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1
Abd, Ghaffar Nur Rinah. "Bioprospecting for extremophile oleaginous yeasts." Thesis, University of Bath, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.760942.
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Palm Oil is the highest produced edible oil globally, with over 66 million tonnes produced annually. It has been estimated that up to 50% of all products sold in the supermarket contain palm oil in some form. Palm oil has attractive properties such as a high melting point and texture due to a balanced ratio of unsaturated and saturated fatty acids. It contains approximately 40% oleic acid (monounsaturated fatty acid), 10% linoleic acid (polyunsaturated fatty acid), 45% palmitic acid and 5% stearic acid (saturated fatty acid), that results in an edible oil that is suitable for use in a variety of food, detergent and cosmetics products. In addition, palm oil is the least expensive oil produced due to its high productivity and extensive production. Due to the high demand for the product, vast amounts of rainforest have been cleared to make way for more plantations, reducing biodiversity and releasing huge levels of carbon dioxide into the atmosphere. There is a clear need for an alternative lipid that can match palm oils properties but can be produced sustainably. Recent work suggests that some yeasts are capable of producing a similar oil to palm oil and can be grown on waste resources. In this thesis a novel bioprospecting protocol was developed to isolate yeasts that can survive the harsh conditions necessary for industrial biotechnology. In this way a vineyard and the local area was sampled for yeasts which were then cultured under extremes of pH, multiple sugars and inhibitors caused from the breakdown of lignocellulose. The wild yeast were cultured in four stages: minimal medium with Lysine; minimal medium with inhibitors; minimal medium with xylose as sole carbon-source; and lastly minimal medium with only arabinose and cellobiose as carbon-sources. Only strains that survived each stage were taken forward to the next, to isolate species that were truly suited to these conditions. Out of the estimated 1000s of strains screened this resulted in 12 strains of yeast, mostly in the Metschnikowia pulcherrima, group being able to cope with the conditions. The 12 strains were further analyzed by culturing them in an array of 4 different model lignocellulosic feedstocks namely wheat straw, corn Stover, sugarcane bagasse, and palm kernel cake hydrolysates. Other conditions incorporated in these analysis were a range of pH from pH 1.5 to pH 7.0; four levels of a mixture of 5 inhibitors; and two different temperatures. All of the 12 strains showed similar behaviour where inhibitor tolerance was only marked at higher pH, and at low pH the strains could not grow at all. Though all strains were able to grow on the hydrolysate models, even those with little glucose and/or xylose content. The lipid profile of the strains was also assessed and proved to be similar to most terrestrial crops, with suitable lipid profiles for a rapeseed oil, and in some cases palm oil substitute. Lastly, to further evaluate the accurate identification of the strains as there are some ambiguity in the Metschnikowia pulcherrima group, we applied an approach only widely used for Pathogenic Bacteria/Yeast identification, Multilocus Sequence Typing (MLST). Using 25 strains (7 of this collection), 6 type species and some isolates from the original culture collection in Bath. Sequences of 6 genes was analysed using the Bayesian statistical method. The result showed grouping of M. pulcherrima into 3-4 groups 9 different for each gene. M. Corniflorae being the outgroup. In all 3 genes successfully sequenced: M. Fruticola; R6; Mp DAH 3; and ICS48 were consistently shown to be clonal. The work presented here demonstrates a new method for bioprospecting strains capable of isolating strains for industrial biotechnology, and for characterisation of the yeast in the Metschnikowia genus. Some of the yeasts identified were oleaginous, and could potentially be used as a novel source of palm oil substitute.
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Lyons, Laura Francis. "Bioprospecting for microorganisms and enzymes with biorefining potential." Thesis, Aberystwyth University, 2015. http://hdl.handle.net/2160/21abce27-49d0-4bcc-8196-cae9f79c8383.
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The rise of biorefining and application of biotechnology to combat climate change and accomplish energy security is a necessity. There have already been steps to produce sustainable biofuels and products throughout the world. However, not all processes are economically viable due to costs of enzymes, pre-treatments, and scale-ups. In this project Miscanthus sp. was the main source of bacterial isolation due to its bioenergy characteristics as a low-input high-output crop. Specifically, due to the high sugar content of harvested senesced Miscanthus. The aim of this project was to discover novel microbes and enzymes with potential to contribute to the generation of fuels and chemicals from plant biomass. Specifically, we aimed to isolate and characterise enzymes capable of efficiently releasing sugars from pre-treated lignocellulosic biomass, for subsequent fermentation to ethanol and platform chemicals. Aerobic bacteria were cultured from harvested chipped Miscanthus and soil surrounding Miscanthus crops and were characterised morphologically, functionally and taxonomically. Bacteria in our collection included amongst others: Pseudomonas sp., Burkholderia sp., Variovorax paradoxus, Luteibacter sp. and Bacillus sp. The collection was screened for carbon utilisation using cellulose (in the form of carboxymethyl cellulose), xylan (from beechwood) and starch by enzymatic activity, at a range of temperatures and pHs. From the bacterial library, 88.5% of cultures showed cellulase activity, 93.2% xylanase activity, 79.7% starch degradation activity over the temperature or pH range, with 66.2% demonstrating activity over all three assays. Proteins were isolated from bacteria that demonstrate effective starch utilisation for further characterisation. Bacterial isolates that exhibited xylan utilisation at high pH and temperature were characterised by whole genome sequencing to identify interesting enzymes and pathways using bioinformatics software CLC genomics and Seed RAST. Finally, homologous proteins have been modelled using Phyre2 and 3DLigandSite to analyse structure and binding sites. This work was part of the wider BEACON project which aimed to establish Wales as a Biorefining Centre of Excellence. BEACON built integrated 'Green Supply Chains' with a focus on developing new routes to functional, cost competitive products using biomass rather than fossil fuels.
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Bessa, Nélita Gonçalves Faria de. "Brazilian savanna forest : conservation, medicinal reservoir and bioprospecting." Doctoral thesis, Universidade de Aveiro, 2014. http://hdl.handle.net/10773/13350.
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Doutoramento em Biologia e Ecologia das Alterações GlobaisThis study aimed to analyse the Brazilian savanna forest from a Legal Reserve (LR) area from a perspective of conservation, reservoir of organic carbon and medicinal biomass for a prospective use of native medicinal plants. An ethnobotanical and ethnopharmacological survey was carried out close to a community settled in the rural area in the south of Tocantins, being selected 9 of the most cited species (cajuí- Anacardium othonianum; inharé-Brosimum gaudichaudii; jatobá-Hymenaeae courbaril; jenipapo-Genipa americana, aroeira-Myracrodruon urundeuva; negramina-Siparuna guianensis; barbatimão- Stryphnodendron obovatum; assa peixe-Vernonia brasiliana, embaúba-Cecropia pachystachya). Crude foliar extracts were subjected to a preliminary phytochemical prospection and triage of secondary metabolites with antimicrobial activity of potential interest in health and familiar agriculture. Phenolic compounds, terpenes and flavonoids were detected in the extracts of most species, which suggests the presence of antimicrobial, antioxidant and anti-insect activities. It was evident the need to better know the LR as a reservoir of medicinal biomass in an area under ecological tension where 35% (610ha) of the property is LR and should be protected by law. Therefore, a forest inventory of live woody species was performed using the allometric or indirect method. This identified a rare remnant of Semidecidual Seasonal Forest amidst the largest world savannah, the Cerrado biome. An analysis of the forest average productivity per basal area (m².ha), aerial live biomass (ton.ha-1) and carbon stock was carried out. The forest fragment was considered relatively rich in species and diversity, although showing signs of disturbance and dominance by a few species. Its horizontal structure suggests biotic regeneration conditions. It is an important reservoir of medicinal plants. Of the families (57.5%) presenting medicinal species, 19 from a total of 33 are represented in the area and contain 44% (27) of the total species (61) and 63% (432) of the total individuals catalogued. Medicinal species have ecological importance for the equilibrium of the local flora and represent 80% of the 10 species with higher Importance Value Index (IVI): Tetragastris altissima, Chrysophyllum marginatum, Oenocarpus distichus, Sclerolobium paniculatum, Simarouba versicolor, Alibertia macrophylla, Siparuna guianensis, Maprounea guianensis, Licania parvifolia e Physocalymma scaberrimum. Medicinal productivity was high for this type of phytophysionomy: 183,2 ton. ha-1 of biomass and 91,51 ton. ha-1 of carbon representing 66% of the total biomass and carbon of this Cerrado forest. From this stage S. guianensis (Siparunaceae) was selected for performing bioassays in order to verify its biological activity against microorganisms of health and agricultural relevance. This is a native aromatic medicinal plant recommended as priority for conservation, with local popular medicinal validation and availability of medicinal feedstock (3300 Kg.ha-1), with the foliar fraction giving 38Kg/ha of crude extract and 5L/ha of essential oil. Foliar crude extracts and essential oil were obtained and tested in vitro using a disk diffusion bioassay. Different concentrations of these natural products were tested against gram-positive bacteria (Staphylococcus aureus ATCC 29213), gram-negative bacteria (Escherichia coli ATCC 25922 and ATCC 35218; Pseudomonas aeruginosa ATCC 10145) and fungi (Candida albicans ATCC 6258 e Fusarium oxysporum). The essential oil inhibited the growth of S. aureus in its crude concentration (380μg.mL-1), as well as diluted to half (190μg.mL-1) and a quarter strength (95μg.mL-1). It’s likely that such action is due to sesquiterpenes major components, such as bisabolol and bisabolene (10.35%), measured by gas chromatography (GC-MS, GC-FID). Extracts did not exhibit any antimicrobial activity against the microorganisms tested. The native medicinal plants prospective market is an alternative that favours the conservation of biodiversity while generating benefits for the development of sustainable family productive activities within local ecosystems instead of the current inappropriate uses. This strengthens conservation policies of Legal Reserve in rural settlements and is in agreement with public policy on global warming and climate changes.
O estudo objetivou analisar floresta de área de Reserva Legal (RL) de savana brasileira na perspectiva da conservação, reservatório de carbono orgânico e biomassa medicinal para uso prospectivo das plantas medicinais nativas. Foi realizado levantamento etnobotânico e etnofarmacológico junto à comunidade assentada em área rural do Estado do Tocantins, sendo eleitas 9 espécies mais citadas (cajuí-Anacardium othonianum; inharé-Brosimum gaudichaudii; jatobá-Hymenaeae courbaril; jenipapo-Genipa americana, aroeira-Myracrodruon urundeuva; negramina-Siparuna guianensis; barbatimão-Stryphnodendron obovatum; assa peixe-Vernonia brasiliana, embaúba-Cecropia pachystachya). Foi feita a prospecção fitoquímica preliminar dos extratos brutos foliares e triagem dos metabolitos secundários potenciais de atividades antimicrobianas. Os compostos fenólicos, terpenos e flavonoídicos apresentaram positividade nos extratos da maioria das espécies, sugerindo atividades antimicrobianas, antioxidantes e contra insetos. A RL é criada por lei e ocupa localmente 35% da propriedade rural, tornando-se importante reservatório de biomassa medicinal, mas está sob tensão ecológica. Nela foi realizado inventário florestal de espécies lenhosas arbórea-arbustivas vivas usando o método alométrico, identificando raro remanescente de Floresta Estacional Semidecídua em meio à maior savana mundial, o Bioma Cerrado. Foi feita a análise da produtividade média da floresta pela área basal (m².ha), biomassa (ton.ha-1) aérea viva e estoque de carbono (ton.ha-1). O fragmento de floresta foi considerado relativamente rico em espécies e diversidade ainda mantida, mas com sinais de distúrbios e dominada por poucas espécies. Sua estrutura horizontal é sugestiva de condições de regeneração biótica. É um importante reservatório de plantas medicinais: mais da metade (57,5%) das famílias são de espécies medicinais, 19 de um total de 33; guardam 44% (27) do total de espécies (61) e 63% (432) do total de indivíduos (686) inventariados. As espécies medicinais têm importância ecológica para o equilíbrio da flora local, onde 80% estiveram representadas dentre as 10 espécies de maior Índice de Valor de Importância (IVI): Tetragastris altissima, Chrysophyllum marginatum, Oenocarpus distichus, Sclerolobium paniculatum, Simarouba versicolor, Alibertia macrophylla, Siparuna guianensis, Maprounea guianensis, Licania parvifolia e Physocalymma scaberrimum. A produtividade medicinal foi alta para este tipo de fitofisionomia: biomassa de 183,2 ton. ha-1 e carbono de 91,51 ton. ha-1 representando 66% de toda biomassa e carbono desta floresta de Cerrado. Desta etapa foi eleita S. guianensis (Siparunaceae) para realização de bioensaio objetivando verificar atividade biológica frente aos microorganismos de interesse da agricultura familiar e da saude, sendo uma espécie medicinal aromática nativa e recomendada como prioritária de conservação, com validação medicinal popular local e disponibilidade de matéria prima medicinal (3300 Kg.ha-1), conferindo a fração foliar 38Kg/ha de extrato bruto e 5L/ha de óleo essencial. Extratos brutos e óleo essencial foliar foram obtidos e testados em bioensaio in vitro feito por difusão em disco, utilizando diferentes concentrações dos produtos naturais frente a bactérias gram-positivas (Staphylococcus aureus ATCC 29213), bactérias gram-negativas (Escherichia. coli ATCC 25922 e ATCC 35218; Pseudomonas aeruginosa ATCC 10145) e fungos (Candida albicans ATCC 6258 e Fusarium oxysporum). O óleo essencial inibiu o crescimento bacteriano de S. aureus nas concentrações brutas (380μg. mL-1), diluído a metade (190μg.mL-1) e a um quarto (95μg.mL-1). É provável que tal ação seja devido aos componentes majoritários sesquiterpenos, como bisabolol e bisaboleno (10,35%), avaliados por cromatografia gasosa (CG-SM; CG-FID). Para os extratos não houve positividade quanto à ação antimicrobiana. Estudos prospectivos envolvendo plantas medicinais nativas podem ajudar na conservação da biodiversidade, gerando subsídios para o desenvolvimento de atividades produtivas familiares sustentáveis no âmbito de ecossistemas locais em lugar dos usos inadequados atualmente praticados. Isto fortalece a politica de conservação de RL em assentamentos rurais e está em sintonia com a política pública de aquecimento global e mudanças climáticas.
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Speda, Jutta. "Methods development for metaproteomics-guided bioprospecting of novel enzymes." Doctoral thesis, Linköpings universitet, Kemi, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-133206.
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Industrial biotechnology has been announced by several organizations and governments as a key enabling technology for the enhanced economic growth in a low-carbon and knowledge-based bioeconomy. An important goal to promote an environment friendly and sustainable industrial biotechnology is the discovery of new enzymes. To date, almost all enzymes used in industry have been discovered by pure culturing of microorganisms, however, it is known that less than 1% of all microorganisms can be obtained in pure cultures. The remaining majority of microorganisms is only viable by close biological interactions provided in microbial communities and is not available for enzyme discovery using the classical pure culture approaches. The investigation of microbial communities, which can be viewed as metaorganisms, has been enabled during the last two decades by refining established methods for the analysis of genes, mRNA or proteins and are called metagenomics, metatranscriptomics and metaproteomics, respectively. To date, these techniques have mostly been used in the field of microbial ecology for the understanding of the composition, function and metabolism of microbial communities but not for the purpose of bioprospecting for novel enzymes. Identification of genes that code for possible enzyme candidates is hindered, due to the fact that 30-40% of the sequenced metagenomes contain genes coding for unidentified proteins. Additionally, the -omics techniques generate large amounts of data that need to be analyzed and the outcome of the analysis does not necessarily lead to the discovery of novel applicable enzymes. The work presented in this thesis describes the establishment of the necessary conditions for a metaproteomics-based method that allows for a straightforward and targeted identification of novel enzymes with desired activity from microbial communities. The approach provides a valuable alternative to the incomplete and inefficient analysis of non-targeting data and laborious workflow, which is typically generated by the established meta-omics techniques. In developing the methods presented in this thesis, microbial communities in constructed environments were established, which allowed for the controlled expression of extracellular hydrolytic enzymes under defined conditions. By combination and modulation of advanced metaproteomics and metagenomics techniques, we were able to directly identify the enzymes and the corresponding gene sequences of several cellulolytic enzymes as a first example for the feasibility of this approach.
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Marques, Lana Grasiela Alves. "Mapping and legal approaches to bioprospecting networks in Brazil." Universidade Federal do CearÃ, 2014. http://www.teses.ufc.br/tde_busca/arquivo.php?codArquivo=12658.
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Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgicoCoordenaÃÃo de AperfeÃoamento de Pessoal de NÃvel Superior
FundaÃÃo de Amparo à Pesquisa do Estado do CearÃ
Este trabalho tem como propÃsito realizar o mapeamento das redes de bioprospecÃÃo no Brasil e as abordagens legais que envolvem o acesso aos recursos genÃticos para fins de bioprospecÃÃo. A BioprospecÃÃo se tornou uma das principais discussÃes nos Ãltimos anos desde que a ConvenÃÃo sobre Diversidade BiolÃgica (CDB) reconheceu a soberania de cada paÃs sobre os recursos genÃticos localizados em seu territÃrio. Ainda, o reconhecimento de cada paÃs signatÃrio da CDB na implementaÃÃo de polÃticas nacionais de biodiversidade. Para atender essas e outras exigÃncias, o Brasil estabeleceu aÃÃes voltadas à conservaÃÃo e ao uso sustentÃvel da biodiversidade por meio de programas e redes de pesquisa. Neste contexto, o trabalho identificou os avanÃos quanto à criaÃÃo e fortalecimento das redes de pesquisa em biodiversidade e os resultados obtidos por meio do desenvolvimento de produtos, em especial, a produÃÃo de biofÃrmacos. No entanto, a Medida ProvisÃria n 2.186-16/2001 considerada o marco regulatÃrio a respeito do acesso aos recursos genÃticos no Brasil estabeleceu normas que tem provocado entraves ao desenvolvimento das pesquisas bioprospectivas. Portanto, hà um senso comum da necessidade de um aprimoramento na Medida ProvisÃria n 2.186-16/2001 que, atualmente està em curso à elaboraÃÃo de um Projeto de Lei. Diante do exposto, espera-se que as redes e programas em bioprospecÃÃo possam transformar os recursos naturais em ganhos econÃmicos, alavancar o desenvolvimento cientÃfico e tecnolÃgico, e agregar valor aos bens e serviÃos provenientes desses recursos naturais.
This work has as its objective to map the bioprospection networks in Brazil, as well as the legal approaches involved in the access to genetic resources for bioprospection purposes. Bioprospection has become one of the main topics of discussion in recent years, since the Biological Diversity Convention (CBD in Portuguese) recognized the sovereignty of each country over genetic resources found within their territories. Moreover, the CDB recognized the right of each signatory country in the implementation of their own national biodiversity policies. In order to fulfil these and other demands, Brazil has established actions focused on the conservation and sustainable use of biodiversity through research programmes and networks. In this context, this work has identified the progress regarding the creation and strengthening of research networks in biodiversity, as well as the results obtained through the development of products, especially bio-pharmaceuticals. However, the Provisional Policy No. 2.186-16/2001, considered to be the regulatory framework regarding the genetic resources in Brazil, has established norms that have caused obstructions to the development of bioprospective research. As a result, there is a common agreement about the need to improve the Provisional Policy No. 2.186-16/2001, which is presently in course through a Law Bill. In face of these facts, it is hoped that the networks and programmes in bioprospection can transform the natural resources in economic gain, leverage the scientific and technological development and add value to the goods and services derived from these natural resources.
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Khera, Smriti. "Drug lead discovery through plant bioprospecting in Latin America." Diss., The University of Arizona, 2005. http://hdl.handle.net/10150/282898.
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The bioassay guided fractionation of two Latin American plants, structure elucidation of pure isolates, and LC/MS studies of six plant extracts is presented here along with the structure determination of two compounds using X-ray diffraction. The bioassay guided fractionation of the antibacterial and antitubercular CH2Cl2-MeOH extracts of the Argentinean plant Caiophora coronata Hook. et Arn. (Loasaceae) and the Chilean plant Myrcianthes coquimbensis (Barn.) Landrum et Grifo (Myrtaceae) respectively led to the isolation and complete structure elucidation of nine compounds from the active fractions. Three of these isolates were determined to be new. Namely, a new triterpene, 1beta,3beta-dihydroxyurs-12-en-27-oic acid, a new iridoid, 1alpha-methoxy-6alpha,10-dihydroxyisoepiiridomyrmecin (caiophoraenin) from C. coronata and a new monoterpene (1S,3 S,4R)-1-methyl-4-(1-methylethenyl)-1,3-cyclohexanediol from M. coqumibensis. All chemical structures were established unequivocally by physical and spectroscopic techniques including-melting point, optical rotation, 1D and 2D NMR, HR-FABMS, and FT-IR. Absolute configuration of the new monoterpene was established by Mosher's esterification. The antibacterial activity of all isolates from C. coronata were determined against methicillin-sensitive (MSSA) and -resistant (MRSA) strains of Staphylococcus aureus, Bacillus subtilis (BS), vancomycin-resistant Enterococcus faecium (VREF), Escherichia coli (EC), E. coli imp (ECimp), and Candida albicans (CA). 1beta,3beta-Dihydroxyurs-12-en-27-oic acid was found active against BS, MSSA, MRSA, VREF, and ECimp with MIC values of 2, 4, 4, 4 and 16 mug/ml respectively, whereas, other isolates were essentially inactive. The antitubercular activity of all isolates from M. coquimbensis was evaluated against M. tuberculosis using the microplate alamar blue assay. Oleanolic acid was determined to be the active principle of the extract with an MIC value of 29.66 mug/mL whereas other isolates were regarded as inactive (MIC > 128 mug/mL). Chemical investigations by LC/MS of species closely related to C. coronata and M. coquimbensis were also conducted. Structure solutions by single crystal X-ray crystallography, of an iridoid (4R,5R,7S,8S,9 S)-(-)-7-hydroxy-8-hydroxymethyl-4-methylperhydrocyclopenta[ c]pyran-1-one, and a fernane (3R,5S,9 R,10S,13S,14S,17 R,18R,21R)-(-)-fern-7-ene-3alpha-ol, isolated from the antitubercular methanolic extracts of Valeriana laxiflora DC (Valerianaceae) and Sebastiania brasiliensis Spreng. (Euphorbiaceae) respectively is also presented. The absolute configuration of these compounds was also determined.
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Flagg, Melissa L. "Bioprospecting, chemical investigations and drug discovery from Chilean plants." Diss., The University of Arizona, 2000. http://hdl.handle.net/10150/284167.
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This dissertation, completed as part of an International Cooperative Biodiversity Group (ICBG) Program, encompasses the field collection, taxonomic determination, bioassay-guided isolation, and chemical characterization of three plants native to Chile, each of which was collected using a distinct collection approach. Chuquiraga ulicina ssp. ulicina , collected by the ecological or environmental strategy, yielded ten compounds including four novel taraxastane-type triterpenoids, 3β-acetoxy-6β-hydroxytaraxasta-20-ene (1), 6β-hydroxytaraxast-20-en-3-one (2), 6β-hydroxytaraxasta-20-ene 3β-palmitate (3), and 3β, 6β-dihydroxytaraxasta-20-ene (4), together with the known triterpenoids lupeol (5), lupenyl acetate (6), lupenone (7), friedelinol ( 8), 3β-acetoxy-30-nor-lupan-20-one (9), and 30-norlupan-3β-ol-20-one (10). Lupeol (5) was the only compound to show antitubercular activity. Sphacele salviae, collected by the ethnobotanical or ethnomedical approach, allowed the isolation of three known compounds, including the two abietane diterpenoids carnosol ( 11) and rosmadial (12), as well as one pentacyclic triterpenoid, ursolic acid (13). Greigia sphacelata, collected according to the random approach, afforded nine compounds. These include the two novel flavanones 5,7,3'-trihydroxy-6,4' ,5'-trimethoxy flavanone (14) and 5,3'-dihydroxy-6,7,4',5 '-tetramethoxy flavanone (15), as well as four known phenylpropanoids, 1,3-O-di-trans-p-coumaroylglycerol (16), 1-O-trans-cournaroylglycerol (17 ), 1-(ω-feruloyldocosanoyl)glycerol and 1-(ω-feruloyltetracosanoyl)glycerol (18), and trans-ferulic acid 22-hydroxy docosanoic acid ester (19), and three known pentacyclic triterpenoids, arborinone (20), arborinol (21), and isoarborinol (22).
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Kagaba, James. "Bioprospecting for novel lipases from indigenous olive wastewater biofilms." Thesis, Cape Peninsula University of Technology, 2019. http://hdl.handle.net/20.500.11838/2828.
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Thesis (MTech (Food Technology))--Cape Peninsula University of Technology, 2018.Lipase-catalysed chemical transformations are today routinely considered by synthetic organic chemists as economical and competitive “green chemistry” alternatives. Although lipases can effortlessly be produced on a large-scale by fermentation, their industrial application was, until recently, limited to the detergent, oleo-chemistry and dairy industry. However, during the last few decades, the biotechnological application of lipases has expanded significantly, becoming indispensable in the manufacture of pharmaceuticals, pesticides, single cell protein production, biosensor preparations and waste management. Similarly, lipases have become a vital ingredient in the contemporary food processing industry with applications ranging from fruit juice production to baked foods, vegetable fermentations and dairy enrichment. Furthermore, lipases are routinely used as flavour development agents in cheese, butter and margarine products. Lipases are also applied in the leather industry for processing hides and skins and for treatment of activated sludge and other aerobic waste product treatments where its action enhances oxygen transfer. While lipases currently account for less than 21 % of the enzyme market, a growing interest in lipases is reflected by the publication of an average of 1000 research papers per year and the growing number of available lipases since the 1980s. There is a sustained interest to bioprospect for novel lipase enzymes from available unexplored biodiversity. This study aimed to screen for lipase-producing microorganisms resident in olive wastewater biofilms. Lipase activity of positive isolates was subsequently also quantitatively determined to select for the highest producers of true lipases. A Geotrichum candidum isolate from olive mill wastewater biofilms was selected for subsequent studies based on its superior lipase production phenotype. Using a yeast mediated ligation approach the G. candidum GCL1 lipase gene was cloned and heterologously expressed in Saccharomyces cerevisiae as an enzyme production host. The recombinant lipase was purified and analysed in terms of substrate specificity, pH optima, temperature optima and stability as well as organic solvent tolerance. The G. candidum gcl1 lipase presented enhanced thermo- and organic solvent-stability that are highly sought after traits for industrial application.
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Takeshita, Chikako. "Coordinates of Control: Indigenous Peoples and Knowledges in Bioprospecting Rhetoric." Thesis, Virginia Tech, 1999. http://hdl.handle.net/10919/41439.
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In this thesis, I draw attention to how representations of indigenous peoples and knowledges in the rhetoric of bioprospecting weave the people into multiple coordinates of discursive control. Bioprospecting, or the exploration of biological resources in search of valuable genetic and chemical material for commercial use, is portrayed by proponents as an ideal project which benefit all of its stakeholders. I challenge such perception by exposing the power relationships underlying bioprospecting proposals as well as the various interests built into their rhetoric. My particular interest lies in exploring the implications for indigenous peoples whose appearances in bioprospecting proposals are less than voluntary. I make three claims: (1) that the representation of indigenous peoples as stewards of the environment is a role assigned to them, which is then circulated and mobilized within the bioprospecting rhetoric in order to support its arguments concerning biodiversity conservation; (2) that indigenous knowledges of the environment, of medicinal plants in particular, are taken out of their original socio-cultural contexts, utilized, appropriated, and valorized by bioprospectors who construct the rhetoric; (3) that the visibility of indigenous peoples and knowledges, which was heightened as a result of the increased interest taken in controlling them, opens up new opportunities for the people to resist misappropriation and struggle for self-definition. In short, this project takes indigenous peoples and knowledges as the intersection of forces and interests comprising an intricate web of power relationships, within which any participant can attempt to empower oneself either by resisting or manipulating the control to which one is exposed.Master of Science
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Davis, Jason Michael. "Reconsidering Antarctic Bioprospecting through Territorialities of Science, Property, and Governance." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1299535648.
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Sweiss, Mais Ahed. "Microalgae for wastewater treatment and biomass production from bioprospecting to biotechnology." Thesis, University of Bath, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.760883.
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Improving wastewater (WW) treatment process is a major issue in different parts of the world. For a developed country like the UK where eutrophication is a problem that causes environmental and economical losses, and for a developing country like Jordan that is considered one of the most water scarce countries in the world, it is crucially important to improve the quality of the WW for safe reuse. Applying microalgae for WW treatment and biomass production is an economical and environmentally friendly method. However, this method has some challenges that need to be addressed, such as microalgae species selection, harvesting of the microalgae and the large area footprint. In this research, the overall aim was to bioprospect for microalgae that are adapted to the wastewater treatment plants (WWTPs) and evaluate the obtained microalgae depending on specific criteria for a successful application in high rate algal ponds (HRAPs), then there were attempts to improve the phosphorus removal in microalgae to increase the efficiency of the treatment process and reduce the area footprint. Bioprospecting for indigenous microalgae to the WW took place from January to May 2014. Water samples were collected from wastewater treatment plants (WWTPs) in the UK and Jordan. Eight different microalgae isolates were identified from each country. The results showed the Chlorella, Scenedesmus and Desmodesmus are common genera between the two countries and dominated the obtained isolates from the UK and Jordan. The isolates were identified using 18S rDNA and ITS1 5.8S ITS2 DNA barcoding markers. It was difficult to identify some of the isolates at the species level, as the 18S rDNA is too conserved to differentiate between the closely related species and due to the relatively poor representation of algae in GenBank. Then the obtained microalgae isolates were evaluated by their growth, efficiency in removing nutrients (i.e. nitrogen and phosphorus) and the settleability of the microalgae by gravity. Depending on the results the microalgae species were ranked to come up with some promising candidates to be applied on large scale. From the UK, Avonmouth_12 (Av_12) and Avonmouth_10 (Av_10) and from Jordan, Jordan_18 (Jo_18) and Jordan_29 (Jo_29) were distinguished in their performance in the WW. Since phosphorus is a major cause of eutrophication in the fresh water and it is important to reduce the level of phosphorus in the released WW to the legally permitted limits, this research aimed to study the possibility of improving phosphorus removal by microalgae. Using Chlamydomonas reinhardtii as a model to optimise the protocol to be applied in parallel with Av_12, which is a promising microalga isolate that has been applied on large scale in HRAPs in Beckington WWTP, the strategy was to overexpress a Phosphorus Starvation Response (PSR1) gene. The transformation process was successful in C. reinhardtii but not in Av_12. There was an enhancement of the specific phosphate removal rate in the transformed microalgae isolate CC 1010_B2 and CC 1010_A6 in comparison to the wild type strain CC 1010.
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Nitsch?Velasquez, Lucia. "Bioprospecting Three Plants from the Tropical Rainforest as Potential Antimicrobial Adjuvants." Thesis, State University of New York at Buffalo, 2019. http://pqdtopen.proquest.com/#viewpdf?dispub=13428243.
Full textAbstract:
The ‘resistance bacteria era’ is intrinsically related to the hospital acquired infections (HAI). The most frequent HAI causal agent in USA is multidrug resistant Staphylococcus aureus (MRSA) a priority of the Center for Disease Control for development for development of new drug treatments. A HAI key treatment is the oto and nephrotoxic aminoglycosides. One strategy is to enhance the antibiotic activity of antibiotics by combination with NPs, e.g., amoxicillin and the beta–lactamase inhibitor clavulanate.
Solutions for this urgent worldwide need can derive from bioprospecting species, specially from biodiversity rich countries e.g., the Guatemalan rainforests. Bioprospecting studies are envisioned under a business framework that are economic, social and eco–sustainable in the long term.
The enhancement of the bactericidal activity of commercially available aminoglycosides (GEN, VAN) by polar extracts from three Guatemalan rain forest plants were evaluated: the cosmetic oil producer palm Attalea cohune (Ac and fraction Ac11k), the Catholic relic Bourreria huanita (Bh), and the food spice Dysphania ambrosioides (Da). The antibiotics’ minimal bactericidal concentration (MBC) against MRSA–USA–300: Ac11k was reduced to 1/16MBCGEN at 9.9 Ac11k mg/mL (synergistic effects), and to 1/2MBCVAN at 94 Ac11k mg/mL (additive effects), and to 1/4MBCGEN at 136 mg/mL ethanolic extract of Bh. The Da–ascaridol–less leaves’ extracts reduced Erwinia carotovora doubling time from 17 min to 12.5 min. hinting out that they may be potentially useful for the probiotics’ industry.
With an emphasis in natural products dereplication by chemoinformatics tools, the experimental data gathered (HR–MS, FTIR, 1H–NMR spectroscopies) and the computational–assisted structure elucidation scheme were applied to derive the proposed structure of a new chemical entity probably present in Ac11k sample, Corozine A: a non–basic alkaloid with several putative ring types: (Z)-4-ethyl-1,2,6,7- tetrahydro-6,9-(methanoazenometheno)pyrrolo[2,1-d][1,5]oxazonine. Two additional potential targets related to sugars metabolism were found with ligand similarity search engines.
Corozine A and the other extracts studied are of interest for further research to improve its commercial exploitation. The vision of new commercial products derived from A. cohune would require a pilot project that re–engineers the extraction of fatty acids, essential oil, and alkaloids from the same raw material.
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Omardien, Soraya. "Bioprospecting for beta-glucosidases and beta-xylosidases from non-Saccharomyces yeast." Thesis, Stellenbosch : Stellenbosch University, 2013. http://hdl.handle.net/10019.1/80152.
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Thesis (MSc)--Stellenbosch University, 2013.ENGLISH ABSTRACT: The argument of whether to use food for biofuel (bioethanol) production prompted the search for an alternative non-food biomass, such as lignocellulose, as feedstock for bioethanol production. However, a hindrance in producing bioethanol from lignocellulose on an industrial scale is the cost associated with hydrolysing the lignocellulose to its respective sugar monomers. Improving enzyme production and enhancement of enzyme cocktails for efficient lignocellulose hydrolysis is, therefore, a necessary prerequisite. In this study, a yeast culture collection from the Wine and Fermentation Technology Division (ARC Infruitec- Nietvoorbij, Stellenbosch, South Africa), isolated from fruit from various regions in South Africa, was screened for β-glucosidase and β-xylosidase enzyme activities. β-glucosidases catalyse the hydrolysis of cellobiose and by doing so prevents end-product inhibition of cellobiohydrolases and endoglucanases during cellulose degradation. Similarly, β-xylosidases hydrolyse xylobiose and prevents end-product inhibition of endoxylanases during hemicellulose degradation. After initially screening 2180 non- Saccharomyces yeasts, two yeast isolates were selected that could potentially serve as enzyme source for lignocellulose hydrolysis; one as a producer of a β-glucosidase and another as a β-xylosidase producer. The yeasts were identified as a β-glucosidase producing Rhodotorula slooffiae-like yeast isolate 131B2 and a β-xylosidase producing Aureobasidium pullulans isolate 23B25, respectively. The production of β-glucosidase by Rhodotorula slooffiae-like yeast isolate 131B2 and of β-xylosidase by Aureobasidium pullulans isolate 23B25 was optimised using response surface methodology according to a central composite design. Subsequently, the crude and partially purified enzymes were characterised based on molecular mass, pH optima and stability, temperature optima and stability and inhibition by lignocellulose hydrolysis end-products, such as glucose, xylose and ethanol. The crude β-glucosidase from Rhodotorula slooffiae-like yeast isolate 131B2 was also compared to the commercial Aspergillus niger βglucosidase preparation (Novozyme 188) based on the characteristics mentioned above and as βglucosidase supplement during Avicel (microcrystalline cellulose) hydrolysis by the commercial cellulase preparation (Celluclast). The crude β-xylosidase by Aureobasidium pullulans isolate 23B25 could not be compared to a commercial β-xylosidase as none was available at the time of the study. During the study, the crude β-glucosidase 131B2 and β-xylosidase 23B25 showed potential as lignocellulose hydrolytic enzymes. Attempts were made to obtain the β-glucosidase and β-xylosidase genes from the respective yeast isolates using PCR-based approaches and by constructing cDNA libraries. However, cloning the β-glucosidase and β-xylosidase genes using these methods proved after several attempts to be unsuccessful, although, during this section of the study valuable information was obtained about the obstacles involved with using these approaches when the desired gene sequence is unknown and novel.
AFRIKAANSE OPSOMMING: Die debat oor die toepaslikheid van voedsel vir bio-brandstofproduksie (bio-etanol), het daartoe gelei dat alternatiewe nie-voedsel grondstowwe, soos lignosellulose, as voermateriaal vir bio-ethanol ondersoek word. Die koste geassosieer met die hidrolise van lignosellulose na die onderskeie suiker monomere belemmer industriële-skaal toepassing van lignosellulose vir bio-etanolproduksie. Verbeterde ensiemproduksie en verhoogde doeltreffendheid van ensiemmengsels vir lignosellulose hidrolise is dus ‘n noodsaaklik voorvereiste. In hierdie studie is 'n giskultuurversameling geisoleer vanaf vrugte van verskillende streke in Suid-Afrika deur die Wyn en Fermentasie Tegnologie Afdeling (ARC Infruitec-Nietvoorbij, Stellenbosch, Suid-Afrika) vir β-glukosidase en β-xilosidase ensiemaktiwiteite gesif. β-glukosidases wat die hidrolise van sellobiose kataliseer voorkom eindprodukinhibisie van sellobiohidrolases en endoglukanases tydens sellulose afbraak. β-xilosidases, op hul beurt, hydroliseer xilobiose en voorkom eindprodukinhibisie van endoxilanases tydens hemisellulose afbraak. Na afloop van die aanvanklike sifting van 2180 nie-Saccharomyces giste, is twee giste wat potensiëel as 'n ensiembron vir lignosellulose hidrolise kan dien geselekteer; een vir β-glukosidase en ‘n ander vir β-xilosidase produksie. Die giste is as ʼn β-glukosidase-produserende Rhodotorula slooffiaeagtige gisras 131B2 en ʼn β-xilosidase-produserende Aureobasidium pullulans gisras 23B25 onderskeidelik geïdentifiseer. Die Rhodotorula slooffiae-agtige gisras 131B2 se produksie van β-glukosidase en die Aureobasidium pullulans gisras 23B25 produksie van β-xylosidase was geoptimiseer met behulp van “response surface methodology” volgens 'n “central composite design”. Daarna was die gedeeltelik-gesuiwerde kru-ensieme volgens molekulêre massa, pH optima en stabiliteit, temperatuur optima en stabiliteit, en inhibisie deur lignocelluloses hidrolise end-produkte soos glukose, xylose en etanol, gekarakteriseer. Die kru βglukosidase van die Rhodotorula slooffiae-agtige gisras 131B2 is ook met die kommersiële Aspergillus niger β-glukosidase (Novozyme 188) volgens die eienskappe vroeër genoem vergelyk en as β-glukosidase aanvulling tydens die kommersiële sellulase (Celluclast) se hidrolise van Avicel (mikrokristalline sellulose). Die kru β-xylosidase van die Aureobasidium pullulans gisras 23B25 kon nie vergelyk word met 'n kommersiële β-xylosidase nie, aangesien daar nie een beskikbaar was tydens die studie nie. Gedurende die studie het altwee, die kru β-glukosidase 131B2 en β-xylosidase 23B25, potensiaal getoon as lignosellulose hidrolitiese ensieme. Pogings was aangewend om die β-glukosidase en β-xilosidase gene vanuit die onderskeie gis isolate met behulp van PKR-gebaseerde tegnieke en die opstel van cDNA biblioteke te kloneer. Hierdie klonering strategieë was egter na verskeie pogings onsuksesvol, maar waardevolle inligting oor die struikelblokke betrokke by die gebruik van hierdie benaderings wanneer die gewenste geen se DNS basispaarvolgorde onbekend en uniek is, was verkry.
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January, Grant Garren. "Bioprospecting for bioactive polysaccharides from marine algae endemic to South Africa." University of the Western Cape, 2016. http://hdl.handle.net/11394/5322.
Full textAbstract:
>Magister Scientiae - MScFucoidan is a marine-derived sulphated polysaccharide with bioactive properties ideal for the food, chemical and pharmaceutical industries. The polysaccharide consists largely of L-fucose, has a highly heterogeneous structure and is of diverse origin. Fucoidan was extracted from Ecklonia maxima, Laminaria pallida and Splachnidium rugosum and the effect of different extraction methods on fucoidan heterogeneity was assessed. Extraction methods employed hot water, hydrochloric acid or calcium chloride salt. Fucoidan yield and purity were determined by various colorimetric assays. Highest fucoidan yield was obtained with the hot water extraction method as seen by highest L-fucose content. Splachnidium rugosum extracts contained ~5 times more L-fucose than Ecklonia maxima and Laminaria pallida extracts. The salt extraction method yielded extracts free of contaminants, however L-fucose content in all extracts was >20 times lower. Acid extraction yielded highest levels of uronic acid contamination and liberated sulphate from the fucoidan polysaccharide. The fucose-to-sulphate ratio for Ecklonia maxima was approximately 1:5, whilst the ratios for Splachnidium rugosum and Laminaria pallida were approximately 1:1 and 1:2, respectively. The acid and salt extraction methods removed all traces of protein contaminants, while the hot water method retained very low levels of protein. The extraction method used to isolate fucoidan was a determining factor in yield and purity. Chemical compositional analyses of hot water extracts were assessed by gas chromatography mass spectroscopy. Splachnidium rugosum and Laminaria pallida extracts consisted largely of L-fucose, while Ecklonia maxima fucoidan was characterized with high glucose abundance. Crude hot water and acid extracts from Splachnidium rugosum tissue were fractionated and purified by (anionic) ion exchange chromatography as bioactivity has been correlated to lower molecular weight forms. In water extracts, ion exchange chromatography resulted in close to 90% decrease in L-fucose, sulphate and uronic acid, while protein content increased by 57%. Similar results were reported for acid extracts; however protein content did not change significantly. These results show that method of extraction may affect the composition of fucoidan post-purification. Hot water extraction is recommended due to higher fucoidan yield, as reflected by L-fucose content, and higher sulphate-to-fucose ratio. High protein content after ion exchange chromatography was however of concern. Since mucilage in Splachnidium rugosum thallus was free of protein, fucoidan was precipitated from mucilage with ethanol. Fucoidan yield of mucilage was >15-fold higher than content in purified hot water extracts with a sulphate-to-fucose ratio of ~1:1. The average molecular weight of native fucoidan in mucilage was estimated at 2367 kDa. The polysaccharide was hydrolysed by gamma-irradiation levels of 10-50 kGy to fractions ranging between 60 and 15.5 kDa. Hot water crude fucoidan extracts from Ecklonia maxima, Laminaria pallida, and Splachnidium rugosum were assessed for anti-oxidant activity by measuring the ability to scavenge free radicals and the capacity to reduce copper ions with 2,2-Diphenyl-1-picrylhydrazyl and Cupric Reducing Anti-oxidant Capacity assays, respectively. Ecklonia maxima crude fucoidan displayed highest anti-oxidant activity and capacity, having the potential to scavenge reactive oxygen species as well as the capacity to reduce copper to less toxic forms in mammalian systems. Splachnidium rugosum showed weakest anti-oxidant activity and lowest reducing capacity. The anti-cancer activity of crude and purified hot water Splachnidium rugosum extracts, as well as non-irradiated (native) and gamma-irradiated fucoidan, and commercially procured fucoidan were assessed for anti-cancer activity against MCF-7 breast cancer cells. Splachnidium rugosum crude and purified fucoidan displayed a half maximal inhibitory concentration of 0.7 mg/mL and 0.029 mg/mL, respectively. Low cytotoxicity of crude and purified Splachnidium rugosum fucoidan against non-cancerous breast epithelial cell line MCF-12A was observed, as seen by half maximal inhibitory concentration values of 2 mg/mL and 0.663 mg/mL, respectively. The cancer specific selectivity of purified Splachnidium rugosum fucoidan was therefore much higher as reflected by 10-fold higher selectivity index than that of crude fucoidan. Native and low molecular weight gamma-irradiated fucoidan also showed bioactive properties including anti-cancer activity as seen by the reduction of cell proliferation in vitro, whereas crude fucoidan showed the ability to scavenge free radicals, and the capacity to reduce copper ions.
National Research Foundation (NRF)
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Cummings, Matthew. "Harnessing synthetic biology for the bioprospecting and engineering of aromatic polyketide synthases." Thesis, University of Manchester, 2018. https://www.research.manchester.ac.uk/portal/en/theses/harnessing-synthetic-biology-for-the-bioprospecting-and-engineering-of-aromatic-polyketide-synthases(e2317dbb-c1b7-4e6e-83d5-03a1453848b2).html.
Full textAbstract:
Antimicrobial resistant microorganisms are predicted to pose an existential threat to humanity inside of the next 3 decades. Characterisation of novel acting antimicrobial small molecules from microorganisms has historically counteracted this evolutionary arms race, however the bountiful source of pharmaceutically relevant bioactive specialised metabolites discovered in the Golden era of drug discovery has long since dried up. The clinicians' arsenal of useful antimicrobials is diminishing, and a fresh perspective on specialised metabolite discovery is necessary. This call to action is being answered, in part, through advances in genome sequencing, bioinformatics predictions and the development of next generation synthetic biology tools aiming to translate the biological sciences into an engineering discipline. To expedite our route to new pharmaceutically relevant specialised metabolites using the synthetic biology toolbox several bottlenecks need to be addressed, and are tackled here in. Biosynthetic gene clusters (BGCs) represent blueprints to pharmaceuticals, however to date the vast wealth of knowledge about biosynthetic gene clusters is inconsistently reported and sporadically disseminated throughout the literature and databases. To bring the reporting of BGCs in line with engineering principles we designed and built a community supported standard, the Minimum Information about a Biosynthetic Gene cluster (MIBiG), for reporting BGCs in a consistent manner, and centralised this information in an easy to operate and open access repository for rapid retrieval of information, an essential resource for the bioengineer. Prioritisation represents the next bottleneck in specialised metabolite discovery. Bioinformatics tools have predicted a cache of thousands of BGCs within publicly available genome sequences, however high experimental attrition rates drastically slows characterisation of the corresponding specialised metabolite. We designed and built an Output Ordering and Prioritisation System (OOPS), to rank thousands of BGCs in parallel against molecular biology relevant parameters, pairing BGCs with appropriate heterologous expression hosts and facilitating a judicious choice of BGCs for characterisation to reduce experimental attrition. To fully realise the potential of synthetic biology in specialised metabolite discovery a genetically amenable heterologous host, capable of completing rapid design-build-test-learn cycles, is necessary. This cannot be achieved for the pharmaceutically important type II polyketides, as their biosynthetic machinery is largely restricted to Actinobacteria. Using MIBiG datasets, antiSMASH and BLASTP we identify 5 sets of soluble type II polyketide synthases (PKS) in Escherichia coli for the first time. We construct and test the robustness of a plug-and-play scaffold for bioproduction of aromatic polyketides using one PKS in E. coli, yielding anthraquinones, dianthrones and benzoisochromanequinones intermediates. Through bioprospecting for biological 'parts' to expand the chemical diversity of our plug-and-play scaffold we describe a new lineage of type II PKSs predominantly from non-Actinobacteria. The standards, softwares, and plug-and-play scaffold and biosynthetic 'parts' described here-in will act as an engine for rapid and automated bioproduction of existing, and novel, pharmaceutically relevant aromatic polyketides in E. coli using the synthetic biology toolbox.
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Tarasova, Yekaterina. "Protein engineering and bioprospecting for selective hydroxyacid production in engineered Escherichia coli." Thesis, Massachusetts Institute of Technology, 2016. http://hdl.handle.net/1721.1/107877.
Full textAbstract:
Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2016.Cataloged from PDF version of thesis.
Includes bibliographical references (pages 103-110).
Engineering of microbes can allow for the sustainable production of a variety of useful chemical compounds upon which we rely in our everyday lives. The many advantages of metabolic engineering include the use of renewable resources, mild reaction conditions, such as ambient pressure, pH and temperature, as well as the avoidance of toxic chemicals in the conversion process. The 3-hydroxyacid (3HA) pathway, also known as coenzyme-A (CoA) dependent chain elongation, can allow for the synthesis of dozens of diverse classes of chemicals. However, this pathway suffers from byproduct formation, which is due to the promiscuous activities of the pathway enzymes. On one hand, this enables the synthesis of such a diverse set of chemicals, but on the other, prohibits commercialization due to high downstream separation costs and low yields of desired longer chain products. The goal of this thesis was to improve this pathway though engineering and bioprospecting of more selective pathway enzymes. Engineering efforts were focused on the thiolase enzyme, which catalyzes the first step of the 3HA pathway, while bioprospecting efforts were focused on the enzyme immediately downstream of the thiolase - the 3-ketoacyl-CoA reductase. Our computationally guided protein engineering efforts identified a thiolase point mutant which allowed for more selective synthesis of longer chain 3HAs. Bioinformatics approaches were used to identify a set of ten diverse 3-ketoacyl-CoA reductases which were then screened and characterized within the context of the 3HA pathway. Two of the reductases led to increased selectivity for longer chain products. Combinations of the most selective thiolase and reductase enzyme pairs almost eliminated C4 byproduct formation by the 3HA pathway. In all, the thiolase and 3-ketoacyl-CoA reductase enzyme variants identified in this work should be applicable to other heterologous pathways where formation of short chain products is undesired, bringing such pathways closer to commercialization.
by Yekaterina Tarasova.
Ph. D.
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Vetter, Henning. "International and selected national law on bioprospecting and the protection of traditional knowledge." Thesis, University of the Western Cape, 2006. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_1427_1183465033.
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This thesis discussed the subjects of bioprospecting and the protection of traditional knowledge. At first the international approach to the subjects was elaborately discussed. The focus was on the respective provisions of the United Nations Convention on Biological Diversity and the related Bonn Guidelines, stressing the matter of access to genetic resources and the fair and equitable sharing of benefits arising from their utilization. Enclosed in this discussion was the examination of different legislatory approaches to tackle the subject with an emphasis on national intellectual property rights laws and the role and potential merit of national registers of and databases for specific traditional knowledge. The way national legislators have implemented the concerned obligations of the convention, and their peculiarities as for example the restriction of scope of law to indigenous biological resources, was exemplified with the respective Bolivian, South African as well as Indian laws.
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Christian, Nigel David. "From biopiracy to bioprospecting : an historical sociology of the search for biological resources." Thesis, University of Warwick, 2007. http://wrap.warwick.ac.uk/1122/.
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With the 1992 United Nations Convention on Biological Diversity humanity's ongoing search for biological resources became subject to global regulation. The collection of biological materials for use in agriculture and medicine by one nation from another became conditional on criteria of informed consent, benefit-sharing, and the preservation of environments. This practice has become known as bioprospecting. Collections of biological materials and/or of 'traditional' knowledge of how to utilize them which did not meet the Convention's requirements henceforth became known as biopiracy. The thesis takes its structure from the Convention, which is treated as marking a shift from historical biopiracy to contemporary bioprospecting. The thesis is that critics of the Convention who oppose it and the forms of bioprospecting which it mandates in terms of neo-colonialism and neo-imperialism have misunderstood the character of contemporary economic and political power. The thesis argues that although contemporary bioprospecting is not practiced, as the Convention requires it to be, in ways that are 'fair and equitable', it cannot be understood as a neo-imperialist practice. Instead, the thesis concludes that the Convention should be understood in the context of new forms of governance and sovereignty. The Convention facilitates planet management and supports the exercise of biopower. Several cases studies of imperialist biopiracy are presented and their social impacts are discussed in contrast to contemporary bioprospecting. A broad range of historical and sociological literature is brought together for the first time. The history of the transition from biopiracy to bioprospecting is described and discussed in terms of several social, epistemological/technological, scientific, political and economic changes, respectively: the transition from imperialism to globalization, a shift away from exploitation of 'nature' toward management of 'biodiversity', the transition from natural history to ecological science, the appearance of environmentalist concerns in national and global politics, the completion of the globalization of capitalist property relations and the demise of the notion of biological resources as the 'common heritage of humankind'.
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Chen, Xingchen. "Using structural biology and bioprospecting to design potent inhibitors of kallikrein-related peptidases." Thesis, Queensland University of Technology, 2019. https://eprints.qut.edu.au/128375/1/Xingchen_Chen_Thesis.pdf.
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Protein proteolysis occupies a pivotal niche in human biology where it controls a plethora of physiological and pathological processes. The goal of the studies in this thesis was to provide novel molecular templates for the design of modulators of proteolytic activity. Three distinct templates were investigated yielding powerful and specific modulators of enzymes with roles in ovarian cancer progression and skin homeostasis. These studies also predicate the utility of these templates in blockade of other proteolytic pathways.
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Corrêa, Wallace Ribeiro 1973. "Prospecção de substâncias bioativas em Pfaffia townsendii e Pfaffia tuberosa (Gomphreneae, Amaranthaceae)." [s.n.], 2014. http://repositorio.unicamp.br/jspui/handle/REPOSIP/317600.
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Orientador: Marcos José SalvadorTese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Pfaffias são plantas da família Amaranthaceae, tribo Gomphreneae, conhecidas como ginseng brasileiro e "Paratudo". São plantas com importância na medicina popular, sendo empregadas no tratamento de várias patologias e algumas espécies de Pfaffia apresentam estudos farmacológicos reportando atividade analgésica, antinociceptiva, anti-inflamatória, antimicrobiana e antitumoral, sugerindo potencial para a busca de agentes biologicamente ativos nestas matrizes vegetais. Assim, este estudo teve por objetivo a prospecção de extratos, frações e substâncias bioativas em Pfaffia townsendii Pedersen e Pfaffia tuberosa (Spreng.) Hicken que apresentam poucos estudos documentados na literatura e são espécies praticamente intocadas sob o ponto de vista fitoquímico e de avaliação de atividades biológicas. O estudo foi monitorado pela avaliação da atividade antioxidante (ensaios Folin-Ciocalteu, redução do radical DPPH e ORACFL); antiproliferativa (frente a linhagens de células tumorais e não tumorais); antimicrobiana (frente a bactérias, leveduras e protozoários); anti-inflamatória (in vitro pelo ensaio da desnaturação da albumina bovina (BSA) e in vivo empregando o modelo de edema de pata e pleurisia em camundongos induzida por carragenina). No estudo fitoquímico, primeiramente, obteve-se o perfil químico dos extratos bioativos (CCDC, CG, CLAE e ESI-MS), sendo possível identificar algumas substâncias conhecidas diretamente nos extratos brutos (desreplicação). Os extratos bioativos com substâncias que não puderam ser identificadas na etapa de desreplicação foram submetidos ao fracionamento biomonitorado e as substâncias isoladas foram identificadas utilizando-se métodos espectroscópicos de análise (UV, RMN 1H e RMN 13C) e espectrometria de massas e as substâncias isoladas foram submetidas à avaliação das atividades biológicas. Os extratos brutos e fases de partição dos extratos etanólicos apresentaram considerável atividade antioxidante nos três ensaios realizados, observando correlação positiva da capacidade antioxidante com o conteúdo de fenólicos totais solúveis. Estas amostras mostraram atividade antiproliferativa frente a linhagens de células tumorais, com valores de concentração mínima necessária para inibição total do crescimento celular, TGI, variando entre 0,42 e 250 ?g/mL para as amostras bioativas e não se mostraram citotóxicas para linhagem de células controle, não tumoral, VERO (TGI ? 250 ?g/mL). Ainda, exibiram atividade antimicrobiana frente a bactérias (com valores de concentração biocida mínima, CBM, entre 0,5 e 2,0 mg/mL), frente a fungos (CBM de 1,0 mg/mL) e frente a protozoários com atividade tripanocida moderada para formas tripomastigotas de Trypanosoma cruzi (com % de tripomastigotas viáveis variando de 4,70 a 69,40% para os extratos brutos a 4,0 mg/mL e para as fases de partição a 2,0 mg/mL) e com atividade leishmanicida para formas amastigotas-like de Leishmania (L.) amazonensis (com % de amastigotas viáveis variando de 0 a 8,10% para os extratos brutos a 1,0 mg/mL e para as fases de partição a 0,5 mg/mL). Os flavonoides isolados patuletina 3-O-B-D-glucopiranosídeo e tilirosídeo apresentaram promissora atividade antiprotozoário com efeito na viabilidade celular tanto para tripomastigotas de T. cruzi, quanto frente à amstigotas de L. (L.) amazonensis. Os extratos apresentaram atividade anti-inflamatória in vitro com inibição da desnaturação da proteína BSA (IC50 entre 41,1 e 400 µg/mL) e em modelo in vivo onde as fases etanólicas da partição do extrato etanólico das duas plantas estudadas e os flavonoides isolados patuletina 3-O-B-D-glucopiranosídeo e tilirosídeo inibiram o edema de pata induzido por carragenina e a migração de leucócitos na cavidade intrapleural induzido por carragenina, sendo também mensurado o extravasamento de proteínas pela reação de Bradford. O estudo fitoquímico monitorado pelas atividades biológicas e antioxidantes mostrou-se efetivo, sendo possível identificar cinco esteroides, três triterpenoides, três acetatos de triterpenoides, quatro ácidos orgânicos e três flavonoides nas amostras bioativas. Pode-se também isolar dois flavonoides (Patuletina 3-O-B-D-glucopiranosídeo e Tilirosídeo). Os resultados obtidos justificam, pelo menos em parte, a utilização popular de espécies do gênero Pfaffia e confirma o potencial da família Amaranthaceae como fontes de moléculas bioativas e insumos para a saúde, sendo importante a continuidade dos estudos com as amostras bioativas mais promissoras com vistas a aplicações farmacológicas, biotecnológicas e clínicas
Abstract: Pfaffias are plants of the Amaranthaceae family, Gomphreneae tribe, known as Brazilian ginseng and "paratudo". Plants have great importance in folk medicine, being used in the treatment of various diseases. Some species of Pfaffia exhibit pharmacological studies reporting anti-nociceptive, anti-inflammatory, antimicrobial, analgesic and antitumor activities and suggesting a potential for biologically active agents in these vegetable matrices. Thus, this study aimed to explore for extracts, fractions and bioactive compounds in Pfaffia townsendii Pedersen and Pfaffia tuberosa (Spreng.) Hicken, which show little documented data in the literature studies and are species virtually untouched from the point of view of phytochemical evaluation of biological activities. The study was monitored by the evaluation of the activities: antioxidant (Folin-Ciocalteu assays, reducing the DPPH and ORAC-FL); antiproliferative (against strains of tumor and non-tumor cells); antimicrobial (against bacteria, yeasts and protozoa); anti-inflammatory (in vitro assay for denaturation of bovine serum albumin (BSA) and in vivo study using the model of paw edema and carrageenan-induced pleurisy in mice). In the first step, the phytochemical, it was obtained the chemical profile of the bioactive extracts (TLC, GC, HPLC and ESI-MS), and it was possible to identify some known substances directly from crude extracts (dereplication). The bioactive extracts with substances that could not be identified in the dereplication step were subjected to bioassay-guided fractionation and isolated substances were identified using spectroscopic analysis methods (UV, 1H and 13C NMR) and mass spectrometry and isolated compounds were subjected to evaluation of bioactivity. Extracts and phases of partition of ethanol extracts showed significant antioxidant activity in the three trials, observing positive correlation between antioxidant capacities with the phenolic content. These samples showed antiproliferative activity against tumor cell lines with minimum concentration values required for complete inhibition of cell growth, TGI, ranging between 0.42 and 250 ug/mL for bioactive samples and were not cytotoxic to the cell line control, non-tumor VERO (TGI ? 250 ug/mL). They also showed antimicrobial activity against bacteria (with values of Minimum Biocidal Concentration, MBC, between 0.5 and 2.0 mg / ml), against fungi (MBC 1.0 mg mL) and trypanocidal moderate activity against protozoans such as trypomastigote forms of Trypanosoma cruzi (% viable with trypomastigotes ranging from 4.70 to 69.40% for the gross 4.0 mg/mL and extract phases partition to 2.0mg/mL), leishmanicidal activity for axenic amastigotes of Leishmania (L.) amazonensis (% viable with amastigotes ranging from 0 to 8.10% for the crude at 1,0 and 0.5 mg/mL extracts). The flavonoids isolated patuletin 3-O-B-D-glucopyranoside and tiliroside showed promising antiprotozoal activity and effect on cell viability for both T. cruzi tipomastigotas, and amastigotas of L. (L.) amazonensis. The extracts exhibited anti-inflammatory activity in vitro with inhibition of denaturation of BSA protein (IC50 between 41.1 and 400 ug/mL), where the ethanolic phases for the ethanol extract partition and flavonoids isolated patuletin 3-O-B-D-glucopyranoside and tiliroside inhibited in vivo the paw edema induced by carrageenan and the migration of leukocytes in the pleural cavity induced by carrageenan. The extravasation of proteins were also measured by Bradford reaction. The phytochemical study was monitored by biological and antioxidants activities, and was possible to identify five steroids, three triterpenoids, three acetates triterpenoids, four phenolic acids and three flavonoids in bioactive samples. It is as well possible to isolate two flavonoids (patuletin 3-O-B-D-glucopyranoside and tiliroside). The results obtained in this study justify, at least in part, the popular use of the genus Pfaffia and confirm the potential of the Amaranthaceae family as a source of bioactive molecules and its inputs to health. It is important to continue on the studies with the most promising bioactive samples, with goals on the pharmaceutical, biotechnological and clinical applications
Doutorado
Fármacos, Medicamentos e Insumos para Saúde
Doutor em Biociências e Tecnologia de Produtos Bioativos
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Kabir, Kamaluddeen. "Bioprospecting surfactants produced by Pseudomonas spp. isolated from soil for potential application in biotechnology." Thesis, Abertay University, 2017. https://rke.abertay.ac.uk/en/studentTheses/1911389c-5bea-4ed8-9ac3-dce4f6ff645b.
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Bacteria produce a range of surface-active compounds called biosurfactants that reduce the surface tension of liquid and exhibit different oil-water behaviours. These are used in various biotechnological applications including agriculture, cosmetics, medical and food. A recent study has predicted a limit to bacterial surface tension-reducing ability. If this limit exists, it has strong negative consequences in surveys for more active compounds. In this work, the aim is to (i) investigate this prediction more robustly by using chemical media and (ii) study the diversity amongst the best-performing surfactants produced by Pseudomonas spp. with the intention of finding novel surfactants that could be used in different biotechnological applications. A total of 251 Pseudomonas spp. were isolated from soil. Strains were first screened for liquid surface tension-reducing ability (LSTRA) using qualitative drop-collapse assay before quantitative surface tension measurement. Of the 58 LSTRA strains, only 46 significantly reduced the surface tension of sterile media. Individual Distribution Identification (IDI) analysis was used to determine the predicted limit for surfactant activity in KB* and M9Glu media, and results were found to be in agreement with earlier studies. To investigate the chemical structural diversity amongst the best performing surfactants, a collection of 25 key strains producing a limited range of very low surface tension in liquid culture media (~24 – 26 mN/m) were examined. Initial phenotypic characterisation including biochemical, metabolic profiling and 16S rDNA sequencing confirmed strains were a diverse collection of Pseudomonas spp. A series of behaviour assays including emulsion formation, foam stabilisation and oil displacement assays to investigate behavioural diversity among surfactants expressed by the key strains were then undertaken. For the oil displacement, diesel, mineral, vegetable, and used lubricating oils were tested with the underlying aqueous layer containing 0 or 200 mM NaCl at pH 6.0 or 8.0 to reflect a range of biotechnological applications and conditions. Analysis of variance of the emulsion indices, foam stabilisation and oil displacement data showed significant difference in surfactant behaviour among the key surfactant-expressing strains (P < 0.001). Moreover, Hierarchical Cluster Analysis (HCA) was used to produce a constellation dendrogram in which isolates were grouped according to similarities in phenotype and surfactant behaviour. Critically, this resulted in more groups (≥ 5 groups) than could be explained by statistically significant differences in mean surface tensions (previously determined by ANOVA and Tukey-Kramer HSD, alpha = 0.05). These findings provide strong evidence that the key strains were expressing structurally more than one type of surfactant with differing air-water and oil-water behaviours. Similarly, in vitro surfactant characterisation within a range of pH and salt concentrations confirmed diversity among strains (P < 0.001). Investigating surfactant potential by a two-way behaviour cluster dendrogram resulted in more diversity among oil types than the conditions used. These findings indicate that bioprospecting surfactants by screening only the more active compounds is likely to reveal a range of functionalities.
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Seini, Monica Michelle, and n/a. "Bioprospecting and Access to Indigenous Flora: Policy Implications of Contested Ways of 'Knowing' and 'Owning'." Griffith University. School of Science, 2005. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20060302.122535.
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This thesis critically explores the issue of access to biological resources and Indigenous knowledge Historically, biological resources collected and documented, and knowledge associated with their use, have been considered the 'common heritage of mankind' The Convention on Biological Diversity (CBD) changed this understanding to tights of states over biological resources, but also gave rise to issues of equity and justice, especially with regard to Indigenous Nations encapsulated within First World states-so-called 'Fourth World Nations', A central concern of Fourth World Peoples is their marginalisation within access negotiations, despite their claims of connate (birth) rights to r esou.r ces and knowledge they identify as their own. Increasing global Indigenous activism over their concerns, has in turn raised an increasingly important policy gap that is becoming recognised in fora and processes with regard to access to biological resources. My thesis addresses this policy gap. I explore some of the complex historical, political and cultural dimensions that led to the emergence and resilience of this policy problem The failure to address the concerns of Indigenous peoples, and Fourth World Nations in particular, is more important and problematic now because of contemporary biotechnological developments and the emergence of bioprospecting. Bioprospecthg refers to the practice of appropriating biological resources, and Indigenous knowledge of those resources, and incorporating them into biopharmaceutical processes. Literature on bioprospecting as a problematic issue for Third World States has been emerging steadily over the last decade under the impact of the commercialisation of biodiversity, which has become big business for biopha.rmaceutical companies. The unique interests and experiences of Fourth World Nations are not recognised within this literature as significantly different to that of the Third World, and of their encapsulating states.. This study has addressed this significant gap by utilising and developing an analytical approach that uses Fourth World theory, synthesised with elements of Foucault's analytics of power. When combined, these two theoretical approaches provide a new and rich under standing of how dominant 'ways of knowing' and 'ways of owning' have been privileged, while other knowledge and ownership systems have been, and continue to be, marginalised, Eoucault's understanding of discursive power as having the capability to be either, or both, dominant and resistant is important to my analysis, as it accommodates the Fourth World as a discursive site of resistance to dominant power. I posit that richer insights are gained through the development and application of this theoretical framework to the issue of fair and equitable access to biological resources, than other approaches offer. I demonstrate the framework's utility by applying it to a case study on bioprospecting in Australia. Important findings have emerged while tracking the activities of Fourth World peoples on the international stage, and their attempts to challenge dominant power/knowledge structures within political institutions For example, participation at the international level has enabled Fomth World peoples to apply pressure on their encapsulating states to accommodate their interests. This has been furthered through forming alliances with, for example, environmentalists, and through the adoption of the language of effective participation within international fora.. Overall, however, the study found that the participation of Eourth World peoples within international, central state and local state policy processes is not always empowering in challenging dominant interests Instead, the more accurate impression is that at this stage of the discursive policy terrain, it may only create an illusion of participation that actually serves to entrench their disempowerment. This places pressule on policy processes to address and resolve this access issue equitably if social turbulence is to subside, justice be served, and certainty provided for all.
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Tomko, Timothy. "Bioprospecting For Genes That Confer Biofuel Tolerance To Escherichia Coli Using A Genomic Library Approach." ScholarWorks @ UVM, 2015. http://scholarworks.uvm.edu/graddis/487.
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Microorganisms are capable of producing advanced biofuels that can be used as 'drop-in' alternatives to conventional liquid fuels. However, vital physiological processes and membrane properties are often disrupted by the presence of biofuel and limit the production yields. In order to make microbial biofuels a competitive fuel source, finding mechanisms of improving resistance to the toxic effects of biofuel production is vital. This investigation aims to identify resistance mechanisms from microorganisms that have evolved to withstand hydrocarbon-rich environments, such as those that thrive near natural oil seeps and in oil-polluted waters.
In this study, screened the genomes of two types of bacteria, Pseudomonas aeruginosa and Marinobacter aquaeolei, looking for genes that impart biofuel tolerance when expressed in Escherichia coli. Both of these microbes have adapted in their respective natural environments to contain mechanisms for dealing with environmental stress. For initial work, P. aeruginosa was used to test our experimental design and procedure, and we validated our methods by identifying a gene, ohr from P. aeruginosa, that increased tolerance to the bio-jet fuel precursor limonene in Escherichia coli.
Using genomic DNA from M. aquaeolei, we constructed a transgenic library that we expressed in E. coli. We exposed cells to inhibitory levels of pinene, a monoterpene that can serve as a jet fuel precursor with chemical properties similar to existing tactical fuels. Using a sequential strategy of a fosmid library followed by a plasmid library, we were able to isolate a region of DNA from the M. aquaeolei genome that conferred pinene tolerance when expressed in E. coli. We determined that a single gene, yceI, was responsible for the tolerance improvements. Overexpression of this gene placed no additional burden on the host. We also tested tolerance to other monoterpenes and showed that yceI selectively improves tolerance.
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Tomko, Timothy. "Bioprospecting For Genes That Confer Biofuel Tolerance To Escherichia Coli Using A Genomic Library Approach." ScholarWorks @ UVM, 2017. http://scholarworks.uvm.edu/graddis/798.
Full textAbstract:
Microorganisms are capable of producing advanced biofuels that can be used as ‘drop-in’ alternatives to conventional liquid fuels. However, vital physiological processes and membrane properties are often disrupted by the presence of biofuel and limit the production yields. In order to make microbial biofuels a competitive fuel source, finding mechanisms for improving resistance to the toxic effects of biofuel production is vital. This investigation aims to identify resistance mechanisms from microorganisms that have evolved to withstand hydrocarbon-rich environments, such as those that thrive near natural oil seeps and in oil-polluted waters.
First, using genomic DNA from Marinobacter aquaeolei, we constructed a transgenic library that we expressed in Escherichia coli. We exposed cells to inhibitory levels of pinene, a monoterpene that can serve as a jet fuel precursor with chemical properties similar to existing tactical fuels. Using a sequential strategy of a fosmid library followed by a plasmid library, we were able to isolate a region of DNA from the M. aquaeolei genome that conferred pinene tolerance when expressed in E. coli. We determined that a single gene, yceI, was responsible for the tolerance improvements. Overexpression of this gene placed no additional burden on the host. We also tested tolerance to other monoterpenes and showed that yceI selectively improves tolerance.
Additionally, we used genomic DNA from Pseudomonas putida KT2440, which has innate solvent-tolerance properties, to create transgenic libraries in an E. coli host. We exposed cells containing the library to pinene, selecting for genes that improved tolerance. Importantly, we found that expressing the sigma factor RpoD from P. putida greatly expanded the diversity of tolerance genes recovered. With low expression of rpoDP. putida, we isolated a single pinene tolerance gene; with increased expression of the sigma factor our selection experiments returned multiple distinct tolerance mechanisms, including some that have been previously documented and also new mechanisms. Interestingly, high levels of rpoDP. putida induction resulted in decreased diversity. We found that the tolerance levels provided by some genes are highly sensitive to the level of induction of rpoDP. putida, while others provide tolerance across a wide range of rpoDP. putida levels. This method for unlocking diversity in tolerance screening using heterologous sigma factor expression was applicable to both plasmid and fosmid-based transgenic libraries. These results suggest that by controlling the expression of appropriate heterologous sigma factors, we can greatly increase the searchable genomic space within transgenic libraries.
This dissertation describes a method of effectively screening genomic DNA from multiple organisms for genes to mitigate biofuel stress and shows how tolerance genes can improve bacterial growth in the presence of toxic biofuel compounds. These identified genes can be targeted in future studies as candidates for use in biofuel production strains to increase biofuel yields.
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Seini, Monica Michelle. "Bioprospecting and Access to Indigenous Flora: Policy Implications of Contested Ways of 'Knowing' and 'Owning'." Thesis, Griffith University, 2005. http://hdl.handle.net/10072/366804.
Full textAbstract:
This thesis critically explores the issue of access to biological resources and Indigenous knowledge Historically, biological resources collected and documented, and knowledge associated with their use, have been considered the 'common heritage of mankind' The Convention on Biological Diversity (CBD) changed this understanding to tights of states over biological resources, but also gave rise to issues of equity and justice, especially with regard to Indigenous Nations encapsulated within First World states-so-called 'Fourth World Nations', A central concern of Fourth World Peoples is their marginalisation within access negotiations, despite their claims of connate (birth) rights to r esou.r ces and knowledge they identify as their own. Increasing global Indigenous activism over their concerns, has in turn raised an increasingly important policy gap that is becoming recognised in fora and processes with regard to access to biological resources. My thesis addresses this policy gap. I explore some of the complex historical, political and cultural dimensions that led to the emergence and resilience of this policy problem The failure to address the concerns of Indigenous peoples, and Fourth World Nations in particular, is more important and problematic now because of contemporary biotechnological developments and the emergence of bioprospecting. Bioprospecthg refers to the practice of appropriating biological resources, and Indigenous knowledge of those resources, and incorporating them into biopharmaceutical processes. Literature on bioprospecting as a problematic issue for Third World States has been emerging steadily over the last decade under the impact of the commercialisation of biodiversity, which has become big business for biopha.rmaceutical companies. The unique interests and experiences of Fourth World Nations are not recognised within this literature as significantly different to that of the Third World, and of their encapsulating states.. This study has addressed this significant gap by utilising and developing an analytical approach that uses Fourth World theory, synthesised with elements of Foucault's analytics of power. When combined, these two theoretical approaches provide a new and rich under standing of how dominant 'ways of knowing' and 'ways of owning' have been privileged, while other knowledge and ownership systems have been, and continue to be, marginalised, Eoucault's understanding of discursive power as having the capability to be either, or both, dominant and resistant is important to my analysis, as it accommodates the Fourth World as a discursive site of resistance to dominant power. I posit that richer insights are gained through the development and application of this theoretical framework to the issue of fair and equitable access to biological resources, than other approaches offer. I demonstrate the framework's utility by applying it to a case study on bioprospecting in Australia. Important findings have emerged while tracking the activities of Fourth World peoples on the international stage, and their attempts to challenge dominant power/knowledge structures within political institutions For example, participation at the international level has enabled Fomth World peoples to apply pressure on their encapsulating states to accommodate their interests. This has been furthered through forming alliances with, for example, environmentalists, and through the adoption of the language of effective participation within international fora.. Overall, however, the study found that the participation of Eourth World peoples within international, central state and local state policy processes is not always empowering in challenging dominant interests Instead, the more accurate impression is that at this stage of the discursive policy terrain, it may only create an illusion of participation that actually serves to entrench their disempowerment. This places pressule on policy processes to address and resolve this access issue equitably if social turbulence is to subside, justice be served, and certainty provided for all.Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Science
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Adeoyo, Olusegun Richard. "Bioprospecting for amylases, cellulases and xylanases from ericoid associated fungi, their production and characterisation for the bio-economy." Thesis, Rhodes University, 2018. http://hdl.handle.net/10962/64327.
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Pereira, Andréia Mara 1976. "Bioprospecção e conhecimentos tradicionais : uma proposta institucional para sua gestão no Brasil." [s.n.], 2013. http://repositorio.unicamp.br/jspui/handle/REPOSIP/286103.
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Orientador: Bastiaan Philip ReydonTese (doutorado) - Universidade Estadual de Campinas, Instituto de Economia
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Resumo: A presente tese de doutorado teve como foco apresentar uma proposta de criação de uma instituição gestora para o uso da biodiversidade e dos conhecimentos tradicionais associados à biodiversidade, para as atividades de bioprospecção no Brasil. Para tanto, identificaram-se alguns dos elementos que fazem parte das práticas de bioprospecção, ressaltando as especificidades dos biomas brasileiros e apresentando as peculiaridades dos povos dos conhecimentos tradicionais, indígenas e não indígenas. A tese abordou algumas distinções entre o conhecimento tradicional e o conhecimento científico, tendo em vista que as práticas de bioprospecção envolvem a gestão de acordos que utilizam recursos da biodiversidade, mas também agentes com interesses nem sempre convergentes. Para melhor visualização deste framework, foram apresentadas as características das atividades econômicas da bioprospecção e foram analisados estudos de casos internacionais e do Brasil, com base na Economia dos Custos de Transação aplicados a estes acordos de bioprospecção. Dentro dos casos analisados, identificou-se que há necessidade da criação de instituições intermediárias e de mecanismos pelo governo federal para facilitar a interação entre os agentes. Neste sentido, como resultado, o estudo propõe o desenho de um modelo institucional para a biodiversidade e conhecimentos tradicionais associados para as práticas de bioprospecção no Brasil
Abstract: This doctoral thesis focused on presenting a proposal to establish an institution for managing the use of biodiversity and traditional knowledge associated with biodiversity, bioprospecting activities in Brazil. For this, we identified some of the elements that compose the bioprospecting practices, emphasizing the specificities of Brazilian biomes and explaining the singularities of traditional indigenous and non-indigenous peoples and knowledge. We have also addressed some distinctive features between traditional knowledge and scientific knowledge. All considering that bioprospecting practices involve the management of agreements, which use biodiversity resources, but also agents who not always show converging interests. To provide a better understanding of this framework, we present the characteristics of economic activities related to bioprospection and analyze national and international case studies based on the Transaction Cost Economics applied to these bioprospecting agreements. Based on the cases analyzed, we realized that the Federal Government should create intermediate institutions and mechanisms to facilitate the interaction between the agents. In this regard, and as a result, the study proposes the design of an institutional model for the use of biodiversity and associated traditional knowledge with bioprospecting practices in Brazil
Doutorado
Desenvolvimento Economico, Espaço e Meio Ambiente
Doutora em Desenvolvimento Econômico
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Orozco, Rousel Antonio. "Characterization of the Entomopathogenic Bacterium Photorhadus Luminescens Sonorensis, and Bioactivity of its Secondary Metabolites." Thesis, The University of Arizona, 2012. http://hdl.handle.net/10150/228614.
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Photorhabdus are motile Gram-negative bacteria that have a mutualistic association with entomopathogenic Heterorhabditis nematodes. Nematodes vector the bacteria from one insect host to another, while the bacterial symbiont produces toxins and secondary metabolites that kill that the insect host. In this study, we characterize the bacterial symbiont of Heterorhabditis sonorensis, recently discovered in the Sonoran desert. Biochemical and molecular methods including sequence data from five genes: 16s rDNA, gyrB, recA, gltX, dnaN were considered. Evolutionary relationships of this new Photorhabdus subsp. were inferred considering maximum parsimony and Bayesian analyses. We also surveyed for secondary metabolites (SM) produced by this microorganism, considering HPLC and mass spectrometry analyses. SM crude extracts showed activity against the corn ear worm Helicoverpa zea, the root-knot nematode (Meloidogyne incognita), the bacterium Pseudomonas syringae, and the fungus Fusarium oxysporum; and were more toxic that those produced by related species. Results from these studies showed that Photorhabdus l. sonorensis' secondary metabolites have potent antagonistic activity against these plant pathogens.
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Andrade, Luísa Carvalho. "Bioprospecção de microrganismos marinhos isolados na Baía de Todos os Santos com atividade antagonista a bactérias patogênicas." Instituto de Ciências da Saúde, 2014. http://repositorio.ufba.br/ri/handle/ri/23505.
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A Baía de Todos os Santos (BTS), segunda maior baía do Brasil, abriga grande diversidade de ambiente, flora e fauna, porém pouco se sabe sobre sua diversidade microbiológica e seu potencial de bioprospecção. Este trabalho teve por objetivo a busca de novas substâncias antimicrobianas, capazes de inibir o crescimento de bactérias patogênicashumanas e de animais marinhos, isolados do solo de cinco praias da BTS. No total foram isoladas 107 bactérias das quais dez apresentaram produção de substâncias antimicrobianas. Após o sequenciamento verificou-se que dos dez isolados, somente oito foram de fato identificadas molecularmente, sendo sete destes membros da família Bacillaceae, grupo ao qual pertencem diversas produtoras de substâncias antimicrobianas. Entre os isolado que apresentaram produção de substâncias inibidoras do crescimento constatou-se que os isolados 21, 50 e 54 tinham maior poder de inibição. O extrato destes três isolados foi confeccionado através da acidificação do sobrenadante livre de células, isolado após o período de incubação. Os extratos do isolado 21 e do isolado 50 foram capazes de inibir a bactéria Klebsiella rizophyla, e o extrato 21 conseguiu ainda inibir a Vibrio vulnificus. Após verificação molecular através de primers específicos de sete substâncias antimicrobianas produzidas por Bacillus, constatou-se que os isolados 21 e 50 são produtores da substância ericina.
The All Saints´s Bay (BTS), the second largest bay in Brazil, where can be founded great diversity of flora and fauna including diversity of landscapes, but unknown about their microbial diversity and potential to bioprospecting. This study aimed to search for new antimicrobial substances that can be able to inhibit the growth of human pathogenic bacterias and marine animals, isolated from sediment from five BTS beaches. Was isolated a total of 107 bacterias where 10 showed antimicrobial substances production. After 16S sequencing, was found that from the ten isolates, only eight were identified by amplification PCR and seuqencing, seven members of the Bacillaceae family, where antimicrobial substances belonging this group are known. It was found that the isolates numbers 21, 50 and 54 had a greater power of inhibition. The extract of these three strains was made by acidification of the cell-free supernatant, isolated after the incubation period. The extracts of the isolate 21 and 50 were able to inhibit the bacteria Kocuria rhizophyla , and the same extract could also inhibit Vibrio vulnificus. In advance, was realized a molecular bioprospecting using specific molecular primers from conserved regions belongs to seven antimicrobial substances produced by Bacillus. It was found that the isolates 21 and 50 are Erycina producing.
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PANZERI, DAVIDE. "A Bioprospecting Multidisciplinary Approach to Valorise Biodiversity: The Case of Bowman-Birk Protease Inhibitors in Vigna unguiculata (L.) Walp." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2023. https://hdl.handle.net/10281/404605.
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La biodiversità naturale è un’importante risorsa per l’uomo sin da tempi antichi. Tuttavia, sta sperimentando un critico declino e molte specie sono a rischio estinzione. Il cambiamento climatico e l’attività umana sono i maggiori drivers di una conservazione e gestione del territorio non sostenibili. L’opportunità di ammortizzare gli effetti del cambiamento climatico è fornita da specie indigene, domesticate in contesti locali ma poco investigate.
In questo contesto, questa tesi mira a sviluppare strategie per valorizzare la biodiversità naturale, considerando diversi e integrati aspetti scientifici. Il primo obiettivo è quello di riscoprire un legume tradizionale africano, Vigna unguiculata (L.) Walp., e verificare la sua adattabilità a condizioni di stress tipiche del cambiamento climatico o pratiche agricole poco esigenti. La ricerca di composti bioattivi è un valore aggiunto al fine di dare coscienza della potenzialità della specie. Per questa ragione, i successivi obiettivi della tesi sono l’esplorazione della diversità genetica di composti bioattivi dei legumi, gli inibitori delle proteasi Bowman-Birk (BBIs), e la valutazione delle proprietà nutraceutiche.
Un design sperimentale multidisciplinare è stato applicato integrando diversi approcci per creare un flusso di lavoro coerente. Un esperimento di campo e analisi di produzione e metaboliti è stato organizzato per dimostrare l’adeguatezza di Vigna unguiculata come coltivazione per il cambiamento climatico. La diversità genetica di questa specie è stata esplorato con tecniche di biologia molecolare e analisi computazionali e filogenetiche. Le proprietà nutraceutiche sono state determinate grazie a procedure biochimiche e test su modelli, cellulari e in vivo, di cancro ed invecchiamento.
Dal punto di vista della coltivazione, Vigna unguiculata può essere considerata una specie poco esigente in termini di richiesta d’acqua e pratiche agronomiche. Ciò rende questo legume adatto a pratiche di agricoltura conservativa in paesi in via di sviluppo o in quei paesi colpiti fortemente dal cambiamento climatico. Questo legume si dimostra importante come risorsa di macronutrienti essenziali, e, per promuovere la sua diffusione a livello globale, abbiamo indagato anche la presenza di molecole con azione dirette per la salute umana. L’esplorazione genetica ha considerato quasi 200 accessioni, tovando13 diverse isoforme di BBI tra accessioni selvatiche e domesticate distribuite sul continente Africa e in altre parti del mondo. In aggiunta, abbiamo sviluppato una metodica estrattiva e purificative che ha permesso l’isolamento e caratterizzazione delle singole isoforme di BBI. La dimostrazione di attività correlate a BBI nei diversi modelli, rende questa famiglia di proteine un valore aggiunto per la salute umana. L’azione verso linee cellulari tumorali suggerisce possibili applicazioni terapeutiche anche in sinergia con farmaci d’elezione. Ciò apre opportunità per la futura ricerca in specie e generi affini e la valutazione di isoforme maggiormente efficaci anche in sistemi in vivo.
Concludendo, questo progetto dimostra che i) la bioprospezione per la ricerca di molecole bioattive in specie regionali è un importante passo per la loro salvaguardia, ii) conoscere evoluzione e diversificazione di piante di interesse è uno strumento per migliorare azioni bioprospettive e identificare migliori varianti di composti bioattivi, iii) analisi di efficacia funzionale in vitro e in vivo è un passaggio fondamentale per dedicare ricerca scientifica al miglioramento della biodiversità in contesti operativi. Quest’ultima è una fase importante per stimolare investitori, sia privati che pubblici, al fine di portare valore economico e sociale alla conservazione della biodiversità.Natural biodiversity is an important source for humans since ancient times. However, biodiversity is experiencing a dramatic decline and many species are at extinction risk. Climate change and human activity are the main drivers of non-sustainable landscape conservation and management. The opportunity to dampen climate change effects is provided by indigenous species, domesticated in local contexts but are little investigated. In this framework, this PhD thesis aims at developing strategies to valorise natural biodiversity, considering different integrative scientific aspects. The first objective of this thesis is the rediscovery of a traditional African legume, Vigna unguiculata (L.) Walp., and assess its adaptability to stressful conditions typically caused by climate change or undemanding agricultural practices. Moreover, the research for bioactive compounds is an added value to give consciousness of the species potential. For this purpose, exploration of genetic diversity of known legume bioactive compounds, the Bowman-Birk protease inhibitors (BBIs) and appraisal of their nutraceutical properties are the second objectives of the project. A multidisciplinary experimental overview has been applied by integrating different approaches to create a coherent workflow. The demonstration of Vigna unguiculata L. as suitable species for climate change was carried out with a field experiment and subsequent laboratory analyses to evaluate production parameters and metabolic features. The genetic diversity of this species was explored through molecular biology techniques and in silico computational and phylogenetic analyses. The nutraceutical features were established by biochemical procedures and cellular biology by testing compounds on different ageing and cancer models. From the point of view of cultivation needs, it is possible to consider V. unguiculata (L.) Walp. as undemanding in terms of water demand and agronomic practices. This makes this legume suitable for conservation agriculture practices in developing countries and where climate change is having a dramatic impact on indigenous crop. This legume is also an important resource of essential macronutrients and to enhance this species and promote its cultivation globally, we also wanted to focus on the presence of bioactive molecules with direct action on humans. The genetic exploration considered almost 200 accessions and found 13 isoforms of BBI were identified in different wild and cultivated accessions, distributed in the African continent and in other areas of the world. Furthermore, we managed to develop an extraction and purification procedure to isolate single isoforms and characterise them. Our data suggest that V. unguiculata BBIs possess a great natural genetic and biochemical diversity. Moreover, the demonstration of BBI-related bioactivities on different models makes them very promising as a high-value natural compound for human wellbeing. The direct action on different tumour cell lines suggests a possible therapeutic application also in synergy with some drugs (i.e. Cetuximab). This opens opportunities for future research on similar related species and genera, and on the analyses to evaluate the most effective isoforms also in in vivo systems. In conclusion, this PhD project demonstrates that i) bioprospection of local species directed to the search for bioactive molecules represents an important lever for safeguarding; ii) the knowledge of evolution and diversification of the plants of interest is a tool to improve bioprospecting actions and identify molecular variants of bioactive compounds; iii) analyses of functional efficacy of bioactive compounds in in vitro and in vivo systems is a fundamental step to dedicate scientific research to the enhancement of biodiversity in an operational context. This is an essential phase to stimulate private investors and businesses to bring economic and social value and biodiversity conservation.
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Smith-Baedorf, Holly D. "Microalgae for the biochemical conversion of CO2 and production of biodiesel." Thesis, University of Bath, 2012. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.564010.
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As the global population rises to an estimated 9.4bn by 2050, the pressure for food, fuel and freshwater will continue to increase. Current renewable energy technologies are not widely applicable to the transport sector, which requires energy dense liquid fuels that drop into our existing infrastructure. Algal biofuels promise significantly higher yields than plants, without the displacement of valuable agricultural resources and have the potential to meet the global demand for transport fuel. Fossil fuel energy is largely ‘a legacy of algal photosynthesis’, with algae accounting for ~50% of global CO2 fixation today. In addition, these curious organisms show remarkable diversity in form, behaviour and composition. Recently there has been a global resurgence of interest in microalgae as a resource of biomass and novel products. With the present level of technology, knowledge and experience in commercial scale aquaculture, the capital cost and energy investment for algal biomass production is high. Culturing, harvesting and disrupting microalgal cells account for the largest energy inputs with more positive energy balances requiring low energy designs for culture, dewatering and extraction, efficient water and nutrient recycling with minimal waste. Little is known about the variable cell wall of microalgae, which presents a formidable barrier to the extraction of microalgal products. Staining, transmission electron microscopy (TEM) and enzymatic digestion were all utilised in an attempt to visualise, digest and characterise the cell wall of stock strains of Chlorella spp. and Pseudochoricystis ellipsoidea. The presence of algaenan, a highly resistant biopolymer, rendered staining and enzymatic digestion techniques ineffective. TEM revealed that algaenan is present in the outer walls of microalgae in a variety of conformations which appeared to impart strength to cells. A preliminary investigation utilising Fusarium oxysporum f.sp. elaeidis as a novel source of enzymes for the digestion of algaenan has also been described. Methods were developed for the mutagenesis of Chlorella emersonii and P. ellipsoidea using EMS and UV with the intent of generating cell-wall mutants. Although no viable cell wall mutants were produced, a viable pale mutant of C. emersonii was recovered 5 from UV mutagenesis. Growth rates of the pale mutant were significantly slower than the wild type, yet FAME profile was largely unaffected. Fluorescence activated cell sorting (FACS) was also investigated as a means for the rapid screening of mutagenized cells for cell wall mutants. In an attempt to reduce cooling costs of closed-culture systems, temperature tolerant species of microalgae were sought by bioprospecting the thermal waters of the Roman Baths. Numerous methods for isolation and purification of microalgae from the Baths were employed, ultimately yielding seven diverse isolates including cyanobacterial, eukaryotic, filamentous and single celled species. Despite some species possessing an increased tolerance to higher temperatures, none showed marked temperature tolerance coupled with high productivity. Further improvements to the culture conditions may have improved the productivity at higher temperatures. All seven isolates were deposited to the Culture Collection of Algae and Protozoa (CCAP). A variety of extraction methods including soxhlet, beadbeating, sonication and microwaving was investigated for efficacy of extracting fatty acid methyl esters (FAMEs) from C. emersonii. Beadbeating proved most effective in the extraction of FAMEs from C. emersonii. Microwaving showed potential as a rapid method of extraction yet was coupled with degradation of FAMEs, requiring further method development to resolve this issue. Method development has been a significant component of the work described in this thesis.
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Fonseca, Cristine da. "Fenologia e caracterização fitoquímica do óleo essencial de Tagetes minuta L. (Asteraceae)." Universidade Federal de Pelotas, 2018. http://guaiaca.ufpel.edu.br:8080/handle/prefix/4209.
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As agriculturas de base ecológica têm como pressupostos o conhecimento, o uso e a preservação da biodiversidade local. O Brasil é um país formado por seis Biomas distintos e detentor de ampla biodiversidade e, assim, é fundamental considerar a influência dos fatores ambientais na composição fitoquímica dos óleos essenciais. Não só como um apelo comercial, mas também como uma estratégia de sustentabilidade dos agroecossistemas, o cultivo de plantas aromáticas para extração de óleo essencial ou uso como técnicas de manejo em sistemas de produção de base ecológica constitui importante estratégia para o desenvolvimento rural sustentável. Neste sentido, o estudo objetivou caracterizar as fases fenológicas de Tagetes minuta L., conforme a época de plantio, correlacionando os fatores edafoclimáticos ao desenvolvimento e composição fitoquímica do óleo essencial da espécie. Para tanto, a pesquisa foi realizada em duas etapas distintas, a primeira referiu-se à bioprospecção de populações de Tagetes minuta L. em dez municípios da região sul do Rio Grande do Sul, onde o método utilizado para extração do óleo essencial foi a destilação água-vapor. A segunda etapa tratou da instalação, condução e avaliação do experimento realizado na Embrapa Clima Temperado, Estação Experimental Cascata, Pelotas, RS (31°37’15” S; 52°31’24” O; 180 m a.n.m.). O plantio foi realizado em quatro épocas distintas para determinação das fases fenológicas da espécie e a extração do óleo essencial realizada pelo método de hidrodestilação. O óleo essencial oriundo de plantas coletadas e cultivadas caracterizou-se pela presença dos compostos majoritários cis-tagetona (37,47%), cis-β-ocimeno (35,76%), dihidrotagetona (14,19%), trans-tagetona (3,24%) e limoneno (3,16%). O ciclo de desenvolvimento da época 2 foi 29 dias mais curto que a época 1. As duas primeiras épocas de plantio 20 de dezembro e 18 de janeiro alcançaram o estadio de desenvolvimento reprodutivo no mesmo período. Considerando o alto rendimento de óleo essencial e o tempo de duração do ciclo, a melhor época para o cultivo de Tagetes minuta L. foi 18 de janeiro.
Ecological-based farms have as their presuppositions the knowledge, use and preservation of local biodiversity. Brazil is a country made up of six distinct Biomes and has a wide biodiversity and, therefore, it is fundamental to consider the influence of environmental factors on the phytochemical composition of essential oils. Not only as a commercial appeal but also as a sustainability strategy for agroecosystems, the cultivation of aromatic plants for extraction of essential oil or use as management techniques in ecologically based production systems is an important strategy for sustainable rural development. In this sense, the study aimed to characterize the phenological phases of Tagetes minuta L., according to the planting season, correlating the edaphoclimatic factors to the development and phytochemical composition of the essential oil of the species. In order to do so, the research was carried out in two distinct stages, the first one referring to the bioprospection of Tagetes minuta L. populations in ten municipalities in the southern region of Rio Grande do Sul, where the method used to extract the essential oil was distillation water vapor. The second stage dealt with the installation, conduction and evaluation of the experiment performed at Embrapa Temperate Climate, Cascata Experimental Station, Pelotas, RS (31 ° 37'15 "S, 52 ° 31'24" W, 180 m a.m.). The planting was carried out in four distinct seasons to determine the phenological phases of the species and the extraction of essential oil by the hydrodistillation method. The essential oil from the collected and cultivated plants was characterized by the presence of cis-tagetone (37.47%), cis-β- ocimene (35.76%), dihydrotagetone (14.19%), trans-tagetone (3.24%) and limonene (3.16%). The development cycle of season 2 was 29 days shorter than season 1. The first two planting seasons December 20 and January 18 reached the stage of reproductive development in the same period. Considering the high yield of essential oil and the duration of the cycle, the best time for the cultivation of Tagetes minuta L. was January 18.
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Júnior, Fábio Lino Soares. "Descrição e caracterização de uma nova ?-N-acetil-hexosaminidase (GH3) por metagenômica de solo de manguezal." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/64/64133/tde-27012016-141443/.
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Bactéria e fungos são as principais fontes de enzimas envolvidas na transformação de compostos chave para o fluxo de carbono em solos de manguezal, caracterizado por alta prevalência de anaerobiose, salinidade e elevado teor de matéria orgânica. A decomposição de plantas ou resíduos de animais nestas condições é muito lenta, devido à pressão seletiva sobre a evolução de enzimas envolvidas nos processos de mineralização de nutrientes. A metagenômica, permiti o acesso a grande maioria da diversidade microbiana no ambiente, por meio da geração de bibliotecas de clones, o que resulta em um cenário promissor para bioprospecção de novas atividades enzimáticas. Neste estudo, foi relatada a descrição e caracterização de uma nova ?-N-acetil-hexosaminidase (EC 3.2.1.52) da família GH3, envolvida na degradação da matéria orgânica em solo de manguezal contaminado por derramamento de óleo localizado no município de Bertioga-SP, por meio de uma triagem de 12.960 clones metagenômicos. O clone positivo para a atividade celulolítica foi sequenciado e um total de 1.175.586 reads foram gerados com tamanho médio de 198 pb. As sequencias foram trimadas com base na qualidade de índice PHRED >= 30.0, e remoção de sequencias do hospedeiro (E. coli) e do vetor (fosmídeo), originando um contig final com 39.586 Kb. Entre as ORF\'s anotadas a partir do contig gerado, uma sequencia de 1.065 nucleotídeos foi identificada como codificante para a enzima ?-N-acetil-hexosaminidase, evidenciando baixa similaridade (32 %) com as demais encontradas no bancos de dados comparativos. A enzima foi expressa e purificada, onde uma banda isolada foi visualizada por SDS-PAGE com massa molecular prevista de 43 kDa. Por fim, as atividades ótimas da enzima (30 °C; pH 5.0; 0.5 M de NaCl; diminuição de atividade após 3hs de incubação) foram caracterizadas por meio do indicador p-nitrophenol (pNP) ligados aos substratos GlNac, GalNac e Glc. A detecção da enzima por meio da metagenômica, evidenciou que os manguezais são reservatórios de novas enzimas com características diferenciadas e altos potenciais de aplicabilidades biotecnológicasBacteria and fungi are major sources of enzymes involved in the transformation of key compounds for the carbon fluxes on mangrove soils, characterized by the high prevalence of anaerobiosis salinity and high content of organic matter. The decomposition of plant or animals residues under these conditions is very slow, acting as a selective pressure on the evolution of enzymes involved in the mineralization process of nutrients. Metagenomics has provided access to the vast majority of the microbial diversity in the environment through the generation of fosmid libraries, resulting in a promising scenario for bioprospection enzymatic activities. In this study, we report the description and characterization of a novel ?-N-acetylhexosaminidase (EC 3.2.1.52) of GH3 family, involved in the degradation of organic matter in mangrove soils contaminated by oil spill located in the city of Bertioga-SP through of a screening of 12.960 metagenomic clones. The positive clone for cellulolytic activitie was sequenced and a total of 1.175.586 reads were generated with measuring size 198 bp. The sequences were trimmed based on the index of quality PHRED >= 30.0 and removing the sequences to host (E. coli) and vector (fosmid) resulting in a contig of 39.586 Kb. Between the anoted ORF\'s from generated contig a sequence of 1.065 nucleotides was identified coding for a ?-N-acetylhexosaminidase showing low similatrity (32 %) with the other found in comparatives databases. The enzyme was expressed and purified where an isolated band can be visualized by SDS-PAGE with molecular mass of 43 kDa. Finally, as optimum activity of the enzyme (30 °C; pH 5.0; 0.5M NaCl; decreased activity after 3 h incubation) were characterized by the indicator p-nitrophenol (pNP) linked to the substrates GlNac, GalNac and Glc. The detection of the enzyme through metagenomics indicated that mangroves are reservoirs of novel enzymes with different characteristics and high potential for biotechnological applicability
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Crivelente, Horta Maria Augusta 1981. "Análise do transcriptoma de Trichoderma harzianum para a bioprospecção de enzimas hidrolíticas = Analysis of Trichoderma harzianum transcriptome for bioprospecting of hydrolytic enzymes." [s.n.], 2014. http://repositorio.unicamp.br/jspui/handle/REPOSIP/316510.
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Orientadores: Anete Pereira de Souza, Sindélia Freitas AzzoniTese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Buscando contribuir com o desenvolvimento da tecnologia de produção do etanol de segunda geração, o presente estudo analisa o transcriptoma de T. harzianum IOC-3844 utilizando técnicas de sequenciamento high-thoughput. O principal objetivo dessas análises foi identificar, caracterizar e catalogar os transcritos expressos por T. harzianum relacionados com a degradação de substratos complexos, como o bagaço de cana de açúcar, revelando o conjunto de genes envolvidos na degradação da biomassa. A análise do transcriptoma do fungo Trichoderma harzianum sob condições que induzem a degradação da biomassa permitiu a identificação de sequências de genes potencialmente eficazes no processo de biodegradação, uma etapa essencial à compreensão do processo de hidrólise enzimática. O sequenciamento resultou em 246 milhões de sequências com 72 pb, o que corresponde a 14,7 GPB analisados. Após a montagem , 32.494 contigs foram gerados, submetidos à identificação e classificados de acordo com sua identidade. Todas as sequências de contigs foram comparados com o banco de dados do NCBI, Gene Ontology (GO terms), Enciclopédia de Genes Kyoto (KEGG), Carbohydrate Active-Enzymes (CAZYmes). Foram identificados 487 CAZymes no transcriptoma, inclusive aquelas ligadas as reações químicas de despolimerização de celulose e hemicelulose. As sequências classificadas como atividade catalítica (6.975) e atividade reguladora (143) podem estar envolvidas com esse tipo de reação.A análise permitiu definir o principal conjunto de genes envolvidos na degradação da celulose e de hemicelulose do T. harzianum , e genes acessórios relativos à despolimerização de biomassa. Uma análise dos níveis de expressão permitiu determinar os conjuntos de genes diferencialmente expressos em diferentes condições de cultivo. Os resultados obtidos acrescentam conhecimento sobre a constituição do genoma, as atividades de expressão gênica do fungo Trichoderma harzianum e fornece informações importantes a respeito dos mecanismos genéticos de degradação de biomassa que o fungo utiliza. As informações obtidas poderão ser utilizadas para outras espécies de fungos filamentosos com potencial para a biodegradação
Abstract: In order to contribute to the development of second-generation ethanol technology, this study analyzes the transcriptome of T. harzianum IOC-3844 using high-thoughput sequencing techniques. The main objective of this analysis was to identify, characterize and catalog the transcripts expressed by T. harzianum related to the degradation of complex substrates such as sugar cane bagasse, revealing the set of genes involved in the degradation of biomass. The analysis of the transcriptome of the fungus Trichoderma harzianum under conditions that induce the degradation of biomass allowed the identification of genes potentially effective in the biodegradation process, an essential step for understanding the enzymatic process. Sequencing resulted in 246 million sequences with 72 bp, which corresponds to 14.7 GBP analyzed. After assembly, 32,494 contigs were generated, identified and classified according to their identity. All sequence contigs were compared with NCBI database, Gene Ontology (GO terms), Kyoto Encyclopedia of Genes (KEGG), Carbohydrate Active-Enzymes (CAZYmes). 487 CAZymes were identified in the transcriptome, including those related to reactions of cellulose and hemicellulose depolymerization. Sequences classified as catalytic activity (6,975) and regulatory activity (143) may be involved with this type of reaction. This analysis define the set of genes involved in the degradation of cellulose and hemicellulose of T. harzianum, and accessories genes related to depolymerization of the biomass. An analysis of expression levels was used to calculate the set of differentially expressed genes in different culture conditions. The results add to knowledge about the composition of the genome and gene expression activity of the fungus Trichoderma harzianum, and provides important information regarding the genetic mechanisms of biomass degradation that the fungus uses. The information obtained may be used for other species of filamentous fungi with potential for biodegradation
Doutorado
Genetica Vegetal e Melhoramento
Doutora em Genética e Biologia Molecular
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Alexandre, Artur Ribeiro de Sá. "Bioprospecção de bactérias quitinolíticas e caracterização da atividade da enzima quitinase." Universidade Federal de Goiás, 2018. http://repositorio.bc.ufg.br/tede/handle/tede/8471.
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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
Bioprospecting is defined as the search for organisms, genes, enzymes, compounds, processes and any pieces from living beings, that could have economic potential, and eventually lead to a product development. Thus, enzymes, protein catalysts, have aroused the interest of industries for its conversion mode specific biomass, low energy cost and not production of toxic waste. Chitinases are enzymes that catalyze the lysis of chitin (biopolymer composed of N-acetylglucosamine monomers), and thus have several applications, among them: obtaining N-acetylglucosamine monomers, used in the production of prebiotics; degradation of chitinous waste from fishing activities; and the use in control of fungi and insects. The present work aimed to bioprospect chitinase producing bacteria from soil samples and to characterize the enzymatic activity patterns of the best bacterial isolate. To do so, a screening with 17 soil samples collected in the states of Minas Gerais, Santa Catarina and Rio Grande do Sul, was carried out using a culture medium with colloidal chitin. Thirteen chitinase producing bacteria were obtained, among them, the isolate Q1 (identified as Paenibacillus illinoisensis), demonstrated to be a good producer of the enzyme and thus was selected for determination and optimization of its chitinase activity evaluating reaction time, temperature and pH. The chitinase produced by the isolate showed 0.098 U of activity, which was subsequently improved to 0.66 U when tested under the optimal conditions of 1 hour of reaction at 37 ºC and pH 4, an increase of 573% over the initial value. The values of chitinase activity from the isolate P. illinoisensis are close to those found in other studies, which also emphasize the potential application of the enzyme, mainly in the control of phytopathogenic pests. Bioprospecting of chitinase producing bacteria is possible and promising.
A Bioprospecção é definida como a busca por organismos, genes, enzimas, compostos, processos e partes provenientes de seres vivos, que possam ter potencial econômico, e eventualmente, levar ao desenvolvimento de um produto. Nesse âmbito, as enzimas, catalisadores biológicos de natureza proteica, têm despertado o interesse de indústrias pelo seu modo de conversão de biomassa específico, de baixo custo energético e que não produz resíduos tóxicos. As quitinases são enzimas que catalisam a quebra da quitina (biopolímero composto por monômeros de N-acetilglicosamina), possuindo assim, diversas aplicações, dentre as quais: a obtenção de monômeros de Nacetilglicosamina, usados na produção de pré-bióticos; a degradação de resíduos quitinosos oriundos da pesca e do consumo de crustáceos; e uso no combate de fungos e insetos. O presente trabalho teve como objetivo bioprospectar bactérias produtoras de quitinase a partir de amostras de solo e caracterizar a enzima do melhor isolado bacteriano quanto aos padrões de atividade enzimática. Para tal, foi realizada uma triagem com 17 amostras de solo coletadas nos estados de Minas Gerais, Santa Catarina e Rio Grande do Sul, utilizando meio de cultura com quitina coloidal. Foram obtidas 13 colônias bacterianas produtoras de quitinase, dentre elas, o isolado Q1 (identificado como Paenibacillus illinoisensis), que se mostrou bom produtor da enzima e foi selecionado para testes de determinação e otimização da atividade enzimática quanto ao tempo de reação, temperatura e pH. A quitinase produzida pelo isolado apresentou atividade de 0,098 U, sendo melhorada posteriormente para 0,66 U quando testada nas condições ótimas de 1 hora de reação, a 37 ºC e em pH 4, um aumento de 573% em relação ao valor inicial. Os valores obtidos da atividade da qutininase produzida pela P. illinoisensis são próximos aos encontrados em outras pesquisas, que destacam também, o potencial de aplicação da enzima, principalmente no combate de pragas fitopatogênicas. A bioprospecção de bactérias produtoras de quitinase é possível e promissora.
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GUERRIERI, MARIA CHIARA. "Bioprospecting di simbionti vegetali con proprietà PBS per lo sviluppo di nuovi prodotti biostimolanti: bridging tra i risultati della ricerca e gli aspetti normativi." Doctoral thesis, Università Cattolica del Sacro Cuore, 2021. http://hdl.handle.net/10280/95717.
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L'agricoltura moderna sta affrontando sfide come la perdita di fertilità del suolo, la variabilità climatica e gli attacchi di agenti patogeni in continuo aumento. Le pratiche agricole si stanno evolvendo verso sistemi sostenibili e rispettosi dell'ambiente. L'uso di biostimolanti (PBS, plant biostimulant) è una soluzione innovativa per affrontare le sfide di un’agricoltura sostenibile che garantisce un assorbimento ottimale dei nutrienti, una resa delle colture e tolleranza agli stress abiotici. In particolare, tra i diversi tipi di biostimolanti presenti sul mercato, i rizobatteri, classificati come Plant Growth Promoting Rhizobacteria (PGPR), offrono un nuovo approccio per promuovere la crescita delle piante, la mitigazione degli stress e l’aumento della resa colturale. Pertanto i PGPR sono considerati come una sorta di "probiotici" vegetali, poiché contribuiscono in modo efficiente alla nutrizione e all'immunità delle piante. L'obiettivo principale di questa tesi è isolare e identificare batteri presenti nella rizosfera di pomodoro (Solanum lycopersicum L.) che mostrano proprietà PBS, nonché valutare i meccanismi coinvolti nell'azione di promozione della crescita delle piante (Capitolo 2) e la genetica alla base di questi meccanismi (Capitolo 3 e 4). Infatti, una profonda comprensione dei meccanismi d’azione dei PGPR potrebbe colmare la mancanza di coerenza del dato di efficacia tra gli studi di laboratorio e gli studi in campo e stimolare la ricerca per la produzione e la commercializzazione di nuovi prodotti biostimolanti microbici.Modern agriculture faces challenges such as loss of soil fertility, fluctuating climatic factors and increasing pathogen and pest attacks. Agricultural practices have been evolving towards organic, sustainable and environmentally friendly systems. The use of natural plant biostimulants (PBS) is an innovative solution to address the challenges in sustainable agriculture, to ensure optimal nutrient uptake, crop yield, quality and tolerance to abiotic stress. In particular, among different types of biostimulants present on the market, plant growth promoting rhizobacteria (PGPR) offer a novel approach for promoting plant growth, mitigate stress and increase crop yield. Hence, PGPR inoculants are now considered as a kind of plant ‘probiotics’, since they efficiently contribute to plant nutrition and immunity. The main goal of this thesis was to isolate and identify bacteria symbionts of tomato (Solanum lycopersicum L.) rhizosphere, which showed PBS properties and evaluate mechanism involved in the action of PGPR (Chapter 2), underlying genetics and physiological pathways (Chapter 3 and 4). Indeed, a deeply understanding of the mechanisms of plant growth promotion, could fulfill the lack of consistency between lab, greenhouse and field studies, and support commercialization of novel plant biostimulant products.
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GUERRIERI, MARIA CHIARA. "Bioprospecting di simbionti vegetali con proprietà PBS per lo sviluppo di nuovi prodotti biostimolanti: bridging tra i risultati della ricerca e gli aspetti normativi." Doctoral thesis, Università Cattolica del Sacro Cuore, 2021. http://hdl.handle.net/10280/95717.
Full textAbstract:
L'agricoltura moderna sta affrontando sfide come la perdita di fertilità del suolo, la variabilità climatica e gli attacchi di agenti patogeni in continuo aumento. Le pratiche agricole si stanno evolvendo verso sistemi sostenibili e rispettosi dell'ambiente. L'uso di biostimolanti (PBS, plant biostimulant) è una soluzione innovativa per affrontare le sfide di un’agricoltura sostenibile che garantisce un assorbimento ottimale dei nutrienti, una resa delle colture e tolleranza agli stress abiotici. In particolare, tra i diversi tipi di biostimolanti presenti sul mercato, i rizobatteri, classificati come Plant Growth Promoting Rhizobacteria (PGPR), offrono un nuovo approccio per promuovere la crescita delle piante, la mitigazione degli stress e l’aumento della resa colturale. Pertanto i PGPR sono considerati come una sorta di "probiotici" vegetali, poiché contribuiscono in modo efficiente alla nutrizione e all'immunità delle piante. L'obiettivo principale di questa tesi è isolare e identificare batteri presenti nella rizosfera di pomodoro (Solanum lycopersicum L.) che mostrano proprietà PBS, nonché valutare i meccanismi coinvolti nell'azione di promozione della crescita delle piante (Capitolo 2) e la genetica alla base di questi meccanismi (Capitolo 3 e 4). Infatti, una profonda comprensione dei meccanismi d’azione dei PGPR potrebbe colmare la mancanza di coerenza del dato di efficacia tra gli studi di laboratorio e gli studi in campo e stimolare la ricerca per la produzione e la commercializzazione di nuovi prodotti biostimolanti microbici.Modern agriculture faces challenges such as loss of soil fertility, fluctuating climatic factors and increasing pathogen and pest attacks. Agricultural practices have been evolving towards organic, sustainable and environmentally friendly systems. The use of natural plant biostimulants (PBS) is an innovative solution to address the challenges in sustainable agriculture, to ensure optimal nutrient uptake, crop yield, quality and tolerance to abiotic stress. In particular, among different types of biostimulants present on the market, plant growth promoting rhizobacteria (PGPR) offer a novel approach for promoting plant growth, mitigate stress and increase crop yield. Hence, PGPR inoculants are now considered as a kind of plant ‘probiotics’, since they efficiently contribute to plant nutrition and immunity. The main goal of this thesis was to isolate and identify bacteria symbionts of tomato (Solanum lycopersicum L.) rhizosphere, which showed PBS properties and evaluate mechanism involved in the action of PGPR (Chapter 2), underlying genetics and physiological pathways (Chapter 3 and 4). Indeed, a deeply understanding of the mechanisms of plant growth promotion, could fulfill the lack of consistency between lab, greenhouse and field studies, and support commercialization of novel plant biostimulant products.
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Silva, Patrícia Mota da, and 92-99202-2595. "Produção de Lipases por fungos isolados de amostras de solo da Floresta Amazônica." Universidade Federal do Amazonas, 2015. https://tede.ufam.edu.br/handle/tede/6389.
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Made available in DSpace on 2018-05-16T15:53:58Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação_Patrícia Mota Silva.pdf: 979608 bytes, checksum: ce07874ce0155d48be4cc47700cab638 (MD5) Previous issue date: 2015-08-31
FAPEAM - Fundação de Amparo à Pesquisa do Estado do Amazonas
Microbial lipases have a great potential for industrial applications, including cosmetic industry. Forward the great diversity of the Amazon forest micro-organisms, it can be said that few bioprospecting work of producing lipases fungi were performed. This study aimed to investigate lipase production by fungi isolated from soil samples of the Amazon rainforest. Samples were collected in Reserva Florestal Adolpho Ducke. The fungi were isolated, purified and identified (macro and micromorphologically). Lipase producing fungi were selected by the ability of those in hydrolyzing pNPP (p-nitrophenylpalmitate). Univariate experiments were conducted in order to investigate the influence of bioprocess variables in the production of lipase-selected isolated Fusarium INPA 4. It was also investigated the optimum temperature and pH of Fusarium lipases INPA 4:01 preliminary analysis as the cytotoxicity of broth cultivation of the fungus. The results revealed that: 100 isolates were obtained and the three most common genera of fungi were Aspergillus (41%), Penicillium (34%) and Trichoderma (13%). The isolated highlighted on lipase production were INPA83 Penicillium, Fusarium INPA 4, Paecilomyces Aspergillus INPA INPA 53 and 59. The levels of the most appropriate factors for production of the Fusarium lipases INPA 4 were 15 g / L Soy oil 1 g / L (NH4) 2SO4, pH 5, the relationship between the culture medium and glassware 1/5, initial inoculum of 105 cell / ml, orbital stirring of 100 rpm and a time of 72 h.Na determination of pH and temperature optimum the lipase of Fusarium INPA 4 was more active in the pH range of 7.0-9.0 and temperature range of -20oC to 30oC. The cultivation broth of Fusarium INPA 4 showed no cytotoxicity from the third dilution having a viability above 100%. The Fusarium INPA 4 showed a good lipase producer, may be a source of this enzyme for future industrial applications, but for this, it is necessary an additional study for the best biotech enzyme potential and his producer are exploited.
Lipases microbianas têm um grande potencial para aplicações industriais, inclusive para indústria de cosméticos. Frente a grande diversidade de micro-organismos da floresta amazônica, pode-se afirmar que poucos trabalhos de bioprospecção de fungos produtores de lipases foram realizados. Este trabalho teve como objetivo investigar a produção de lipases por fungos isolados de amostras de solo da floresta amazônica. Foram coletadas amostras na Reserva Florestal Adolpho Ducke. Os fungos foram isolados, purificados e identificados (macro e micromorfologicamente). Foram selecionados fungos produtores de lipase pela capacidade desses em hidrolisar o pNPP (p-nitrofenilpalmitato). Foram realizados experimentos univariados com a finalidade de investigar a influência das variáveis de bioprocesso na produçao de lipases pelo isolado selecionado Fusarium INPA 4. Foi investigada também a temperatura e pH ótimos das lipases de Fusarium INPA 4 e uma análise preliminar quanto a citotoxicidade do caldo de cultivo deste fungo. Como resultados observou-se que: foram obtidos 100 isolados e os três generos fúngicos mais frequentes foram Aspergillus (41%), Penicillium (34%) e Trichoderma (13%). Os isolados destacados na produção de lipases foram: Penicillium INPA83, Fusarium INPA 4, Paecilomyces INPA 53 e Aspergillus INPA 59. Os níveis dos fatores mais adequados para produção das lipases pelo Fusarium INPA 4 foram 15 g/L de óleo de soja, 1 g/L de (NH4)2SO4, pH 5, relação entre meio de cultura e vidraria de 1/5, inóculum inicial de 105 cell/mL, agitação orbital de 100 rpm e o tempo de 72 h.Na determinação do pH e temperatura ótimos, a lipase do Fusarium INPA 4 foi mais ativa na faixa de pH entre 7,0-9,0 e na faixa de temperatura de 20oC a 30oC. O caldo de cultivo do Fusarium INPA 4 não apresentou citotoxicidade a partir da terceira diluição, tendo uma viabilidade acima de 100%. O Fusarium INPA 4 se mostrou um bom produtor de lipase, podendo ser uma fonte dessa enzima para futuras aplicações industriais, mas para isso, torna-se necessário um estudo complementar para que o melhor potencial biotecnológico da enzima e seu produtor sejam explorados.
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Silveira, Lucas de Abreu. "Produção de fitase por fungos endofíticos dos manguezais do Estado de São Paulo." Universidade Federal de São Carlos, 2017. https://repositorio.ufscar.br/handle/ufscar/9320.
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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
A source of great biotechnological potential for the production of enzymes are endophytic microorganisms. These microorganisms are capable of producing a wide variety of enzymes, among them phytase, which is responsible for phytate hydrolysis. The objective of this work was to evaluate the potential of endophytic fungi of the mangroves of the state of São Paulo for phytase production. Initially, a qualitative selection of 33 isolates with PSM (Phytase Screening Medium) and a second quantitative selection was carried out, in which the preselected fungi were submitted to solid state fermentation (SSF) for phytase production. Among the isolates evaluated, the fungus Aspergillus awamori 9(4) was selected, which presented phytase activity of 43.98 U/100g when submitted to SSF culture for 72 h with soybean meal as substrate. In order to improve phytase production, the use of wheat bran as substrate was evaluated, reaching an activity of 82.77 U/100g, under the same conditions of cultivation. The use of KH2PO4 as an inducer in phytase production and the use of dialysis for the removal of interfering ions from the crude extract in phytase activity were also evaluated. For the experiments performed without the inducer, the dialysis of the extract resulted in activities of 85.42 U/100g with soybean meal and 55.44 U/100g with wheat bran. For the experiments of cultivation with the inductor and then dialysed resulted in activities of 132.35 U/100g with soybean meal and 115.52 U/100g with wheat bran, indicating that the effect of the inducer is positive in the production of Phytase and that dialysis is important for enzyme activity. These results indicate that the endophytic fungus of the mangrove of the State of São Paulo is promising for the production of phytase enzyme, being unpublished the use of endophytic micro-organisms for the production of this enzyme.
Uma fonte de grande potencial biotecnológico para a produção de enzimas são os microrganismos endofíticos. Estes microrganismos são capazes de produzir grande variedade de enzimas, dentre elas a fitase, que é responsável pela hidrólise do fitato. O objetivo deste trabalho foi avaliar o potencial de fungos endofíticos dos manguezais do estado de São Paulo para a produção de fitase. Inicialmente foi realizada uma seleção qualitativa de 33 isolados com meio diferencial PSM (Phytase Screening Medium), e uma segunda seleção quantitativa, em que os fungos pré-selecionados foram submetidos à fermentação em estado sólido (FES) para produção de fitase. Dentre os isolados avaliados, selecionou-se o fungo Aspergillus awamori 9(4), tendo este apresentado atividade de fitase de 43,98 U/100g quando submetido a cultivo FES por 72 h com farelo de soja como substrato. A fim de melhorar a produção de fitase, avaliou-se o uso de farelo de trigo como substrato, atingindo assim atividade de 82,77 U/100g, sob as mesmas condições de cultivo. Avaliou-se também o uso de KH2PO4 como indutor na produção de fitase e o uso de diálise para a remoção de íons interferentes do extrato bruto na atividade de fitase. Para os experimentos realizados sem o indutor, a diálise do extrato resultou em atividades de 85,42 U/100g com farelo de soja e 55,44 U/100g com farelo de trigo. Já para os experimentos de cultivo com o indutor e em seguida dialisados resultaram em atividades de 132,35 U/100g com farelo de soja e 115,52 U/100g com farelo de trigo, indicando que o efeito do indutor é positivo na produção de fitases e que a diálise é importante para a atividade enzimática. Esses resultados indicam que o fungo endofítico do manguezal do Estado de São Paulo é promissor para a produção da enzima fitase, sendo inédito o uso de micro-organismos endofíticos para a produção desta enzima.
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Egan, Laurie K. "Community Control and Compensation: An Analysis for Successful Intellectual Property Right Legislation for Access and Benefit Sharing in Latin American Nations." Scholarship @ Claremont, 2012. http://scholarship.claremont.edu/hmc_theses/25.
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Abstract: Indigenous communities have worked for centuries to develop systems of knowledge pertaining to their local environments. Much of the knowledge that has been directly acquired or passed down over generations is of marketable use to corporations, especially in the pharmaceutical industry. Upon gaining the necessary information to convert traditional knowledge into a marketable entity, the corporation will place a patent on the product of their research and development and reap the monetary benefits under the protection of intellectual property legislation. Without appropriate benefit sharing, indigenous communities are robbed of their cumulative innovation and development and denied access to the very medicines that they assisted in development. This study will examine the efforts made by indigenous communities to develop benefit-sharing agreements under national ‘sui generis’ legislation and the international legislation of the Agreement on Trade-Related Aspects of Intellectual Property Rights (TRIPS) and the Convention on Biological Diversity (CBD).
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Oliveira, Bruno Francesco Rodrigues de. "Análise das atividades antimicrobiana e citotóxica de actinobactérias isoladas de diversos habitats." Universidade Federal de Goiás, 2015. http://repositorio.bc.ufg.br/tede/handle/tede/4835.
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Fundação de Amparo à Pesquisa do Estado de Goiás - FAPEG
The actinobacteria constitute a wide and diverse phylum of gram-positive bacteria with a high content of guanine and cytosine in their DNA, standing out by the production of secondary metabolites with antibiotic activity. The treatment of multi-drug resistant pathogens infections and cancer represents a major challenge for modern medicine and bioactive microbial metabolites constitute a rich source of antimicrobial and antitumor drugs. The main aim of this study was to analyse the potential antimicrobial and cytotoxic activities of actinobactéria isolated from various habitats. The evaluation of these bioactivities was accomplished by the application of a set of qualitative screening in vitro assays. From a total of 19 morphospecies, isolated from the goiano Cerrado soil, 16 were evaluated in the primary antimicrobial activity tests, of which 5 (31,25%) antagonized the growth of indicator microorganisms. The ECL of the morphospecie BC-A22 and the crude extracts of the strain ADU 1.3 stood out for their strong antibiotic action against MRSA. The most of the crude extracts exhibited lytic action on the staphylococcal cell wall. Only ECLs of morphospecies GUARA 1 and PEG 23 and extracts of the strain PEG 30 were cytotoxic against A. salina in low concentration. None of the extracts showed an intercalation effect on the DNA molecule. With exception of BC-A22, the other six isolates were morphologically characterized as members of the Streptomyces genus. The morphospecie ADU 1.3 was molecularly identified as Streptomyces sp., highly likely to consist of a new species. The results of this initial phase of microbial bioprospecting highlight that all the isolates are potential producers of antibiotics biomolecules, excelling the morphospecie BC-A22, which exhibited a strong antimicrobial activity against the clinical isolates of MRSA and most of gram-negative bacteria.
As actinobactérias compõem um amplo e diverso filo de bactérias gram-positivas com elevado conteúdo de guanina e citosina em seu DNA, destacando-se pela produção de metabólitos secundários com atividade antibiótica. O tratamento das infecções por patógenos multirresistentes aos antimicrobianos e do câncer representa um dos maiores desafios da medicina moderna e moléculas microbianas bioativas ainda constituem uma fonte rica de drogas antimicrobianas e antitumorais. O objetivo principal do presente estudo foi analisar as potenciais ações antimicrobiana e citotóxica de actinobactérias isoladas de vários habitats. A avaliação dessas bioatividades foi efetuada mediante aplicação de uma série de ensaios qualitativos de triagem in vitro. Do total de 19 morfoespécies, isoladas do solo de Cerrado goiano, 16 foram avaliadas nos testes primários de atividade antimicrobiana, das quais 5 (31,25%) antagonizaram o crescimento dos micro-organismos indicadores. O ECL da morfoespécie BC-A22 e os extratos brutos do isolado ADU 1.3 se sobressaíram quanto a sua alta ação antibiótica frente à MRSA. A maioria dos extratos brutos exibiu ação lítica sobre a parede celular estafilocóccica. Apenas os ECLs das morfoespécies GUARA 1 e PEG 23 e os extratos do isolado PEG 30 foram citotóxicos a A. salina em baixa concentração. Nenhum dos extratos mostrou efeito de intercalação sob a molécula de DNA. Com exceção de BC-A22, os seis isolados foram caracterizados morfologicamente como membros do gênero Streptomyces. A morfoespécie ADU 1.3 foi identificada molecularmente como Streptomyces sp., com alta probabilidade de consistir em uma espécie nova. Os resultados obtidos dessa etapa de bioprospecção inicial evidenciam que todos os sete isolados são potenciais produtores de biomoléculas antibióticas, notabilizando-se a morfoespécie BC-A22, que exibiu uma alta ação antimicrobiana frente aos MRSA hospitalares e a maior parte das bactérias gram-negativas. Palavras-chave: actinobactérias; bioprospecção microbiana; metabólitos bioativos; antibióticos.
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Soares, Enio Saraiva. "Identificação e seleção de bactérias produtoras de quitinases." Universidade Federal de Goiás, 2016. http://repositorio.bc.ufg.br/tede/handle/tede/6349.
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Currently there are different approaches to synthesize and discover new compounds, but the pursuit of these products on biodiversity is still advantageous. In bioprospecting microorganisms, which often are seeking their properties that can be exploited in biotechnology products. This is the case of chitinases, enzymes that degrade chitin. Chitinases (EC 3.2.1.29) are glycosyl hydrolases type enzymes that specifically cleave β-1,4 bonds between N-acetylglucosamines units of chitin with sizes ranging from 20 kDa to 90 kDa. The main producers of chitinase are the bodies that have chitin in their cell wall or exoskeleton, such as insects, crustaceans, fungi, algae, among others. This study aimed to select and identify producing bacteria chitinase in soil samples from different coastal regions of southern Brazil. Seventeen soil samples, collected close to fishing for shellfish waste disposal sites, were prepared and seeded in four minimum culture medium containing colloidal chitin as the sole source of carbon and energy, incubated and the colonies were isolated and purified. After yielded a total of thirteen isolates that were submitted to enzymatic index test, stressed that four isolates. The four isolated genomic DNA was extracted, amplified and purified, and sequenced region encoding 16S rRNA of these organisms. Bacteria were then pooled and identified by construction of a phylogenetic tree. The results showed the presence of the species Paenibacillus illinoisensis and Paenibacillus chitinolyticus and two members of the genus Bacillus. Future studies may indicate the possibility of its use as a source of genes for biotechnological applications such as the production of new biopesticides.
Existem atualmente diferentes abordagens para se sintetizar e descobrir novos compostos, mas a busca desses produtos na biodiversidade ainda é vantajosa. Na bioprospecção de microrganismos, o que muitas vezes se busca são as suas propriedades que possam ser aproveitadas em produtos biotecnológicos. Esse é o caso das quitinases, enzimas capazes de degradar a quitina. As quitinases (EC 3.2.1.29) são enzimas do tipo glicosilhidrolases, com tamanhos que variam de 20 kDa até 90 kDa, que clivam especificamente as ligações β-1,4 entre unidades de N-acetilglicosaminas da quitina. Os principais produtores de quitinases são os organismos que possuem quitina no seu exoesqueleto ou parede celular, como insetos, crustáceos, fungos, algas, bactérias entre outros. O presente estudo teve como objetivo selecionar e identificar bactérias produtoras de quitinases em amostras de solos de diferentes locais litorâneos da região Sul do Brasil. Dezessete amostras de solo, coletadas próximo a locais de descarte de resíduos de crustáceos por pescadores, foram preparadas e semeadas em meio de cultura mínimo contendo quitina coloidal como única fonte de carbono e energia, incubadas e as colônias foram isoladas e purificadas. Ao fim obteve-se um total de treze isolados de bactérias, que foram submetidas ao teste de índice enzimático, que destacou desses quatro isolados. O DNA genômico de quatro isolados foi extraído, amplificado e purificado, sendo sequenciada a região codificadora do gene 16S rRNA destes microrganismos. As bactérias foram então agrupadas e identificadas pela construção de uma árvore filogenética. Os resultados mostraram a presença das espécies Paenibacillus illinoisensis e Paenibacillus chitinolyticus além de dois membros do gênero Bacillus. Estudos futuros poderão indicar a possibilidade de seu uso como fonte de genes para aplicação biotecnológica, como a produção de novos bioinseticidas.
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Godinho, Rosemary de Sampaio. "Contribuição ao estabelecimento de marcos jurídicos sobre o acesso, repartição de benefícios e proteção dos conhecimentos tradicionais associados à biodiversidade e à bioprospecção." Universidade do Estado do Rio de Janeiro, 2014. http://www.bdtd.uerj.br/tde_busca/arquivo.php?codArquivo=8536.
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Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorOs conhecimentos produzidos por comunidades tradicionais, quilombolas e povos indígenas são cada vez mais reconhecidos como importante fonte de informação para as atividades de bioprospecção, especialmente àqueles conhecimentos associados à biodiversidade. Em uma sociedade denominada por muitos autores como a sociedade do conhecimento, torna-se urgente a discussão sobre a titularidade e a forma de proteção de tais conhecimentos tradicionais, que geram lucros de forma direta ou indireta às empresas do setor biotecnológico sem, contudo possuírem ainda sistemas eficazes de proteção. A presente tese analisa a forma de produção desses conhecimentos tradicionais associados à biodiversidade e à bioprospecção, a fim de estudar os mecanismos legais regulatórios incidentes sobre a sua proteção, acesso e repartição de suas informações, conforme estabelecido pela Convenção sobre a Diversidade Biológica. O objetivo central é contribuir ao estabelecimento de marco jurídico, através da discussão sobre a viabilidade, os benefícios e as limitações para a sua elaboração. Através de uma metodologia qualitativa são estabelecidas inicialmente a importância, as características e as diversas espécies dos conhecimentos tradicionais. Em seguida é abordada a interface desses conhecimentos com os conhecimentos científicos, os direitos de propriedade intelectual e a natureza jurídica dos direitos sobre tais conhecimentos que passam a ser denominados COTABIOs. Após essa etapa cognitiva, é apresentado o arcabouço institucional-legal da repartição de benefícios, acesso e proteção dos COTABIOs em diversos fóruns internacionais e em alguns países, com especial ênfase ao panorama nacional brasileiro. Ao término desse processo são formuladas propostas de medidas especificas que poderão contribuir para a proteção legal dos COTABIOs. A tese é concluída com considerações gerais sobre as propostas formuladas, a fim de contribuir para o preenchimento da lacuna existente em decorrência da esparsa legislação nacional e internacional sobre a repartição de benefícios, acesso e proteção dos COTABIOs sem, contudo ter a pretensão de esgotar o tema e sim contribuir para a reflexão e discussão sobre a necessidade de mudanças nesse setor
The knowledge generated by traditional communities, quilombolas and indígenous people, have been even more recognized as an important source of information for bioprospecting activities, in special those associated with biodiversity knowledge. In a knowledges society, as understood for several authors, it is mandatory to discuss about the ownership and the way of protection of such traditional knowledges, that are giving profits to biotechnologic sector corporations, however without any effective protection mecanism. This thesis examines these traditional knowledges associated with biodiversity and bioprospecting are produced, studying the legal mechanisms usually applyied for protecting them, and accessing the available informations as preconized by Biologic Diversity Convention. Our main objective is to make a contribution to the establishment of legal framework, by discussion about its viability, benefits and limitations. Adopting a qualitative methodology, it establish first the traditional knowledges importance, characterístics and the categories envolved. Afterwards it is approached the connection among traditional and cientific knowledges, the rights about the intellectuak property and the legal nature will be called here after as COTABIOs. After this cognitive stage, it is submitted the legal and institutional framework of benefit sharing, access and protection from COTABIOs on many international foruns and some countries, emphasizing mainly the national brazilian panorama. Subsequently are proposed specific mechanisms that could contribute to the legal COTABIOs protection. The thesis is concluded doing general considerations about the proposals presented, believing to be contributing to close the gap resulting from the termous national and international legislation about benefit sharing, access and protection from COTABIOs, without however understanding the discussion is been finalized, but only a new horizon for reflexion and discussion about changes on this area was proposed
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SILVA, Flaviana Gonçalves da. "Bactérias halotolerantes associadas a plantas de atriplex nummularia L. e sua inoculação em mudas." Universidade Federal Rural de Pernambuco, 2014. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6148.
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Flaviana Goncalves da Silva.pdf: 1354445 bytes, checksum: 5abe875dd282d6291759e442aef4933a (MD5)Made available in DSpace on 2016-12-14T18:24:04Z (GMT). No. of bitstreams: 1 Flaviana Goncalves da Silva.pdf: 1354445 bytes, checksum: 5abe875dd282d6291759e442aef4933a (MD5) Previous issue date: 2014-07-11
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
Salinity is a limiting factor for agriculture and a frequent problem in arid and semi-arid region which rainfall is low and poorly distributed. In these areas most of plants can not grow due soils unprotected and become degraded with time. The use of bacteria with plant growth promoting and tolerance to salt stress may indicate biotechnological alternative that allows the use of plants associated with these microorganisms as inoculants may provide beneficial effects on soil-plant interaction. The cultivation of Atriplex nummularia L. has been conducted in order revegetate soils, promoting the improvement of their physical and chemical properties as a phytoremediation technique of salinized soils. In order to isolate and select bacteria promoter of plant growth associated with Atriplex nummularia L. plants cultivated in two experiments were carried out in the Pernambuco state and evaluated the effects of its inoculation in Atriplex plants grown in greenhouse. The population density of the bacteria was determined and then the same were tested in respect to plant growth promotion in vitro solubilization of inorganic phosphate (SFI), biological nitrogen fixation (BNF), synthesis of indole acetic acid (IAA), exopolysaccharides production (EPS) and quorum sensing molecule. Some bacteria to plant inoculation Atriplex grown in a protected environment were also selected in order to analyses content of chlorophyll a, b and total; stomatal conductance (gs); leaf temperature; green matter, dry and total fractional parts (root, stem and leaf) of plants; content and levels of sodium, potassium, calcium and magnesium; total nitrogen, crude protein of leaves and total organic carbon. Through the insulation, it was obtained 107 halotolerant bacterial isolates with positive results to plant growth promotion. Regarding the content of chlorophyll a, b and total, stomatal conductance and crude protein in plants, there was no effect of treatments. Inoculation with halotolerant bacteria and plant growth promoters influenced total nitrogen and total organic carbon in plants of Atriplex. Therefore, there are halotolerant bacteria associated with Atriplex plants able to solubilize inorganic phosphate, N2 fixation, IAA production, EPS and quorum sensing molecule, with the possibility of these micro-organisms contribute positively to plant growth. Bacterial isolates are promising on vegetative and nutritional development of Atriplex. However, it requires further explore the effect of bacterial inoculants associated with halophytes, giving improved conditions for phytoremediation process of salinized soils.
A salinidade constitui um fator limitante à agricultura e tem se tornado um problema frequente em áreas sob clima árido e semiárido, onde as precipitações são reduzidas e mal distribuídas. Nessas áreas, a maioria das plantas não consegue se desenvolver, por isso, os solos ficam desprotegidos e tornam-se degradados com o tempo. A utilização de bactérias promotoras de crescimento vegetal com tolerância ao estresse salino pode indicar alternativa biotecnológica que possibilite o uso de plantas associadas a esses micro-organismos como inoculantes, podendo proporcionar efeitos benéficos na interação solo-planta. O cultivo da Atriplex nummularia L. tem sido realizado com o objetivo de revegetar estes solos, promovendo a melhoria de suas propriedades físicas e químicas, como técnica de fitorremediação de solos afetados por sais. Com isso, objetivou-se isolar e selecionar bactérias promotoras de crescimento vegetal associadas às plantas de Atriplex nummularia L. cultivadas em dois experimentos instalados no estado de Pernambuco e avaliar os efeitos da inoculação destas bactérias em plantas de Atriplex cultivadas em ambiente protegido. Foi determinada a densidade populacional das bactérias e em seguida as mesmas foram testadas quanto às características de promoção de crescimento vegetal in vitro: solubilização de fosfato inorgânico (SFI), fixação biológica de nitrogênio (FBN), síntese de ácido indol acético (AIA), produção de exopolissacarídeo (EPS) e molécula quorum sensing. Foram também selecionadas algumas bactérias para inoculação em plantas de Atriplex cultivadas em ambiente protegido, analisando-se nas plantas, aspectos como teor de clorofila a, b e total; condutância estomática (gs); temperatura foliar; fitomassa verde, seca e total das partes fracionadas (raiz, caule e folha) das plantas; conteúdos e teores de sódio, potássio, cálcio e magnésio; nitrogênio total, proteína bruta de folhas e carbono orgânico total. Por meio do isolamento, foi possível obter 107 isolados bacterianos halotolerantes, com resultados positivos quanto às características de promoção de crescimento vegetal. Em relação ao teor de clorofila a, b e total, condutância estomática e proteína bruta nas plantas, não houve efeito dos tratamentos aplicados. A inoculação com bactérias halotolerantes e promotoras de crescimento vegetal influenciou o nitrogênio total e carbono orgânico total em plantas de Atriplex. Portanto, é possível afirmar que existem bactérias halotolerantes associadas às plantas de Atriplex, capazes de solubilizar fosfato inorgânico; fixar N2; produzir AIA, EPS e molécula quorum sensing, havendo a possibilidade destes micro-organismos, quando associados às plantas, contribuírem de forma positiva em relação à promoção de crescimento vegetal. Os isolados bacterianos são promissores no desenvolvimento vegetativo e nutritivo da Atriplex. No entanto, necessita-se explorar melhor o efeito dos inoculantes bacterianos associados às plantas halófitas, dando condições para melhoria no processo de fitorremediação de solos salinos.
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Ribeiro, Maycon Carvalho. "Bioprospecção e caracterização de bactérias produtoras de ciclodextrina glicosiltranferase em solos de biomas brasileiros." Universidade Federal de Goiás, 2014. http://repositorio.bc.ufg.br/tede/handle/tede/4730.
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Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq
Cyclodextrin glycosyltransferase (CGTase, EC 2.4.1.19) is an important industrial enzyme for being the only one able to convert starch and related glucans in cyclic oligosaccharides called cyclodextrins (CDs). The arrangement of the glucose units in the formation of CD results in a molecule with the shape of a cone, with hydrophobic interior and hydrophilic surface. This arrangement of glucose molecules in CDs allows its use as a host molecule in the formation of inclusion complexes with organic and inorganic compounds. This mechanism is advantageous in protecting the guest molecule from light, heat and oxidizing conditions and also enable the "dissolution" of compounds of low solubility in aqueous media. Cyclodextrins are used from the food industry to the pharmaceutical, in controlled drug delivery systems and immobilization of toxic compounds for environmental protection. The CGTases are mainly produced by bacteria of the genus Bacillus, found degrading starch rich substrates. The aim of this study was to identify, isolate, select and characterize strains of CGTase-producing bacteria from soil samples from different regions of Brazil as well as calculate the enzymatic production of these bacteria on low-cost substrates. With this work, it was possible to identify 17 bacteria producing cyclodextrin glycosyltransferase enzyme, with nine of them had values above 1.5 for enzymatic production. Of these, all were characterized as gram positive Bacillus. Bioprospecting of bacteria in soils of different cultures led to the identification of bacteria that may be used in studies for the production of cyclodextrin glycosyltransferase and subsequent implementation by various industries.
Ciclodextrina glicosiltransferase (CGTase, EC 2.4.1.19) é uma enzima industrial importante, sendo a única capaz de converter o amido e glucanos afins em oligossacarídeos cíclicos chamados ciclodextrinas (CDs). O arranjo das unidades de glicose na formação da CD resulta em uma molécula com a forma de cone, com interior hidrofóbico e superfície hidrofílica. Este arranjo das moléculas de glicose permite seu uso como molécula hospedeira na formação de complexos de inclusão com compostos orgânicos e inorgânicos. Este mecanismo é vantajoso na proteção de moléculas contra luz, calor e condições oxidantes e também possibilita melhorar a solubilidade de compostos hidrofóbicos. Ciclodextrinas são utilizadas desde a indústria de alimentos até a farmacêutica, em sistemas de liberação controlada de drogasse e imobilização de compostos tóxicos para proteção ambiental. As CGTase são principalmente produzidas por bactérias do gênero Bacillus, encontradas degradando substratos ricos em amido. O objetivo deste estudo foi identificar, isolar, selecionar e caracterizar linhagens de bactérias produtoras de CGTase a partir de amostras de solo de diferentes regiões do Brasil bem como calcular o índice enzimático destas bactérias em substratos de baixo custo. Com a realização deste trabalho, foi possível identificar 17 bactérias produtoras da enzima ciclodextrina glicosiltransferase, sendo que nove delas apresentaram valores para índice enzimático acima de 1,5. Destas, todas foram caracterizadas como sendo Bacillus gram positivos. A bioprospecção de bactérias em solos de diferentes culturas possibilitou a identificação de bactérias que poderão ser usadas em estudos para a produção da ciclodextrina glicosiltransferase e posterior aplicação pelas mais diversas indústrias.
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Conti, Raphael. "Micro-organismos de interesse farmacêutico e agrícola: estudo químico e biossintético." Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/60/60138/tde-04092012-162732/.
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A biodiversidade microbiana de diferentes ecossistemas tem incentivado estudos químicos e biológicos com micro-organismos dos mais variados habitats, os quais têm conduzido à obtenção de moléculas bioativas com aplicações na medicina, indústria química e agricultura, proporcionando melhorias na qualidade de vida ao homem. O presente trabalho teve como objetivos a bioprospecção por actinobactérias endofíticas e seus metabólitos, além do estudo da via biossintética dos sesquiterpenos aristoloquenos produzidos pelo fungo fitopatogênico Botrytis cinerea. No estudo de bioprospecção foram isoladas 41 linhagens de actinobactérias endofíticas de duas espécies de Asteraceae (Thitonia diversifolia e Lychnophora ericoides). A identificação através do sequenciamento de DNAr indicou predominância do gênero Streptomyces. As linhagens foram cultivadas em meio de arroz e os extratos etanólicos submetidos aos ensaios de citotoxicidade frente a células tumorais e antimicrobiano. Um total de 58,5% dos extratos apresentou atividade em pelo menos um dos ensaios realizados. Foram selecionadas as linhagens Streptomyces cattleya RLe 4 e Streptomyces sp. RLe 8 para cultivo em escala ampliada, isolamento e identificação de metabólitos bioativos. O isolamento dos compostos foi realizado através de diferentes técnicas cromatográficas e a identificação estrutural foi baseada em dados de ressonância magnética nuclear de 1H e 13C e espectrometria de massas. De S. cattleya RLe 4 foram isolados quatro compostos: 2-hidroxibenzamida, desferrioxamina E, 1-(3\',4\'-dimetoxifenil)-1-propanona e 1-(3\',4\'-dimetoxifenil)-1-etanona. Dos extratos de Streptomyces sp. RLe 8 foram isolados dez compostos: benzamida, 3- hidroxibenzamida, 3-hidróxi-4-metoxibenzamida, 4-hidróxi-3-metoxibenzamida, 3,4- dimetoxibenzamida, 2-fenilacetamida, dois isômeros de 3,4-diidro-3,4,6,8-tetraidróxi-1(2H)- naftalenona, 2,3-diidro-2,2-dimetil-4(1H)-quinazolinona e desferrioxamina B. O composto 2,3-diidro-2,2-dimetil-4(1H)-quinazolinona apresentou elevada atividade frente as células de câncer de cólon (HCT-8) e glioblastoma (SF295), com 93,9 % e 87.0 % de inibição, respectivamente. O outro enfoque da tese envolveu a otimização da produção de sesquiterpenos aristoloquenos por linhagens de B. cinerea, seguido de estudo biossintético destes produtos naturais através de experimentos de incorporação de precursores isotopicamente enriquecidos com 2H (deutério) e 13C (carbono treze). As análises dos dados obtidos de RMN de 2H e de 13C do sesquiterpeno majoritário indicaram que a biossíntese desta substância ocorre pela via do mevalonato (MVA). Os resultados também sugeriram o possível envolvimento da via do metil-eritritolfosfato ou 1-desoxi-D-xilulose-fosfato (MEP/DPX) na biossíntese deste sequiterpeno. Estes resultados podem contribuir para o planejamento racional de fungicidas seletivos com aplicação na agricultura. O trabalho desenvolvido mostrou o grande potencial de actinobactérias endofíticas para a obtenção de moléculas bioativas e que estudos usando precursores isotopicamente marcados fornecem informações precisas acerca da origem biossintética de produtos naturais.The microbial biodiversity from different ecosystems has incited chemical and biological studies with microorganisms from several habitats, leading to the isolation of bioactive natural products with applications in medicine, chemical industry and agriculture, and thus contributing to a better quality of life. This thesis aimed the biopropecting on endophytic actinobacteria and their natural products, and also the biosynthetic study of aristolochene sesquiterpenes in the phytopathogenic fungus Botrytis cinerea. A total of 41 actinobacterial strains were isolated of two Asteraceae species (Thitonia diversifolia and Lychnophora ericoides) for the bioprospecting study. The rDNA sequencing showed predominancy of Streptomyces genus. All the strains were cultured on rice medium, and the ethanolic extracts were screened in cytotoxity and antimicrobial assays. As a result, 58.5% of the extracts showed activity in al least one bioassay. The strains Streptomyces cattleya RLe 4 and Streptomyces sp. RLe 8 were selected for scale up cultures, isolation and identification of bioactive compounds. Different chromatographic methods were applied for the isolation of compounds, and structural analysis were based on 1H and 13C nuclear magnetic resonance and mass spectrometry data. Four compounds were isolated from S. cattleya RLe 4: 2- hydroxybenzamide, desferrioxamine E, 1-(3\',4\'-dimethoxyphenyl)-1-propanone, and 1-(3\',4\'- dimethoxyphenyl)-1-etanone. Ten compounds were isolated from Streptomyces sp. Rle 8: benzamide, 3-hydroxybenzamide, 3-hydroxy-4-methoxybenzamide, 4-hydroxy-3- methoxybenzamide, 2-phenylacetamide, two isomers of 3,4-dihydro-3,4,6,8-tetrahydroxy- 1(2H)-naphthalenone, 2,3-dihydro-2,2-dimethyl-4(1H)-quinazolinone, and desferrioxamine B. Compound 2,3-dihydro-2,2-dimethyl-4(1H)-quinazolinone showed high antiproliferative activity against colon cancer cells (HCT-8) and glioblastoma cells (SF295), with 93.9 and 87.0% of inhibition, respectively. The second focus of the thesis involved the optimization of aristolochene sesquiterpenes production by two strains of B. cinerea, followed by the biosynthetic study through feeding experiments with 2H (deuterium) and 13C isotopically labeled precursors. The 2H and 13C NMR obtained data showed that the biosynthesis of the sesquiterpene proceeds by the mevalonate pathway (MVA). The results also suggested the possible participation of the non mevalonate pathway, methylerytritol phosphate ou 1-deoxy- D-xylulose phosphate (MEP/DXP), in the biosynthesis. These results might contribute to the rational design of selective fungides with application in agriculture. This thesis showed the endophytic actinobacteria as promising sources of bioactive natural products, and also showed that the isotopically labeled feeding experiments give reliable information about the natural products biosynthetic pathways.
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Silva, Leonardo José da. "Actinobactérias da Antártica produtoras de compostos anticâncer." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-17012019-165858/.
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A utilização de produtos naturais para a terapêutica do câncer foi iniciada com a actinomycina D, obtida a partir de culturas de Streptomyces e desde então, a busca por compostos bioativos de origem natural constitui uma importante linha de pesquisa. Estima-se que aproximadamente 60% dos agentes antineoplásicos, introduzidos para a terapia do câncer nas últimas décadas, tem origem vegetal ou microbiana. Dentre os micro-organismos proeminentes para produção de compostos ativos, as actinobactérias se destacam pela versatilidade metabólica, praticidade para cultivo in vitro e eficiência para produção de compostos com atividade anticâncer. Em seu último relatório, a Organização Mundial da Saúde reportou 8,8 milhões de mortes em decorrência de câncer, no ano de 2017. O índice representa um em cada seis óbitos em todo o mundo, sendo mais expressivo em países de média e baixa renda. Vale ressaltar que avanços significativos foram alcançados nos últimos anos para o tratamento de leucemia aguda infantil e tumores derivados de células germinais. Contudo, tumores sólidos de pulmão, próstata, mama e cólon ainda representam altos índices de mortalidade. Frente a isso, torna-se evidente a necessidade de identificar e desenvolver estratégias para o tratamento da doença. Com intuito de acessar novos recursos microbianos com potencial biotecnológico, a prospecção avança para áreas pouco exploradas, como por exemplo, o Continente Antártico. A Antártica foi o último dos continentes a ser acessado pelo homem e apresenta características edafoclimáticas favoráveis ao endemismo. Em vista da problemática e da potencialidade do Continente Antártico, os recursos microbiológicos associados à rizosfera de Deschampsia antarctica Desv. foram acessados e avaliados para a produção de compostos com propriedade antitumoral. Em resultado foram obtidos 42.528 clones metagenômicos e 72 linhagens de actinobactérias, dentre as quais Streptomyces sp. CMAA 1527, que apresentou pronunciada atividade antiproliferativa in vitro, para tumores de mama, pulmão, rim e sistema nervoso central, através da produção de cinerubina B. A análise taxonômica das actinobactérias isoladas revelou a presença de linhagens com baixo índice de similaridade, com as linhagens tipo conhecidas, o que pode significar a presença de novas espécies para os gêneros Nocardia, Rhodococcus e Streptomyces, reconhecidos pela capacidade de produzir metabólitos ativos e enzimas de interesse biotecnológico. A análise taxonômica polifásica da linhagem CMAA 1533 possibilitou a descrição da espécie Rhodococcus psychrotolerans sp. nov. (TaxoNumber TA00191; NRRL B-65465T = DSM 104532T), grupo bacteriano promissor como agente de biorremediação e produção de compostos bioativos. Com isso, o Continente Antártico foi considerado um ambiente promissor para a busca de novos micro-organismos, dentre eles actinobactérias, eficientes na produção de compostos antitumorais e outras substâncias com potencial biotecnológico.The use of natural products for cancer therapy was initiated with the actinomycin D, obtained from Streptomyces. Since then, the search of bioactive from natural sources represent an essential line of research. It is estimated that approximately 60% of the antineoplasic agents inserted for the cancer therapy in recent decades have vegetal and microbial origin. Among the prominent microorganisms used to produce active compounds, actinobacterias are known by their metabolical versatility, convenience related to in vitro culture, and efficiency on the production of anticancer compounds. The Health World Organization, on its last review, reported 8.8 million of deaths in 2017, caused by cancer. Those numbers represent one out of six deaths worldwide, being more expressive in middle and low income countries. It is worth pointing out that meaningful advances were established in recent years for the treatment of childhood acute leukemia and germ cell-derived tumors. However, solid tumors of the lung, prostate, breast and colon still represent high mortality rates. For this reason, it is necessary to identify and develop strategies for the treatment of the disease. With the aim of accessing new microbial resources that contain biological potential, the prospection advance to areas barely explored, such as the Antarctic Continent. Antarctica was the last of the continents to be accessed by man and presents edaphoclimatic characteristics favorable to endemism. In light of the problematic and the potentiality of the Antarctic Continent, the microbiological resources associated with the rhizosphere of Deschampsia antarctica Desv. were accessed and evaluated for the production of compounds with antitumor properties. The results obtained had shown 42,528 metagenomic clones and 72 strains of actinobacteria, among them Streptomyces sp. CMAA 1527, which had presented anti-proliferative activity in vitro to breast, lung, kidney and central nervous system tumors, through the production of cinerubin B. The taxonomic analysis of the actinobacteria isolated revealed the presence of strains with low rate of similarity, with known type strains, which may mean the presence of new species for the genera Nocardia, Rhodococcus and Streptomyces, recognized for the ability to produce active metabolites and enzymes of biotechnological interest. The polyphasic approach of the CMAA 1533 strain made possible the description of the species Rhodococcus psychrotolerans sp. nov. (TaxoNumber TA00191; NRRL B-65465T = DSM 104532T), promising bacterial group as a bioremediation agent and production of bioactive compounds. As a result, the Antarctic Continent was considered a promising environment to search new microorganisms, among them, the actinobacteria, which is efficient on the production of antitumor compounds and other substances with biotechnological potential.
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Nunes, Rosangela Silva Gonçalves. "ISOLAMENTO E INOCULAÇÃO DE BACTÉRIAS DIAZOTRÓFICAS E PROMOTORAS DE CRESCIMENTO EM ARROZ IRRIGADO." Universidade Federal de Santa Maria, 2013. http://repositorio.ufsm.br/handle/1/4865.
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Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorBrazil is one of the largest producers and consumers of rice (Oryza sativa) in the world, and the largest portion of its production comes from floodplain ecosystems, and Rio Grande do Sul (RS), the largest national producer. Rice production is dependent on nitrogen fertilizers, which have high costs and high capacity to pollute the environment. An alternative to reducing the application of nitrogen (N) is inoculated Diazotrophic able to promote biological fixation of nitrogen (BNF) and or, in rice plant growth. The aim of this study was to select and evaluate the efficiency of diazotrophic bacteria isolated from different rice varieties, and the potential contribution of BNF these isolates in the culture in question, under flooding conditions. Three studies were performed. In the first, we performed the isolation of diazotrophic bacteria present in 20 varieties of rice grown in RS. In the second, the isolates were tested for their ability to fix atmospheric nitrogen, produce and release acid-indole-3-acetic acid (IAA) and promote growth ten rice cultivars under conditions gnotobiotics. In the third study, an experiment was conducted in a greenhouse with four isolates that showed improved plant growth and the three cultivars responded more inoculation in vitro test. In tests with inoculation was used strain BR11417 (ZAE94 - Herbaspirillum seropedicae) as a reference. 35 isolates were obtained from roots and shoots of rice cultivars tested, and all showed the ability to fix atmospheric nitrogen and produce and release IAA in vitro. In the experiment under conditions gnotobiotics cultivars IRGA 409, Puita Inta-CL and Pampa cultivars proved more responsive to inoculation and isolates 12, 13, 29 were those who had higher plant growth promotion. In the greenhouse, the IRGA 409 was the one that responded to inoculation and isolated 12, who provided the greatest benefits in relation to plant tillering and dry matter production in rice cultivars studied.
O Brasil é um dos maiores produtores e consumidores de arroz (Oryza sativa) do mundo, e a maior parcela de sua produção é proveniente dos ecossistemas de várzeas, sendo o Rio Grande do Sul (RS) o maior produtor nacional. A produção de arroz é dependente de fertilizantes nitrogenados, os quais apresentam custos elevados e alta capacidade de poluir o meio ambiente. Uma alternativa para a redução na aplicação de nitrogênio (N) é a inoculação de bactérias diazotróficas capazes de promover a fixação biológica de N (FBN) e, ou, o crescimento vegetal em arroz. O objetivo deste trabalho foi selecionar e avaliar a eficiência de bactérias diazotróficas endofíticas isoladas de diferentes variedades de arroz, e o potencial de contribuição da FBN destes isolados na cultura em questão, sob condição de inundação. Foram realizados três estudos. No primeiro, foi realizado o isolamento de bactérias diazotróficas presentes em 20 variedades de arroz irrigado cultivadas no RS. No segundo, os isolados obtidos foram testados quanto à capacidade de fixar nitrogênio atmosférico, produzir e liberar ácido-3-indol-acético (AIA) e promover o crescimento de dez cultivares de arroz irrigado sob condições gnotobióticas. No terceiro estudo, foi realizado um experimento de casa de vegetação com os quatro isolados que proporcionaram melhor crescimento vegetal e as três cultivares que mais responderam a inoculação no teste in vitro. Nos testes com inoculação foi utilizada a estirpe BR11417 (ZAE94 Herbaspirillum seropedicae) como referência. Foram obtidos 35 isolados da raiz e da parte aérea das cultivares de arroz irrigado testadas, e todos apresentaram capacidade de fixar nitrogênio atmosférico e produzir e liberar AIA in vitro. Em experimento sob condições gnotobióticas as cultivares IRGA 409, Puitá Inta-CL e Pampa demonstraram ser as cultivares mais responsivas a inoculação, e os isolados 12, 13, 29 foram aqueles que apresentaram maior promoção de crescimento vegetal. Em casa de vegetação, a cultivar IRGA 409 foi a que mais respondeu a inoculação e o isolado 12, aquele que proporcionou os maiores benefícios em relação ao perfilhamento de plantas e produção de matéria seca nas cultivares de arroz irrigado estudadas.
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Vaz, Marcelo Gomes Marçal Vieira. "Cianobactérias de ambiente costeiro: filogenia, prospecção gênica e química de moléculas bioativas." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-17092014-163006/.
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O filo Cyanobacteria constitui um grupo filogeneticamente coerente, embora, apresente grande diversidade morfológica, sendo sua sistemática constantemente revisada. Esses micro-organismos são, ainda, alvos de estudos biotecnológicos em razão da produção de toxinas e na busca por substâncias de interesse farmacológico. Dentre as linhagens analisadas neste estudo, sete sequências do gene rRNA 16S foram geradas e avaliadas com sequências previamente obtidas. Ao menos dois grupos podem representar novos gêneros de cianobactérias, sendo que um grupo demonstra ser endêmico de manguezais brasileiros. Os genes de inibidores de proteases, aeuruginosina, cianopeptolina e microviridina, foram detectados e a produção de aeruginosina foi confirmada por LC-MS nos gêneros Cyanobium e Nostoc. Sequências de aminoácidos do precursor de microviridina indicaram a produção de três novas variantes em quinze linhagens de cianobactérias dos gêneros Cyanobium, Synechococcus, Cyanobacterium, Nodosilinea e Nostoc. O potencial genético para produção de cilindrospermopsina (cyrJ) foi confirmado em vinte e seis linhagens. Em cinco linhagens dos gêneros Cyanobium e Nostoc foram encontrados os genes mcyD, mcyE e mcyG, envolvidos na biossíntese de microcistina. A sequência McyG da linhagem Nostoc sp. CENA175 agrupou-se filogeneticamente com outras de linhagens produtoras de microcistina. Os genes sxtA e sxtI, envolvidos na biossíntese de saxitoxina, foram encontrados em nove linhagens dos gêneros Cyanobium, Oxynema, Leptolyngbya, Nodosilinea e Nostoc. A sequência de SxtI da linhagen Leptolyngbya sp. CENA134 apresentou similaridade >= 70 % com proteínas hipotéticas enquanto as de Nostoc sp. CENA159 e Nostoc sp. CENA160 apresentaram similaridade >= 82 % com O-carbamoiltransferase. Na análise filogenética, a sequência de SxtI da linhagem Nostoc sp. 160 agrupou-se com sequências de linhagens produtoras de saxitoxina. Nas análises químicas, a fração 3 do extrato da linhagem Oxynema sp. CENA135 revelou uma substância com características de ácidos graxos poli-insaturados e a fração 2 do extrato da linhagem Nostoc sp. CENA175 apresentou uma estrutura aromática ligada a uma cadeia alifática. Outros três extratos, obtidos das linhagens Cyanobium sp. CENA157, Nodosilinea sp. CENA183 e Nostoc sp. CENA184 mostraram-se promissores quanto à presença de substâncias nitrogenadas. Os ensaios de bioatividade revelaram que 48 % dos extratos metanólicos inibiram o crescimento de ao menos um isolado de bactéria e/ou levedura. Os extratos das linhagens Cyanobium sp. CENA142 e Cyanobacterium sp. CENA169 foram eficientes contra o crescimento de seis bactérias patogênicas. Nos ensaios de inibição de células tumorais, o extrato de DCM da linhagem Cyanobium sp. CENA154 (100 ?gomL-1) inibiu moderadamente culturas de células 3LL. Os extratos etanólicos de Oxynema sp. CENA135 (20 ?gomL-1) e Cyanobium sp. CENA154 (100 ?gomL-1) inibiram as células CT-26. Em ensaios conduzidos com linhagens de células de glioma (U251), câncer de mama (MCF-7) e câncer de pulmão (NCI-H460), o extrato de DCM da linhagem Cyanobium sp. CENA136 inibiu 50 % do crescimento das respectivas células tumorais nas concentrações 7,8; 27,1 e 14,0 ?gomL-1. Desta forma, além de filogeneticamente diversas, as cianobactérias isoladas de ambiente marinho do Estado de São Paulo constituem fonte promissora de inibidores de proteases, cianotoxinas e substância bioativas com ação antibacteriana, antifúngica e antitumoral.The phylum Cyanobacteria is a phylogenetically coherent group, although presenting great diversity, and its systematic have been constantly reviewed. These microorganisms are also targets of biotechnological studies due to the production of toxins and the search for novel substances of pharmacological interest. Among the strains analyzed in this study, sequences of the 16S rRNA gene were generated for seven and, than, analyzed with sequences previously obtained. At least two groups may represent new cyanobacterial genera, while a group of Cyanobium proves to be endemic of Brazilian mangroves. Genes of the proteases inhibitors, aeuruginosin, cyanopeptolin and microviridin, were detected and the production of aeruginosin was confirmed by LC-MS for Nostoc and Cyanobium. The amino acid sequences of microviridin precursor indicated the production of three new variants in fifteen cyanobacterial strains of the genera Cyanobium, Synechococcus, Cyanobacterium, Nostoc and Nodosilinea. The genetic potential for production of cylindrospermopsin (cyrJ) was confirmed in twenty-six strains. In five strains of the genera Nostoc and Cyanobium the mcyD, mcyE and mcyG genes, which are involved in the microcystin biosynthesis, were found. The McyG sequence of Nostoc sp. CENA175 was phylogenetically grouped with sequences of microcystin-producing strains. The sxtA and sxtI genes, from saxitoxin biosynthesis, were found in nine strains of the genera Cyanobium, Oxynema, Leptolyngbya, Nodosilinea and Nostoc. The SxtI sequence of Leptolyngbya sp. CENA134 showed similarity >= 70 % with hypothetical proteins, while the sequences of Nostoc sp. CENA159 and Nostoc sp. CENA160 showed similarity >= 82% with O-carbamoyltransferase. In the phylogenetic analysis, the SxtI sequence of Nostoc sp. CENA160 grouped with sequences of strains that produce saxitoxin. In chemical analysis, the fraction 3 of the Oxynema sp. CENA135 extract revealed a substance with poly-unsaturated fatty acids characteristics and the fraction 2 of Nostoc sp. CENA175 extract indicated an aromatic structure, attached to an aliphatic chain. Other three extracts obtained from Cyanobium sp. CENA157, Nodosilinea sp. CENA183 and Nostoc sp. CENA184 were promising for the presence of nitrogenous substances. Bioactivity assays revealed that 48 % of the methanolic extracts inhibited the growth of at least one isolate of bacteria and/or yeast. The extracts of Cyanobium sp. CENA142 and Cyanobacterium sp. CENA169 were efficient against six pathogenic bacteria. In the inhibition assays of tumor cells, the DCM extract of Cyanobium sp. CENA154 (100 mgomL-1) moderately inhibited the growth of 3LL cells. Ethanol extracts of Oxynema sp. CENA135 (20 mgomL-1) and Cyanobium sp. CENA154 (100 mgomL-1) were able to inhibit cultures of CT- 26 cells. In tests conducted with glioma cell lines (U251), breast cancer (MCF-7) and lung cancer (NCI-H460), the DCM extract of Cyanobium sp. CENA136 caused 50 % of growth inhibition, respectively, when used at concentrations of 7.8, 27.1 and 14.0 mgomL-1. Thus, besides their phylogenetically diversity, the cyanobacteria strains from marine environment of the São Paulo state are a promising source of protease inhibitors, cyanotoxins and bioactive compounds with antibacterial, antifungal and antitumor activities.
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OLIVEIRA, João Tiago Correia. "Caracterização fisiológica e genética de bactérias potencialmente diazotróficas associadas a capim braquiária." Universidade Federal Rural de Pernambuco, 2012. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6118.
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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
The genus Brachiaria grasses are widely used throughout Brazil, enduring a series of restrictive limits and conditions of use, which corrobora para that is widely cultivated throughout the country. The changes in agricultural systems, mostly aiming at the improvement of environmental quality, in order to know the microbial diversity associated with these systems has been useful for ecosystem sustainability and improvement of production, where the combination of grasses with diazotrophs can provide plant growth and represents a promising alternative for the better development and plant performance. Given the above, the objective of this study was to isolate diazotrophic bacteria potentially associated grasses B. decumbens and B. humidicola cultivated in Agreste region of Pernambuco State, niches in the rhizosphere and endophytic root, evaluating functional and genetic variability. After isolation on semi selective fixation of atmospheric nitrogen (NFB semisolid medium) and in a non-functional selection was the selection of bacterial isolates for characteristics involved with the promotion of plant growth and genetic variability, using the technique of BOX-PCR and PCR-DGGE of nifH. The phenotypic characteristics were analyzed: biological nitrogen fixation (BNF), production of indole acetic acid (IAA) by different biochemical pathways, inorganic phosphate solubilization, production of extracellular enzymes and the quorum sensing molecule. The results obtained evidenced the high and variable functionality of the bacteria associated with these grasses in both media (especially the ones from the middle NFB, with the highest frequency of positive isolates for the evaluated tests), plant species and niches evaluated. In bacteria associated with the grass was observed fixation of atmospheric nitrogen in different salt concentrations in the culture medium, producing different biochemical pathways by EIA with and without the presence of precursor L-tryptophan, solubilization of inorganic phosphate in the presence of different sources of carbon production of extracellular enzymes (cellulase, pectinase and amylase) and quorum sensing molecule. Regardless of the technique used to study the genetic variability was observed high genetic diversity for isolates from both the root and the rhizoplane of grasses B. decumbens and B. humidicola. Thus we conclude that this association bacteria-plant-soil can provide several benefits to the plant, not just the biological nitrogen fixation. Therefore, investments in research using the association grasses/bacteria are essential for the high value they represent in animal feed.
As gramíneas do gênero Brachiaria são largamente utilizadas por todo o Brasil, tolerando uma série de limites e condições restritivas de utilização, o que corrobora para que seja largamente cultivada em todo território nacional. As mudanças ocorridas nos sistemas agrícolas, em sua maioria, têm por finalidade a melhoria da qualidade ambiental, neste sentido conhecer a diversidade microbiana associada a estes sistemas tem sido útil para sustentabilidade dos ecossistemas e o aperfeiçoamento da produção, onde a associação de gramíneas com bactérias diazotróficas pode proporcionar crescimento vegetal e representa uma alternativa promissora para o melhor desenvolvimento e desempenho vegetal. Diante do exposto, o objetivo deste trabalho foi isolar bactérias potencialmente diazotróficas associadas as gramíneas B. decumbens e B. humidicola cultivadas na Região Agreste do Estado de Pernambuco, nos nichos rizosfera e endofítico de raiz, avaliando-se características funcionais e variabilidade genética. Após o isolamento em meio semi seletivo para fixação de nitrogênio atmosférico (meio NFb semi-sólido) e em meio não seletivo ocorreu a seleção funcional dos isolados bacterianos quanto a características envolvidas com a promoção de crescimento vegetal, e a variabilidade genética, através da técnica de BOX-PCR e PCR-DGGE do gene nifH. As características fenotípicas analisadas foram: fixação biológica de nitrogênio (FBN), produção de ácido indol acético (AIA) por diferentes rotas bioquímicas, solubilização de fosfato inorgânico, produção de enzimas extracelulares e da molécula quorum sensing. Nos resultados obtidos ficou evidente a elevada e variável funcionalidade das bactérias associadas a estas gramíneas em ambos os meios (com destaque para os isolados provenientes do meio NFb, que apresentou maior frequência de isolados positivos para os testes avaliados), espécies vegetais e nichos avaliados. Nas bactérias associadas as gramíneas se observou fixação de nitrogênio atmosférico em diferentes concentrações de sal no meio de cultura, produção de AIA por diferentes rotas bioquímicas com e sem a presença do precursor L-triptofano, solubilização de fosfato inorgânico com a presença de diferentes fontes de carbono, produção de enzimas extracelulares (celulase, pectinase e amilase) e molécula quorum sensing. Independente da técnica utilizada para o estudo da variabilidade genética foi observado elevada diversidade genética para os isolados tanto da raiz como do rizoplano das gramíneas B. decumbens e B. humidicola. Desta forma é possível concluir que tal associação bactéria-planta-solo pode proporcionar vários benefícios à planta, não apenas a fixação biológica do nitrogênio. Portanto, investimentos em pesquisas utilizando a associação gramíneas forrageiras/bactérias são de fundamental importância pelo elevado valor que estas representam na alimentação animal.