Lignocellulosic biomasses are considered promising feedstocks for the next generation of biofuels and chemicals; however, the recalcitrance of lignocellulose remains a barrier to its utilisation over conventional sources. Pyrolysis is the heating of biomass to several hundred degrees Celsius in the absence of oxygen, which can thermally depolymerise lignocellulose. Products of pyrolysis are a solid biochar, liquid bio-oil and syngas. Biochar has roles in both carbon sequestration and soil amendment however bio-oil has no defined use, despite a high concentration of fermentable sugars. Bio-oil is a complex organic microemulsion with a host of biocatalyst inhibitors that makes its microbial degradation a challenge. In this work, the use of aerobic cultures using microbial communities isolated from natural environments saw limited potential; however, the use of anaerobic digestion (AD) successfully generated a higher volume of biogas from reactors with bio-oil than controls. Biogas yield test reactors were set up with anaerobic digestate from a wastewater treatment plant as the substrate for degradation and conversion of bio-oils. Next-generation 16S rRNA gene sequencing was utilised to characterise the communities in the reactors while the ultrahigh resolution mass spectrometry technique of Fourier transform ion cyclotron resonance (FT-ICR) was used for characterisation of the chemical changes occurring during AD. Both sets of high-resolution data were additionally combined for multivariate analysis and modelling of the microbial genera that correlated best with the changes in digestate chemistry. This represents a novel analysis method for the microbial degradation of complex organic products. Bio-oil from common lignocellulosic feedstock was the most easily degradable by the AD communities, with significant inhibition observed when bio-oils from anaerobic digestate and macroalgae were used. Additionally it was found that the inclusion of biochars that were pre-incubated in anaerobic digestate prior to use in AD were capable of significantly reducing the lag time observed for biogas production in bio-oil-supplemented reactors. The addition of biochars that were not pre-incubated had no effect on biogas production. Specific inhibition of methanogenesis was also capable of causing the digestates to accumulate volatile fatty acids (VFAs) as a product of greater value than biogas. Scale-up experiments will be required to confirm the precise practicalities of the addition of bio-oil to AD as well as to establish the potential for isolation and purification of VFAs.

腐乳是中國傳統豆類發酵製品，具有綿軟的口感和特殊的風味。其是豆腐通過真菌固態發酵，并加入鹽，米酒和香料等進行後期熟化而成的產品。本文的研究分為兩部份，第一部份對腐乳發酵過程中的毛胚，鹽胚，熟化第一天，熟化一個月以及熟化六個月的腐乳樣本進行採樣，并採用傳統微生物培養法和克隆文庫法對每個階段真菌和細菌的生態結構和動態變化進行研究。第二部份重點比較了四株腐乳產品中分離的微生物和購自台灣生物資源保存及研究中心的四株細菌的產揮發性含硫化合物能力，并挑選了最高產的一株微生物進行紫外誘變，最後獲得理想的突變株。本研究的結論如下： / 1. 真菌和細菌的總數均是在毛胚階段為最高，在進入熟化階段后開始下降。在傳統微生物培養方法下分別分離出了三株真菌和九株細菌，通過18S rDNA和16 rDNA測序，發現絲孢酵母屬（Trichosporon spp.）是真菌中的優勢菌種，蠟狀芽孢桿菌（Bacillus cereus）和解澱粉芽孢桿菌（Bacillus amyloliquefaciens）為細菌中的優勢菌種； / 2. 本研究建立了五個真菌18S rDNA克隆文庫和五個細菌16 rDNA克隆文庫用于研究真菌和細菌的生態結構和動態變化。通過聚合酶鏈式反應-限制性片段長度多態性（PCR-RFLP）的研究，分別在真菌和細菌克隆文庫中發現23和38種圖譜類型，并計算其相應比例。在進行真菌細菌測序之後，對優勢菌群進行了定性和定量分析； / 3. 在對比傳統微生物培養方法和克隆文庫技術的結果后發現，二者的結果存有差異，有些在克隆文庫中鑒定到的微生物在傳統培養方法中未能分離鑒定，而有些微生物則只能在傳統培養方法中被分離鑒定。因此，本研究中將這兩種方法結合有助於我們更為全面、客觀地研究腐乳發酵過程中真菌和細菌的生態結構和多樣性。 / 4. 對四株腐乳中分離純化的微生物和四株外來購入細菌的產揮發性含硫化合物能力進行比較，結果發現，從腐乳產品中分離純化的B-1菌株擁有最高的產揮發性含硫化合物能力，通過紫外誘變后，突變株#3在產揮發性含硫化合物以及L-蛋氨酸代謝酶活力都比初始菌株有了顯著的提升。B-1菌經測序比對后鑒定為絲孢酵母（Trichosporon sp.）。 / 本研究結果對于傳統腐乳發酵的有效控制和現代腐乳生產工藝的建立有一定指導意義，並且對於腐乳產品中的風味物質，特別是揮發性含硫化合物的產生和優化提供信息。 / Sufu (fermented soybean curd) is a soft creamy cheese-type product with a pronounced flavor and is made by fungal solid state fermentation of tofu (soybean curd) followed by aging in brine containing salt and alcohol. In first part of this research, the eco-structure and the dynamic changes of microbes during sufu production process (Pehtze, Salted pehtze, 0 Month sufu, 1 Month sufu and 6 Month sufu sample) were studied by combined microbiology techniques, including plate culture, 16S rDNA and 18S rDNA clone library and restriction fragment length polymorphism (RFLP) analysis. The second part of this research focus on the comparison of volatile sulfur-containing compounds (VSCs) production ability within isolated strains in sufu product and bacteria purchased commercially, the strain that possessed highest ability was selected and followed by a UV mutation experiment, finally obtained the desired mutant. The results of this research are as followed: / 1. The population of both fungi and bacteria were all at highest number in Pehtze stage and started to decrease in ripening stages. A combined total of three and nine living strains of fungi and bacteria were obtained from the plate culture, respectively. Through 18S rDNA and 16S rDNA sequencing, Trichosporon spp. was the dominant fungi and Bacillus cereus and Bacillus amyloliquefaciens were the dominant bacteria; / 2. Five 18S rDNA clone libraries and five 16S rDNA clone libraries from different stages of sufu production were constructed to analyze the structure and dynamic changes of fungi and bacteria. A total of 23 and 38 RFLP patterns were found, and the ratio of each pattern were calculated. After sequencing, qualitative and quantitative analysis on the dynamic changes of dominant strains was performed; / 3. After comparing the results of plate culture and clone library, it was found that there were some differences between the two. Some strains were only found in clone library while some only found in plate culture approach. Therefore, the combination of the two microbiology methods will help us to objectively and completely analyze the structure and dynamic changes of microbes in the sufu production process; / 4. The ability to produce VSCs within four strains (B-1, B-2, B-3 & B-4) isolated from a commercial sufu manufacturing process and four commercial strains (B. acetylicum, L. Lactics, S. thermophilus and L. Paracasei) were compared. Results showed that B-1 possessed both the highest VSCs production ability and L-methionine metabolism enzymatic activities among the eight strains. After UV light mutagenesis of B-1 strain, its mutant #3 significantly increased in DMDS and DMTS production and all four L-methionine-related enzymatic activities in reference to that of the starting strain (B-1). B-1 was identified as Trichosporon sp. by sequencing. / These results would make a profound significance on the control of traditional sufu production and the development of new technology for modern sufu manufacturing. They will also help to provide some important information of optimal production of VSCs in sufu ripening and the overall flavor in sufu product. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Huang, Ruolan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 106-117). / Abstracts also in Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgement --- p.v / Table of contents --- p.vi / List of Figures --- p.x / List of Tables --- p.xiii / Chapter Chapter 1 --- : Introduction --- p.1 / Chapter 1.1 --- Sufu --- p.1 / Chapter 1.1.1 --- Classification --- p.1 / Chapter 1.1.1.1 --- Classified by processing technology --- p.1 / Chapter 1.1.1.2 --- Classified by color and flavor --- p.1 / Chapter 1.1.1.3 --- Other classifications --- p.2 / Chapter 1.1.2 --- Typical commercial manufacturing process --- p.2 / Chapter 1.1.2.1 --- Production process of naturally fermented sufu --- p.2 / Chapter 1.2.2.2 --- Production process of traditional mold-based sufu --- p.5 / Chapter 1.2.2.3 --- Production process of traditional bacteria-based sufu --- p.5 / Chapter 1.2.2.4 --- Acceleration of sufu ripening process --- p.6 / Chapter 1.1.3 --- Important ingredients in sufu production --- p.6 / Chapter 1.1.4 --- Flavor components in sufu --- p.7 / Chapter 1.1.4.1 --- Volatile flavor components --- p.7 / Chapter 1.1.4.2 --- Essential odor in sufu product --- p.8 / Chapter 1.1.4.3 --- Volatile sulfur compounds in sufu --- p.9 / Chapter 1.1.4.4 --- Using Head Space-Solid phase Microextraction (HS-SPME) to analyze the volatile sulfur components --- p.9 / Chapter 1.1.5 --- Relationship between microbes and sufu --- p.12 / Chapter 1.1.5.1 --- Microbes involved in fermentation process --- p.13 / Chapter 1.1.5.2 --- Microbial changes during the production of sufu --- p.14 / Chapter 1.1.6 --- Study on microbial ecology in food product --- p.15 / Chapter 1.1.6.1 --- PCR-based molecular techniques --- p.16 / Chapter 1.1.6.2 --- Non-PCR based molecular techniques --- p.16 / Chapter 1.1.6.3 --- The common techniques used in microbial ecology research --- p.17 / Chapter 1.1.6.4 --- Microbial ecology study by molecular biological techniques --- p.18 / Chapter 1.2 --- Objectives --- p.19 / Chapter Chapter 2 --- : Analysis of fungi diversity during sufu fermentation process --- p.21 / Chapter 2.1 --- Introduction --- p.21 / Chapter 2.2 --- Materials and methods --- p.21 / Chapter 2.2.1 --- Sample collection and preparation --- p.22 / Chapter 2.2.2 --- Plate count of fungi during sufu fermentation process --- p.22 / Chapter 2.2.3 --- Change of pH values and moisture content --- p.22 / Chapter 2.2.4 --- Total DNA extraction from fungi --- p.23 / Chapter 2.2.5 --- Preparation of competent cell --- p.23 / Chapter 2.2.6 --- 18S rDNA PCR amplification and construction of 18S rDNA clone library --- p.24 / Chapter 2.2.7 --- RFLP analysis of 18S rDNA clone library --- p.25 / Chapter 2.2.8 --- DNA sequencing for fungi identification --- p.26 / Chapter 2.2.9 --- Analysis of the diversity of 18S clone library --- p.26 / Chapter 2.2.10 --- Frequency percentage analysis --- p.27 / Chapter 2.2.11 --- Enzyme Solutions --- p.27 / Chapter 2.2.12 --- Determination of protease activity --- p.28 / Chapter 2.2.13 --- Determination of lipase activity --- p.29 / Chapter 2.2.11 --- Microtox test --- p.30 / Chapter 2.2.12 --- Statistical analysis --- p.30 / Chapter 2.3 --- Results and discussion --- p.30 / Chapter 2.3.1 --- Fungi growth on plate counting result --- p.30 / Chapter 2.3.2 --- Changes in pH and moisture content of sufu during production --- p.33 / Chapter 2.3.3 --- Construction and selection of 18S rDNA clone library --- p.35 / Chapter 2.3.4 --- Fungal diversity based on 18S rDNA clone library analysis --- p.38 / Chapter 2.3.5 --- Protease and lipase activities in Actinomucor elegans and Trichosporon japonicum --- p.45 / Chapter 2.3.5.1 --- Protease activity --- p.46 / Chapter 2.3.5.2 --- Lipase activity --- p.47 / Chapter 2.3.6 --- Toxicity of Actinomucor elegans and Trichosporon japonicum --- p.49 / Chapter 2.3.7 --- Analysis of fungi eco-structure and function during sufu fermentation process --- p.50 / Chapter 2.3.8 --- The influence of PCR bias and artifact --- p.53 / Chapter 2.2 --- Summary --- p.55 / Chapter Chapter 3 --- : Analysis of bacteria diversity during sufu fermentation process --- p.57 / Chapter 3.1 --- Introduction --- p.57 / Chapter 3.2 --- Materials and methods --- p.57 / Chapter 3.2.1 --- Sample collection and preparation --- p.57 / Chapter 3.2.2 --- Plate count of bacteria during sufu fermentation process --- p.57 / Chapter 3.2.3 --- Total DNA extraction from bacteria --- p.58 / Chapter 3.2.4 --- Preparation of competent cell --- p.58 / Chapter 3.2.5 --- 16S rDNA PCR amplification and construction of 16S rDNA clone library --- p.58 / Chapter 3.2.6 --- RFLP analysis of 16S rDNA clone library --- p.59 / Chapter 3.2.7 --- DNA sequencing for bacteria identification --- p.60 / Chapter 3.2.8 --- Analysis of the diversity of 16S rDNA clone library --- p.60 / Chapter 3.3 --- Results and discussion --- p.60 / Chapter 3.3.1 --- Bacteria growth on plate counting result --- p.60 / Chapter 3.3.2 --- Construction and selection of 16S rDNA clone library --- p.63 / Chapter 3.3.3 --- 16S rDNA clone library analysis of bacteria diversity --- p.65 / Chapter 3.3.4 --- Analysis of bacteria eco-structure and function during sufu fermentation process --- p.74 / Chapter 3.4 --- Summary --- p.77 / Chapter Chapter 4 --- : Screening the mutant possess higher capacity of forming volatile sulfur compounds (VSCs) from non-starter microbes of sufu product --- p.80 / Chapter 4.1 --- Introduction --- p.80 / Chapter 4.2 --- Materials and methods --- p.82 / Chapter 4.2.1 --- Strains and culture conditions --- p.82 / Chapter 4.2.2 --- Head space-solid phase microextraction (HS-SPME) analysis --- p.83 / Chapter 4.2.3 --- Gas Chromatography-Mass Spectrometry (GC-MS) analysis --- p.84 / Chapter 4.2.4 --- UV mutation --- p.85 / Chapter 4.2.5 --- Ellman’s method --- p.86 / Chapter 4.2.6 --- Preparation of cell-free extracts (CFE) for enzymatic assays --- p.86 / Chapter 4.2.7 --- Enzymatic assay --- p.86 / Chapter 4.2.7.1 --- L-methionine aminotransferase activity assay --- p.86 / Chapter 4.2.7.2 --- L-methionine demethiolase activity assay --- p.87 / Chapter 4.2.7.3 --- α-keto acid decarboxylase activity assay --- p.87 / Chapter 4.2.7.4 --- C-S lyase activity --- p.88 / Chapter 4.2.8 --- Statistical analysis --- p.88 / Chapter 4.3 --- Results and discussion --- p.89 / Chapter 4.3.1 --- Optimization of SPME extraction condition --- p.89 / Chapter 4.3.2 --- Selecting the start strain --- p.90 / Chapter 4.3.4.1 --- Comparison of VSCs production ability --- p.90 / Chapter 4.3.4.2 --- Comparison of enzymatic activity in L-methionine metabolism --- p.92 / Chapter 4.3.3 --- Optimization of UV exposure time --- p.95 / Chapter 4.3.4 --- Screening the mutants --- p.96 / Chapter 4.3.4.1 --- Comparison of VSCs production ability among the mutants --- p.96 / Chapter 4.3.4.2 --- Comparison of the L-methionine related enzymatic activities among the mutants --- p.99 / Chapter 4.3.4.3 --- Identified of strian B-1 --- p.101 / Chapter 4.4 --- Summary --- p.102 / Chapter Chapter 5 --- : General conclusions and future work --- p.103 / References --- p.106

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O presente estudo teve como objetivo quantificar as emissões de Compostos Orgânicos Voláteis do Aterro Controlado Morro do Céu localizado na cidade de Niterói, Rio de Janeiro, Brasil. Para tanto, vinte amostras foram coletadas, usando uma bomba de ar operada a bateria durante dois dias de dezembro de 2009. Uma câmara de fluxo cilíndrica de PVC de 30L foi inserida 5 cm no solo do aterro, e as amostras foram coletadas através de uma válvula na parte superior da câmera. Os resultados indicaram um valor de 1.980 Kg Km-2 h-1. O modelo Gaussiano de dispersão atmosférica ISCST3 foi utilizado para calcular a difusão e transporte dos poluentes a fim de estimar as concentrações de COV no bairro, usando dados topográficos, meteorológicos e de emissões. Valores de 525 μg m-3 de COV foram encontrados a 500 metros do aterro. As emissões do aterro foram usadas em conjunto com dados meteorológicos, utilizando o modelo de trajetória OZIPR e o mecanismo químico SAPRC para demonstrar o impacto na formação do ozônio troposférico na região. É conhecido que o ozônio é formado pela reação entre COV, NOx e luz solar. A contribuição de valores elevados de COV provenientes das emissões do aterro conduzirá a uma nova situação com valores mais elevados de ozônio na região. Os resultados da modelagem indicaram um aumento maior que 1000% nos níveis de ozônio na região do aterro, se comparado com a modelagem do ozônio para a região metropolitana do Rio de Janeiro. Os resultados mostram ser necessário que maior atenção seja dada à política de gerenciamento de RSU no Brasil, incluindo a escolha adequada para o local de instalação, o monitoramento da área durante e após o período de operação e técnicas mais adequadas de disposição dos resíduos sólidos urbanos / The purpose of this work was to quantify the Volatile Organic Compounds emissions by a landfill (Morro do Céu), located in Niterói, Rio de Janeiro State, Brazil. Twenty 10-minute samples were collected using a battery-operated air pump operated at 500 mL min-1, over a period of four days in May and December, 2009. A cylindrical 30 L PVC flux chamber was used, the open, bottom side of which was inserted into the landfill soil to a depth of 5 cm. The samples were collected using a valve at the upper, closed side of the chamber. The results indicated an elevated value of 1,980 kg km-2 h-1. A Gaussian plume dispersion model, implemented by Aermod software, was used to calculate the diffusion and transport of pollutants, and the pollutant concentrations in the surroundings were estimated using the emission, meteorological, and topographical data. Maximum values of 525 μg m-3 for VOC were found at approximately 500 m from the landfill. The landfill emissions were also compiled with meteorological data, using the OZIPR trajectory model coupled with SAPRC chemical mechanism, to demonstrate their impact on the formation of tropospheric ozone. It is well known that ozone is formed by the reactions of VOC, NOx and sunlight. The contribution of high VOC levels from the landfill emissions was expected to lead to higher ozone values. The results indicated a 1000% increase in ozone in the region of the landfill, compared to the ozone values for the metropolitan region of Rio de Janeiro. The results show that it is both necessary and desirable that greater attention be given to the political management of municipal solid waste in the Brazil, including the location choice, the monitoring of landfill areas during and after their period of operation and techniques more appropriate of disposition of municipal solid waste

O ozônio (O3) é um dos poluentes que representa grande preocupação em termos de qualidade do ar na Região Metropolitana de São Paulo (RMSP). No ano de 2012 foram observados 98 dias de ultrapassagens do padrão horário da qualidade do ar para este poluente na RMSP. A exposição aos poluentes atmosféricos como O3 e outros está associada ao prejuízo da saúde respiratória. O enfoque deste estudo é determinar os principais Compostos Orgânicos Voláteis (COV) precursores de O3 para auxiliar no controle deste poluente. Neste trabalho foram realizadas 78 amostragens durante a semana de hidrocarbonetos no ano de 2006 e 66 amostragens de hidrocarbonetos, 62 de aldeídos e 42 de etanol durante o ano de 2011/2012, 7:00 h às 9:00 h, na estação CETESB IPEN/USP. Medidas de COV também foram realizadas no ano de 2006 e 2008, na Estação CETESB Cerqueira César. Adicionalmente, foram efetuados testes de emissões veiculares durante o ano de 2009 de 5 veículos a diesel, 3 a etanol, 2 a gasolina C e 1 motocicleta. O modelo de trajetórias OZIPR foi utilizado para determinar os principais precursores de O3. Durante o ano de 2011/2012, na Estação CETESB IPEN/USP, a classe de aldeídos representou 35,3% dos COV analisados em concentração na atmosfera, seguido pelo etanol 22,6%, compostos aromáticos 15,7%, alcanos 13,5%, cetonas 6,8%, alcenos 6,0% e alcadienos < 0,1%. Considerando a concentração dos compostos e sua reatividade, as simulações executadas com o modelo OZIPR mostraram que o acetaldeído contribuiu com 61,2% da formação do O3 na atmosfera da RMSP no ano de 2011/2012. Dos COV analisados, a classe dos aldeídos contribui com 74% da produção de O3, aromáticos 14,5%, alcenos 10,2%, alcanos 1,3% e alcadienos (isopreno) 0,03%. O estudo de emissão veicular mostrou que 39% dos aldeídos foram provenientes de veículos a etanol, 28% a diesel, 26% a gasolina C e 7% de motocicletas. As emissões dos COV por veículos a gasolina contribuíram com 44% da formação de O3, a diesel 22%, a etanol 19% e motocicletas 15%. / Ozone (O3) is a pollutant that represents great concern in terms of air quality in the metropolitan area of São Paulo (MASP). In 2012 were observed 98 days of exceedances of the standard time air quality for this pollutant in the MASP. Exposure to air pollutants such as O3 and others is associated with the injury of respiratory health. The focus of this study is to determine the main Volatile Organic Compounds (VOCs) precursors of O3 to auxiliary in control this pollutant. In this work were made 78 samples during the week of hydrocarbons in 2006 and 66 samples of hydrocarbons, 62 of aldehydes and 42 of ethanol during the year 2011/2012, 7:00 am to 9:00 am, at the CETESB IPEN/USP station. Measurements of VOCs also were realized in 2006 and 2008, in the CETESB Cerqueira Cesar station. Additionally, were performed tests vehicle emissions during the year 2009 of 5 diesel vehicles, 3 ethanol, 2 gasool and 1 motorcycle. The OZIPR trajectory model was used to determine the main O3 precursors. During the year 2011/2012, in the CETESB IPEN/USP station the class of aldehydes represented 35.3% of VOCs analyzed in concentration in the atmosphere, followed by ethanol 22.6%, aromatics 15.7%, alkanes 13.5%, ketones 6.8%, alkenes 6.0% and alkadienes <0.1%. Considering the concentration of the compounds and their reactivity, the simulations executed with the model showed that acetaldehyde OZIPR contributed with 61.2% to the formation of O3 in the atmosphere of MASP in the year 2011/2012. VOCs analyzed, the class of aldehydes contributed with 74% of the production of O3, aromatics 14.5%, alkenes 10.2%, alkanes 1.3% and alkadienes (isoprene) 0.03%. The study of vehicular emission showed that 39% of aldehydes were obtained from ethanol vehicles, 28% diesel, 26% gasoline and 7% of motorcycles. Emissions of VOCs by gasoline vehicles contributed with 44% of the formation of O3, diesel 22%, ethanol 19% and motorcycles gasoline 15%.

O Brasil destaca-se como um dos maiores produtores mundiais de mamão, fruto apreciado em todo o mundo pelo sabor e polpa delicada. Os compostos voláteis contribuem para a formação do aroma do fruto, o que faz deles essenciais para o desenvolvimento de características sensoriais que definem sua apreciação pelos consumidores. Os principais compostos de aroma do mamão incluem, principalmente, terpenos, ésteres, aldeídos, álcoois, ácidos orgânicos e cetonas, com destaque para o monoterpeno linalool que é o composto mais abundante na cultivar \'Golden\'. Embora a biossíntese de compostos voláteis seja particularmente afetada pelos tratamentos para consevação pós-colheita, estes são essenciais para a comercialização do mamão, em vista de sua alta perecibilidade. O uso de baixas temperaturas tem sido um dos métodos mais empregados para extensão de vida pós-colheita do mamão. Precedentes da literatura indicam que o tratamento pós-colheita com metil jasmonato (MJ) é capaz de reduzir possíveis efeitos prejudiciais decorrentes do armazenamento a baixa temperatura. Além disso, a aplicação do metil jasmonato em frutos é capaz de estimular a atividade de enzimas das vias de produção de compostos voláteis. Desta forma, o presente trabalho tem por objetivo avaliar os efeitos da aplicação pós-colheita do MJ em mamão (Carica papaya L. cv \'Golden\'), focando os efeitos sobre as vias de biossíntese de componentes voláteis do aroma, em frutos amadurecidos a temperatura de 22ºC, assim como em outros armazenados a 10ºC seguido de transferência a 22ºC para o pleno amadurecimento. Além dos perfis de compostos voláteis, também foram avaliados os perfis de respiração, produção de etileno, cor da casca, quantificação de MJ e expressão gênica de linalool sintase (LIS). Dada a importância do linalool para o aroma característico do mamão, as variações na transcrição de um gene da linalool sintase foram avaliadas buscando correlacioná-las aos efeitos dos tratamentos na produção dos compostos voláteis. O tratamento com MJ influenciou a produção de compostos voláteis, particularmente de linalool e hexanal nos grupos mantidos a 10ºC. Os frutos tratados com o hormônio apresentaram maior abundância do composto quando comparados ao grupo controle. Embora o protocolo de tratamento empregado não tenha conseguido recuperar os altos níveis encontrados nos frutos armazenados a 22ºC, não se exclui a possibilidade de que outros desenhos experimentais possam responder de maneira ainda mais satisfatória a este mesmo tratamento / Brazil stands out as one of the world\'s largest producers of papaya, fruit appreciated worldwide for it taste and delicate pulp. The volatile compounds contribute to the formation of the flavor of the fruit, making them essential for the development of sensory characteristics that define their appreciation by consumers. The main papaya flavor compounds include mainly terpenes, esters, aldehydes, alcohols, organic acids and ketones, especially the monoterpene linalool which is the most abundant compound in the cultivar \'Golden\'. Although the biosynthesis of volatile compounds is particularly affected by postharvest treatments for conservation, those are essential for the marketing of papaya, in view of their high perishability. The use of low temperatures has been one of the most used methods for extension of papaya postharvest life. Previous literature indicate that postharvest treatment with methyl jasmonate (MJ) is capable of reducing possible adverse effects arising from storage at low temperature. Furthermore, the application of methyl jasmonate in fruits is able to stimulate the activity of the enzymes of volatiles production pathways. Thus, this study aims to evaluate the effects of postharvest application of MJ in papaya (Carica papaya L. cv \'Golden\'), focusing on the effects in the biosynthetic pathways of volatile aroma compounds in fruit ripened to temperature of 22ºC, as well as other stored at 10ºC followed by transfer to 22ºC for the complete maturation. In addition to the volatile compounds profiles were also evaluated respiration profiles, ethylene production, peel color, quantification of MJ and gene expression of linalool synthase (LIS). Given the importance of linalool to the characteristic aroma papaya, changes in the transcription of a linalool synthase gene were evaluated seeking to correlate them to the treatment effects on the production of volatile compounds. Treatment with MJ influence the production of volatile compounds, particularly linalool and hexanal in the groups kept at 10ºC. Fruits treated with the hormone had higher abundance of the compound when compared to the control group. Although the treatment protocol employee has not been able to recover the high levels found in fruits stored at 22ºC, do not exclude the possibility that other experimental designs can respond even more satisfactorily to this same treatment.

Submitted by Rosivalda Pereira (mrs.pereira@ufma.br) on 2017-06-13T19:49:13Z No. of bitstreams: 1 ClariceDias.pdf: 1740990 bytes, checksum: d4150e4fe41ba8404104c64d6b4fe6d3 (MD5) / Made available in DSpace on 2017-06-13T19:49:13Z (GMT). No. of bitstreams: 1 ClariceDias.pdf: 1740990 bytes, checksum: d4150e4fe41ba8404104c64d6b4fe6d3 (MD5) Previous issue date: 2013-02-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Dengue is considered the most important mosquito-borne viral disease in the world; it is endemic in over 100 countries, where 50-100 million are infected by dengue virus per year. As the dengue control is based on the combat of the primary vector of dengue, the mosquito Aedes aegypti L. (Diptera: Culicidae), one of the most effective ways to achieve this combat is by the use of synthetic chemicals, such as organophosphate temephos, against the mosquito larvae. However, the continued and repeated use of these products has contributed to the development of resistant strains. The occurrence of resistance can be figured out by researches to find out news plant-base insecticides, such as essential oils. Thus, this study aimed to perform a review of the essential oils with studies on the evaluation of the larvicidal activity against A. aegypti and to analyze the chemical composition and the larvicidal activity of essential oils from six plant species from Amazônia Legal maranhense. This dissertation was divided into two chapters; the chapter 1 consists of a review of the scientific papers which showed that 210 essential oils from 159 plant species have already been tested by their larvicidal potential and that more than 75% of these essential oils were considered active (LC50 < 100 mg/L). This study has further information such as the factors affecting the activity, structure-activity relationships, mechanisms of action, as well as the studies with the main compounds of these oils. The chapter 2 reports the experimental research to evaluate the larvicidal activity and chemical composition of essential oils extracted from six plants collected in the “Parque Nacional da Chapada das Mesas”, Maranhão, Brazil (Amazônia Legal maranhense). The oils were extracted by hydrodestilation, using a Clevenger apparatus and their compositions were analyzed by gas chromatography coupled to mass spectrometry (GC/MS). The larvicidal activity was performed according to the guidelines from the World Health Organization. The sesquiterpenes hydrocarbons were the major fraction of the essential oils from Eugenia patrisii Vahl, Eugenia piauhiensis Vellaff., Myrcia erythroxylon O. Berg, Psidium myrsinites DC., and Siparuna camporum (Tul.) A. DC.; whereas, Lippia gracilis Schauer essential oil showed the oxygenated monoterpenes as major compounds. Five of the six tested oils showed promising results for the development of products larvicides against A. aegypti larvae, showing LC50 value among 230 and 417 mg/L. From these results, it was possible to observe the need for standardization of methodologies for evaluation of larvicides against A. aegypti, the low number of patents and potential of oils from the “maranhense” flora to obtain products that can contribute to the control of dengue. / A dengue é considerada a arbovirose mais importante do mundo, sendo endêmica em mais de 100 países, onde anualmente, 50 a 100 milhões de pessoas são infectadas pelo seu vírus. Como o seu controle depende essencialmente do combate ao seu principal vetor, o mosquito Aedes aegypti L. (Diptera: Culicidae), uma das formas mais eficientes de realizar este combate é através do uso de produtos químicos sintéticos, como o organofosforado temefós, contra as larvas deste mosquito. No entanto, o uso contínuo e repetido destes produtos tem contribuído para o desenvolvimento de cepas resistentes. A solução para a ocorrência de resistência pode ser encontrada na busca por novos compostos inseticidas obtidos a partir de produtos do metabolismo das plantas, como os óleos essenciais. Sendo assim, o presente trabalho objetivou realizar um levantamento dos óleos essenciais com estudos de avaliação da atividade larvicida contra A. aegypti e analisar a composição química e a atividade larvicida de óleos obtidos de seis espécies vegetais da Amazônia Legal maranhense. Esta dissertação foi divida em dois capítulos; o capítulo 1 consiste no artigo de revisão de literatura, onde foi possível observar que 210 óleos essenciais de 159 espécies vegetais foram avaliados quanto à atividade larvicida e que mais de 75% destes óleos mostraram-se ativos (CL50 < 100 mg/L). Foram apresentados, ainda, os fatores que afetam a atividade larvicida, a relação estrutura atividade, os mecanismos de ação, bem como trabalhos com os constituintes isolados desses óleos. Já o capítulo 2 relata a parte experimental da avaliação da atividade larvicida e a composição química de óleos essenciais extraídos de seis espécies vegetais coletadas na região do Parque Nacional da Chapada das Mesas (Amazônia Legal maranhense), Maranhão, Brasil; os óleos foram extraídos por hidrodestilação, utilizando aparelho de Clevenger e sua composição analisada por cromatografia a gás acoplada a espectrometria de massas (CG/EM). A atividade larvicida foi avaliada segundo metodologia preconizada pela Organização Mundial de Saúde. Os sesquiterpenos hidrocarbonados constituíram a maior fração dos óleos essenciais de Eugenia patrisii Vahl, Eugenia piauhiensis Vellaff., Myrcia erythroxylon O. Berg, Psidium myrsinites DC. e Siparuna camporum (Tul.) A. DC.; enquanto que, Lippia gracilis Schauer apresentou como constituintes majoritários, monoterpenos oxigenados. Cinco dos seis óleos testados demonstraram resultados promissores para o desenvolvimento de produtos larvicidas contra as larvas de A. aegypti, apresentando CL50 entre 230 e 417 mg/L. A partir desses resultados foi possível observar a necessidade de padronização de metodologias para avaliação larvicida contra A. aegypti, o baixo número de patentes e o potencial de óleos essenciais da flora maranhense para obtenção de produtos que possam contribuir com o controle da dengue.

O uso de produtos anti-inflamatórios e antioxidantes sintéticos representa uma grande preocupação nos dias atuais para a população. Importante destacar alguns aspectos relevantes quanto às dificuldades encontradas no tratamento, como o aumento da resistência aos fármacos e os efeitos colaterais da terapia convencional, que apresenta como consequência o desencadeamento de doenças graves, prejudicando e comprometendo a saúde humana. Os óleos voláteis têm sido alvo na busca de novas substâncias, por apresentarem como potencial fonte de novos agentes antioxidantes e anti-inflamatórios. Dentre as espécies vegetais caracterizadas pela produção deste metabólito secundário, estão aquelas pertencentes ao gênero Schinus, da família Anacardiaceae, de ampla ocorrência do Rio Grande do Sul. Nesse contexto, o presente trabalho objetivou analisar e identificar a composição química e avaliar as atividades biológicas a partir da extração do óleo volátil das espécies de Schinus weinmannifolius e Schinus polygamus, nativas do Rio Grande do Sul. Assim, o óleo volátil foi obtido por hidrodestilação de diversas coletas de folhas e frutos de S. weinmannifolius e folhas de S. polygamus. A análise química, realizada por cromatografia à gás acoplada à espectrometria de massas (CG-EM), demonstrou grande variedade de compostos presentes nas amostras, com predominância dos sesquiterpenos β-cariofileno, germacreno D, biciclogermacreno, germacreno A, δ-cadineno, espatulenol, óxido de cariofileno e α-cadinol para folhas e frutos de S. weinmannifolius e predominância do composto alifático n-nonano para S. polygamus. Os óleos obtidos foram submetidos aos ensaios de reação com 2,2-difenil-1-picrilidrazila (DPPH) e substâncias reativas ao ácido tiobarbitúrico (TBARS) para avaliação da atividade antioxidante, sendo esta ação apenas observada para as amostras de folhas de S. weinmannifolius coletadas no outono e inverno de 2016 pelo ensaio de TBARS. No método de DPPH as amostras de folhas de S. weinmannifolius coletadas no outono de 2015 e 2016 apresentaram baixa atividade antioxidante, na concentração de 100 μg/mL. A ação anti-inflamatória foi inicialmente avaliada pela técnica in vitro da câmara de Boyden, nas concentrações de 0,001 a 10 μg/mL. Todas as amostras apresentaram inibição significativa da migração leucocitária em relação ao controle, avaliada no ensaio de antiquimiotaxia. A avaliação da atividade anti-inflamatória in vivo, realizada pelo método de edema de pata, foi conduzida apenas para a amostra que melhor resultado obteve no ensaio in vitro, folhas de S. weinmannifolius, verificando-se inibição significativa do edema na segunda hora de tratamento, na dose de 100 mg/kg. Os resultados encontrados suportam o uso tradicional dos óleos de Schinus como agentes anti-inflamatórios. / The use of synthetic anti-inflammatory and antioxidant products represents a major worry these days for the population. It is important to highlight some aspects relevant concerning the difficulties of the treatment, such as the increase in drug resistance and the side effects of conventional medication, leading to the development of serious diseases, impairing and compromising human health. Volatile oils have been the target of this search for new substances, as they are potential sources of new antioxidant and anti-inflammatory agents. Among the plant species characterized by the production of this chemical group, are those belonging to the Schinus genus, from the Anacardiaceae family, of large occurrence in Rio Grande do Sul. In this context, the present work aimed to analyze and identify the chemical composition and to evaluate the biological activities from the extraction of volatile oil from species of Schinus weinmannifolius and Schinus polygamus, native from Rio Grande do Sul. Thus, the volatile oil was obtained by hydrodistillation of several leaves and fruit collections of S. weinmannifolius and S. polygamus leaves. The chemical analysis, performed by gas chromatography coupled with mass detector (GC-MS), demonstrated great variety of compounds present in the samples, with predominance β-caryophyllene, germacrene D, bicyclogermacrene, germacrene A, δ-cadinene, espatulenol, caryophyllene oxide and α-cadinol for leaves and fruits of S. weinmannifolius and predominance of the compound aliphatic n-nonane for S. polygamus. The obtained oils were submitted to the reaction with 2,2-diphenyl-1-picrylhydrazyl (DPPH) and thiobarbituric acid reactive substances (TBARS) assays to evaluate the antioxidant activity, being this action only observed for leaves samples of S. weinmannifolius collected in autumn and winter of 2016 by the TBARS assay. In the DPPH method, leaves samples of S. weinmannifolius collected in the autumn of 2015 and 2016 presented low antioxidant activity at the concentration of 100 μg/mL. The anti-inflammatory potential was initially performed by the in vitro technique of the Boyden chamber, at concentrations 0,001 to 10 μg/mL. All samples showed significant inhibition of leukocyte migration in relation to the control, evaluated in the antichemotactic assay. The evaluation of in vivo anti-inflammatory activity by the paw edema method was conducted only for the sample that obtained the best result in the in vitro test, S. weinmannifolius oil, with significant inhibition of edema in the second hour of treatment, at dose 100 mg/kg. The results found support the traditional use of Schinus oils as anti-inflammatory agent.

A indústria alimentícia utiliza variados materiais para embalagens, sendo o polietileno de baixa densidade linear (PELBD) um dos materiais de maior importância, por possuir características únicas e adequadas à produção de embalagens. A geração de espécies químicas em embalagens de alimentos vem sendo foco de estudos no mundo. Este controle de compostos é denominado como estudo de Substâncias Não Intencionalmente Adicionadas (NIAS) e tem sua importância justificada pela preocupação com a saúde humana devido à capacidade de contaminação do alimento embalado. Para este estudo foram escolhidas duas resinas PELBD amplamente aplicadas na produção de embalagens alimentícias, analisadas na forma de pellets. As amostras foram nomeadas como PELBD1 e PELBD2 e analisadas antes e após exposição natural e acelerada (estufa a 50°C) de um, dois e três meses. Ambas apresentaram grande aumento no número de NIAS detectadas por Cromatografia Gasosa com detecção de Massas (GC-MS) após envelhecimentos, quando comparadas à resina virgem, chegando a 1100% em PELBD1 e 100% em PELBD2, com surgimento de substâncias oxigenadas e tóxicas. O aditivo antioxidante ativo foi sendo consumido e analisado via Cromatografia Líquida de Alta Eficiência (HPLC) ao longo das exposições, confirmando os efeitos do envelhecimento. Através do Infravermelho por transformada de Fourier (FTIR) foi verificada degradação inicial em PELBD1 exposto por três meses à estufa. Porém, até mesmo em períodos menores de exposição natural, a formação de grupos cromóforos foi comprovada pela análise de cor, onde houve pequeno e gradual aumento do amarelecimento e diminuição da brancura principalmente em PELBD1. Por Cromatografia de Permeação à Gel (GPC), as amostras apresentaram pequena tendência para diminuição de M̅z. Já nas análises de Reometria Rotacional com variação de Frequência (DSR), Índice de Fluidez (IF) e Calorimetria Exploratória Diferencial (DSC), foram observadas mínimas tendências de degradação. Pode-se concluir a partir destes resultados que a maior degradação e produção de NIAS ocorreram em ambiente acelerado. Esta pesquisa trouxe grandes contribuições para futuros trabalhos que envolvam o estudo de NIAS e suas condições de formação. / The food industry uses various packaging materials being linear low density polyethylene (LLDPE) one of the most important materials, have unique features suitable for the production and packaging. The generation of chemical species in food packages has been the focus of research in the world. This control compounds is referred to as study Non-Intentionally Added Substances (NIAS) and has its importance justified by concern human health because of capacity contamination the food packaging. For this study it was chosen two LLDPE resins widely applied in the production of food packaging, analyzed in the form of pellets. The samples were named as PELBD1 and PELBD2 and analyzed before and after natural and accelerated exposure (oven at 50°C) of a two and three months. Both showed huge increase in the number of NIAS detected by Gas Chromatography with Mass detection (GC-MS) after ageing, when compared to virgin resin, reaching of 1100% in PELBD1 and 100% in PELBD2 with the appearance of oxygen substances and toxic substances. The active antioxidant additive was being consumed and analyzed via High-Performance Liquid Chromatography (HPLC), during the exposures, confirming the effects of ageing. For Fourier Transform Infrared (FTIR) was observed in initial degradation PELBD1 exposed for three months in an oven. However, even at under natural exposure periods, the formation of chromophoric groups were confirmed by analysis of color where there was a slight and gradual increase in yellowing and brightness decreased mainly PELBD1. For the Gel Permeation Chromatography (GPC), the samples showed a slight tendency to decrease M̅z. Already in the analysis of Rheometry Rotational Varying Frequency (DSR), Melt Flow Index (MFI) and Differential Scanning Calorimetry (DSC), were observed minimum trends of degradation. It can concluded from these results that the greatest degradation and NIAS production occurred in an accelerated environment. This research has brought great contributions to future work involving the study of NIAS and their conditions of training.

Esta tese apresenta os resultados obtidos com a utilização de efluente produzido em digestor hidrolítico de lodo primário, para a remoção de nutrientes em reator em bateladas seqüenciais (RBS) tratando efluente de reator UASB. Assim, foram realizados dois estudos. O primeiro consistiu na verificação da melhor condição operacional para que se alcançasse a maior produção de ácidos graxos voláteis (AGV) em digestor hidrolítico de lodo (DHL), enquanto que, o segundo estudo, consistiu na avaliação do desempenho do RBS na remoção de nitrogênio e fósforo diante da aplicação dos AGV presentes no sobrenadante de lodo hidrolisado, do DHL. Os resultados do primeiro estudo permitiram a verificação de que a produção de AGV no DHL foi proporcional à taxa de aplicação. Foi verificado, também, que a maior produção destes ácidos se deu com a aplicação de tempos de detenção de lodo da ordem de 2 dias. Os resultados do segundo estudo permitiram a verificação de que a relação C/N igual, ou próxima a 3, se mostrou como a mais viável para o processo de desnitrificação, onde, neste caso foram observadas desnitrificações superiores a 70%. Nesta mesma condição, foram observadas remoções de fósforo da ordem de 25%. Entretanto, a utilização do sobrenadante de lodo hidrolisado, e os AGVs nele contido como fonte de carbono, ficou comprometida devido a alta concentração de nitrogênio (predominantemente sob a forma amoniacal) nele presente. O nitrogênio introduzido no RBS, pela adição de sobrenadante de lodo hidrolisado, se manteve inalterado fazendo com que o RBS acabasse por apresentar altas concentrações deste nutriente ao final do ciclo. Portanto, a aplicação deste tipo de fonte de carbono em escala plena para remoção de nutrientes, acabou ficando inviável da forma como se propôs, nesta pesquisa, que fosse realizada. / This thesis presents the results obtained with the use of effluent produced in primary sludge hydrolytic digester for nutrient removal in sequential batch reactor (SBR) treating effluents from an up-flow anaerobic sludge blanket (UASB) reactor. Two bench scale studies were developed. The first one consisted in verifying the best operational condition to produce volatile fatty acids (VFA) in the primary sludge hydrolytic reactor; the second one aimed to evaluate the sequential batch reactor (SBR) performances in the removal of nitrogen and phosphorus. In this operational condition, VFA from the liquid phase of the hydrolytic reactor was applied as external carbon source. The results from the first study permitted to verify that the VFA production in the HSR was proportional to the application rate (AR). The best VFA production was obtained for sludge retention time (SRT) of two days. In the second study, the best C/N ratio for the denitrification process was found to be close to 3, with denitrification efficiency higher than 70%. In this condition, phosphorus removal efficiency was approximately 25%. However, nitrogen removal was deeply affected by the high concentration of ammonium nitrogen in the liquid phase of the hydrolyzed sludge that remains unaltered in the SBR. Consequently, the use of such endogenous carbon source for denitrification needs further development in order to be applied in full-scale treatment systems.

Lactic acid bacteria are essential as flavor producers of cheese and fermented products. They are capable of catabolizing aromatic, branched chain, and sulfur amino acids to flavor compounds. During cheese ripening the numbers of lactococcal colonies decrease, but lactococci survive without replication in culture. This prompted an investigation into possible mechanisms of catabolism of branched chain amino acids into branched chain fatty acids and the physiological relevance of amino acid catabolism to the bacteria. We hypothesized that lactococci catabolize branched chain amino acids to branched chain fatty acids during nonculturability. Lactococci, lactobacilli, and brevibacteria catabolized both branched chain amino acids and keto acids into branched chain fatty acids. Lactococci survived carbohydrate-limited conditions for over 4 yrs. Their survival was represented by maintaining intracellular ATP, enzyme activity, membrane integrity, capability of ATP- and PMF-dependent substrate transport, transcription, and catabolism of amino acids to fatty acids. Assays conducted with NMR spectroscopy coupled with in silico analysis showed that branched chain substrates are catabolized via keto acids, HMG-CoA, and acetyl-CoA to branched chain fatty acids. A short list of candidate genes was identified for the pathway by gene expression analysis coupled to NMR analysis. The expression of these genes and the presence of the related catabolites were identified in long-term starved cultures of nonculturable lactococci. This verified that catabolism of branched chain amino acids to branched chain fatty acids occurred during the nonculturable state only and in conditions of carbohydrate deprivation. The pathway also facilitated fixation of carbon by lactococci, revealing the mechanism of survival of lactococci over 4 yrs in culture without the addition of external carbon sources. Between strains the availability of carbohydrate and acid stress played significant roles in modulating their ability to produce branched chain catabolites. The ability of lactococci to catabolize branched chain amino acids during sugar starvation represents a shift in carbon catabolic routes. The identified pathway also represented a balance between catabolism and anabolism, suggesting that the bacteria were in a homeostatic state during nonculturability. We accepted the hypothesis that nonculturable lactococci catabolized branched chain amino acids to branched chain fatty acids during starvation./p>