HPLC) and UV/ visible methods were used to analyze the flavonoid fractions. Morphoanatomic characterization of leaves and stems of the species was done to applied in the quality control of the crude drug. In the tested model, P. edulis extract did not show its potential gastroprotective property. At this point we can not discard its use to treat gastric ulcers. The flavonoid content quantified in the extract were 1,2%. The freeze¬dried extract presented antioxidant activity, demonstrated with DPPH (EC50 de 160&#181;g/mL) and ORAC (775,35 &#177; 36,12 &#181;mol eq Trolox/g) methods. The chemical profile of flavonoid fractions showed predominant concentration of flavones. Important morphoanatomic characters of the crude drug were documented.

In this dissertation a computer-aided automation design methodology for biotechnology applications is proposed that leads to several design guidelines. Because of the biological nature of the samples that propagate in the automation line, a very specific set of environmental and maximum allowed shelf time conditions have to be followed to obtain good yield. In addition all biotechnology protocols require precise sequence of steps, the samples are scarce and the reagents are costly, so no waste can be afforded. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2014. / FAU Electronic Theses and Dissertations Collection

Thesis (MTech (Chemistry))--Cape Peninsula University of Technology, 2016. / In Africa, herbal medicines are often used as primary treatments for a variety of ailments and diseases including HIV/AIDS and for HIV-related problems. In general, traditional medicines are not well researched scientifically in controlled studies, and are poorly regulated. Since the pharmacological effectiveness of natural products is affected by several native and foreign factors, studies on the variations of chemical composition and biological activity of these medicines are necessary. The processes of investigating plants to identify chemical substances are of great interest to natural product researchers because there is a need to discover new drugs for treating old and new diseases. These facts underscore an urgent need to develop new anti HIV and AIDS drugs with fewer or no side effects. Research into drug discovery and development using natural products is increasingly becoming better established. Marine organisms as a source of natural products delivered numerous novel compounds with multiple pharmacological properties. Natural products give endless opportunities for discovering novel compounds that can be used as drugs or backbones of drug leads. Manayi is a natural product that has been used to treat and manage people with HIV, but no scientific studies have been done to prove its efficacy on the HIV under controlled conditions. For the purposes of this study, cooked and uncooked Manayi product was evaluated for its efficacy on HIV in vitro. Manayi samples were collected in East London, Eastern Cape and sequentially extracted with hexane, chloroform, dichloromethane, butanol, methanol, and water as a series of increasingly polar solvents for its bioactive chemical constituents.

Syzygium cumini (L.) Skeels (Myrtaceae) encontra uso na medicina tradicional como hipoglicemiante. Folhas mostram emprego adicional , no tratamento de úlceras pépticas, diarréias e hemorróidas. A dissertação apresenta descrições macro e microscópicas das folhas, acompanhadas de fotografias. Teores de flavonóides, saponinas e taninos foram determinados no extrato hidroetanólico liofilizado e na droga coletada no verão e outono. O óleo volátil foi analisado. O extrato foi avaliado quanto à atividade antimicrobiana, antioxidante e toxicidade aguda. Os flavonóides (0,53%-0,62%), saponinas (4,22%-8,64%) e taninos (3,47-4,15%) apresentaram teores elevados no outono. No extrato, teores de flavonóides: saponinas e taninos foram de 2,19%, 10,92% e 13.97%, respectivamente.&#945;-pineno, &#945;-terpineol, &#946;-pineno e limoneno foram os componentes majoritários do óleo. O extrato não revelou sinais de toxicidade aguda, e não apresentou atividade contra bactérias e fungos até 1.000&#181;-g/mL. O extrato apresentou atividade antioxidante. Os dados obtidos contribuem para melhor conhecimento da espécie e no controle de qualidade. / Syzygium cumini (L.) Skeels (Myrtaceae) is a hipoglycemic in folk medicine. Its leaves are also used as anti-ulcer, anti-diarrheic and antihemorrhoid. This work brings macro and microscopic descriptions of the leaves, with photos. Flavonoids, saponins and tannins contents were determinated in the liophylizated hydroethanolic extract and in the drug collected in summer and falI. The volatile oil was analysed. The antimicrobial and antioxidant activities and the acute toxicity were avaliated . The flavonoids (0.53%-0.62%), saponins (4 .22%-8.64%) and tannins (3.47%-4-15%) showed their highest values in the fall sample. The contents of flavonoids, saponins and tannins in the extract were, 2.19%, 10.92% and 13.97%, respectively. &#945;-pinene, &#945;-terpineol, &#946; and limonene were the major constituents of the oil. The extract showed no signs of acute toxicity nor was it active against bacteria and funghi up to 1,000&#181;g/mL . The extract showed antioxidant activity. These data contribute for a better knowledge of the specie and quality control.

The aim of this dissertation was to elaborate the exploration of biologically active secondary metabolites from the marine sponge Cacospongia cf. linteiformis collected from the Bahamas and the soft coral Briareum asbestinum collected from two different sites in Florida State, Boca Raton and Dry Tortugas. In chapter one, a review on previous chemical and biological studies of the marine sponge C. cf. linteiformis and soft coral B. asbestinum is provided. Particular attention is given to spongianolides and briarellins, two important classes of compounds isolated from C. cf. linteiformis and B. asbestinum, respectively, and their structural features and diverse bioactivities. In chapter two, the isolation and relative configuration determination of four epimeric sesterterpenoids, spongianolides E & F (18c, 18d, 19c, 19d) from C. cf. linteiformis collected from the Bahamas are discussed. Thanks to chemical modification (acetylation), diastereomeric 18c&18d and 19c&19d, respectively, were able to be isolated using chromatographic techniques for the first time, and then the relative configurations of 18c, 18d, 19c, 19d were determined based on NOESY NMR experiments. The bioactivity of mixture of compounds 18c, 18d, 19c, 19d were tested and it exhibited inhibition against Schnurri-3 (a regulator of postnatal bone mass). In chapter three, the isolation and structural elucidation of four new compounds, florellins A-D (49-52), from B. asbestinum collected off the coast of Boca Raton, FL are discussed. The molecular structures of these compounds were established by spectroscopic analysis. Compounds 49-52 are the first briarellins containing an acyl group at C-13, while 49 and 50 are the first briarellins possessing acylation at C-15. Florellins A–C (49-51) were screened and found cytotoxic against three human cell lines, BT474, WM266−4 and HEK293. In chapter four, the isolation and structural elucidation of four new compounds, florellins E-H (57-60), from B. asbestinum collected in Dry Tortugas, FL are discussed. The molecular structures of these compounds were established by spectroscopic analysis. Florellins F (58) and H (60) were screened against three human cell lines, BT474, WM266−4 and HEK293, but no cytotoxicity was exhibited. In chapter five, all the experimental procedures are described, including analytical instruments, animal materials, extraction and isolation processes, spectroscopic data and protocols of bioassays. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2017. / FAU Electronic Theses and Dissertations Collection