Considerando o crescimento do setor sucroenergético na última década e a busca por combustíveis renováveis, a fim de reduzir a dependência aos combustíveis fósseis, pesquisas nas áreas de açúcar e álcool estão comumente ligadas ao desenvolvimento de tecnologias que visem obtenção de processos e produtos com qualidade, lucro, altos rendimentos, minimização de perdas e gastos. O desenvolvimento de equipamentos e procedimentos analíticos que busquem vantagens competitivas em relação às metodologias convencionais está cada dia mais evidente e no processo de produção de açúcar e álcool, faz-se cada vez mais usuais. Uma dessas técnicas analíticas é a cromatografia. Por isso, a necessidade de trabalhos que avaliem técnicas cromatográficas aumenta diariamente, visto a importância que essas têm dentro do processo produtivo, no controle de qualidade e para a tomada de decisões. Por esses motivos, o objetivo deste trabalho foi avaliar o uso da técnica de cromatografia de íons para o controle analítico e de qualidade em fermentações para a produção de etanol. Para a realização deste estudo, inicialmente foi realizada validação da metodologia para análise de açúcares e depois foram realizados ensaios de fermentação. Os ensaios consistiram em fermentações com três tipos de mostos diferentes (três tratamentos), sendo os mostos obtidos a partir de caldo ou de xarope ou misto ( caldo + xarope), com teores de ART (açúcares redutores totais, respectivamente de 146,26 (±1,53) g L-1, 148,61 (±3,62) g L-1 e 158,35 (±4,67) g L-1, resultados estes obtidos pela cromatografia iônica e 107,75 (±3,19) g L-1, 115,90 (±10,41) g L-1 e 134,53 (±8,33) g L-1 para os mostos a base de caldo, xarope e misto, quando analisados pela metodologia convencional. Cada tratamento foi feito em cinco repetições, utilizando 300 mL de mosto cada, onde foram inoculados 9g de levedura Saccharomyces cerevisiae. Os pârametros analisados foram: viabilidade celular, brotamento, brotos vivos das leveduras, teores de Brix, teores de AR e ART por cromatografia iônica e por DNS, eficiência e rendimento fermentativo. Os resultados obtidos para AR final no caldo, xarope e misto na análise cromatográfica foram de 0,197 (±0,04) g L-1, 0,283 (±0,12) g L-1 e 0,259 (±0,08) g L-1 , respectivamente. Por outro lado, nas análises por DNS foram obtidos valores de: 0,352 (±0,14) g L-1, 0,811 (±0,03) g L-1 e 0,572 (±0,03) g L-1, respectivamente. Os valores de ART no vinho encontrados no caldo, xarope e misto por cromatografia foram 0,231 (±0,05) g L-1, 0,358 (±0,09) g L-1 e 0,335 (±0,19) g L-1 enquanto por DNS os teores foram 0,389 (±0,04) g L-1, 1,521 (±0,20) g L-1 e 0,906 (±0,21) g L-1. Estes valores geraram rendimentos fermentativos médios nos mostos fermentados obtidos a partir de caldo de cana, xarope e xarope + caldo de cana, foram 89,77 %, 83,59% e 76,65 %, respectivamente. Nos mostos fermentados obtidos a partir de caldo de cana, xarope e xarope + caldo de cana, os rendimento fermentativos foram 121,88 %, 106,82 e 90,23% para as análises cromatográficas e DNS, respectivamente. Conclui-se que, a técnica de cromatografia de íons quando utilizada para o controle analítico da fermentação alcoólica permite ter como vantagens em relação às técnicas convencionais analisadas, precisão, exatidão e robustez do método, rapidez de análise e facilidade no preparo das amostras, além de versatilidade, sendo capaz de analisar outros compostos de uma mesma amostra analítica, tais como ácidos, cátions e ânions, sendo capaz de fornecer resultados confiáveis para o controle analítico e de qualidade e monitoramento em fermentações para a produção de etanol. / Considering the growth of the sugar-energy sector in the last decade and the search for renewable fuels, in order to reduce dependence on fossil fuels, research in the areas of sugar and alcohol are commonly linked to the development of technologies that aim at obtaining processes and products with quality, high profit, and loss and expense minimization. The development of equipment and analytical procedures that seek competitive advantages over conventional methodologies is increasing, and arriving at the plants and distilleries for the production of sugar and alcohol, becoming more and more usual. Such is the case of chromatography. The necessity for research evaluating chromatographic techniques increases daily, considering the importance they have within the production process and in quality control. Thus, this work aims to evaluate the use of chromatographic techniques for analytical and quality controls in fermentations for ethanol production, through sugar analysis by traditional methods and ion chromatography. For the accomplishment of this study, initially the sugar analysis methodology was validated, and then fermentation tests were carried out. Such tests consisted of three treatments, each one of them composed of broth, syrup and mixed content (broth + syrup), with the respective Total Reducing Sugars (ART) of 146,26(±1,53) g L-1, 148,61 (±3,62) g L-1 and 158,35 (±4,67) g L-1, wen obtained by ionic chromatography, and, respectively, of 107,75 (±3,19) g L-1, 115,90 (±10,41) g L-1 and 134,53 (±8,33) g L-1 through the conventional methodology. Each treatment had five repetitions, with 300ml of wort each, where 9g of Saccharomyces cerevisae yeast were inoculated. The parameters evaluated were: cell viability, budding, living buds of yeast, °Brix levels, AR and ART levels through ionic chromatography and traditional method, efficiency and fermentative yield. The AR obtained results were 0,197 (±0,04) g L-1, 0,283 (±0,12) g L-1 and 0,259 (±0,08) g L-1 respectively for broth, syrup and mixture in the ionic chromatographic analysis, and 0,352 (±0,14) g L-1, 0,811 (±0,03) g L-1 and 0,572 (±0,03) g L-1 in the traditional analysis method. The ART obtained results were 0,231 (±0,05) g L-1, 0,358 (±0,09) g L-1 and 0,335 (±0,19) g L-1 through the chromatographic method and 0,389 (±0,04) g L-1, 1,521 (±0,20) g L-1 e 0,906 (±0,21) g L-1 through the traditional method. These values generated mean fermentative yields of respectively 89,77%, 83,59% and 76,65% for broth, syrup and mixed samples, in the chromatographic analysis and 121,88%, 106,82% and 90,23%, in the conventional analysis. The fermentative efficiency was satisfactory for the chromatographical methodology, with means of 83,25%, versus 106,31% of the conventional methodology. It was concluded that the ionic chromatography technique, when used for the analytical control of alcoholic fermentation, has advantages when compared to the analyzed conventional technique, such as precision, accuracy and methodological robustness, faster analysis, shorter sample preparation time, and versatility, being capable of analyzing different substances in the same source, such as acids, cations and anions, being capable of supplying trustworthy results for analytical and quality control and the surveillance of fermentations for ethanol production.

Na primeira etapa deste trabalho avaliou-se o desempenho fermentativo de 6 cepas de leveduras comerciais utilizadas na produção de vinhos e cerveja, em mosto de mel 30°Brix suplementado com 0,5 g/L de peptona e 1 g/L de extrato de levedura. Como resposta determinou-se o crescimento das leveduras em número de células e as concentrações de glicose, frutose e etanol por HPLC, que permitiram determinar os parâmetros rendimento, eficiência de fermentação e produtifidade em etanol, bem como a produção de sulfeto de hidrogênio. Assim, selecionou-se a cepa Saccharomyces bayanus - Pasteur Champagne - Red Star, que teve seu comportamento avaliado em mosto de mel suplementado com diferentes nutrientes. Os resultados demonstraram que a suplementação do mosto de mel com sais ou com suplemento comercial Enovit contribuiu para o bom desempenho da referida cepa. Na etapa seguinte, estudou-se a produção de hidromel em escala piloto a 18°C em reatores de polipropileno contendo 130 L de mosto sem suplementação (controle), suplementado com Enovit e suplementado com pedaços de maçãs (10% m/v). Terminada a fermentação, após 60 dias, o hidromel foi devidamente caracterizado físico-quimicamente quanto aos padrões de qualidade e identidade para hidromel estabelecidos pela Legislação Brasileira. Os resultados demonstraram que, com exceção da acidez total, todos os parâmetros avaliados se encontravam em consonância com os referidos padrões. Assim, as formulações estudadas foram adequadas para a obtenção de hidromel. Terminada a fermentação o hidromel foi submetido ao processo de envelhecimento a 20°C, por 216 dias, em tonel de carvalho (50L); galão de plástico (20 L); e frasco de vidro tipo bolha (20L), sendo neste período, caracterizado físico-químicamente, bem como quanto aos teores de compostos fenólicos totais, capacidade antioxidante, presença de aminas bioativas e compostos aromáticos derivados do tonel de carvalho. O hidromel envelhecido em galão de plástico apresentou teores de extrato seco reduzido abaixo do mínimo estabelecido nos padrões da legislação. A utilização de maçãs como suplemento do mosto de mel resultou em hidromel com maior teor de compostos fenólicos e maior capacidade antioxidante. As concentrações de aminas bioativas encontradas não comprometem a qualidade do hidromel, nem oferecem risco à saúde do consumidor. A análise de compostos fenólicos dos hidroméis envelhecidos em tonel de carvalho revelou que o processo de envelhecimento promoveu a extração de furfural, vanilina e ácido gálico, compostos que contribuem para o perfil sensorial da bebida, ficaram abaixo do limiar de percepção sensorial, fato que pode ser alterado pelo aumento do tempo de envelhecimento. Quanto a intenção de compra, os hidroméis avaliados apresentaram mais de 50% de notas variando entre 3 (tenho dúvidas se compraria) e 5 (certamente compraria), sendo os de melhor aceitação envelhecidos em recipientes de plástico e vidro. A análise sensorial revelou pouca influência do recipiente de envelhecimento sobre os atributos avaliados e a intensão de compra mostrou que os provadores preferiram os hidroméis envelhecidos em recipientes inertes (vidro e plástico). Diante do exposto, conclui-se que a produção de hidromel e seu posterior envelhecimento, por 216 dias, em recipientes inertes ou em tonéis de carvalho recondicionados, resultam em bebidas que apresentam boa aceitação, sendo uma alternativa viável para apicultores diversificarem a sua produção. / In the first stage of this work it was evaluated the fermentation performance of six commercial strains of yeast used in the of wine and beer production in honey wort 30 °Brix supplemented with 0.5 g / L peptone and 1 g / L yeast extract. In response, it was determined the yeast growth by number of cells and concentrations of glucose, fructose and ethanol by HPLC, as well as fermentation parameters as yield, efficiency and productivity in ethanol, and the production of hydrogen sulfide. Saccharomyces bayanus -Pasteur Champagne - Red Star showed better fermentation performance and it was evaluated in honey wort supplemented with different nutrients. The results showed that must supplementation with salts or commercial supplement Enovit contributes to the yeast performance. In the next step, it was studied the production of mead on a pilot scale at 18 ° C in polypropylene reactors containing 130 L of wort without supplementation (control), supplemented with Enovit and supplemented with apple pieces (10% m/v). By the end of fermentation, after 60 days, mead was properly characterized physico-chemically regarding the standards of quality and identity for mead, established by Brazilian legislation. The results showed that, with the exception of total acidity, all parameters were in accordance with the standards. After the fermentation, mead was subjected to aging process at 20 ° C by 216 days in oak barrel (50L) plastic gallon (20 L) and glass bottle (20L), and was characterized physico-chemically, as well as regarding the content of total phenolics, antioxidant capacity, presence of bioactive amines and aromatic compounds derived from the oak barrel. The results showed that the mead aged in plastic gallon presented content of \"reduced dried extract\" below the minimum patterns stated. It was also observed that the use of apples as the must supplement resulted in honey mead with a higher content of phenolic compounds and antioxidant capacity. The concentrations of bioactive amines found do not compromise the quality of mead, neither consumer health. The analysis of phenolic compounds in meads aged in oak barrel revealed that the aging process promoted the extraction of furfural, vanillin, gallic acid, compounds which contribute to the sensory profile of the beverage. Concentrations of these compounds were below the threshold of sensory perception, which can be changed by increasing aging time. Regarding purchase intent, all meads evaluated obtained more than 50% of positive scores, ranging between 3 (I doubt if buy) and 5 (definitely would buy), and meads aged in plastic and glass containers showed better acceptance. Sensory analysis showed little influence of the aging container on the attributes evaluated and the tasters preferred the meads aged in inert containers. So,it is concluded that the production of mead and its subsequent aging in inert containers or reconditioned oak casks, resulted in beverages that have good acceptance and are a viable alternative for beekeepers to diversify their products.

O objetivo do presente trabalho foi avaliar os efeitos de diferentes concentrações de quitosana nas dietas de novilhos Nelore canulados no rúmen sobre o consumo e digestibilidade aparente total da matéria seca e nutrientes, fermentação e síntese de proteína microbiana ruminal, concentrações de parâmetros sangüíneos, e os balanços de energia e de nitrogênio. Foram utilizados 8 novilhos canulados da raça Nelore. Os animais foram distribuídos aleatoriamente em 2 quadrados latinos 4 x 4 balanceados e contemporâneos, para receber as seguintes rações experimentais: 1) Controle (Q0), composta por ração sem a inclusão de quitosana; 2) Q50, com a inclusão de 50 mg/kg de peso corporal de quitosana; 3) Q100, com a inclusão de 100 mg/kg de peso corporal de quitosana; e 4) Q150, com a inclusão de 150 mg/kg de peso corporal de quitosana. Diariamente foram realizadas pesagens das quantidades dos volumosos e concentrados fornecidos e das sobras de cada animal, para estimativa do consumo. As amostras de sobras, silagem e fezes foram coletadas do 15° ao 18° dias de cada período experimental, armazenadas em sacos plásticos em freezer à 20°C, e posteriormente submetidas a análises químico-bromatológicas dos principais nutrientes. Na determinação da digestibilidade aparente total dos nutrientes a quantidade total de matéria seca fecal excretada foi estimada pela concentração de fibra em detergente ácido indigestível (FDAi). As amostras de líquido ruminal foram coletadas no último dia de cada período, sendo uma coleta realizada antes da alimentação (0 hora), e seis coletas com intervalos de 2 horas após a alimentação (2, 4, 6, 8, 10 e 12 horas). Foram determinados no líquido ruminal o pH, as concentrações de nitrogênio amoniacal e as concentrações dos ácidos graxos de cadeia curta. As amostras spot de urina foram obtidas de no 16º dia de cada período experimental, quatro horas após a alimentação matinal, durante micção espontânea. As amostras de sangue foram coletadas em tubos vacuolizados (vacutainer) por punção da veia jugular. Houve efeito quadrático com menores valores sobre o consumo de FDN, expressos em kg/dia e porcentagem de peso vivo (PV) para o tratamento Q150 (P<0,05). A inclusão de quitosana na dieta proporcionou aumento linear crescente (P<0,05) da digestibilidade da matéria seca (MS), matéria orgânica (MO), proteína bruta (PB), carboidratos totais (CT), FDN e nutrientes digestíveis totais (NDT). Houve efeito quadrático (P<0,05) sobre a concentração de N-NH3 com redução no tratamento Q150. A inclusão de quitosana na dieta não causou diferenças nas concentrações totais de AGCC (P>0,05), porém alterou as proporções molares de AGCC individualmente. Houve efeito linear crescente (P>0,05) das concentrações e porcentagens molares de propionato (mmol/L) à medida que se elevou as concentrações de quitosana na dieta. Houve diminuição (P>0,05) da relação acetato: propionato, principalmente para o tratamento Q150. Foi observado efeito linear decrescente para as proporções molares de acetato e butirato com a inclusão de quitosana. Houve efeito linear crescente sobre as concentrações plasmáticas de glicose com os tratamentos. As concentrações de quitosana utilizadas não influenciaram a síntese de proteína microbiana. O balanço de energia não foi influenciado pelos tratamentos. Não houve efeito dos tratamentos sobre o balanço de nitrogênio, porém ocorreu decréscimo na excreção do nitrogênio total (NT) nas fezes em porcentagem de NT. A quitosana quando utilizada como aditivo modulador da fermentação ruminal, resultou em alterações que possibilitam sua utilização como alternativa ao uso de ionóforos para bovinos. / The aim of this study was to evaluate the effect of different levels of chitosan on intake, digestibility, ruminal fermentation, ruminal microbial protein, balance of energy and nitrogen and blood parameters. Eight Nellore steers cannulated in the rumen were divided into two 4 x 4 balanced Latin squares. The daily doses of chitosan were 0, 50, 100 and 150 mg/kg BW respectively the treatments Q0 (Control), Q50, Q100 and Q150. The diets consisted of corn silage and concentrate in ratio 60:40 and the chitosan were inserted directly through the ruminal cannula, twice a day before feeding. Daily weights of the amounts of corn silage and concentrated supplied, and the orts refused of each animal, were recorded for estimate the nutrient intake. Samples of orts and feedstuffs were analyzed for composition and subsequent nutrient intake calculation. For determination of total apparent digestibility of nutrients, the total amount of fecal dry matter excreted was estimate by indigestible detergent acid fiber (ADFi). The feces were collected in the 15th until the 18th day of each experimental period, and frozen in freezer at -20°C. In the end of the collection period it was made composed sample by animal with base in the dry matter, and analyzed. Samples of ruminal fluid were collected at 0 (before feeding) and 2, 4, 6, 8, 10 and 12 hours after feeding. Spot urine samples were collected on day 16 of the experimental period. The estimation of microbial protein synthesis was performed by the method of total excretion of purine derivatives. Blood samples were collected by puncture of jugular vein. Intakes (kg/d) of DM, OM, CP, ether extract , total carbohydrates , non-fiber carbohydrates and TDN were not different, however, NDF intake decresead quadratically (P<0,05) when expressed as kg/d and percent of BW. Digestibility in the total digestive tract increased linearly to NDF, DM, OM as well as improved the digestibility of CP and TC, accordingly there was a positive effect on TDN with the inclusion of chitosan. All the ruminal fermentation parameters were influenced by the time after feeding. The NH3-N concentration decreased quadractically with Q150 treatment and there were no difference in total VFA concentration, however the individual VFA proportions were affected. Propionate concentration was higher with Q150 and similarly increasing linearly the proportion of propionate (P<0,05) which means an increase of 7.47% (Q0 vs. Q150). Chitosan decreased linearly the molar proportion of acetate and butyrate .Was observed linear and quadratic decrease effect on acetate: propionate ratio. The plasmatic metabolites and enzymes had no effect by the treatments, nevertheless the glucose concentration was markedly superior with the supplementation corresponding to an increase of 18.58%, 26.35%, 23.68% for Q0 versus Q50, Q100 e Q150 respectively. The energy and nitrogen balance had no effect with the treatments, however there was a decrease of fecal total nitrogen (%) excretion. Chitosan when used as modulator of ruminal fermentation resulted in changes that allow its use as an alternative to the use of ionophores for cattle without showing damage to the health of the animal.

No presente trabalho, primeiramente avaliou-se o desempenho fermentativo de 10 cepas de leveduras isoladas de frutas, de mel centrifugado e de resíduo do processamento de mel após a etapa de sedimentação (borra do mel), em tubo de ensaio contendo mosto constituído de Mel 15 °Brix suplementado, individualmente, com diferentes nutrientes, a saber: extrato de levedura comercial (10,0 g/L), farelo de arroz (20,0 g/L), mistura (1,0 g/L) constituída de farelos de milho, arroz e soja na proporção 5:2:5 e nutriente comercial (D&R) (0,15 g/L). Os parâmetros avaliados coexistiram da determinação do número de células em câmara de Neubauer e das concentrações de açúcares e etanol por HPLC, o que permitiu a determinação dos parâmetros rendimento, eficiência de fermentação e produtividade. Estes resultados sustentaram a seleção do mosto suplementado com extrato de levedura comercial e da melhor cepa de levedura que foram posteriormente avaliados em escala piloto. A cepa selecionada foi a levedura isolada a partir da borra de mel, a qual foi posteriormente identificada como Saccharomyces cerevisiae e nomeada Saccharomyces cerevisiae EEL 2009. A avaliação da produção de aguardente de mel em escala piloto foi realizada nas instalações do Alambique da Associação Rural de Canas, Canas-SP e o destilado obtido armazenado em tonel de carvalho de 200 L por 6 meses. Amostras foram coletadas a cada 30 dias para caracterização físico-química em conformidade com os parâmetros estabelecidos na Legislação Brasileira para aguardente de frutas. Em paralelo, as respectivas amostras foram devidamente avaliadas sensorialmente por 120 provadores não treinados, por meio de testes de aceitação, em escala hedônica, considerando os quesitos aparência, aroma, sabor, corpo e impressão global, além da atitude de compra. Estes resultados foram analisados estatisticamente por ANOVA e teste de Tukey, sendo os resultados de aceitação em relação à impressão global, analisados por meio da análise multivariada o que permitiu traçar o Mapa de Preferência Interno - MDPREF. Os resultados referentes à caracterização físico-química das respectivas amostras demonstraram que todas apresentaram os parâmetros de avaliação em conformidade com os padrões estabelecidos pela legislação. Os resultados da análise sensorial revelaram que o tempo de armazenamento de 180 dias em tonel de carvalho não foi suficiente para a ocorrência de reações desejadas, o que influenciaria nas características sensoriais da bebida, tornado-a mais agradável e suave. No entanto, apesar do pouco tempo de armazenamento, a aguardente de mel apresentou uma boa aceitação por parte dos provadores, cuja maioria manifestou sua aceitação em termos de \"gostei ligeiramente\" e \"não gostei, nem desgostei\". No tocante ao MDPREF, este revelou que a amostra referente a 180 dias de armazenamento foi preferida por um maior número de provadores. Em relação à atitude de compra, as amostras armazenadas por 60 (79,17%) e 180 dias (75,83%) apresentaram os melhores resultados. Diante do exposto, conclui-se que o mel se apresenta como uma alternativa viável para a formulação de mosto para produção de aguardente no período de entre safra de cana-de-açúcar, contribuindo para melhor aproveitamento das instalações e como fonte alternativa de renda para o produtor rural. / In this work, first it was evaluated the fermentation performance of 10 yeast strains isolated from fruits, centrifuged honey and residue of honey processing after the sedimentation (sludge honey) step. This experiment was carried out in test tube containing wort made from honey 15 °Brix supplemented individually with different nutrients, namely: commercial yeast extract (10.0 g/L), rice bran (20.0 g/L), mixture (1.0 g/L) consisting of bran corn, rice and soy in the ratio 5:2:5 and commercial nutrient (D&R) (0.15 g/L). The parameters evaluated included the cell number determination in Neubauer chamber, and sugars and ethanol concentrations by HPLC, which allowed to determine the fermentation parameters as yield, fermentation efficiency and productivity. These results supported the selection of the wort supplemented with commercial yeast extract and the best yeast strain, which were subsequently evaluated in a pilot scale. The selected strain was the yeast isolated from the sludge honey, which was later identified as Saccharomyces cerevisiae and named Saccharomyces cerevisiae EEL 2009. The production evaluation of honey spirit on a pilot scale was conducted at the Canas Rural Association Pilot Plant for cachaça production - Canas-SP and distillate was stored in oak barrel of 200 liters per 6 months. Samples were collected every 30 days for physic-chemical characterization in accordance with the guidelines established in the Brazilian Legislation for fruit spirit. Beyond that, the respective samples were properly sensory evaluated by 120 untrained consumers regarding to acceptance testing employing a hedonic scale, considering characteristics as appearance, aroma, flavor, and overall impression, and the attitude of purchase, as well. These results were statistically analyzes by ANOVA and Tukey`s test and the results of acceptance regarding to the overall impression, were analyzed by multivariate analysis which allowed tracing the Internal Preference Map - MDPREF. The results concerning the physic-chemical characterization of the respective samples showed that all presented the evaluation parameters in accordance with standards established by the legislation. The results of sensory analysis revealed that the storage time of 180 days in oak barrel was not enough for the occurrence of desired reactions, which influence the sensory characteristics of the beverage, making it nicer and smoother. However, despite short time storage, honey spirit showed good acceptance by the consumers. Most of them expressed acceptance in terms of \"liked slightly\" and \"not liked nor disliked\". Regarding the MDPREF results, these proved that the sample relating to 180 days of storage was preferred by majority of consumers. Regarding the attitude of purchase, the samples stored for 60 days (79.17%) and 180 days (75.83%) showed the best results. Therefore, we conclude that honey can be considered as a viable alternative for wort formulation for the production of spirit, mainly in the period between harvests of sugar cane, contributing to better utilization of the facilities and as alternative source of income for farmers.

Thesis (PhD. (Microbiology)) --University of Limpopo, 2013 / The current study investigated four fungal species namely Aspergillus niger FGSC A733, Aspergillus versicolor EF23, Penicillium citrinum AZ01 and Trichoderma harzianum NCGR 0509 for their abilities to produce cellulases and xylanases in submerged and solid state fermentations. Five different substrates (carboxymethyl cellulose, xylan, common thatch grass, wheat bran and Jatropha curcas seed cake) were examined for their potential use as low cost feedstock for fermentation by the fungal species. Aspergillus niger FGSC A733 produced the highest titres of cellulase and xylanase in solid state fermentations using wheat bran as a substrate. However, because of the need to lower the cost of enzyme production, Jatropha seed cake a relatively underutilised oilseed cake was used. Supplementation of the Jatropha seedcake with 10% common thatch grass (Hyperrhenia sp) resulted in a fivefold increase in the levels of xylanase produced. Cellulase production was not affected by this supplementation. Addition of ammonium chloride increased production of xylanase while cellulase production was not affected nitrogen supplementation. Maximum xylanase was produced on Jatropha seed cake at 25 °C after 96 hours while cellulase was maximally produced at 40 °C after 96 hours of solid state fermentations. Peak production of xylanase was obtained at an initial pH of 3 whilst cellulase was maximally produced at an initial pH of 5. The crude xylanase was most active at pH 5 and cellulase at pH 4. The optimum temperature for cellulase activity was 65 °C and that of xylanase was 50 °C. Under optimized conditions, 6087 U/g and 3974 U/g of xylanase and cellulase per gram of substrate used were obtained respectively. The diversity of cellulases was investigated so as to determine the most appropriate enzyme mixture for saccharification of the common thatch grass. Proteins from the four species under investigation were partially purified by affinity chromatography on swollen Avicel. The proteins were analysed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis SDS-PAGE and zymography. Potential cellulase bands from SDS-PAGE were sequenced by mass spectrometry. The basic logical alignment tool (BLAST) and Clustal W were used for matching and identifying the sequences with closely related ones in the databases. The identified proteins from Penicillium citrinum AZ01 and Aspergillus versicolor EF23 were found to closely resemble a catalytic domain of cellobiohydrolase from Trichoderma sp. The three proteins obtained from Aspergillus niger showed resemblance to 1,4-beta glucan cellobiohydrolase A precursor from Aspergillus niger FGSC A733 was also found to have cellobiase and endoglucanase activity was determined using cellobiase and carboxymethyl cellulose as substrates. Cellulase and xylanase zymograms of proteins from A. niger FGSC A733 demonstrated six active bands ranging from 20 kDa to 43 kDa for cellulase and a 31 kDa active band for xylanase. The cellulase produced by Aspergillus niger FGSC A733 on Jatropha seed cake under optimised conditions was used for saccharification of 2% (w/v) common thatch grass (CTG) in combination with Celluclast™. Celluclast™ and Aspergillus niger cellulase were mixed at different ratios and the amount of glucose produced over time was monitored using high performance liquid chromatography (HPLC). A ratio of 2 volumes Celluclast™ to one volume Aspergillus niger cellulase was chosen for the saccharification process. The main enzymes in the mixture were identified using peptide mass fingerprinting as endoglucanases from the Celluclast™ and cellobiase from the Aspergillus niger cellulase. Concentration of the Celluclast™ tenfold times (164 FPU) improved the yield of glucose by 42.8 and 37.8% in acid and alkali pre-treated CTG, respectively. Concentrating Aspergillus niger cellulase (13.2 FPU) decreased the production of glucose by 4.8% in acid pre-treated CTG while in alkali pre-treated CTG, a 5% increase in glucose production was observed. Increasing the substrate loading of acid pre-treated CTG from 2% to 10% (w/v) resulted in a two and a half times increase in glucose production while an increase of 1.5 g/l glucose was obtained from 7% (w/v) alkali pre-treated CTG. Addition of xylanases from Aspergillus niger to the Celluclast™-Aspergillus niger cellulase mixture decreased glucose production by 16.3% on acid pre-treated CTG while there was an increase of 18.3% glucose in alkali pre-treated CTG. Addition of enzyme preparations from Aspergillus versicolor EF23, Penicillium citrium AZ01 and Trichoderma harzianum NCGR 0509 to the Celluclast™-Aspergillus niger cellulase mixture resulted in lower glucose production both in acid and alkali pre-treated CTG. Addition of Pentopan™ improved glucose production by 8 and 25% on 10% acid and 7.5% alkali loading of pre-treated CTG respectively. The optimal conditions for the production of the glucose rich hydrolysate in 10% (w/v) acid and 7% (w/v) alkali pre-treated CTG was found to be the use of Celluclast™-Aspergillus niger cellulase-Pentopan™ mixture (164 FPU Celluclast™ and 13 FPU Aspergillus niger cellulase, 7178 IU) Pentopan™ at 50 °C for 32 hours. The fermentability of the glucose in glucose-rich CTG hydrolysates to ethanol using Saccharomyces cerevisae WBSA 1386 and Candida shehatae CSIR Y-0492 was investigated. The highest yield of ethanol produced by S. cerevisae WBSA 1386 was 9.8 g/l in the alkali pre-treated CTG hydrolysate and 8.7 g/l in acid pre-treated CTG. C. shehatae CSIR Y-0492 produced 9 g/l of ethanol in alkali pre-treated CTG within 48 hours while acid pre-treated CTG hydrolysate produced 8.8 g/l of ethanol within 24 hours of the fermentation process. Addition of the nutrient supplement boosted the ethanol yield in the acid pre-treated hydrolysates. The consumption of glucose during fermentation by S. cerevisae WBSA 1386 and C. shehatae CSIR Y-0492 on average was 97%. The C. shehatae CSIR Y-0492 was expected to produce much higher ethanol yield than the Saccharomyces because of its ability to utilize xylose for ethanol production. This however was not observed in this investigation. The conclusion of this study is that it is possible to produce bioethanol from Hyperrhenia spp. (CTG) using a combination of fungal enzymes for the production of fermentable sugars.

Two methods of single-stage fermentation were used to produce methane from whey. The first method involved batch fermentation in which the pH was automatically controlled at 7.0. In addition to pH control, the second method was characterized by introducing the substrate continuously into the fermenter along with dilution water to keep the organic acids at a non-toxic level. The first method did not improve the production of methane compared to the batch fermentation in which a pH control system was not used (41). However, the second method showed that the concentration of organic acids has a major effect on limiting the production of methane from whey, and that by diluting the substrate with water the concentration of these acids was kept at levels not toxic to methanogenic bacteria until the population of methanogenic bacteria was such that they could convert organic acids to methane as rapidly as the acids were produced by non-methanogenic bacteria in a continuous process. This continuous process was capable of producing .179 m3 methane per kilogram of lactose in cheese whey.

In the beef cattle industry, sustainable beef production is a primary focus, as it has direct effects on environmental stewardship, farm profitability, and public concerns. Research has been and is continually being conducted to evaluate alternative forages such as Brown Midrib Corn Silage (BMRCS) as a major component in growing beef cattle diets, to improve animal performance, ruminal fermentation, and economic returns. The objective of this study was to determine growth performance, ruminal fermentation characteristics, and economic returns of growing beef steers when fed a brown midrib corn silage-based TMR (BMRT) compared with a conventional corn silage-based TMR (CCST). This growing beef study was performed in a completely randomized design with 24 Angus crossbred steers (initial body weight (BW) = 258 ± 23.2 kg) to test 2 treatments: CCST vs. BMRT. All animals were placed in individual pens, and 12 animals allocated to each treatment (n = 12). All steers were adapted to the CCST for a 2-wk period prior to start of the trial. The CCST contained 48.1% CCS whereas the BMRT consisted of 49.0% BMRCS on a dry matter (DM) basis. All steers were fed once per day, and feed bunks assesed each afternoon and prior to morning feeding, which was used to determine the amount of feed to deliver to each pen the following day. The experiment lasted 84 d. For all steers, BW and ruminal fermentation characteristics were measured on wk 4, 8, and 12. Intake of DM averaged 9.54 kg/d across the treatments and was similar between the treatments. Steers fed the BMRT tended to increase average daily gain (ADG) compared to those fed the CCST (1.54 vs. 1.42 kg/d; P = 0.09). In addition, feeding the BMRT tended to increase G:F compared with the CCST (0.165 vs. 0.146; P = 0.07). Feeding the BMRT decreased ruminal pH (6.42 vs. 6.67; P < 0.01), whereas it increased total VFA concentration (P = 0.01) compared with the CCST. Feeding the BMRT decreased molar proportion of acetate (P < 0.01), but increased propionate proportion (P = 0.01), resulting in decreased acetate-to-propionate ratio compared with the CCST (P < 0.02). Steers fed BMRT increased feed margin (P = 0.05) and net return (P = 0.02) compared to those fed CCST throughout the trial. Overall data in this study indicate that feeding the BMRT to growing beef steers enhanced ruminal fermentation and beneficially shifted VFA profiles, which contributed to improved growth performance and economic performance of steers fed the BMRT.

Mathematical modelling, simulation, and optimisation are core methodologies for future developments in engineering, natural, and life sciences. This work aims at applying these mathematical techniques in the field of biological processes with a focus on the wine fermentation process that is chosen as a representative model. In the literature, basic models for the wine fermentation process consist of a system of ordinary differential equations. They model the evolution of the yeast population number as well as the concentrations of assimilable nitrogen, sugar, and ethanol. In this thesis, the concentration of molecular oxygen is also included in order to model the change of the metabolism of the yeast from an aerobic to an anaerobic one. Further, a more sophisticated toxicity function is used. It provides simulation results that match experimental measurements better than a linear toxicity model. Moreover, a further equation for the temperature plays a crucial role in this work as it opens a way to influence the fermentation process in a desired way by changing the temperature of the system via a cooling mechanism. From the view of the wine industry, it is necessary to cope with large scale fermentation vessels, where spatial inhomogeneities of concentrations and temperature are likely to arise. Therefore, a system of reaction-diffusion equations is formulated in this work, which acts as an approximation for a model including computationally very expensive fluid dynamics. In addition to the modelling issues, an optimal control problem for the proposed reaction-diffusion fermentation model with temperature boundary control is presented and analysed. Variational methods are used to prove the existence of unique weak solutions to this non-linear problem. In this framework, it is possible to exploit the Hilbert space structure of state and control spaces to prove the existence of optimal controls. Additionally, first-order necessary optimality conditions are presented. They characterise controls that minimise an objective functional with the purpose to minimise the final sugar concentration. A numerical experiment shows that the final concentration of sugar can be reduced by a suitably chosen temperature control. The second part of this thesis deals with the identification of an unknown function that participates in a dynamical model. For models with ordinary differential equations, where parts of the dynamic cannot be deduced due to the complexity of the underlying phenomena, a minimisation problem is formulated. By minimising the deviations of simulation results and measurements the best possible function from a trial function space is found. The analysis of this function identification problem covers the proof of the differentiability of the function–to–state operator, the existence of minimisers, and the sensitivity analysis by means of the data–to–function mapping. Moreover, the presented function identification method is extended to stochastic differential equations. Here, the objective functional consists of the difference of measured values and the statistical expected value of the stochastic process solving the stochastic differential equation. Using a Fokker-Planck equation that governs the probability density function of the process, the probabilistic problem of simulating a stochastic process is cast to a deterministic partial differential equation. Proofs of unique solvability of the forward equation, the existence of minimisers, and first-order necessary optimality conditions are presented. The application of the function identification framework to the wine fermentation model aims at finding the shape of the toxicity function and is carried out for the deterministic as well as the stochastic case. / Mathematische Modellierung, Simulation und Optimierung sind wichtige Methoden für künftige Entwicklungen in Ingenieurs-, Natur- und Biowissenschaften. Ziel der vorliegende Arbeit ist es diese mathematische Methoden im Bereich von biologischen Prozessen anzuwenden. Dabei wurde die Weingärung als repräsentatives Modell ausgewählt. Erste Modelle der Weingärung, die man in der Literatur findet, bestehen aus gewöhnlichen Differentialgleichungen. Diese modellieren den Verlauf der Populationszahlen der Hefe, sowie die Konzentrationen von verwertbarem Stickstoff, Zucker und Ethanol. In dieser Arbeit wird auch die Konzentration von molekularem Sauerstoff betrachtet um den Wandel des Stoffwechsels der Hefe von aerob zu anaerob zu erfassen. Weiterhin wird eine ausgefeiltere Toxizitätsfunktion benutzt. Diese führt zu Simulationsergebnissen, die im Vergleich zu einem linearen Toxizitätsmodell experimentelle Messungen besser reproduzieren können. Außerdem spielt eine weitere Gleichung für die zeitliche Entwicklung der Temperatur eine wichtige Rolle in dieser Arbeit. Diese eröffnet die Möglichkeit den Gärprozess in einer gewünschten Weise zu beeinflussen, indem man die Temperatur durch einen Kühlmechanismus verändert. Für industrielle Anwendungen muss man sich mit großen Fermentationsgefäßen befassen, in denen räumliche Abweichungen der Konzentrationen und der Temperatur sehr wahrscheinlich sind. Daher ist in dieser Arbeit ein System von Reaktion-Diffusions Gleichungen formuliert, welches eine Approximation an ein Modell mit rechenaufwändiger Strömungsmechanik darstellt. Neben der Modellierung wird in dieser Arbeit ein Optimalsteuerungsproblem für das vorgestellte Gärmodell mit Reaktions-Diffusions Gleichungen und Randkontrolle der Temperatur gezeigt und analysiert. Variationelle Methoden werden benutzt, um die Existenz von eindeutigen schwachen Lösungen von diesem nicht-linearen Modell zu beweisen. Das Ausnutzen der Hilbertraumstruktur von Zustands- und Kontrolraum macht es möglich die Existenz von Optimalsteuerungen zu beweisen. Zusätzlich werden notwendige Optimalitätsbedingungen erster Ordnung vorgestellt. Diese charakterisieren Kontrollen, die das Zielfunktional minimieren. Ein numerisches Experiment zeigt, dass die finale Konzentration des Zuckers durch eine passend ausgewählte Steuerung reduziert werden kann. Der zweite Teil dieser Arbeit beschäftigt sich mit der Identifizierung einer unbekannten Funktion eines dynamischen Modells. Es wird ein Minimierungsproblem für Modelle mit gewöhnlichen Differentialgleichungen, bei denen ein Teil der Dynamik aufgrund der Komplexität der zugrundeliegenden Phänomene nicht hergeleitet werden kann, formuliert. Die bestmögliche Funktion aus einem Testfunktionenraum wird dadurch ausgewählt, dass Abweichungen von Simulationsergebnissen und Messungen minimiert werden. Die Analyse dieses Problems der Funktionenidentifikation beinhaltet den Beweis der Differenzierbarkeit des Funktion–zu–Zustand Operators, die Existenz von Minimierern und die Sensitivitätsanalyse mit Hilfe der Messung–zu–Funktion Abbildung. Weiterhin wird diese Funktionenidentifikationsmethode für stochastische Differentialgleichungen erweitert. Dabei besteht das Zielfunktional aus dem Abstand von Messwerten und dem Erwartungswert des stochastischen Prozesses, der die stochastische Differentialgleichung löst. In dem man die Fokker-Planck Gleichung benutzt wird das wahrscheinlichkeitstheoretische Problem einen stochastischen Prozess zu simulieren in eine deterministische partielle Differentialgleichung überführt. Es werden Beweise für die eindeutige Lösbarkeit der Vorwärtsgleichung, die Existenz von Minimierern und die notwendigen Bedingungen erster Ordnung geführt. Die Anwendung der Funktionenidentifikation auf die Weingärung zielt darauf ab die Form der Toxizitätsfunktion herauszufinden und wird sowohl für den deterministischen als auch für den stochastischen Fall durchgeführt.

Yeast are exposed to many physical and chemical stresses when used in large-scale industrial fermentations, particularly the initial stages in which yeast are shifted from anaerobic storage to aerated wort. This work investigated the transcriptional and physiological responses of yeast that had been shifted from anaerobic to aerobic growth conditions. Microarray technology was employed to determine the transcriptional changes that occurred in the first hour of a pilot-plant fermentation compared to the 23rd hour. It was found that over 100 genes were up-regulated initially including genes involved in the synthesis of the essential membrane sterol ergosterol and genes for the protection of cells against oxidative stress. It was also determined that cells which accumulate ergosterol precursors in the absence of ergosterol were more sensitive to exogenous oxidative stresses, indicating a role for ergosterol in oxidative stress tolerance. Aeration of anaerobically grown cells did not affect their growth kinetics or viability. However, anaerobically grown cells were hypersensitive to exogenous oxidative stress compared to their aerobic counterparts. Anaerobic cells that underwent a short period of aeration prior to treatment with hydrogen peroxide generated a tolerance to the oxidant, indicating that the period of aeration produced an adaptive-like response. Microarray analysis of the cells during the period of aeration showed that representative genes from the oxidative stress response family were up-regulated rapidly and it was determined that the response was controlled by the Yap1p and Skn7p transcription factors. Deletion of the transcription factor genes indicated that they were responsible for the creation of tolerance to oxidant. Target gene products of the two transcription factors (Gpx2p, Gsh1p and Trx2p) were shown to be induced during the shift to aeration; however, the glutathione redox balance did not seem to be affected as the cells were shifted from highly reduced to oxidising environments. Unexpectedly, it was discovered that genes involved in the synthesis of amino acids were up-regulated during anaerobic growth and stringently downregulated upon aeration of cells. The transcriptional activator of those genes (Gcn4p) was essential for growth in anaerobic media which included amino acid supplementation.

Includes bibliographical references (leaves [259]-290) Investigates nitrogen assimilation and metabolism in rumen bacteria with the object of understanding the basic process and their controls.