Le vin est un milieu peu propice à la croissance de la levure mais il est néanmoins possible de la faire croître sur base de vin enrichit en nutriments et dilué pour diminuer la concentration en éthanol. En vue de l’élaboration des vins effervescents par une seconde fermentation, produire la levure Saccharomyces cerevisiae dans ces conditions est indispensable pour l’acclimater mais il s’agit d’un enjeu complexe qui doit prendre en compte de nombreux paramètres physico-chimiques mais aussi économiques. En effet, les paramètres opératoires peuvent induire des conditions de croissance pouvant affecter le développement de la levure. Seule la levure S. cerevisiae (Fizz+) a été utilisée car elle est spécialement sélectionnée pour cette seconde fermentation en vase clos. Le principal enjeu était donc d’obtenir une bonne adaptation de la levure à croître dans un milieu hydro-alcoolique, conditions contraignantes pour elle, mais aussi d’obtenir une production maximale.Nous avons tout d’abord étudié en fioles Erlenmeyer (250 mL) l’influence de divers paramètres : conditions physico-chimiques, concentrations en nutriments, concentration minimale en levure sèche active nécessaire à une bonne activité ainsi que son temps de réhydratation.Dans un deuxième temps, nous avons effectué des propagations en mode batch dans un bioréacteur (5 L) pour valider les conclusions réalisées à la suite de l’étude en Erlenmeyer et ainsi étudier l’influence de différentes aérations sur la production de S. cerevisiae. Les données obtenues ont servi de base pour comparer les améliorations apportées par le procédé développé en mode fed-batch. Les concentrations en levures obtenues suite à l’optimisation des conditions du milieu de culture en cinq litres sont supérieures d’un facteur cinq à celles obtenues dans la pratique en cave.Ensuite l’étude s’est concentrée sur le développement d’un nouveau procédé d’alimentation en nutriments pour cultiver S. cerevisiae en métabolisme respiratoire dans des cuves réalisées par la société partenaire du projet, OEno Concept. La nouveauté réside dans la façon de réguler la température de la culture qui se fait simultanément à l’apport des nutriments suite au dégagement de chaleur lors de la croissance de S. cerevisiae. Un brevet a été déposé sur cette technologie. Ce nouveau procédé a permis une augmentation de la productivité cellulaire, d’un facteur supérieur à quatre, car il a permis aux levures de s’adapter à cet environnement stressant et a favorisé l’oxydation du glucose au détriment de la fermentation. / Wine is an aggressive/stressful growth medium; it is depleted of micronutrients, rich in ethanol and very poor in assimilable nitrogen. Despite all these difficulties, it is possible to grow yeast in a medium largely based on wine by diluting the ethanol concentration and enriching the medium with micronutrients, a carbon source and assimilable nitrogen. It is, desirable to propagate Saccharomyces cerevisiae in such environment in order to produce a culture of yeast adapted to a second fermentation of alcoholic beverages. Production of microorganism in wine growing environment, is a complex issue that must take into account many, physicochemical and economic parameters. Indeed, the operating parameters can affect the development of yeast in a bioreactor. Therefore, it is important to know the most influential parameters on growth. The strain S. cerevisiae (Fizz+), a commercial strain that has been selected for the second fermentation in bottles, was used during this project. The propagation process served to increase the amount of yeast as well as to adapt the yeast to grow in an alcoholic environment. We first studied in shake-flasks cultures various physicochemical conditions such as nutrients concentration, the rehydration time and the minimum concentration of active dry yeast necessary for good yeast activity.In a second step, we performed batch fermentations in bioreactors (5 L) to confirm the conclusions from the shake-flask cultures and additionally to study the influence of aeration on S. cerevisiae production. The data obtained served as a basis for performing fed-batch cultures. The yeast concentrations obtained as a result of the optimization of the conditions of the culture medium in five liters were five times greater than those obtained in actual industrial production processes. The next step was to develop an automated fed-batch culture to grow S. cerevisiae respiratively in partnership with the industrial partner of the project, OEno Concept. The novelty of the process is the way in which the growth medium feed-rate is linked to the heat produced by the growing S. cerevisiae.This research has allowed an increase in cell productivity, by a factor greater than four, thanks to the novel process in stressful growth environment promoting respiration with regard to fermentation.

No presente trabalho foram estudados sistemas pilotos de tratamento integrado de lodos ativados com sistemas de separação por membranas de ultrafiltração visando o reúso de água. O esgoto bruto utilizado foi originado do Conjunto Residencial associado ao esgoto do restaurante universitário da Universidade de São Paulo. Este esgoto possui características físico-químicas e biológicas similares ao esgoto doméstico. O esgoto utilizado nos sistemas de tratamento foi submetido à pré-tratamento: gradeamento e caixa de areia. Foram montados dois sistemas pilotos: I) sistema piloto recebendo esgoto doméstico primário com sistema de tratamento composto por reator biológico de lodos ativados (500 L de volume) e sistema de separação por membranas de ultrafiltração tipo tubular (1,4 m² de área superficial) externa ao tanque de aeração. Este sistema apresentou como resultados principais: taxas médias de produção de permeado de 22,9 ±2,7 L.h-¹.m-² e 17 ±2,7 L.h-¹.m-².bar-¹; valores característicos do permeado em relação à variável turbidez média de 0,3±0,1 UNT, cor aparente média de 31,2±4,6 mg de PtCo.L-¹, sólidos dissolvidos totais de 201±47mg.L-¹ e sólidos suspensos totais não detectável. O sistema de lodos ativados operou em regime de aeração prolongada. II) sistema piloto recebendo esgoto doméstico, após tratamento anaeróbio em reator anaeróbio de fluxo ascendente com manta de lodo (UASB), composto por reator biológico de lodos ativados (1.500 L) e sistema de separação por membrana de ultrafiltração tipo espiral (14,4m² de área superficial) interna ao tanque de aeração. Este sistema apresentou como resultados principais: taxas médias de produção de permeado de 16,1 ± 4,1 L.h-¹.m-²; valores característicos do permeado em relação à variável turbidez média 0,2±0,1 UNT, cor aparente média de 25±5 mgPtCo.L-¹ e sólidos suspensos totais não detectável. Em ambos os sistemas as eficiências de remoção de nitrogênio e fósforo podem ser consideradas pequenas. Desta forma, foram adicionados sulfato de alumínio e cloreto férrico no tanque de aeração do sistema piloto II. As dosagens variaram de 40 a 80 mg.L-¹. Os melhores resultados de eficiência de remoção de fósforo solúvel foram obtidos com o sulfato de alumínio (em torno de 79%) na dosagem de 80 mg.L-¹. / A pilot plant integrating an extended aeration activated sludge unit and an ultrafiltration membrane system was constructed and operated aiming at the production of an effluent to be reused in industrial activities. Raw wastewater was collected from a student residential building and from one of the University of São Paulo\'s restaurants. The wastewater characteristics have shown to be very close to conventional domestic wastewaters. This wastewater was submitted to preliminary treatment by screening and subsequent grit removal. Two pilots systems were studied. The first one, treating the preliminary treated wastewater by the activated sludge unit followed by the ultrafiltration membrane system of the tubular type (1,4 m² of surface area). In this case, the membrane was located external to the aeration tank. This system has shown the following main results: average rates of percolate production of 22,9 ±2,7 L.h-¹.m-².bar-¹ and 17 ±2,7 L.h-¹.m-².bar-¹; average characteristic values of the percolate as: turbidity of 0,3±0,1 UNT, apparent Color of 31,2±4,6 mg of PtCo.L-¹, total dissolved solids of 201±47mg.L-¹ and total suspended solids not detectable. The second pilot unit received the effluent from an Upflow Anaerobic Sludge Blanket (UASB) reactor and was composed by the extended aeration activated sludge reactor and an ultrafiltration membrane system of the spiral type (14,4m² of surface area), internal to the aeration tank of the activated sludge unit. This system has shown the following main results: average rates of percolate production of 16,1 ± 4.1 L.h-¹.m-²; average characteristic values of the percolate as: turbidity of 0,2±0,1 UNT, apparent color of 25±5 mgPtCo.L-¹ and total suspended solids not detectable. In both systems, the efficiencies of Nitrogen and Phosphorus removal can be considered as small. To improve the efficiencies of soluble phosphorus removal it has been added aluminum sulphate and ferric chloride to the aeration tank of the second pilot unit. The dosages had varied from 40 to 80 mg.L-¹. The best soluble phosphorus removal (about 79%) was achieved with the dosage of 80 mg.L-¹ of aluminum sulphate.

Foram estudados a composição e o pH do meio de cultivo para a produção do fragmento de anticorpo (scFv) anti-LDL(-), expresso em Pichia pastoris recombinante. Os experimentos que definiram a composição e pH do meio assim como a concentração inicial de células na fase de indução foram realizados em agitador orbital a 250 rpm, com temperatura de 30 ºC na fase de crescimento e 20 ºC na fase de indução, durante 72 horas, com adição diária de 1% (v/v) de metanol. Para modificação do meio foi realizado um planejamento experimental empregando como variáveis independentes: extrato de soja, casaminoácidos e ureia, os quais substituíram o YNB e a biotina presentes no meio padrão (BMMY). Apesar de haver maior produção no meio com extrato de soja, o meio contendo 10 g.L-1 de casaminoácidos foi selecionado, uma vez que este favoreceu a etapa de purificação. A partir da faixa de pH estudada entre 3,0 e 8,0, determinou-se que o pH 8,0 no início da fase de indução favorece a maior produção. Finalmente, o meio BMMY-CA (pH 8,0) foi utilizado para cultivo em biorreator com volume de trabalho de 10L e a partir deste cultivo foram calculados os parâmetro cinéticos (velocidades de crescimento, de consumo de substrato e de produção, bem como produtividade). Diante do conjunto de experimentos realizados, foi possível otimizar a composição do meio de cultivo e as condições operacionais, que possibilitaram um aumento do rendimento bem como aumento do volume de produção do scFv anti-LDL(-) em biorreator. / Composition and pH culture medium for the production of anti- LDL(-) antibody fragment (scFv) were studied in recombinant yeast Pichia pastoris. The experiments that defined medium composition, pH and the initial cell concentration in the induction phase were carried out in baffled shaker flasks at 250 rpm, with temperature at 30°C for growth phase and 20°C for induction phase, during 72 hours with daily addition of 1% (v/v) methanol. A design of experiments employing for medium modification with independent variables: soy extract, casamino acids and urea, which replaced the YNB and biotin present in the standard medium (BMMY) was conducted for medium formulation. Even though, there was an increase in medium production with soy extract, the medium containing 10 g.L-1 casamino acids was selected since it favors the purification step. Through a pH range study (3.0 to 8.0), it was determined that pH 8.0 during induction phase offered higher levels. Then, the best results from shaker flask cultivation (BMMY-CA with casamino acids and pH 8.0) were carried out in 1L bioreactor, showing a biomass and a scFv production increase compared to the standard, BMMY (pH 6.0). Finally, the medium BMMY-CA (pH 8,0) was submitted to a volume scale-up into a 10L bioreactor, which was also used to evaluate kinetic parameters (rates of growth, substrate consumption and production as well as productivity). In conclusion, by optimizing culture medium and operating conditions it was possible to increase yield and scale-up the production of scFv anti-LDL(-) in bioreactor.

CAPES / Glutathione (GSH) is a tripeptide, soluble in water; it consists of three amino acids: L-glutamate, L-cysteine and glycine. It is the most efficient intracellular antioxidant present in the all biologics systems (animals and vegetables). Currently, it is reported many applications of GSH in different segments, like in food industries as food additive; in the sports segment, in the pharmaceutical field; it is used as food supplement; and is also used in the treatment of industrial effluents due to ability to absorb heavy metals. The application in magnetic fields (MF) inside the fermentative process can cause stimulants or inhibitory effects. However, it is not that reported in the literature. Thus, the main objective of this work is to study the influence of magnetic fields in the production of glutathione in reactor ?air-lift?? pressurized and the microorganism used was Saccharomyces cerevisiae ATCC 7754 During the fermentations were analyzed the pH, concentrations of cell (g/L), glucose (g/L) and GSH (mg/L). Firstly, it was evaluated at different pressures (0; 0.25; 0.50; 0.75; 1.00; 1.25 e 1.50 kgf/cm2). In this context, conditions fermentation were 20?C, total time of 96 h e pHinitial 5.0. The results showed that the best two pressures were 1.50 kgf/cm2, concentration of cell 17.25 g/L and 141.30 mg/L of GSH (72 h) and 0.50 kgf/cm2, with 13.51 g/L concentration of cell and 178.21 mg/L of GSH (96 h). Secondly, was realized experimental factorial design 22 with the two best results of pressures obtained in the first step. In the planning seven tests were done including three central points, in which the variables investigated were: pressure and velocity of cell recycling. The percent of variation of 90% (p < 0.1) was used to statistical analysis of results of concentration of cell and GSH. Therefore, according to the results of the second step, the highest concentration of cell (7.26 g/L) was obtained with 1.5 kgf/cm2 and recycle of 15 cm/s at 72 h of fermentation. In relation to production of GSH, the highest result was 67.24 mg/L at 0.50 kgf/cm2 and recycle of 3.0 m/s with 67.24 mg/L at 94 h. There was no significant response in both responses to the variables (concentration of cell and GSH), according to the table of estimated effects. The production of GSH by S. cerevisiae ATCC 7754 on a pressurized system presented as an efficient and promising technology. However the influence of MF and the velocity of cell recycling led to mechanisms that promoted particular synthesis inhibition leading to a decrease in its final concentration. / A glutationa (GSH) ? um tripept?deo hidrossol?vel composto por tr?s amino?cidos: ?cido glut?mico, ciste?na e glicina. ? o mais eficiente antioxidante intracelular presente em todos os sistemas biol?gicos (animais e vegetais). Atualmente, relatam-se in?meras aplica??es da GSH nas ind?strias em diferentes segmentos, sendo eles: nas ind?strias de alimentos como aditivo alimentar; no campo farmac?utico; no segmento desportivo como suplemento alimentar e ainda no tratamento de efluentes industriais, pois tem a capacidade de adsorver metais pesados. A aplica??o de campos magn?ticos (CM) em processos fermentativos pode causar efeitos estimulantes ou inibit?rios, or?m ainda ? pouco reportada na literatura. Desta forma o principal objetivo deste trabalho foi estudar a influ?ncia de CM na produ??o de glutationa em reator ?air-lift? pressurizado, o micro-organismo utilizado foi Saccharomyces cerevisiae ATCC 7754. Durante as fermenta??es foi feito o acompanhamento do pH, concentra??o celular (g/L), GSH (mg/L) e glicose (g/L). Na primeira etapa foi estudada apenas a varia??o da press?o no reator (0,00; 0,25; 0,50; 0,75; 1,00; 1,25 e 1,50 kgf/cm2). As condi??es das fermenta??es foram: 20?C, tempo total 96 h e pHinicial 5,0. De acordo com os resultados desta etapa, as duas melhores press?es em estudo foram 1,50 kgf/cm2, apresentando 17,25 g/L de concentra??o celular e 141,30 mg/L de GSH (72 h) e 0,50 kgf/cm2, com 13,51 g/L de concentra??o celular e 178,21 mg/L de GSH (96 h). Na segunda etapa, na qual se aplicou os CM, foi realizado um delineamento fatorial 22, onde foram realizados 7 ensaios, incluindo 3 pontos centrais, nos quais as vari?veis independentes testadas foram: press?o (kgf/cm2) e velocidade de reciclo celular (cm/s). O n?vel de signific?ncia de 90% (p < 0,1) foi utilizado na an?lise estat?stica dos dados de concentra??o celular e de GSH. De acordo com os resultados desta etapa, o maior resultado quanto ao crescimento celular (7,26 g/L) foi obtido a 1,50 kgf/cm2 e reciclo de 15 cm/s ap?s 72 h de fermenta??o. Em rela??o a concentra??o de GSH, o maior resultado foi a 0,50 kgf/cm2 e reciclo de 3,0 m/s com 67,24 mg/L a 94 h. A an?lise estat?stica mostrou que n?o houve vari?veis significativas em ambas as respostas (concentra??o celular e GSH). A produ??o de GSH a partir de S. cerevisiae ATCC 7754 em sistema pressurizado apresentou-se como uma tecnologia eficiente e promissora, por?m a influ?ncia de CM juntamente com a velocidade de reciclo celular, levou a mecanismos que promoveram determinada inibi??o em sua s?ntese, levando a diminui??o na sua concentra??o final.

O mercado de biorreatores com membranas (BRMs) têm exibido alto crescimento. Contudo, o fouling diminui o desempenho desses sistemas drasticamente. A aeração promove a mitigação do fouling, mas possui alto custo operacional (de até 70% do total da demanda energética) e é utilizada de forma otimizada apenas 10% das vezes, gerando a necessidade de estudos sobre a hidrodinâmica em BRMs. Ferramentas de dinâmica de fluidos computacional (CFD) são úteis para esse tipo de análise. Um dos primeiros passos para encontrar uma solução apropriada em simulações numéricas é propor uma modelagem correta. Dentre os fenômenos a serem modelados, os efeitos da turbulência são provavelmente um dos mais importantes; porém, nenhum artigo que comparasse predições com base na escolha de modelo de turbulência para simulações de sistemas BRM foi encontrado. Dessa forma, o objetivo desse trabalho foi verificar a importância da escolha do modelo de turbulência para simulações de biorreatores com membranas através de CFD. Para isso, as predições obtidas de velocidade local próxima às superfícies das membranas e de tensão cisalhante nessas superfícies para duas taxas de aeração, 5 e 15 m³/h, empregando dois modelos de turbulência, k-ϵ com funções de parede para alto (aR) e para baixo número de Reynolds (bR) e k-ω SST (Shear Stress Transport) com funções de parede automáticas, na análise de um sistema BRM foram comparadas entre si e com dados experimentais e simulados disponíveis na literatura. Os perfis temporais da velocidade e da tensão cisalhante exibiram alta variabilidade no período das oscilações em certos pontos, exigindo um longo tempo de escoamento para a convergência das variáveis locais. Identificou-se a necessidade de outorgar maior importância à definição do intervalo de tempo de coleta de dados experimentais, de modo a adquirir médias representativas do perfil dinâmico das variáveis e destes perfis serem também analisados para comparações mais definitivas entre resultados de simulações e medições experimentais. As diferenças, entre as medições experimentais da literatura e predições, obtidas pelas simulações deste trabalho foram, no geral, de ordem similar ou menores que as obtidas pelas simulações na literatura. Além disso, maior atenção deve ser dada à escolha da estratégia de modelagem de turbulência, visto que houve alta sensibilidade das predições, que variaram em até 21,6% dependendo dessa escolha. / Membrane bioreactors (MBR) market has been showing high growth rates over recent years. However, membrane fouling drastically reduces MBR overall performance. Aeration promotes fouling mitigation, but at a high operational cost (up to 70% of the MBR energy demand) and it is optimally employed only in 10% of the cases. This created the need of studies focused on MBR hydrodynamic. Computational fluid dynamics (CFD) is a useful tool for hydrodynamic analysis. One of the first steps in finding a proper solution for numerical simulation is proposing a correct modelling. Among the phenomena to be modelled for MBR simulations, turbulence effects are probably one of the most important; nevertheless, no paper comparing the predictions based on the turbulence model choice for MBR simulations was found. In sight of that, this work aimed to verify the relevance of the choice of turbulence model for MBR simulations through CFD. Predictions of local velocities near membranes surfaces and of local shear stress on those surfaces, for two aeration rates (5 and 15 m³/h), employing k-ϵ with wall functions for high (aR) and low (bR) Reynolds number and k-ω SST with automatic wall functions, on the analysis of a MRB system, were compared between each other and with experimental and simulated data available in the literature. The velocity and shear stress temporal profiles showed oscillations with highly variable periods in some points, which required a long process real time to verify the local variables convergence. It was identified the need to give more importance to the definition of the time interval for experimental data collection in order to acquire reliable temporal means; also, one must properly analyze the temporal profiles for more definitive comparisons between predictions and experimental measurements. The differences, between experimental data and predictions, obtained through this work simulations were, in general, of similar order or smaller than the ones reported in the literature. Besides, more attention must be given to the turbulence modelling choices, since the predictions obtained here were highly sensitive to those choices, showing differences up to 21,6% among them.

The prominence of insect cell culture has grown rapidly due to its ability to produce baculovirus biopesticides and recombinant proteins using the Baculovirus Expression Vector System. A critical problem in the mass production of these products is CO2 accumulation to inhibitory levels within the bioreactor. The current research investigated the effect of elevated CO2 concentrations on insect cell growth and metabolism and the roles of oxidative stress and intracellular pH (pHi) in CO2 inhibition. Spodoptera frugiperda Sf-9 insect cells were cultured in a 3 L bioreactor (1.2 L working volume) controlled at 20% air saturation, 27oC and a pH of 6.2. The cells were exposed to a constant CO2 concentration by purging the medium with CO2 and the headspace with air. The experiments were repeated for different CO2 concentrations and samples were taken every 24 h to determine cell density, viability, metabolism and oxidative stress. The population doubling time (PDT) of Sf-9 cells increased with increasing CO2 concentration. Specifically, the PDT for 0-37, 73, 147, 183 and 220 mm Hg CO2 concentrations were 23.2 ± 6.7, 32.4 ± 7.2, 38.1 ± 13.3, 42.9 ± 5.4 and 69.3 ± 35.9 h (n = 3 or 4; 95% confidence level), respectively. An 80 mL working volume shaker flask was maintained as a control and had an average PDT of 24.9 ± 3.1 h (n = 7; 95% confidence level). The viability of cells in all experiments was above 90%. The osmolality for all bioreactor experiments was observed to be 300 - 360 mOsm/kg, a range that is known to have a negligible effect on insect cell culture. Elevated CO2 concentration did not alter the cell specific glucose consumption rate (2.5 to 3.2 x 10-17 mol/cell-s), but slightly increased the specific lactate production rate from -3.0 x 10-19 mol/cell-s to 10.2 x 10-19mol/cell-s. Oxidative stress did not contribute to CO2 inhibition in uninfected Sf-9 cells as no significant increase in the levels of lipid hydroperoxide and protein carbonyl concentrations was discovered at elevated CO2 concentration. The experiments conducted to determine the effect of CO2 on pHi were not successful and different experimental methods tested were well documented.

Se(VI) and Se(IV), as the two major species of selenium in water, are toxic to aquatic lives and may cause adverse health effects to humans at high levels. Biological reduction of Se(VI) is a two-stage process first from Se(VI) to Se(IV) and then from Se(IV) to Se(0) with potential accumulation of the more toxic Se(IV) due to the slower rate of the second stage. Selenium reduction was first evaluated with batch cultures of Shigella fergusonii strain TB42616 (TB) and Pantoea vagans strain EWB32213-2 (EWB) isolated in our laboratory from sludge and coal slurry sediment samples, respectively. In order to facilitate Se(VI) reduction and reduce Se(IV) accumulation, the Se(VI)-reducing strain TB was co-cultured with a Se(IV)-reducing strain EWB. Although Se(VI) reduction rate was not affected, Se(IV) reduction was significantly enhanced with low Se(IV) accumulation in the defined co-culture. Effects of culture composition as well as nitrate and arsenate on Se(VI) reduction were also investigated. A co-culture composition of 10:1 (EWB:TB) ratio was observed to achieve the best total selenium reduction. In addition, nitrate at 50 mg/L was observed to inhibit Se(IV) reduction but not Se(VI) reduction, while arsenate at 200 mg/L exhibited slight inhibition on both Se(VI) and Se(IV) reduction. Biokinetic parameters were optimized with a Monod-type kinetic model using batch pure culture data through the Robust Global Optimization Algorithm embedded in a computer package. Se(VI) reduction by the defined co-culture was then simulated and verified over a range of culture compositions and initial Se(VI) concentrations, respectively. An inter-species inhibition term was incorporated into the model to illustrate the competition for Se(IV) during Se(VI) reduction in the co-culture. The model showed a significant increase of Se(IV) accumulation with higher initial Se(VI) concentration. However, Se(IV) accumulation can be reduced with increasing population ratio of EWB to TB in the defined co-culture. The relatively high correlation coefficients suggested that the model was robust and applicable in simulating Se(VI) reduction by the defined co-culture. Since activated alumina was reported to be more effective for Se(IV) adsorption than Se(VI), the effect of biological activities on selenium removal was investigated using continuous-flow reactors packed with alum-impregnated activated alumina (AIAA) and cultured with a Se(VI)-reducing strain TB under various influent Se(VI) concentrations and hydraulic retention times (HRTs). A selenium removal efficiency of 92% was achieved in a bioreactor with initial biomass of 2.2×106 cells/g-AIAA after a 70-day operation period. Little improvement was observed by lowering the influent Se(VI) concentration from 50 to 10 mg/L while the removal efficiency was significantly enhanced by either extending the hydraulic retention time from 3.2 to 5.0 days or increasing the attached biomass during the startup. An increase in mass ratios of Se(VI) reduction by immobilized cells to adsorption by AIAA was also observed with increasing cell mass during the operation. Se(VI) reduction using continuous-flow reactors packed with strain TB immobilized Ca2+-alginate beads was investigated under various hydraulic retention times (HRT) and influent Se(VI) concentrations. A high removal efficiency up to 98.7% was achieved under an HRT of 5 days and an influent Se(VI) concentration of 400 mg/L. The results showed that the overall selenium removal was positively correlated to the bed height of the reactor and the HRT but not related to the influent Se(VI) concentration. The steady state was analyzed using a mathematical model based on Monod-type equations with four biokinetic parameters optimized including the half-velocity constants and maximum specific reduction rates. The relatively high correlation coefficients indicate that the model is robust and valid to simulate Se(VI) reduction in the gel-beads-packed continuous-flow system.

Silage bunker runoff is a form of agricultural pollution that contributes to aquatic ecosystem degradation. Current handling and treatment methods for this process wastewater are often ineffective or expensive. A woodchip bioreactor is an emerging treatment technology designed to facilitate denitrification through the provision of an anaerobic, carbon rich environment. A wood chip bioreactor treatment system, consisting of three pre-treatment tanks, two wood chip bioreactors, and one infiltration basin, was constructed at the Miller Research Complex in South Burlington, Vermont in 2016. Runoff and leachate from an adjacent silage storage bunker is directed into the system. The pre-treatment tanks include two settling tanks and one aeration tank. The former allows for sedimentation of organic matter, while the latter is designed to allow for nitrogen transformations that will help maximize nitrogen removal in the bioreactors. During the summer and fall of 2017, sampling occurred at four points within the system in order to determine the efficacy of various treatment steps. Samples were analyzed for nitrate (NOx—N), ammonium (NH4+-N), total nitrogen (TN), soluble reactive phosphorus (SRP), and total phosphorus (TP) in order to compare inflow and outflow pollutant concentrations and loads. Results indicate that this treatment system significantly reduced nutrient loads in the runoff. Over the entirety of the sampling period, the influent TN and TP mass load were both reduced by approximately 44%.

Le travail présenté porte sur le couplage d’un procédé membranaire à extraction liquide-liquide avec un bioréacteur impliquant des molécules hydrophobes. La bioconversion modèle utilisée est la production de cis-2-methyl-5-isopropylhexa-2,5-dienal (isonovalal) à partir d’α-pinène oxyde, instable en phase aqueuse, par des cellules entières perméabilisées de Pseudomonas rhodesiae (CIP 107491). La production d’isonovalal en milieu biphasique eau (tampon phosphate)/hexadécane présente des verrous technologiques importants, dont une inactivation de l'enzyme à l'interface eau-solvant organique ainsi que l'apparition d'une émulsion stable. L’intérêt de la membrane porte sur l'absence de formation d'émulsion et sur l’augmentation de la durée de vie du biocatalyseur en raison de l'absence de contact direct du biocatalyseur avec l'interface liquide-liquide. La nature de la membrane a été choisie à partir de l'analyse des propriétés physico-chimiques du matériau et de l’étude des affinités entre membrane et composés d’intérêt (solutés, solvants). Il a été montré que les conditions d'écoulement au voisinage de la membrane, notamment du côté aqueux, jouent un rôle prépondérant sur les vitesses de transfert. Ce résultat souligne l'importance du design et des conditions d'opération du module membranaire sur les capacités de transfert. Le couplage de l’extraction membranaire liquide-liquide et de la réaction biologique a conduit à la mise en place d’un système bi-membranaire. Le prototype développé a permis de doubler les capacités catalytiques (+ 100 % d’isonovalal par gramme de biomasse) ainsi que de la durée de vie du biocatalyseur (160 h contre 80 h) par rapport à la même bioconversion réalisée en système biphasique conventionnel. / The study deals with the combination of a membrane process based on liquid/liquid extraction with a bioreactor producing hydrophobic molecules. The bioconversion used is the production of cis-2-methyl-5-isopropylhexa-2,5-dienal (isonovalal) from α-pinene oxide (unstable in aqueous phase) by whole cells of Pseudomonas rhodesiae (CIP 107491). The production of isonovalal in two-phase medium water/organic is known about but presents important technological brakes. Membrane interest concerns the stabilization of liquid/liquid interface and capacity to increase the biocatalyst life-time. Membrane nature is chosen from the analysis of physical and chemical properties of membrane material and study of the affinities between membrane and interest compounds (solutes, solvents). Two membrane contactors are designed and implemented on laboratory scale to study transfers between liquid phases. It is shown that the hydrodynamic conditions in the membrane neighborhood, in particular on aqueous side, play a major role on transfer speeds. This result underlines the importance of design and operation conditions in membrane module about the transfer capacities. The combination of liquid/liquid membrane extraction and biological reaction with unstable substrate had been studied and lead to the implementation of a serial bi-membrane system. The developed prototype, equipped with a PTFE membrane (polytetrafluoroethylene) with 0.22 μm pores’ diameter, highlights a doubling of catalytic capacities (+ 100 % of isonovalal per gram of biomass) as well as biocatalyst life-time (160 hours against 80 hours) compared with the same bioconversion realized in conventional two-phase medium system.

As nontoxic biobased surfactants derived from plant oils and cellulose or starch, saccharide-fatty acid esters are widely used in cosmetics, food, and pharmaceutical industries due to their biocompatibility, biodegradability as well as antimicrobial activity. Generally, saccharide-fatty acid esters are synthesized chemically under high pressure, temperature and the presence of alkaline or acid catalysts leading to low-quality products (chemo-degradation of double bonds and oxygenated moieties) and large amounts of byproducts. In contrast, biocatalytic synthesis enhances sustainability: near-ambient pressure and temperature, the absence of toxic, acids and bases catalysts, and improved selectivity of products. For lipase-catalyzed synthesis under nearly anhydrous conditions, the major hurdle to be overcome is the poor miscibility of the acyl donor and acceptor substrates, resulting in slow reaction rates. Although several approaches such as, the employments of organic solvents, complexation agents, and ionic liquids, have been reported in the literature, a robust solution is desperately needed. This study focused on employing immobilized lipases under completely solvent-free conditions to synthesize saccharide-fatty acid esters using the ester products to enhance miscibility. Experimentally, metastable saccharide particles with a diameter of 10-100 micron-sized suspensions of saccharide were formed in oleic acid-rich ester mixtures initially for synthesis of saccharide-fatty acid esters in packed bed bioreactor containing immobilized lipases. Water, a by-product that limits ester yield by promoting hydrolysis, was removed via free evaporation. In this dissertation, a bioreactor system was developed for the eco-friendly solvent-free, immobilized lipase-catalyzed synthesis of biobasaed surfactants utilizing suspensions as reaction medium with 88 wt% in 6 days; the performance of the bioreactor systems developed for Objective 1 was optimized through water concentration control and interval time with 91 wt% in 4.8 days; and to improve design of bioreactor system developed in Objective 1 by in-line filter and derive a mathematical model to describe the production of esters by the bioreactor systems developed. Finally, 84 wt% ester content was achieved in 8.4 days.