Brewer's spent grain (BSG) is the most abundant by-product of the brewing industry and its main application is limited to low-value cattle feed. Since BSG contains 20 to 25% of proteins, it has the potential to provide a new protein source to the food industry. In this research, an ultrasound-assisted enzymatic extraction was designed to extract protein hydrolysates from BSG. Original BSG and ultrasound pretreated BSG were hydrolyzed under different enzyme (Alcalase) loadings and incubation times. Centrifugation was applied to separate solubilized proteins from insoluble BSG residue. When the enzyme loading increased from 1 to 40 uL /g BSG, the solubilized proteins increased from 34% to 64.8%. The application of ultrasound further increased the solubilized proteins from 64.8% to 69.8%. Solubilized proteins from ultrasound pretreated BSG was significantly higher (p < 0.05) than that from the original BSG. Particle size distribution analysis showed that the application of ultrasound pretreatment reduced the BSG particle size from 331.2 to 215.7 um. Scanning electron microscopy images revealed that the BSG particle surface was partially ruptured by the ultrasound pretreatment. These two phenomena might have contributed to the increased protein separation efficiency with ultrasound pretreatment. The solubility (pH 1.0 to 11.0) of protein hydrolysate increased by the application of ultrasound and the ultrasound did not lead to the change of the amino acid composition of the separated protein hydrolysates. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis profile, the protein was degraded to peptides which had molecular weights lower than 15 kDa. The color of the separated protein hydrolysates by enzymatic hydrolysis was brighter and lighter than the original BSG. The application of ultrasound did not affect the color of the separated protein hydrolysates. Overall, the ultrasound pretreatment prior to enzymatic hydrolysis enhanced the extraction of proteins from BSG in terms of higher protein separation efficiency, lower enzyme loadings, and reduced incubation time. This study developed a novel and green method to effectively extract value-added protein hydrolysates from the low-value food processing byproducts. / MSLFS / Brewer’s spent grain (BSG) is the most abundant waste generated by beer industries after beer production and it is mainly used to feed cattle. Since BSG contains 20 to 25% proteins, it has the potential to provide a new protein source to food industries. The aim of this research is to study if the ultrasound technology can assist the enzymatic extraction of proteins from BSG. If it can, the cost of the protein extraction from BSG can be reduced. In this research, the original BSG and the BSG pretreated with ultrasound were incubated under different enzyme loadings and incubation times. The protein-rich liquid was separated from fiber-rich solids using a centrifuge. When the enzyme loading increased from 1 to 40 L /g BSG, 34% to 64.8% of proteins were separated from the original BSG. The application of ultrasound further increased the solubilized proteins from 64.8% to 69.8%. For the BSG pretreated with ultrasound, there were significantly more proteins separated from BSG compared to the original BSG. Particle size of the original BSG and the ultrasound pretreated BSG was measured, and the results showed that the application of ultrasound pretreatment decreased the BSG particle size from 331.2 to 215.7 µm. Scanning electron microscopy images were taken to investigate the effect of ultrasound on the surface of BSG particles. Based on the photos, we found that the BSG particle surface was partially broken by the ultrasound pretreatment. The surface was rough and contained large amounts of holes instead of being flat and smooth observed without ultrasound. Therefore, there were more locations for the enzyme to attack. These two phenomena might have contributed to extracting more proteins from BSG. Protein solubility (pH 1.0 to 11.0) increased by the application of ultrasound. The nutritional value of the protein extracted was not altered by the ultrasound. The extracted protein hydrolysates had a small molecular weight and the application of ultrasound did not affect the color of the extracted protein hydrolysates. The ultrasound pretreatment prior to enzymatic hydrolysis increased the extraction of proteins from BSG, decreased the enzyme consumption and incubation time. This study developed a novel and green method to effectively extract value-added protein hydrolysates from the low-value food processing byproducts.

Aquaculture, the cultivation of aquatic organisms in a controlled environment, offers both economic and nutritional benefits to human society. As there is an increased demand to feed a growing human population, many wild-caught fisheries have struggled due to the overexploitation of resources. Currently, production relies heavily on wild-caught fish to produce fishmeal to feed farm-raised fish. The demand for alternative materials in fish feeds has grown rapidly as fishmeal resources have become limited. Antibiotic resistance emergence in aquaculture systems is another area of concern. Reducing antibiotic use via alternate prophylactic measures to increase host health is an essential area of research; modulation of the host intestinal bacterial community via prebiotics is one possibility. Prebiotics refer to non-digestible food ingredients that are thought to stimulate the growth of beneficial bacteria, consequently benefiting host health by indirectly reducing the possibility of bacterial pathogen proliferation. This occurs through various measures such as competition for space and resources. The intestinal bacterial community has a significant impact on a variety of host factors that include host development, physiology, immunity, and nutrient acquisition. In turn, there are multiple factors impacting the bacterial community, including the presence of pathogens and/or antibiotics, environmental conditions, host genetics, and the diet consumed. To promote environmental sustainability and improve production and animal health in aquaculture, a collaboration was created with Anheuser-Busch of the brewing industry and Maltento, a functional ingredient company. With breweries around the globe, Anheuser-Busch produces consistent, food grade byproducts that are safe for human consumption. Two of the most prevalent brewery byproducts are brewer's spent yeast (BSY) and brewer's spent grain (BSG). BSY contains a variety of beneficial nutrients such as proteins, essential amino acids, and carbohydrates. BSG is high in fiber but low in protein; however, black soldier fly larvae can be cultured on BSG to convert the low-value product into insect biomass to be used in fish feed, as insects themselves are full of beneficial lipids and proteins. The objective of the work presented in this thesis was to evaluate the efficacy of using low-value brewery waste products, converted into high-value feed additives, for aquaculture practices. Specifically, the effects of dietary feed additives on the production, health, and intestinal bacterial community of aquaculture-raised rainbow trout were examined. Inadvertently, benefits of the feed additives on fish subjected to chronic and acute thermal stress were also assessed. Overall, the results of the study found that the feed additives did not significantly change the production efficiency of the rainbow trout, though some increase in growth was observed. When subjected to chronic thermal stress conditions, fish fed the experimental diets outperformed those fed the control diet regarding growth parameters. The intestinal bacterial community of the fish was significantly altered from the beginning of the trial compared to the end of the trial, though differences were not attributed to the feed additives. Instead, the resulting intestinal dysbiosis is believed to have stemmed from the physiological response of the fish to thermal stress conditions. When the fish underwent an acute thermal stress event, causing mortality, fish fed three of the five experimental diets were found to have higher survival rates compared to the control. Ultimately, results of this project suggest that the BSY and BSG-fed insect feed additives may have increased the health and robustness of the fish during a period of thermal stress. However, further research under controlled conditions is needed to evaluate if the observed host health benefits can directly be attributed to the feed additives. / Master of Science / Aquaculture refers to the method of rearing aquatic organism such as fish and shellfish under controlled conditions. Within the food industry, aquaculture is one of the fastest growing sectors, and provides important economic and nutritional benefits to humans. Additionally, aquaculture is an important alternative to fisheries that rely on catching fish from the natural environment. Wild-caught fisheries have struggled due to the overfishing, and unfortunately, many aquaculture practices still rely on wild-caught fisheries to produce fishmeal used in feed for carnivorous fish. Research into alternate protein sources to use in fish feed has been on the rise. Additionally, as the emergence of multi-drug resistant bacteria continues to increase, reducing antibiotic use has become a priority across all fields whether it be healthcare or the food industry. Within aquaculture, using alternative prophylactic measures such as prebiotics to increase animal health and disease resistance could lead to the overall reduction of antibiotic use. Prebiotics are non-digestible food ingredients believed to help the beneficial bacteria within the intestinal track to grow. In turn, the increased numbers of beneficial bacteria reduce the possibility of pathogenic bacteria invading and establishing a presence in the intestinal track. The intestinal microbiome refers to the various organisms, such as bacteria, viruses, and fungi, that live commensally within the host digestive tract. The bacterial community within the intestinal microbiome has many important roles, including effects on host development, physiology, immunity, and nutrient acquisition. Many factors also impact the bacterial community, including the presence of pathogens and/or antibiotics, environmental conditions, host genetics, and the diet consumed. To promote environmental sustainability and improve production and animal health in aquaculture, a collaboration was created with Anheuser-Busch of the brewing industry and Maltento, a functional ingredient company. With breweries around the globe, Anheuser-Busch produces consistent, food grade byproducts that are safe for human consumption. Two of the most prevalent brewery byproducts are brewer's spent yeast (BSY) and brewer's spent grain (BSG). These low-value waste products can consequently be converted into high-value feed additives for use in aquaculture. The objective of the work presented in this thesis was to evaluate the effects of BSY and BSG-fed insect dietary feed additives on the production, health, and intestinal bacterial community of aquaculture-raised rainbow trout. Unintentionally, benefits of the feed additives on fish subjected to chronic and acute high-temperature thermal stress were also explored. Overall, the results of the study found that while the feed additives did not significantly increase the growth of the rainbow trout, benefits were still observed. When subjected to chronically high-water temperatures, fish fed the experimental diets outperformed those fed the control diet regarding growth parameters. The intestinal bacterial community of the fish was significantly altered from the beginning of the trial compared to the end of the trial, though differences are not believed to be caused by the feed additives. Instead, the resulting shift in the bacterial community is believed to have stemmed from the stress-response of the fish triggered by high water temperature. When the fish underwent an acute thermal stress event, which caused mortality, fish fed three of the five experimental diets were found to have higher survival rates compared to the control. Ultimately, results of this project suggest that the feed additives may have increased the health and robustness of the fish while undergoing thermal stress. However, further research under controlled conditions is needed to evaluate if the observed host health benefits can be attributed directly to the feed additives.

Brewer's spent grain (BSG) is a main by-product of beer manufacturing and is rich in nutrients including 15-30% protein, making it a potentially valuable protein source for human food. Current challenges of extracting protein from BSG include low yields and high manufacturing costs, but the rising trend of plant-based diets for environmental and health reasons increases BSG's appeal. This research proposes an innovative extraction process utilizing wet fractionation and enzymatic purification that targets fiber to effectively separate proteins form BSG. Additionally, the feasibility of BSG protein as a food ingredient is explored, offering a unique approach that limits harsh processing commonly used for protein extraction. The choice of the enzyme (CTec 2 and Viscozyme L), enzyme dose, and incubation time (1, 3, 6, 9, and 24 hrs) were investigated to maximize protein content and recovery and further evaluate processing effects on protein functionality. Following wet fractionation, there was a notable reduction in fiber from 48.6% in the BSG to 22.5% in the filtrate (PRF) on a dry basis. Additionally, the protein content increased from 22.8 to 40.0% from the BSG to the PRF, respectively. The coarse fiber (CF) had an average protein content of 6.30%, highlighting the effectiveness of wet fractionation in enhancing protein recovery from BSG. The optimal enzymatic treatments condition identified was using CTec 2 to degrade fiber at a dose of 6.00% (g enzyme/g solid) for 24 hours. The enzymatic fiber hydrolysis proved to be able to remove lignocellulosic biomass from BSG resulting in a defatted protein concentrate (DPC) with a high protein content (52.8%) along with a high recovery rate (63%). All treatments resulted in protein functionality similar to the two control treatments. Varying fiber hydrolysis conditions did not have a significant effect on the functional properties. Several functional properties were improved using limited hydrolysis (LH) using proteases. LH had negative impacts on foaming stability and emulsifying properties; however, it improved the solubility, foaming capacity, WHC, and OHC of the BSG protein. Potential applications for the DPC include low moisture applications such as protein bars or granola. The DPCs could also have potential use in meat alternatives due to the high WHC and OHC and the need for varying protein solubility in meat alternatives. The LH protein has greater potential in applications such as protein beverages due to the high solubility. This process presents a promising protein extraction approach from BSG, offering producers the flexibility to tailor it to their specific application needs. After obtaining the protein concentrate, additional steps like defatting or limited hydrolysis can be applied to improve the purity and functionality of the result protein. Notably, this approach contributes to sustainable food production by addressing food waste and meeting the rising demand for sustainable protein sources to support the nutritional needs of a growing global population. / Master of Science in Life Sciences / Brewer's spent grain (BSG) is a by-product of beer manufacturing with rich nutrients, including 15-30% protein. It holds potential as a valuable protein source for human food. Current challenges of extracting protein from BSG include low yields and high costs, but the rising trend of plant-based diets for environmental and health reasons increases BSG's appeal. Its abundant availability year-round makes it an attractive option for human food. In this study, an innovative process to extract protein from BSG was explored using wet fractionation to separate the BSG by size. Enzymes were then used to remove remaining fiber. Various enzymes and times were tested to optimize extraction and achieve a high protein content and recovery rate. The protein's physicochemical and functional properties were then evaluated for potential uses in human food. After wet fractionation, nearly half the fiber was removed, and the protein content increased by 17.2%. Using an enzyme called CTec 2 for 24 hours worked best, resulting in a protein concentrate with a high protein content (52.8%) and good recovery (63.0%). Changes in enzyme treatments did not have a significant effect on the protein functional properties. Potential applications for the protein concentrates include low moisture applications such as protein bars or granola due to the poor solubility. The protein concentrates could also have potential use in meat alternatives due to the water and oil holding properties. The low solubility of the protein concentrates was improved using a process called limited hydrolysis making it possible to apply to products like protein beverages. This research highlights BSG's potential as a valuable protein source for protein bars, meat alternatives, and protein beverages. These findings provide a promising approach to utilize BSG as a valuable plant protein source for a healthier and more environmentally friendly food production.

No description available.

O presente estudo teve como objetivo avaliar a produção de etanol pela levedura Pichia stipitis, a partir do hidrolisado hemicelulósico do bagaço de malte (HHBM). Primeiramente estudou-se o efeito da suplementação nutricional do hidrolisado adicionando-se extrato de farelo de arroz (0 a 20% v/v), uréia (0 a 3 g/L) e extrato de levedura (0 a 3 g/L). Os resultados mostraram que o hidrolisado suplementado apenas com extrato de levedura proporcionou os melhores resultados de conversão (YP/S = 0,44 g/g) e produtividade em etanol (QP = 0,33 g/L.h). Em seguida foi avaliado o nível ótimo deste nutriente sobre a bioconversão, sendo confirmada a suplementação do HHBM com 3,0 g/L de extrato de levedura. Após o estabelecimento das condições nutricionais, a levedura foi cultivada em HHBM após o crescimento de 24 horas em meio semissintético, visando aclimatar as células aos inibidores presentes no hidrolisado. Este estudo resultou no aumento das velocidades do processo, sendo 16% sobre a produtividade volumétrica em etanol (QP) e 10% sobre a velocidade de consumo de substrato (QS), quando comparado ao cultivo das células somente em meio semissintético. Na etapa seguinte utilizou-se um planejamento fatorial 22 com face centrada para otimização das condições de pH e concentração inicial de células (X0) na fermentação em HHBM não destoxificado e após a destoxificação com carvão ativado. Foram obtidos modelos para descrever os valores de YP/S e QP na região estudada, sendo que para o HHBM não destoxificado os valores destes parâmetros foram 0,40 g/g e 0,65 g/L.h, respectivamente, em pH 6,4 e X0 = 5,0 g/L (modelo 1). Em HHBM destoxificado, as condições ótimas foram obtidas em pH 6,0 e 1,36 g/L de células (modelo 2), obtendo-se YP/S de 0,40 g/g e QP de 0,46 g/L.h. Nas condições otimizadas foram então realizados ensaios que confirmaram a validade dos modelos 1 e 2, obtendo-se a máxima concentração de etanol (23,4 g/L), YP/S de 0,41 g/g e QP de 0,65 g/L.h em HHBM não destoxificado. Realizouse ensaios para avaliação do efeito do ácido acético sobre a fermentação em meio semissintético por P. stipitis, empregando-se as condições de pH e X0 otimizadas em HHBM não destoxificado (modelo 1) e destoxificado (modelo 2). Este estudo mostrou que nas condições do modelo 1, o ácido acético favoreceu a bioconversão sendo os melhores resultados obtidos na presença deste ácido (YP/S = 0,47 g/g e QP = 1,08 g/L.h). Por outro lado, nas condições do modelo 2, os valores de YP/S foram similares, enquanto que com a adição de ácido acético ao meio de fermentação, o valor de QP foi reduzido em 53%. Na fermentação em biorreator, o emprego das condições otimizadas em frascos (pH 6,4 e 5,0 g/L de células) resultaram em valores de QP 48% inferiores ao obtido em frascos (0,65 para 0,44 g/L.h), entretanto YP/S foi apenas 10% inferior (0,41 para 0,37 g/g). No presente estudo, conclui-se que a suplementação nutricional do HHBM e a otimização das condições de pH e X0 resultaram em valores promissores para os principais parâmetros da fermentação por P. stipitis, ressaltando o potencial deste hidrolisado em processos biotecnológicos para produção de etanol. / This study aimed to evaluate the ethanol production by Pichia stipitis in brewer\'s spent grain hemicellulosic hydrolysate (BSGHH). Initially, the effect of nutritional supplementation was evaluated by adding rice bran extract, urea and yeast extract. The results showed that supplementation only with yeast extract promoted the highest conversion values (YP/S = 0.44 g/L) and ethanol productivity (QP = 0.33 g/L.h). Additional assays showed that the optimal concentration of this nutrient was 3.0 g/L. To acclimate the cells to inhibitors present in BSGHH the yeast was cultivated in hydrolysate after growth for 24 hours in semisynthetic medium. This study resulted in increased of the process rates, 16% of the ethanol productivity (QP) and 10% on the substrate consumption (QS) when compared to growing cells only in the semisynthetic medium. In the second step a 22 full factorial centeredface design was employed to optimized the conditions of pH and initial cells concentration (X0) in fermentation of hydrolysate undetoxified and after detoxification with activated charcoal. Mathematical models that relate the YP/S and QP were obtained. For non-detoxified BSGHH (model 1) the optimal conditions of pH (6.4) and X0 (5.0 g/L) showed values parameters of 0.41 g/g and 0.65 g/L.h, respectively. In detoxified BSGHH (model 2) the optimum conditions of pH (6.0) and X0 (1.36 g/L), resulted in YP/S and QP values of 0.40 g/g and 0.46 g/L.h, respectively. Under these conditions, the the validity of the models were confirmed. The effect of acetic acid on fermentation by P. stipitis in semisynthetic medium, employing optimized conditions of pH and X0 in model 1 and model 2 was evaluated. The results showed that under the conditions of model 1 and in a concentration of 2,9 g/L, the acetic acid favored the bioconversion by P. stipitis, increasing the YP/S (15 %) and QP (66 %). On the other hand, in the conditions of the model 2 the YP/S values were similar, whereas the QP values were reduced by 53% when the acetic acid was added. By using these optimized conditions in bioreactor fermentation it was obtained the ethanol productivity was approximately 48% lower (0.65 to 0.44 g/L.h), however the ethanol production was similar as compared to fermentation flasks. It is possible conclude that the HHBSG requires nutritional supplementation and that the optimized conditions of pH and initial cells concentration can be used as a strategy in order to raising the fermentation parameters.

A utilização de bioinseticida à base de Bacillus thuringiensis tem se mostrado eficaz no combate às larvas de insetos veiculadores de doenças como a dengue, elefantíase e malária. O bioinseticida apresenta alta especificidade, ausência de resistência nos insetos alvos e baixo impacto ambiental comparado aos inseticidas químicos. Porém, a difusão do uso de bioinseticida apresenta alguns desafios como o custo da produção e da formulação. O levedo excedente da indústria cervejeira pode tornar a produção do bioinseticida economicamente viável devido ao seu baixo custo, assim como o glicerol, resíduo da produção de biodiesel. A escolha adequada dos componentes do meio de cultivo é essencial para o sucesso de um produto e deve ser realizada buscando-se obter uma alta produtividade com o menor custo. Além disso, no caso do bioinseticida, os componentes do meio exercem uma grande influência no crescimento e na síntese de toxinas pela bactéria. No presente trabalho, propôs-se formular um meio de cultivo para Bacillus thuringiensis var. israelensis utilizando levedo de cerveja e glicerol proveniente da fabricação de biodiesel, visando obter um meio fermentado com elevada toxicidade e de menor custo, comparado aos obtidos atualmente. Inicialmente foi definido um procedimento de preparo do extrato de levedo de cerveja. Constatou-se que o melhor método consiste da esterilização da suspensão de levedo de cerveja (121 ºC por 20 minutos) seguida de filtração qualitativa. Foi definida também a faixa de concentração de extrato de levedo de cerveja que seria utilizada no estudo de composição do meio (10 g/L a 30 g/L). Os ensaios foram realizados em frascos Erlenmeyer de 1000 mL em incubadora de movimento recíproco. Os componentes do meio de cultivo (glicerol residual, extrato de levedo de cerveja, KNO3, CaCl2.2H2O, MgSO4.7H2O, MnSO4, K2HPO4 e KH2PO4) foram avaliados segundo um planejamento experimental e, nas condições estudadas, não exerceram influência significativa sobre a produção de toxinas. Definiu-se então a composição do meio de fermentação em termos de fonte de carbono e nitrogênio. O glicerol e o extrato de levedo de cerveja não exerceram influência significativa sobre o consumo de substrato pela bactéria. Quanto à concentração celular máxima, o aumento da concentração de glicerol apresentou efeito negativo, enquanto o aumento da concentração de extrato de levedo de cerveja apresentou efeito positivo. Embora as concentrações das fontes de carbono e nitrogênio não tenham sido significativas para a atividade larvicida, a interação entre os fatores foi significativa em seu nível negativo. O melhor resultado quanto à atividade larvicida do meio foi apresentado pelo meio composto por 10 g/L de glicerol residual, 30 g/L de extrato de levedo de cerveja, 5 g/L de KNO3, 0,12 g/L de CaCl2.2H2O, 1,5 g/L de MgSO4.7H2O, 0,09 g/L de MnSO4, 1,5 g/L de K2HPO4 e 1,5 g/L de KH2PO4. Essa mesma composição de meio de cultivo foi testada em um ensaio em biorreator, obtendo-se um meio fermentado mais eficiente que o ensaio realizado em incubadora, e com CL50 de 2,43 &#181;L. / The use of bioinsecticide based on Bacillus thuringiensis has been shown to be effective against larvae of insects able to spread diseases such as dengue, malaria and elephantiasis. The bioinsecticide has high specificity, absence of resistance in target insects and low environmental impact compared to chemical insecticides. However, there are some challenges in the way of widespread use of bioinsecticide, such as the costs of production and formulation. The surplus yeast from the brewing industry can make the production of bioinsecticide economically feasible due to its low cost, as well as the residual glycerol from the biodiesel production. The proper choice of the components of the culture medium is essential for the success of a product and must be carried out in order to obtain a high productivity at the lowest cost. Furthermore, in the case of bioinsecticide, medium components have a major influence on the growth and synthesis of toxins by the bacteria. In the present study it was proposed the formulation of a fermentation medium for Bacillus thuringiensis var. israelensis using brewer\'s yeast and glycerol from the biodiesel manufacturing, seeking to obtain a fermented broth with high toxicity and lower cost compared to those currently obtained. Initially it was defined a procedure to prepare the extract of brewer\'s yeast. It was found that the best method is the sterilization of the brewer\'s yeast suspension (121 °C for 20 minutes) followed by qualitative filtration. The range of brewer\'s yeast extract concentration to be used in the study of the medium composition was also defined (10 g/L to 30 g/L). Assays were performed in 1000 mL Erlenmeyer flasks in a reciprocal motion shaker. The components of the medium (glycerol waste, brewer\'s yeast extract, KNO3, CaCl2.2H2O, MgSO4.7H2O, MnSO4, K2HPO4 e KH2PO4) were evaluated according to an experimental design and, considering the studied conditions, they did not have significant influence on the toxin production. Thus, the composition of the fermentation medium was defined in terms of nitrogen and carbon source. Glycerol and brewer\'s yeast extract did not have any significant influence on the substrate consumption by the bacteria. For the maximum cell concentration, the increase of glycerol concentration showed a negative effect, while increasing the concentration of brewer\'s yeast extract showed a positive effect. Although the concentrations of carbon and nitrogen sources were not significant for the larvicidal activity, the interaction between such factors was significant in its negative level. The best result of larvicidal activity was shown by the following medium: 10 g/L of residual glycerol, 30 g/L of brewer\'s yeast extract, 5 g/L of KNO3, 0.12 g/L of CaCl2.2H2O, 1.5 g/L of MgSO4.7H2O, 0.09 g/L of MnSO4, 1.5 g/L of K2HPO4 and 1.5 g/L of KH2PO4. This same composition of the culture medium was tested in a bench fermenter, resulting in a more efficient fermented broth than the one obtained in shaker flask, and with LC50 of 2.43 &#181;L.

A utilização de bioinseticida à base de Bacillus thuringiensis tem se mostrado eficaz no combate às larvas de insetos veiculadores de doenças como a dengue, elefantíase e malária. O bioinseticida apresenta alta especificidade, ausência de resistência nos insetos alvos e baixo impacto ambiental comparado aos inseticidas químicos. Porém, a difusão do uso de bioinseticida apresenta alguns desafios como o custo da produção e da formulação. O levedo excedente da indústria cervejeira pode tornar a produção do bioinseticida economicamente viável devido ao seu baixo custo, assim como o glicerol, resíduo da produção de biodiesel. A escolha adequada dos componentes do meio de cultivo é essencial para o sucesso de um produto e deve ser realizada buscando-se obter uma alta produtividade com o menor custo. Além disso, no caso do bioinseticida, os componentes do meio exercem uma grande influência no crescimento e na síntese de toxinas pela bactéria. No presente trabalho, propôs-se formular um meio de cultivo para Bacillus thuringiensis var. israelensis utilizando levedo de cerveja e glicerol proveniente da fabricação de biodiesel, visando obter um meio fermentado com elevada toxicidade e de menor custo, comparado aos obtidos atualmente. Inicialmente foi definido um procedimento de preparo do extrato de levedo de cerveja. Constatou-se que o melhor método consiste da esterilização da suspensão de levedo de cerveja (121 ºC por 20 minutos) seguida de filtração qualitativa. Foi definida também a faixa de concentração de extrato de levedo de cerveja que seria utilizada no estudo de composição do meio (10 g/L a 30 g/L). Os ensaios foram realizados em frascos Erlenmeyer de 1000 mL em incubadora de movimento recíproco. Os componentes do meio de cultivo (glicerol residual, extrato de levedo de cerveja, KNO3, CaCl2.2H2O, MgSO4.7H2O, MnSO4, K2HPO4 e KH2PO4) foram avaliados segundo um planejamento experimental e, nas condições estudadas, não exerceram influência significativa sobre a produção de toxinas. Definiu-se então a composição do meio de fermentação em termos de fonte de carbono e nitrogênio. O glicerol e o extrato de levedo de cerveja não exerceram influência significativa sobre o consumo de substrato pela bactéria. Quanto à concentração celular máxima, o aumento da concentração de glicerol apresentou efeito negativo, enquanto o aumento da concentração de extrato de levedo de cerveja apresentou efeito positivo. Embora as concentrações das fontes de carbono e nitrogênio não tenham sido significativas para a atividade larvicida, a interação entre os fatores foi significativa em seu nível negativo. O melhor resultado quanto à atividade larvicida do meio foi apresentado pelo meio composto por 10 g/L de glicerol residual, 30 g/L de extrato de levedo de cerveja, 5 g/L de KNO3, 0,12 g/L de CaCl2.2H2O, 1,5 g/L de MgSO4.7H2O, 0,09 g/L de MnSO4, 1,5 g/L de K2HPO4 e 1,5 g/L de KH2PO4. Essa mesma composição de meio de cultivo foi testada em um ensaio em biorreator, obtendo-se um meio fermentado mais eficiente que o ensaio realizado em incubadora, e com CL50 de 2,43 &#181;L. / The use of bioinsecticide based on Bacillus thuringiensis has been shown to be effective against larvae of insects able to spread diseases such as dengue, malaria and elephantiasis. The bioinsecticide has high specificity, absence of resistance in target insects and low environmental impact compared to chemical insecticides. However, there are some challenges in the way of widespread use of bioinsecticide, such as the costs of production and formulation. The surplus yeast from the brewing industry can make the production of bioinsecticide economically feasible due to its low cost, as well as the residual glycerol from the biodiesel production. The proper choice of the components of the culture medium is essential for the success of a product and must be carried out in order to obtain a high productivity at the lowest cost. Furthermore, in the case of bioinsecticide, medium components have a major influence on the growth and synthesis of toxins by the bacteria. In the present study it was proposed the formulation of a fermentation medium for Bacillus thuringiensis var. israelensis using brewer\'s yeast and glycerol from the biodiesel manufacturing, seeking to obtain a fermented broth with high toxicity and lower cost compared to those currently obtained. Initially it was defined a procedure to prepare the extract of brewer\'s yeast. It was found that the best method is the sterilization of the brewer\'s yeast suspension (121 °C for 20 minutes) followed by qualitative filtration. The range of brewer\'s yeast extract concentration to be used in the study of the medium composition was also defined (10 g/L to 30 g/L). Assays were performed in 1000 mL Erlenmeyer flasks in a reciprocal motion shaker. The components of the medium (glycerol waste, brewer\'s yeast extract, KNO3, CaCl2.2H2O, MgSO4.7H2O, MnSO4, K2HPO4 e KH2PO4) were evaluated according to an experimental design and, considering the studied conditions, they did not have significant influence on the toxin production. Thus, the composition of the fermentation medium was defined in terms of nitrogen and carbon source. Glycerol and brewer\'s yeast extract did not have any significant influence on the substrate consumption by the bacteria. For the maximum cell concentration, the increase of glycerol concentration showed a negative effect, while increasing the concentration of brewer\'s yeast extract showed a positive effect. Although the concentrations of carbon and nitrogen sources were not significant for the larvicidal activity, the interaction between such factors was significant in its negative level. The best result of larvicidal activity was shown by the following medium: 10 g/L of residual glycerol, 30 g/L of brewer\'s yeast extract, 5 g/L of KNO3, 0.12 g/L of CaCl2.2H2O, 1.5 g/L of MgSO4.7H2O, 0.09 g/L of MnSO4, 1.5 g/L of K2HPO4 and 1.5 g/L of KH2PO4. This same composition of the culture medium was tested in a bench fermenter, resulting in a more efficient fermented broth than the one obtained in shaker flask, and with LC50 of 2.43 &#181;L.

Sustainable development is a growing concern for inhabitants of the planet earth. Consumption of fossil sources keeps up the depletion of nature’s capital and causes environmental impacts. One solution to have a sustainable society is to reduce the dependency on fossil fuels and substitute them by renewable energy sources. Among different types of renewable energy, biofuels have great potential for development and improvement. Though, the production of biofuels is criticized by many experts from the energy efficiency, environmental and economical points of view. Biogas as one type of first generation biofuels is achieved from the wastes and by-products of other industries, and can be used as a transportation fuel in the form of biomethane. The use of by-products may give added value as inputs to the biogas production process, a process which may be called "upcycling." The aim of upcycling is to convert wastes into new materials with higher quality or higher environmental value in order to reduce the consumption of raw materials which results in decreasing of energy usage and environmental impacts The aim of this thesis is to study the possibility of a biogas plant to act as an upcycler of wastes and by-products through anaerobic digestion process by the use of exergy analysis and economic evaluation. An imaginary biogas plant which uses a major by-product of brewing industry, i.e. Brewer’s Spent Grain (BSG), is considered to quantify the added value by biogas production process. The results of the exergy analysis show that the exergy of the input BSG (78,320 MJ) is upgraded into two main products as biomethane (47,430 MJ) and biofertilizer (37,026 MJ) with a total exergy amount of 84,456 MJ. On the other hand, the economic analysis of the studied biogas production process indicates that the biogas plant has the added value for the input material. In the economic analysis, the annual costs and benefits of the biogas production is calculated. The results show that the production of biomethane and biofertilizer from the by-product of brewing industry is profitable. However, the price of input BSG and also the variation price of the biofertilizer in different seasons, have great impact on the economy of a biogas plant. The outcomes from exergy and economic analysis are indicated that the biogas production process is an "upcycling" process which has the added value for the inputs, from both economic and quality points of view. The exergy and economic evaluation may be used as indicators of the sustainable development, but only increasing the exergy and the economic value of a production process alone should not be considered as the sustainability of a system.

O presente estudo teve como objetivo avaliar a produção de etanol pela levedura Pichia stipitis, a partir do hidrolisado hemicelulósico do bagaço de malte (HHBM). Primeiramente estudou-se o efeito da suplementação nutricional do hidrolisado adicionando-se extrato de farelo de arroz (0 a 20% v/v), uréia (0 a 3 g/L) e extrato de levedura (0 a 3 g/L). Os resultados mostraram que o hidrolisado suplementado apenas com extrato de levedura proporcionou os melhores resultados de conversão (YP/S = 0,44 g/g) e produtividade em etanol (QP = 0,33 g/L.h). Em seguida foi avaliado o nível ótimo deste nutriente sobre a bioconversão, sendo confirmada a suplementação do HHBM com 3,0 g/L de extrato de levedura. Após o estabelecimento das condições nutricionais, a levedura foi cultivada em HHBM após o crescimento de 24 horas em meio semissintético, visando aclimatar as células aos inibidores presentes no hidrolisado. Este estudo resultou no aumento das velocidades do processo, sendo 16% sobre a produtividade volumétrica em etanol (QP) e 10% sobre a velocidade de consumo de substrato (QS), quando comparado ao cultivo das células somente em meio semissintético. Na etapa seguinte utilizou-se um planejamento fatorial 22 com face centrada para otimização das condições de pH e concentração inicial de células (X0) na fermentação em HHBM não destoxificado e após a destoxificação com carvão ativado. Foram obtidos modelos para descrever os valores de YP/S e QP na região estudada, sendo que para o HHBM não destoxificado os valores destes parâmetros foram 0,40 g/g e 0,65 g/L.h, respectivamente, em pH 6,4 e X0 = 5,0 g/L (modelo 1). Em HHBM destoxificado, as condições ótimas foram obtidas em pH 6,0 e 1,36 g/L de células (modelo 2), obtendo-se YP/S de 0,40 g/g e QP de 0,46 g/L.h. Nas condições otimizadas foram então realizados ensaios que confirmaram a validade dos modelos 1 e 2, obtendo-se a máxima concentração de etanol (23,4 g/L), YP/S de 0,41 g/g e QP de 0,65 g/L.h em HHBM não destoxificado. Realizouse ensaios para avaliação do efeito do ácido acético sobre a fermentação em meio semissintético por P. stipitis, empregando-se as condições de pH e X0 otimizadas em HHBM não destoxificado (modelo 1) e destoxificado (modelo 2). Este estudo mostrou que nas condições do modelo 1, o ácido acético favoreceu a bioconversão sendo os melhores resultados obtidos na presença deste ácido (YP/S = 0,47 g/g e QP = 1,08 g/L.h). Por outro lado, nas condições do modelo 2, os valores de YP/S foram similares, enquanto que com a adição de ácido acético ao meio de fermentação, o valor de QP foi reduzido em 53%. Na fermentação em biorreator, o emprego das condições otimizadas em frascos (pH 6,4 e 5,0 g/L de células) resultaram em valores de QP 48% inferiores ao obtido em frascos (0,65 para 0,44 g/L.h), entretanto YP/S foi apenas 10% inferior (0,41 para 0,37 g/g). No presente estudo, conclui-se que a suplementação nutricional do HHBM e a otimização das condições de pH e X0 resultaram em valores promissores para os principais parâmetros da fermentação por P. stipitis, ressaltando o potencial deste hidrolisado em processos biotecnológicos para produção de etanol. / This study aimed to evaluate the ethanol production by Pichia stipitis in brewer\'s spent grain hemicellulosic hydrolysate (BSGHH). Initially, the effect of nutritional supplementation was evaluated by adding rice bran extract, urea and yeast extract. The results showed that supplementation only with yeast extract promoted the highest conversion values (YP/S = 0.44 g/L) and ethanol productivity (QP = 0.33 g/L.h). Additional assays showed that the optimal concentration of this nutrient was 3.0 g/L. To acclimate the cells to inhibitors present in BSGHH the yeast was cultivated in hydrolysate after growth for 24 hours in semisynthetic medium. This study resulted in increased of the process rates, 16% of the ethanol productivity (QP) and 10% on the substrate consumption (QS) when compared to growing cells only in the semisynthetic medium. In the second step a 22 full factorial centeredface design was employed to optimized the conditions of pH and initial cells concentration (X0) in fermentation of hydrolysate undetoxified and after detoxification with activated charcoal. Mathematical models that relate the YP/S and QP were obtained. For non-detoxified BSGHH (model 1) the optimal conditions of pH (6.4) and X0 (5.0 g/L) showed values parameters of 0.41 g/g and 0.65 g/L.h, respectively. In detoxified BSGHH (model 2) the optimum conditions of pH (6.0) and X0 (1.36 g/L), resulted in YP/S and QP values of 0.40 g/g and 0.46 g/L.h, respectively. Under these conditions, the the validity of the models were confirmed. The effect of acetic acid on fermentation by P. stipitis in semisynthetic medium, employing optimized conditions of pH and X0 in model 1 and model 2 was evaluated. The results showed that under the conditions of model 1 and in a concentration of 2,9 g/L, the acetic acid favored the bioconversion by P. stipitis, increasing the YP/S (15 %) and QP (66 %). On the other hand, in the conditions of the model 2 the YP/S values were similar, whereas the QP values were reduced by 53% when the acetic acid was added. By using these optimized conditions in bioreactor fermentation it was obtained the ethanol productivity was approximately 48% lower (0.65 to 0.44 g/L.h), however the ethanol production was similar as compared to fermentation flasks. It is possible conclude that the HHBSG requires nutritional supplementation and that the optimized conditions of pH and initial cells concentration can be used as a strategy in order to raising the fermentation parameters.

This master‘s thesis deals with the influence of disintegration of brewer's grains for the production of biogas by anaerobic fermentation. The first part introduces us with the issue of biodegradable wastes. There are given their characteristics, ways of handling with them and legislative restrictions associated with them. In other sections there is a detailed description of anaerobic fermentation itself, including characteristics of biogas. Then there is issue of disintegration during fermentation including a list and characteristic of some disintegration methods. The main part of the thesis consists of description of the experiments on which it is observed increment of biogas based on the disintegration of brewer's grains and the evaluation of the individual experiments including the calculation of economic of the disintegration in laboratory conditions.