Os biossurfactantes são substâncias que podem apresentar características tensoativos, sendo denominadas de biossurfactantes, as quais podem ser produzidas por bactérias e fungos quando cultivados em diferentes fontes de carbono como resultado de seu metabolismo. No presente trabalho foi investigado o potencial biotecnológico de Bacillus licheniformis UCP 1016, bactéria isolada de solo do Porto do Recife contaminado por petróleo, na produção de biossurfactante utilizando a casa de abacaxi (Ananas comosus L.), como meio alternativo. Ensaios iniciais foram realizados para avaliar a produção de bioemulsificante pela detecção da atividade hemolítica e degradação do diesel e querosene. A partir desses resultados, estudos foram realizados com a produção do bioemulsificante no meio de abacaxi controle e com diferentes concentrações dos substratos hidrofóbicos (diesel e querosene a 5 e 10%), como suplementos. Os frascos foram incubados à temperatura de 35C, 150rpm, durante 72horas, avaliando a cinética de crescimento, índice de emulsificação e tensão superficial. Com a melhor condição de obtenção do bioemulsificante foi utilizado o líquido metabólico livre de células foi utilizado para realizar a remoção do petróleo e óleo motor, como também os testes de estabilidade e fitotoxicidade. Os resultados iniciais com o B. licheniformis demonstraram atividade hemolítica e habilidade para crescer novo meio formulado com casca de abacaxi (9,5x106 UFC/mL), como também com os suplementos diesel (9,8x106 UFC/mL) e querosene ( 8,0x105 UFC/mL), com 72 h de cultivo. A menor tensão superficial apresentou uma redução de 57,73 0,15 mN/m para 38,32 0,16 mN/m, no meio de produção contendo 5% de diesel. Os testes de estabilidade demonstraram que o bioemulsificante foi estável em pH 2 e 4, em 8% de NaCl e ausência de toxicidade. O líquido metabólico livre de células foi capaz de remover 95,15% do petróleo e 85,55% do óleo queimado de motor, respectivamente. Os resultados obtidos sugerem a formulação de um meio alternativo e de baixo custo para a produção de bioemulsificante. A bactéria B. licheniformis mostrou ser um microrganismo promissor, com potencial para aplicação nos processos de biorremediação / The biosurfactants are substances that can have characteristics surfactants, being called biosurfactants, which can be produced by bacteria and fungi when grown on different carbon sources as a result of their metabolism. In the present study explores the potential of biotechnology of Bacillus licheniformis UCP 1016, soil bacteria isolated from the Port of Recife contaminated by oil, biosurfactant production using the house of pineapple (Ananas comosus L.) as an alternative. Initial trials were conducted to evaluate the production of bioemulsificante by the detection of hemolytic activity and degradation of diesel and kerosene. From these results, studies were conducted with the production of pineapple in the middle bioemulsificante control and different concentrations of hydrophobic substrates (diesel and kerosene to 5 and 10%) as supplements. The vials were incubated at 35 C, 150rpm for 72 hours, evaluating the kinetics of growth, emulsification index and surface tension. With the best condition for obtaining the bioemulsificante was used cell-free metabolic net was used for the removal of oil and motor oil, as well as stability tests and phytotoxicity. Initial results with B. licheniformis showed hemolytic activity and ability to grow new medium formulated with pineapple peel (9.5 x 106 CFU / mL), as well as supplements diesel (9.8 x 106 CFU / mL) and kerosene (8.0 x105 CFU / mL), 72 h of culture. The lowest surface tension was reduced by 57.73 0.15 mN / m to 38.32 0.16 mN / m, in the middle of production with 5% diesel. Stability tests showed that bioemulsificante was stable at pH 2 and 4 in 8% NaCl and no toxicity. The cell-free metabolic net was able to remove 95.15% and 85.55% of oil burned motor oil, respectively. Results suggest the formulation of a medium and low-cost alternative for producing bioemulsificante. The bacterium B. licheniformis was found to be a promising microorganism, with potential for application in bioremediation processes

Thesis submitted in fulfilment of the requirements for the degree of Doctor Technologiae: Chemical Engineering, Faculty of Engineering at the Cape Peninsula University of Technology - Cape Town, South Africa / Polycyclic aromatic hydrocarbons (PAHs) are one of the most common and recalcitrant environmental contaminants – known for their potential toxicity, mutagenicity, and carcinogenicity to humans. Biosurfactant application can enhance the biodegradation of PAHs. The main object of this work was to explore the novelty of biosurfactant produced by the isolated strains of Bacillus sp and Pseudomonas aeruginosa grown exclusively on Beta vulgaris, and the modification of the zeolites nanoparticles by the biosurfactant, for enhanced biodegradation of PAHs in soil. Novel biosurfactant-producing strains were isolated from hydrocarbon-contaminated environments, while several agrowaste were screened as primary carbon sources for the expression of biosurfactants, which were quantified using various standardized methods......

O cianeto é um composto tóxico, que pode ser encontrado no ambiente de maneira natural ou como resultado de atividades antropogênicas tais como a mineração de ouro e a indústria da galvanoplastia. Dentre as diferentes espécies do composto, o cianeto de hidrogênio (HCN) é considerado o mais tóxico, mesmo concentrações de 100ppm são letais para os seres humanos. Para a degradação destes compostos de cianeto a compostos menos tóxicos existem diferentes métodos de tratamento que podem ser químicos, físicos ou biológicos. O presente trabalho estudou a capacidade de degradação bacteriana de cianeto com cepas nativas isoladas de rejeito de mineração de ouro. Para isto, foram escolhidas três cepas dentro de um grupo de vinte cepas isoladas previamente. As cepas foram identificadas mediante as técnicas de MALDI-TOF e sequenciamento do gene 16s. Posteriormente realizou-se o processo de ativação e crescimento bacteriano no qual foram determinados os parâmetros de crescimento para cada uma das cepas, tais como pH, agitação, temperatura e meio de cultura. Após a etapa de ativação bacteriana, realizou-se a adaptação das três cepas em ambientes alcalinos. Nesta etapa, foram feitos ensaios em frascos agitados e avaliou-se o crescimento celular em função da formação de células viáveis para diferentes condições de pH (7; 8; 9,10 e 11). Com as cepas adaptadas ao pH 10 foram realizados ensaios de degradação bacteriana de cianeto em frascos agitados contendo 100mL de solução sintética de cianeto de potássio e 0,2mL de inoculo bacteriano. A concentração da solução de cianeto foi de 500mgL-1 e o pH de 10. Foram avaliadas três condições de temperatura (37, 32 e 27oC). Durante os ensaios foi estudado o crescimento bacteriano, o comportamento do pH e a degradação de cianeto. A quantificação do cianeto livre foi determinada pelo método polarográfico com eletrodo de mercúrio. Através dos resultados obtidos, foi possível identificar que as três cepas isoladas pertencem ás espécies Bacillus pumilus, Bacillus licheniformis e Bacillus subtilis. As cepas bacterianas apresentaram maior produção celular quando cultivadas em meio L.B a pH 7, velocidade de rotação de 190rpm e 37oC de temperatura. A máxima faixa de adaptação a ambientes alcalinos aconteceu em valores de pH 10. As melhores taxas de degradação de cianeto para a cepa B. subtilis e B. pumilus ocorreram em temperatura de 27oC e 65rpm, conseguindo degradar 100% do cianeto. A cepa B. licheniformis apresentou a melhor taxa de degradação de cianeto em temperatura de 32oC e 190rpm obtendo 99,5% de degradação. Através dos resultados obtidos no presente trabalho, foi possível avaliar o potencial de degradação de cianeto para as bactérias B. pumilus, B. licheniformis e B. subtilis as quais podem ser utilizadas como alternativa de tratamento em efluentes contaminados com cianeto. / Cyanide is a highly toxic compound that can be naturally found in the environment or as a result of anthropogenic activities such as gold mining and electroplating industry. Among the different species of the compound, hydrogen cyanide (HCN) is considered the most toxic, even concentrations of 100 ppm are lethal to humans. Degradation of this cyanide compound to less toxic compounds can be carried out through different methods such as chemical, physical and biological treatments. The present work investigated the bacterial degradation capacity of cyanide by isolated native strains from gold mining tailings. Three strains were selected from a group of twenty previously isolated strains which were identified using MALDI-TOF and 16s gene sequencing techniques. The bacterial activation and cellular growth were performed to determine the growth parameters for the strains, such as pH, temperature, rotation speed and culture medium. After the activation, the strains were adapted to grow in alkaline environments. During this phase, cellular growth was carried out in agitated flask at different pHs (7, 8, 9, 10 and 11). The adapted strains were used to perform cyanide degradation tests at pH 10 in agitated flask by using 100mL of a synthetic solution. The solution was composed of 500 ppm of potassium cyanide and 0,2mL of bacterial inoculum. During the experiments, three temperatures were evaluated (37, 32 and 27°C). Bacterial growth, cyanide degradation and pH behaviour were studied Free cyanide quantification was determined by polarographic method with a mercury electrode. It was found that the three isolated strains belong to Bacillus pumilus, Bacillus licheniformis and Bacillus subtilis groups. The highest bacterial growth was observed when the strains were cultivated in a L.B media at pH 7,0, 37°C by using 190 rpm of rotation velocity. The maximum range of adaptation to alkaline environments occurred in pH values of 10. The best cyanide degradation rate (100%) were achieved at 27°C for the strain B. suptilis and B. pumilus. The strain. B. licheniformis showed the best cyanide degradation rate (99,5%) at 37°C. The results obtained in the present work were able to evaluate the potential of cyanide degradation for the bacteria B. pumilus, B. licheniformis and B. subtilis. These results can be used as an alternative to treat wastewaters that are polluted with cyanide.

Made available in DSpace on 2017-06-01T18:20:29Z (GMT). No. of bitstreams: 1 pre-textuais.pdf: 1799292 bytes, checksum: c3fdd9c640c31d402a23a54796793a70 (MD5) Previous issue date: 2010-03-26 / The biosurfactants are substances that can have characteristics surfactants, being called biosurfactants, which can be produced by bacteria and fungi when grown on different carbon sources as a result of their metabolism. In the present study explores the potential of biotechnology of Bacillus licheniformis UCP 1016, soil bacteria isolated from the Port of Recife contaminated by oil, biosurfactant production using the house of pineapple (Ananas comosus L.) as an alternative. Initial trials were conducted to evaluate the production of bioemulsificante by the detection of hemolytic activity and degradation of diesel and kerosene. From these results, studies were conducted with the production of pineapple in the middle bioemulsificante control and different concentrations of hydrophobic substrates (diesel and kerosene to 5 and 10%) as supplements. The vials were incubated at 35 ° C, 150rpm for 72 hours, evaluating the kinetics of growth, emulsification index and surface tension. With the best condition for obtaining the bioemulsificante was used cell-free metabolic net was used for the removal of oil and motor oil, as well as stability tests and phytotoxicity. Initial results with B. licheniformis showed hemolytic activity and ability to grow new medium formulated with pineapple peel (9.5 x 106 CFU / mL), as well as supplements diesel (9.8 x 106 CFU / mL) and kerosene (8.0 x105 CFU / mL), 72 h of culture. The lowest surface tension was reduced by 57.73 ± 0.15 mN / m to 38.32 ± 0.16 mN / m, in the middle of production with 5% diesel. Stability tests showed that bioemulsificante was stable at pH 2 and 4 in 8% NaCl and no toxicity. The cell-free metabolic net was able to remove 95.15% and 85.55% of oil burned motor oil, respectively. Results suggest the formulation of a medium and low-cost alternative for producing bioemulsificante. The bacterium B. licheniformis was found to be a promising microorganism, with potential for application in bioremediation processes / Os biossurfactantes são substâncias que podem apresentar características tensoativos, sendo denominadas de biossurfactantes, as quais podem ser produzidas por bactérias e fungos quando cultivados em diferentes fontes de carbono como resultado de seu metabolismo. No presente trabalho foi investigado o potencial biotecnológico de Bacillus licheniformis UCP 1016, bactéria isolada de solo do Porto do Recife contaminado por petróleo, na produção de biossurfactante utilizando a casa de abacaxi (Ananas comosus L.), como meio alternativo. Ensaios iniciais foram realizados para avaliar a produção de bioemulsificante pela detecção da atividade hemolítica e degradação do diesel e querosene. A partir desses resultados, estudos foram realizados com a produção do bioemulsificante no meio de abacaxi controle e com diferentes concentrações dos substratos hidrofóbicos (diesel e querosene a 5 e 10%), como suplementos. Os frascos foram incubados à temperatura de 35°C, 150rpm, durante 72horas, avaliando a cinética de crescimento, índice de emulsificação e tensão superficial. Com a melhor condição de obtenção do bioemulsificante foi utilizado o líquido metabólico livre de células foi utilizado para realizar a remoção do petróleo e óleo motor, como também os testes de estabilidade e fitotoxicidade. Os resultados iniciais com o B. licheniformis demonstraram atividade hemolítica e habilidade para crescer novo meio formulado com casca de abacaxi (9,5x106 UFC/mL), como também com os suplementos diesel (9,8x106 UFC/mL) e querosene ( 8,0x105 UFC/mL), com 72 h de cultivo. A menor tensão superficial apresentou uma redução de 57,73 ± 0,15 mN/m para 38,32 ± 0,16 mN/m, no meio de produção contendo 5% de diesel. Os testes de estabilidade demonstraram que o bioemulsificante foi estável em pH 2 e 4, em 8% de NaCl e ausência de toxicidade. O líquido metabólico livre de células foi capaz de remover 95,15% do petróleo e 85,55% do óleo queimado de motor, respectivamente. Os resultados obtidos sugerem a formulação de um meio alternativo e de baixo custo para a produção de bioemulsificante. A bactéria B. licheniformis mostrou ser um microrganismo promissor, com potencial para aplicação nos processos de biorremediação

L’objectif de ce projet de recherche était de déterminer les effets d’une supplémentation de probiotiques du genre Bacillus sur les performances de production de vaches de race holstein en mi-lactation et sur le profil en acides gras de leur lait. Pour ce faire, six vaches multipares porteuses d’une canule ruminale ont été utilisées selon un dispositif en chassé croisé répliqué et randomisé. Les vaches ont ainsi reçu soit 200 g/j de poudre de lactosérum comme traitement témoin ou 200 g/j de BioPlus 2B (Chr Hansen, Milwaukee, WI), un probiotique commercial de Bacillus subtilis et de Bacillus licheniformis, représentant un apport quotidien de 6,4 × 10¹¹ UFC et utilisant la poudre de lactosérum comme diluant. La production laitière, la composition du lait et son profil en acides gras, les paramètres de fermentation ruminale et le microbiote ruminal ont été évalués. Les traitements n’ont pas affecté les performances laitières. L’utilisation du probiotique a cependant augmenté la concentration relative d’anteiso 13:0 et d’anteiso 15:0 et tendait à augmenter la concentration totale d’acides gras à chaîne ramifiée dans les matières grasses du lait lorsque comparé au traitement témoin. Les traitements n’ont pas modifié le pH, l’azote ammoniacal ou les concentrations en acétate, propionate et butyrate du rumen. Cependant,le probiotique a augmenté la concentration ruminale d’isovalérate et tendait à augmenter celle de l’isobutyrate. Cette expérience a permis d’observer la sensibilité du profil en acides gras du lait face aux modifications du microbiote ruminal. Les bactéries probiotiques du genre Bacillus pourraient donc être utilisées dans un protocole de maximisation des acides gras à chaîne ramifiée du lait pour permettre une augmentation du niveau de ces acides gras dans les produits laitiers. / The aim of the study was to determine the effect of a Bacillus-based direct-fed microbial on dairy performance of mid-lactating Holstein dairy cows and on their milk fatty acid composition. To this end, six multiparous cows fitted with a rumen cannula were used in a randomized replicated crossover design. Cows received 200 g/d of either whey powder, as a control, or BioPlus 2B (Chr Hansen, Milwaukee, WI), a commercial direct-fed microbial providing Bacillus subtilis and Bacillus licheniformis, representing a daily dose of 6.4 ×10¹¹ cfu, and using whey powder as a carrier. Milk production, composition and fatty acid profile, as well as ruminal parameters and microbiota were evaluated. Bacillus concentrationin the rumen increased with the addition of the direct-fed microbial supplement. Treatmentdid not affect ruminal pH, NH3-N and concentrations of acetate, propionate, and butyrate.However, direct-fed Bacillus increased ruminal concentration of isovalerate and tended to increase the concentration of isobutyrate. Treatments did not affect milk performance. The direct-fed Bacillus increased the relative concentration of anteiso 13:0 and anteiso 15:0 in milk fat, and tended to increase total concentration of branched-chain fatty acids as compared with control. This trial indicates that milk fatty acid composition is sensitive to ruminal microbiota modifications. Direct-fed Bacillus could be used as part of a standard protocol aiming to produce milk enriched in branched-chain fatty acids.

Proteins are important biological products with unique functions, annually produced at the hundreds of millions of dollars value on a worldwide basis. The application of crystallization for these materials primarily was led by structural biologists and crystallographers who are keen on obtaining large and well-ordered crystals for protein structure determination via X-ray diffraction. Usually for this, crystallization is done on a small scale by vapor diffusion using a supersaturated solution of the material. In the past decades, production crystallization has slowly received increasing attention for the large-scale recovery of proteins. Among the numerous products, an industrial enzyme (alpha-amylase) that is extensively involved in food processing and laundry products was chosen for examination due to the lack of relevant data in the literature and the potential industrial interest in crystallizing this material. The chosen alpha-amylase is a product of Genencor International (the Danisco division) and is derived from a genetically modified Bacillus licheniformis. In parallel to the underlying principles that govern the bulk crystallization of small molecules, the broad topics of investigation for this macromolecular material included determination of solubility, studies of nucleation thresholds, and investigation of crystal growth kinetics and special phenomena accompanying the crystallization process. All these studies were preceded by a series of characterization tests conducted for the material. On the whole, this study aimed to extend the existing fundamental knowledge of bulk crystallization for biological macromolecules. Specifically, it intended to enrich the solubility and crystallization kinetic data for the alpha-amylase. The experimental data of this study were all obtained at conditions in line with industrial practice, which included the use of moderate temperatures, mild pH conditions and simple inorganic salts ((NH4)2SO4, Na2SO4 and NaCl) in order for the findings to be transferred to the industry directly. In a 20 mM sodium phosphate buffer (with no added salts), alpha-amylase solubility increased with solvent temperature and had a minimum at pH between 6.4 and 7.1. A generalized equation (as a function of pH and temperature) was obtained to correlate the data. The three inorganic salts examined affected the alpha-amylase solubility in a different manner, both qualitatively and quantitatively. Evidently, the interaction effect of a salt varied with solution pH. This confirms the importance of studying solubility with the two or more condition parameters at the same time. With relevance to crystal growth, the metastable region of the material was relatively wide at (NH4)2SO4 and Na2SO4 concentrations corresponding to maximum solubility. For example, σSNT was 1.2  0.2 in solutions with 5 wt% ammonium sulfate at pH 7.0 and 25oC. A wide metastability range is useful for the practical operation of batch crystallizers as nucleation can be minimized. This range, however, diminished as the salt concentration increased beyond the maximum solubility points, imposing a limit on the range of salt concentration favorable for growth processes. In systems with no added salts at pH 7.0, the solution metastability was slightly higher at 10oC than at 40oC. This would suggest a future further examination of the salt system at a lower temperature, say of 10oC. To develop a batch crystallizer, the growth kinetic data of the material have to be known. Throughout the growth studies, the alpha-amylase crystals obtained were lozenge-shaped thin plates. Apparently, habit was not influenced by the crystallization conditions chosen. Similar to other proteins crystallized in bulk, the growth rate of alpha-amylase demonstrated a second-order dependence upon supersaturation. Importantly, the alpha-amylase demonstrated crystal growth rate dispersion (GRD) under all the conditions tested. To simplify the analysis of growth kinetic results, the seed crystals used were common history (CH) seeds whose growth rates are proportional to their sizes. The spread of growth rates (CV) was 0.54 for the unsieved CH seeds used. Due to GRD, growth rate coefficient data varied with crystal size. For instance, in solutions containing 5 wt% ammonium sulfate at pH 7.0 and 25oC, the growth rate coefficient for seed crystals initially at 20 m was 2.47 m/hr. This order of magnitude was equivalent to that of many other proteins. Although being small, industrial crystallization was feasible with these kinetics, as demonstrated by the sample design calculations included. To improve the design, it is recommended to further examine the solubility, metastability and growth kinetics of the above system at other temperatures to obtain a wider growth rate range. As the important phenomenon of growth rate dispersion has seldom been examined for protein and enzyme materials in the crystallization literature, this study is a significant contribution to this area.

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