Submitted by Neusa Fagundes (neusa.fagundes@unioeste.br) on 2017-09-15T14:04:06Z No. of bitstreams: 1 Cristiane_Andreani2017.pdf: 1929530 bytes, checksum: 1b502a71b320a47e157f59e19b8978c7 (MD5) / Made available in DSpace on 2017-09-15T14:04:06Z (GMT). No. of bitstreams: 1 Cristiane_Andreani2017.pdf: 1929530 bytes, checksum: 1b502a71b320a47e157f59e19b8978c7 (MD5) Previous issue date: 2017-02-16 / Fundação Araucária de Apoio ao Desenvolvimento Científico e Tecnológico do Estado do Paraná (FA) / Anaerobic sequencing batch biofilm reactors (AnSBBR) have been used successfully in wastewater treatment. Currently, this setting has been used for biological production of hydrogen from synthetic and complexes substrates with reasonable results in stability and productivity. Considering the good performance achieved by such configuration, an AnSBBR reactor with a 6.0-L total capacity and a 4.3-L useful volume (2.3-L residual volume and 2.0-L treated volume by cycle) was evaluated, since its goal was hydrogen production, whose substrate was cassava wastewater. Eight experimental conditions were evaluated, in which four volumetric organic loads were applied (5.0; 12.0; 14 and 18 gCarb.L-1 d-1) calculated according to influent concentration (2,000; 3,200; 4,200; 4,700 and 5,000 mgCarb.L-1) and cycle time (4; 3 and 2 h). The AnSBBR feeding of the system was carried out by varying the reactor filling time. Thus, assays were carried out in: i) batch, in which the reactor filling occurs in 20 minutes; and ii) fed- batch, in which the reactor filling occurs during 50% of the total cycle time. Reactor inoculation was carried out with mixed cultures from autofermentation of cassava wastewater and anaerobic sludge that was thermally treated. The highest molar productivities were 36 molH2.m-3.d-1, while volumetric productivity was 2.2 LH2.L-1.d-1 (applied volumetric organic load of 14 g.L-1.d-1) and molar yield was 4.9 molH2.kg-1Carb (applied volumetric organic load of 12 g.L-1.d-1) in assays with the highest cycle times (4 h), inoculated with anaerobic sludge and autofermented cassava wastewater. Lactic acid and bacteriocin nisin were recorded in all the studied assays, consequently, it is evident the presence of lactic bacteria. Therefore, the total hydrogen absence can be explained in both assays, inoculated with autofermented cassava wastewater. / Reatores anaeróbios operados em bateladas sequenciais com biomassa imobilizada (AnSBBR) têm sido utilizados com sucesso no tratamento de águas residuárias. Atualmente, essa configuração vem sendo utilizada para produção biológica de hidrogênio a partir de substratos sintéticos e complexos com bons resultados em termos de estabilidade e produtividade. Tendo em vista os bons desempenhos alcançados por tal configuração, foi realizada a avaliação de um reator AnSBBR, com capacidade total de 6,0 L e volume útil de 4,3 L (2,3 L volume residual e 2,0 L volume tratado por ciclo), destinado à produção de hidrogênio, cujo substrato foi água residuária de fecularia de mandioca. Foram avaliadas oito condições experimentais, nas quais foram aplicadas quatro cargas orgânicas volumétricas (5,0; 12,0; 14,0 e 18 gCarb.L-1.d-1), calculadas em função da concentração afluente (2000; 3200; 4200; 4700 e 5000 mgCarb.L-1) e do tempo de ciclo (4, 3 e 2 h). A alimentação do sistema foi realizada pela variação do tempo de enchimento do reator. Dessa forma, foram conduzidos ensaios em: i) batelada, em que a alimentação ocorre em 20 minutos; e ii) batelada alimentada, na qual, a alimentação ocorre durante 50% do tempo total de ciclo. Na inoculação do reator, foram utilizadas culturas mistas provenientes da água residuária autofermentada e do lodo anaeróbio termicamente tratado. Foram obtidas as maiores produtividade molar de 36 molH2.m-3.d-1, produção volumétrica de 2,2 LH2. L-1.d-1 (carga orgânica volumétrica aplicada de 14 g.L-1.d-1) e rendimento molar de 4,9 molH2.kg-1Carb (carga orgânica volumétrica aplicada de 12 g.L-1.d-1) nos ensaios com os maiores tempos de ciclo (4 h) e inoculados com lodo anaeróbio e água residuária autofermetada. As presenças de ácido lático e da bacteriocina nisina foram verificadas em todos os ensaios, logo, há indícios da existência de bactérias láticas. Assim, é possível se justificar a total ausência de hidrogênio, verificada em dois dos ensaios, ambos inoculados com água residuária autofermentada.

Submitted by Neusa Fagundes (neusa.fagundes@unioeste.br) on 2018-09-06T14:03:20Z No. of bitstreams: 2 Angelo_Mari2018.pdf: 2243304 bytes, checksum: 694a2b64869c2cc929922f0f08581cd4 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-09-06T14:03:20Z (GMT). No. of bitstreams: 2 Angelo_Mari2018.pdf: 2243304 bytes, checksum: 694a2b64869c2cc929922f0f08581cd4 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-01-31 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Cassava starch industries have produced wastewater, which is their main environmental problem. Therefore, two-phase anaerobic digestion systems represent a newsworthy treatment alternative due to their efficiency in organic load removal and biogas production capacity. Thus, this study aimed at evaluating methane production from cassava starch wastewater in a sequential batch biofilm reactor (AnSBBR) as part of a two-phase system. The evaluation was composed of two main parts: i) analysis of a methanogenic reactor performance according to the increasing organic loads; and ii) the economic availability analysis regarding the reactor scaling-up for industrial scale. In the first part of this study, some previously acidified cassava starch wastewater was used to feed a methanogenic AnSBBR reactor on laboratory scale. The reactor has been submitted to increasing applied organic volumetric loads from 3.7 to 12.0 g COD.L-1.d-1, which were established by variation between affluent concentration and cycle time of a sequential batch reactor. The affluent concentration ranged from 2.8; 4.1 and 6.0 g COD-L-1, in sequential batches with cycle time of 6, 8 and 12 hours. The reactor was carried out at 30±1oC; it was stirred by a liquid phase recirculation and has been filled with support medium composed of polyurethane foam. The results of this first stage have shown that there was an increase in the organic loading rate (OLR) due to an increase on the affluent concentration and, mainly, a decrease on the cycle time duration showed an affect on the reactor performance. Methane productivity increased due to the increase in OLR and has reached 2.7 L CH4 L-1d-1 at OLR 12.0 g COD-L-1.d-1. COD removal has also been affected by OLR increase and ranged from 94 to 60%. And, the lowest values of COD removal were ascribed to the shortest cycle times. All the experimental conditions were constant. In the second part of this study, a two-phase anaerobic digestion system was designed on an industrial scale, based on the results previously obtained. Finally, an economic feasibility analysis was developed to settle down this project, which considered a twenty-year useful life term. The results of the second phase evinced that the simple payback was 8.4 years and the discounted payback was 15.9 years. Thus, it was recorded that this project can bring forth a profit similar to the net present value (R$ 1.9 million) at the end of the project lifetime, while internal rate of return was calculated in 11.1%. These results indicated that the two-stage system is economically viable to produce methane from starch wastewater. / As indústrias de produção de fécula de mandioca têm nas águas residuárias seu principal passivo ambiental, e o sistema de digestão anaeróbia em duas fases representa uma alternativa relevante de tratamento, devido à eficiência em remover a carga orgânica e a capacidade de produção de biogás. Assim, este estudo teve como objetivo avaliar a produção de metano a partir de água residuária de fecularia em um reator em batelada sequencial com biomassa imobilizada (AnSBBR) como parte de um sistema em duas fases. A avaliação desenvolvida foi composta por duas partes principais: i) análise do desempenho do reator metanogênico em função de cargas orgânicas crescentes; ii) análise de viabilidade econômica da ampliação do reator para a escala industrial. Na primeira parte do estudo, utilizou-se água residuária de fecularia previamente acidificada para abastecer um reator AnSBBR metanogênico em escala de laboratório. O reator foi submetido a cargas orgânicas volumétricas aplicadas (COVa) de 3,7 a 12,0 g DQO L-1d-1, estabelecidas pela variação entre a concentração do afluente e o tempo de ciclo do reator em batelada sequencial. As faixas de concentração testadas foram de 2,8; 4,1 e 6,0 g DQO.L-1, em bateladas sequenciais com tempos de ciclo de 6, 8 e 12 horas. O reator foi operado na temperatura de 30±1 oC, e contou com agitação por recirculação da fase líquida e meio suporte composto por espuma de poliuretano. Os resultados da primeira etapa demonstraram que o aumento da carga orgânica volumétrica pelo incremento da concentração do afluente e, principalmente, pela redução do tempo de ciclo, influenciou no desempenho do reator. A produtividade de metano aumentou com o acréscimo da COVa e atingiu 2,7 LCH4 L-1d-1 na COVa de 12,0 g DQO L-1d-1. A remoção de DQO também foi afetada pelo aumento da COVa, tendo variado de 94 e 60%, logo, os menores valores de remoção de DQO são atribuídos aos menores tempos de ciclo. Todas as condições experimentais se mostraram estáveis. Na segunda parte do estudo, dimensionou-se um sistema de digestão anaeróbia em duas fases em escala industrial, a partir dos resultados obtidos anteriormente. Por fim, foi desenvolvida uma análise de viabilidade econômica da implantação do projeto, que considerou um período de vida útil de vinte anos. Os resultados da segunda etapa demonstraram que o payback simples foi de 8,4 anos e o payback descontado de 15,9 anos. Ao final do tempo de vida útil, o projeto gerou lucro igual ao valor presente líquido de R$ 1,89 milhão e a taxa interna de retorno foi de 11,1%. Os resultados indicaram que o sistema em duas fases para a produção de metano a partir de água residuária de fecularia é economicamente viável.

Submitted by Marilene Donadel (marilene.donadel@unioeste.br) on 2018-11-23T16:43:21Z No. of bitstreams: 1 Andre_Souza_2018.pdf: 3532438 bytes, checksum: 75b2d4c04117567967dd3496b7c4c303 (MD5) / Made available in DSpace on 2018-11-23T16:43:21Z (GMT). No. of bitstreams: 1 Andre_Souza_2018.pdf: 3532438 bytes, checksum: 75b2d4c04117567967dd3496b7c4c303 (MD5) Previous issue date: 2018-09-28 / The development of societies, has always been interconnected energy sources, this way the H2 appears as a possible source. Nowadays, the biological pathway, the bacterial fermentation, has been highlighted, as it presents good productivity and low cost. The present work proposes to evaluate the potential of microbial consortia, which are easily accessible in the production of BioH2, by fermentation using the whey as a culture medium. This was done by productive complexes called PUBC (Pure Bacterial Complex), PIBC ( Pig Bacterial Complex), LACB (Land Bacterial Complex) and OXBC (Ox Bacterial Complex), where the fermentative medium was formed with 1ml of each complex and 6ml of culture medium, kept in a greenhouse with a temperature of 33ºC for a period of 07 days, where the concentration and production of the organic acids and gases were verified, resulting in 35% PUBC and 700 mLBioH2 / L (substrate) and with higher productivity of acetic acid, PIBC 36% and 1594 mLBioH2 / L, with higher concentration of butyric acid,LACB 37% and 1691 mLBioH2 / L, with higher productivity of succinic acid and OXBC 51% and 4371 mLBioH2 / L, with the productivity of organic acids divided into butyric and succinic. The results obtained showed a productive capacity of BioH2 and of organic acids, which in the case of OXBC obtained higher productive yield than works found in the current literature, making possible a new research and analysis to be done later. / O desenvolvimento das sociedades, sempre esteve interligadas as fontes energéticas, desta forma o H2 aparece como possível fonte. Atualmente a via biológica, a fermentação bacteriana, vem apresentando destaque, por apresentar boa produtividade e baixo custo. O presente trabalho propõem-se a avaliar o potencial de consórcios microbianos, de fácil acesso na produção de BioH2, por via fermentativa, utilizando o soro de leite como meio de cultivo. Esta foi realizada por complexos produtivos denominados de CBPU (Complexo Bacteriano Puro), CBPO (Complexo Bacteriano Porco), CBTE (Complexo Bacteriano Terra) e CBBO (Complexo Bacteriano Boi), onde o meio fermentativo foi formado com um 1ml de cada complexos e 6ml de meio de cultivo, mantidos em uma estufa com a temperatura em 33ºC, por um período de 07 dias, onde foi verificado a concentração e produção dos gases e ácidos orgânicos, tendo como resultado no CBPU 35% e 700 mLBioH2/L(substrato) e com maior produtividade de ácido acético, o CBPO 36% e 1594 mLBioH2/L, com maior concentração de ácido butírico, o CBTE 37% e 1691 mLBioH2/L, com maior produtividade de ácido succínico e o CBBO 51% e 4371 mLBioH2/L, com a produtividade de ácidos orgânicos dividida em butírico e succínico. Os resultados obtidos demonstraram ter capacidade produtiva de BioH2 e de ácidos orgânicos, que no caso do CBBO obteve rendimento produtivo maior que trabalhos encontrados na literatura atual, possibilitando um direcionamento para novas pesquisas e análises a serem feitas posteriormente.

A presente pesquisa teve como proposta avaliar o reator anaeróbio, operado de forma descontínua ou descontínua alimentada, contendo biomassa imobilizada em suporte inerte e com recirculação da fase líquida (AnSBBR) aplicado à produção de biohidrogênio a partir da codigestão de glicerina (efluente da produção de biodiesel) e soro de leite (efluente da produção de laticínios). A estabilidade, os índices de desempenho (referentes à produtividade e rendimento molar do hidrogênio) e o fator de conversão (entre biogás produzido e matéria orgânica consumida) foram analisados em função da composição afluente (porcentagem de cada substrato alimentado ao sistema), da variação da carga orgânica, do tempo de enchimento e da temperatura (20, 25, 30 e 35ºC). Os ensaios foram realizados em diferentes proporções dos substratos utilizando-se variadas cargas orgânicas volumétricas (10,3; 17,1 e 24,0 gDQO.L-1.d-1), as quais foram modificadas em função: (i) da concentração afluente (3, 5 e 7 gDQO.L-1) e (ii) do tempo de ciclo (4, 3 e 2 h, ou seja, 6, 8 e 12 ciclos diários). Também foram realizados ensaios para a produção de biometano a partir da codigestão proposta nesta pesquisa (com COAV de 7,6 gDQO.L-1.d-1) em diferentes proporções de mistura. Para a produção de biometano, a condição com 75% de soro e 25% de glicerina (base DQO) obteve os melhores resultados: produtividade molar de 101,8 molCH4.m-3.d-1 e rendimento por carga aplicada de 13,3 molCH4.kgDQO-1; o que representa um aumento de produtividade de cerca de 9% e 30% quando comparado com a digestão anaeróbia de soro e glicerina puros, respectivamente. A produção de metano no melhor ensaio aconteceu predominantemente pela rota hidrogenotrófica. Para a produção de biohidrogênio, a maior produtividade e rendimento do reator foram obtidas no ensaio operado com razão de mistura de 75% soro e 25% glicerina, com 7 gDQO.L-1 de concentração afluente, tempo de ciclo de 3 h e tempo de enchimento de 1,5 h (modo batelada alimentada - COAV de 23,9 kgDQO.m-3.d-1), a 30°C: foi obtida uma produtividade molar de 129,0 molH2.m-3.d-1 e rendimento de 5,4 molH2.kgDQO-1. Esses resultados representam um aumento de produtividade de 145% em relação a mono-digestão do soro na condição inicial, o que indica o benefício significativo da adição de glicerina ao afluente, provavelmente devido à sua capacidade tamponante, e a otimização das condições operacionais. A adição de glicerina e o aumento da COAV balancearam as rotas de produção de hidrogênio, sendo produzido de forma mais equilibrada pelas vias do ácido acético, butírico e valérico. A caracterização do consórcio microbiano desse ensaio indicou que a comunidade microbiana presente no AnSBBR foi dominada por Ethanoligenens e Megasphaera. / The current research evaluated an anaerobic reactor, operated in batch or fed-batch mode, containing immobilized biomass in inert support and with recirculation of the liquid phase (AnSBBR), applied to the production of biohydrogen co-digesting glycerin (effluent from biodiesel production process) and whey (effluent from dairy industry). Stability, performance (regarding productivity and molar hydrogen yield) and conversion factor (between biogas produced and organic matter consumed) were analyzed according to the percentage of each substrate fed to the system, organic loading rate, filling time and temperature (20, 25, 30 and 35ºC). Assays were carried out using different substrates proportions and organic loading rates (10.3; 17.1 and 24.0 gCOD.L-1.d-1), which have been modified in function of: (i) influent concentration (3, 5 and 7 gCOD.L-1) and (ii) cycle length (4, 3 and 2 h, i.e. 6, 8 and 12 cycles daily). Assays were also carried out aiming for biomethane production using the proposed co-digestion (with AVOL of 7.6 gDQO.L-1.d-1) with different proportions of substrate mixture. For biomethane production, the assay conducted with 75% whey and 25% glycerin (COD basis) obtained the best results: molar productivity of 101.8 molCH4.m-3.d-1 and yield per applied load of 13.3 molCH4. kgCOD-1; which is an increase in productivity of about 9% and 30% when compared with the anaerobic mono-digestion of whey and glycerin, respectively. Methane production in this assay came mainly from the hydrogenotrophic route. For biohydrogen production, the highest productivity and yield were achieved in the assay operated with 75% whey and 25% glycerin, with 7 gCOD.L-1 of influent concentration, 3 h of cycle time and filling time of 1.5 h (fed batch mode - AVOL of 23.9 kgCOD.m-3.d-1), at 30°C: a molar productivity of 129.0 molH2.m-3.d-1 and yield of 5.4 molH2.kgCOD-1 were obtained. These results represent a productivity increase of 145% in relation to whey mono-digestion at its initial condition, which indicates the significant benefit of glycerin addition to the influent, probably due to its buffering capacity, and improvement of operational conditions. The addition of glycerin and the increase in AVOL balanced the hydrogen production routes, since hydrogen was produced similarly by the acetic, butyric and valeric acid routes. The characterization of the microbial consortium of this assay indicated that the microbial community present in the AnSBBR was dominated by Ethanoligenens and Megasphaera.

Cette thèse de doctorat porte sur la valorisation du déchet solide issu de la préparation de la mélasse de caroube libanaise pour la production de bioénergie et de molécules à valeur ajoutée. L’analyse de la composition de ce déchet a montré qu’il contient 45 % (g/g) de sucres, substrats exploitables pour la fermentation alcoolique ou lactique, la production de biohydrogène, ou comme source de carbone pour la croissance d’une algue dans un procédé de phycoremédiation (traitement des eaux par les algues) pour l’élimination de micropolluants pharmaceutiques. Les résultats obtenus ont montré que la fermentation alcoolique en phase liquide à partir d’extraits de déchet de caroube peut conduire à des rendements élevés en bioéthanol sous réserve d’enrichir le milieu de culture par les éléments nécessaires pour la croissance de la levure Saccharomyces cerevisiae (en particulier l’azote), tandis qu’il est possible de produire directement le bioéthanol sur le déchet par fermentation en milieu solide sous réserve de contrôler précisément l’humidité. Nous avons également démontré que la fermentation lactique par immobilisation de Lactobacillus rhamnosus sur des billes d’alginate constitue une alternative possible au bioéthanol pour les sucres extraits du déchet. Cependant, un enrichissement du milieu de culture, ainsi que l’utilisation d’une invertase en prétraitement sont nécessaires afin de maximiser le rendement et la productivité de l’acide lactique. L’immobilisation des microorganismes a permis de réutiliser les mêmes billes d’alginate au cours de cinq cycles successifs de production. Une autre alternative potentielle aux voies précédentes est la fermentation sombre pour la production de biohydrogène directement à partir du déchet. Si des rendements intéressants ont été atteints, il faut noter que comme précédemment dans le cas de la fermentation solide du déchet, une forte dépendance aux conditions initiales de broyage a été mise en évidence ; de plus, à la carence en azote qui obligeait à supplémenter les milieux en fermentation en phase liquide s’ajoutent des besoins en fer. Enfin, l’utilisation du déchet comme substrat carboné dans un procédé de phycoremédiation avec l’algue Ankistrodesmus braunii a montré que l’élimination de 90% du diclofénac initial pouvait être atteinte en conditions de mixotrophie, même si seulement un tiers du diclofénac éliminé est effectivement métabolisé par l’algue. / This PhD thesis deals with the valorization of the solid waste generated by the Lebanese carob molasse process with the aim to produce bioenergy or high added-value platform molecules. The chemical analysis of this waste has highlighted that it contains 45% (w/w) sugars, which can be used as a substrate for alcoholic or lactic fermentation, the production of biohydrogen by dark fermentation, or as a carbon source for the growth of microalgae in a phycoremediation process (water treatment process based on algae/microalgae) for the removal of pharmaceutical micropollutants. Experimental results have shown that high ethanol yield and productivity could be achieved through alcoholic fermentation in the liquid phase using extracts from carob waste as the substrate, provided the culture medium was enriched by complementary nutrients (especially a nitro-gen source) to enhance the growth of the yeast Saccharomyces cerevisiae; in parallel, the direct production of ethanol from the solid waste based on solid-state fermentation was also proved to be efficient, provided humidity was accurately controlled. In addition, lactic fermentation per-formed with immobilized Lactobacillus rhamnosus on alginate beads was shown to constitute a potential alternative to bioethanol for the extracts from carob waste. An enriched culture medium was, however, necessary, together with the use of an invertase enzyme as a pretreatment so as to maximize the yield and the productivity of lactic acid. Using microorganism immobilization lead to the opportunity to recycle at least five times alginate beads in successive culture cycles. Another potential alternative to the above-mentioned valorization pathways was proved to lie in the dark fermentation process for biohydrogen production, directly using the solid waste. While attractive yields were reached, a strong dependence on the particle size from grinding pretreatment was emphasized, as in solid-state fermentation; moreover, iron supplementation in the culture medium was compulsory, in addition to nitrogen supplementation already described when fermentation was carried out in the liquid phase. Finally, when the carob waste was used as a substrate for the growth of the algae Ankistrodesmus braunii in a phycoremediation process, a removal yield of the initial diclofenac content about 90% could be reached under mixotrophic conditions, even though only one third of the diclofenac removed was effectively metabolized.

A ce jour, le contexte énergétique mondial est dominé par une utilisation massive des énergies fossiles non-renouvelables et épuisables par nature. La production de biohydrogène de 2ème génération issu de déchets organiques par le procédé de fermentation sombre constitue donc une solution attractive pour diversifier le mix énergétique actuel. Dans ce cadre, l’objectif de ce travail est d’étudier l’influence de la qualité du mélange sur l’efficacité de la voie fermentaire sombre. En effet, les conditions d’agitation mécanique (type d’agitateur, vitesse d’agitation) et la viscosité du digestat (fonction des intrants en cours de culture), comptent parmi les paramètres abiotiques les moins étudiés à ce jour dans ce procédé. Or, l’agitation joue un rôle clé puisqu’elle doit permettre non seulement d’homogénéiser la phase liquide riche en bactéries, en substrats organiques, en métabolites et en biogaz soluble, mais aussi de favoriser les échanges de matière liquide-bactéries et liquide-gaz. Cependant, pour atteindre la qualité de mélange requise, il faut faire face à deux contraintes : d’une part il faut maintenir un niveau acceptable de stress mécanique pour les bactéries du consortium ; d’autre part, la puissance mécanique consommée par l’agitation doit rester limitée pour assurer la viabilité économique du procédé. Dans ce travail, les effets combinés de la viscosité du digestat et de la vitesse d’agitation des mobiles sur la production de biohydrogène dans un bioréacteur ont été étudiés dans un premier temps. Les résultats ont montré une influence significative de ces deux facteurs sur la productivité en biohydrogène qui a pu être reliée au nombre adimensionnel de Reynolds et au régime d’écoulement du digestat. Un maximum de productivité a été observé lors de la transition laminaire-turbulent. Dans un deuxième temps, des méthodes de détermination du temps de mélange (conductimétrie, décoloration chimique, Fluorescence Induite par Nappe Laser) et du transfert de matière liquide-gaz (désoxygénation/oxygénation) ont été mises en oeuvre dans les mêmes conditions de viscosité et d’agitation afin de rechercher les étapes limitantes pouvant expliquer les évolutions observées lors des essais de fermentation. Les résultats ont montré que transfert interfacial et mélange ne sont limitants qu’en régime laminaire, alors que les faibles productivités en régime turbulent résultent vraisemblablement d’une interaction entre la turbulence et les agrégats bactériens. Ensuite, l’écoulement dans le bioréacteur a été modélisé par une approche de type Mécanique des Fluides Numérique (CFD) et analysé par une méthode de Vélocimétrie par Images de Particules (PIV) afin de déterminer les échelles spatiales locales de la turbulence et de pouvoir les comparer à la dimension caractéristique des agrégats bactériens. Les mesures locales confirment les hypothèses émises à partir des valeurs moyennes observées. Finalement, un modèle de type ADM1 (Anaerobic Digestion Model N°1) standard a été modifié en prenant en compte les ions lactate et un modèle hydrodynamique de type « cascade de cellules » dans le but de simuler la production de biohydrogène en systèmes batch et continu. Les simulations sont en bon accord avec les résultats expérimentaux dans les deux modes de culture en supposant un réacteur parfaitement mélangé. En conclusion, l’ensemble de ce travail confirme que la viscosité du digestat et les conditions de mélange sont effectivement des paramètres essentiels à prendre en compte pour l’optimisation et l’extrapolation du procédé de fermentation sombre. / The global energy trends are currently dominated by a massive use of fossil non-renewable energy sources which are progressively depleting. In this way, the production of second-generation biohydrogen production from organic wastes by the dark fermentation process offers, therefore, an attractive solution to diversify the present energy mix. Within this framework, the aim of this work is to investigate the effect of the efficiency of the mixing process on dark fermentation. The conditions of mechanical agitation (mixer type, mixing speed) and the viscosity of the digestate (which depends on the variability of influent substrate concentration) are, indeed, among the abiotic factors that have been the most disregards up to now in this bioprocess. For example, mixing plays a key role because agitation conditions must ensure on the one hand the homogenization of the liquid phase enriched in bacteria, in organic substrate, in soluble metabolites, and in soluble biogas, and in the other hand promote liquid-to-bacteria and liquid-to-gas mass transfer. However, to reach the desired degree of mixing, two constraints must be faced: firstly, an acceptable level of mechanical stress must be maintained on the microbial consortium, and secondly, mechanical power input due to mixing must comply with the economic sustainability of the process. In this work, the combined effects of digestate viscosity and agitation conditions on the fermentative biohydrogen production in the bioreactor were studied first. Experimental results highlighted a significant effect of these factors on biohydrogen productivity which could be expressed as function of the purely hydrodynamic dimensionless Reynolds number and of the prevailing flow regime. Hydrogen production was maximized in the transition region between laminar and turbulent flow conditions. Secondly, experimental measuring methods of mixing time (conductimetric, chemical decolorization and Planar Laser Induced Fluorescence techniques) and mass transfer (dynamic deaeration/aeration) were implemented in the same conditions of viscosity and agitation conditions so as to investigate the possible limiting steps that could explain the trends observed in the mixed cultures. The results proved that mixing and liquid-gas transfer was slower than hydrogen production rate only in the laminar flow regime, while low production rate under turbulent flow conditions might stem from an interaction between turbulent eddies and bacterial aggregates. Then, the flow field in the bioreactor was simulated using a CFD (Computational Fluid Dynamics) methodology and analyzed experimentally using PIV (Particle Image Velocimetry) to determine the characteristic turbulent length scales and to compare them to the characteristic size of the bacterial aggregates. Local measurements confirmed the assumptions made from average values derived from power input data. Finally, a modified ADM1 model (Anaerobic Digestion Model N°1) was developed to simulate the biohydrogen production, accounting for lactate ions and non-ideal mixing, under batch and continuous culture conditions. Simulations fairly agree with experimental data in both modes of cultures assuming perfect mixing condition. As a conclusion, the present work as a whole confirms that digestate viscosity and mixing conditions constitute key parameters that must be considered for process optimization and for the scale-up of dark fermentation.

Limitation of fuels reserves and contribution of fossil fuels to the greenhouse effect leads to develop a new, clean and sustainable energy. Among the various options, biohydrogen appears as a promising alternative energy source. The fermentative hydrogen production process holds a great promise for commercial processes. Hydrogen production by fermentative bacteria is a very complex and greatly influenced by pH. This paper presents biohydrogen production by bacterial strain Clostridium sp. Tr2. Operational pH strongly affected its hyrogen production. Its gas production rate as well as obtained gas product were roughly increase twice under controlled pH at 6 than non-controlled condition. Dark fermentation for hydrogen production of strain Tr2 was performed under bottle as well as automatic fermenter scale under optimal nutritional and environmental conditions at 30oC, initial pH at 6.5, then pH was controlled at 6 for bioreactor scale (BioFlo 110). Bioreactor scale was much better for hydrogen production of strain Tr2. Clostridium sp. Tr2 produced 0.74 L hydro (L medium)-1 occupying 72.6 % of total gas under bottle scale while it produced 2.94 L hydro (L medium)-1 occupying 95.82 % of total gas under fermenter scale. Its maximum obtained hydrogen yield of Clostridium sp. Tr2 under bioreactor scale Bioflo 110 in optimal medium with controlled pH 6 was 2.31 mol hydro (mol glucose)-1. / Dự trữ nhiên liệu có giới hạn và việc sử dụng nhiên liêu hoá thạch góp phần không nhỏ gây hiệu ứng nhà kính dẫn đến cần phải phát triển năng lượng mới, sạch và bền vững. Trong số các giải pháp, hydro sinh học xuất hiện như một nguồn năng lượng thay thế đầy hứa hẹn. Quá trình lên men sản xuất hydro có tiềm năng lớn để áp dụng trong sản xuất thương mại. Tuy nhiên qúa trình này rất phức tạp và chịu ảnh hưởng lớn bởi pH. Nghiên cứu này trình bày sản xuất hydro sinh học do chủng vi khuẩn Clostridium sp. Tr2. Quá trình sản xuất hydro của chủng này bị ảnh hưởng mạnh mẽ bởi pH thay đổi trong quá trình lên men. Tốc độ tạo khí cũng như lượng khí thu được của chủng này tăng gần gấp đôi trong môi trường có duy trì pH ở pH 6 so với môi trường không kiểm soát pH. Quá trình lên men tối sản xuất hydro của chủng Tr2 được thực hiện ở quy mô bình thí nghiệm cũng như bình lên men tự động trong điều kiện môi trường tối ưu ở 30oC, pH ban đầu 6.5, ở qui mô bình lên men tự động (BioFlo 110), pH môi trường sau đó được duy trì ổn định ở pH 6. Lên men sản xuất hdyro của chủng Tr2 trong bình lên men tự động tốt hơn rất nhiều so với lên men trong bình thí nghiệm. Clostridium sp. Tr2 chỉ tạo ra được 0,74 L hydro (L medium)-1 chiếm 72,6 % tổng thể tích khí thu được ở điều kiện lên men bình thí nghiệm trong khi chủng này sản xuất được 2,94 L hydro (L medium)-1 chiếm 95,82 % tổng thể tích khí ở điều kiện lên men tự động. Sản lượng hydro thu được lớn nhất của chủng này trong bình lên men tự động BioFlo 110 trong trong môi trường tối ưu có kiểm soát pH tại pH 6 là 2,31 mol hydro (mol glucose)-1.

La recherche de sources d’énergie fiables ayant un faible coût environnemental est en plein essor. L’hydrogène, étant un transporteur d’énergie propre et simple, pourrait servir comme moyen de transport de l’énergie de l’avenir. Une solution idéale pour les besoins énergétiques implique une production renouvelable de l’hydrogène. Parmi les possibilités pour un tel processus, la production biologique de l’hydrogène, aussi appelée biohydrogène, est une excellente alternative. L’hydrogène est le produit de plusieurs voies métaboliques bactériennes mais le rendement de la conversion de substrat en hydrogène est généralement faible, empêchant ainsi le développement d’un processus pratique de production d’hydrogène. Par exemple, lorsque l’hydrogène est produit par la nitrogénase sous des conditions de photofermentation, chaque molécule d’hydrogène constituée requiert 4 ATP, ce qui rend le processus inefficace. Les bactéries photosynthétiques non sulfureuses ont la capacité de croître sous différentes conditions. Selon des études génomiques, Rhodospirillum rubrum et Rhodopseudomonas palustris possèdent une hydrogénase FeFe qui leur permettrait de produire de l’hydrogène par fermentation anaérobie de manière très efficace. Il existe cependant très peu d’information sur la régulation de la synthèse de cette hydrogénase ainsi que sur les voies de fermentation dont elle fait partie. Une surexpression de cette enzyme permettrait potentiellement d’améliorer le rendement de production d’hydrogène. Cette étude vise à en apprendre davantage sur cette enzyme en tentant la surexpression de cette dernière dans les conditions favorisant la production d’hydrogène. L’utilisation de résidus organiques comme substrat pour la production d’hydrogène sera aussi étudiée. / The search for alternative energy sources with low environmental impact is in great expansion. Hydrogen, an elegant and simple energy transporter, could serve as means of transporting energy in the future. An ideal solution to the increasing energy needs would imply a renewable production of hydrogen. Out of all the existing possibilities for such a process, the biological production of hydrogen, also called biohydrogen, is an excellent alternative. Hydrogen is the end result or co-product of many pathways in bacterial metabolism. However, such pathways usually show low yields of substrate to hydrogen conversion, which prevents the development of efficient production processes. For example, when hydrogen is produced via nitrogenase under photofermentation conditions, each hydrogen molecule produced requires 4 molecules of ATP, rendering the process very energetically inefficient. Purple non-sulfur bacteria are highly adaptive organisms that can grow under various conditions. According to recent genomic analyses, Rhodospirillum rubrum and Rhodopseudomonas palustris possess, within their genome, an FeFe hydrogenase that would allow them to produce hydrogen via dark fermentation quite efficiently. Unfortunately, very little information is known on the regulation of the synthesis of this enzyme or the various pathways that require it. An overexpression of this hydrogenase could potentially increase the yields of substrate to hydrogen conversion. This study aims to increase our knowledge about this FeFe hydrogenase by overexpressing it in conditions that facilitate the production of hydrogen. The use of organic waste as substrate for hydrogen production will also be studied.

O aproveitamento da energia luminosa por bactérias que produzem hidrogenases poderia aumentar a eficiência do processo de produção de biohidrogênio. Neste trabalho, foi realizada a clonagem do gene que codifica a proteorrodopsina (PR) do isolado metagenômico SAR86 num plasmídeo de expressão para bactérias Gram-negativas. PR é uma proteína ligada ao cromóforo retinal, que, sob iluminação, promove o efluxo de prótons através da membrana celular. O excesso de prótons na face externa da membrana pode servir como substrato para a hidrogenase, resultando em maior eficiência na produção de hidrogênio (2H+ + 2e→ H2). O plasmídeo contendo o gene da PR foi utilizado na transformação genética das bactérias Cupriavidus necator e Escherichia coli, que produzem diversas hidrogenases. Enquanto a PR não se mostrou funcional em C. necator, na linhagem recombinante de E. coli, cultivada em presença de luz e retinal, foi obtido um aumento de até 2,17 vezes na produção de H2 em relação ao cultivo no escuro, desde que a linhagem estivesse produzindo a hidrogenase endógena HYD-4. / The utilization of light energy by hydrogenase producing bacteria could increase the efficiency of the biohydrogen production process. In the present work, the gene coding for proteorhodopsin (PR) of the SAR86 metagenomic lineage was cloned in an expression plasmid for Gram-negative bacteria. PR is an apoprotein linked to the chromophore retinal, which, upon illumination, promotes proton efflux across the cell membrane. The excess of protons on the plasma membrane surface may serve as a substrate for hydrogenases, resulting in a higher efficiency of hydrogen production (2H+ + 2e→ H2). The plasmid containing the PR gene was used to transform the Gram-negative bacteria Cupriavidus necator and Escherichia coli which produce several hydrogenases. Whereas PR did not display functionality in C. necator, in the recombinant E. coli cells, grown under illumination in the presence of retinal, an enhancement up to 2.17 fold in H2 production was found, relative to cells grown under darkness, provided that the cells were expressing the endogenous HYD-4 hydrogenase.

Este estudo investigou a aplicação de um reator anaeróbio operado em bateladas sequenciais com biomassa imobilizada (AnSBBR) com agitação na produção de biohidrogênio tratando água residuária sintética a base de sacarose, sendo o desempenho do biorreator avaliado de acordo com a influência conjunta do tempo de alimentação, do tempo de ciclo, da concentração afluente e da carga orgânica volumétrica aplicada (COVAS). O biorreator, com capacidade útil de 5,6 L, foi dividido em 3 partes: volume de meio tratado por ciclo de 1,5 L, volume residual de meio de 2,0 L e volume de suporte inerte com biomassa de 2,1 L. Foram aplicadas 6 condições experimentais de COVAS de 9,0 a 27,0 gDQO.L-1.d-1, combinado diferentes concentrações afluentes (3500 e 5400 mgDQO.L-1), tempos de ciclo (4, 3 e 2h), sendo tempo de enchimento do reator (tC) correspondente a 50% ao tempo de ciclo. Os resultados mostraram que o aumento COVAS contribuiu para a queda no consumo de sacarose de 99% para 86% e para o aumento do rendimento molar por carga removida (RMCRC,n) de 1,02 molH2.molSAC-1 na COVAS de 9,0 gDQO.L-1.d-1 até atingir o valor máximo de 1,48 molH2.molSAC-1 na COVAS de 18,0 gDQO.L-1.d-1 com queda a partir desse ponto. O aumento da COVAS resultou no aumento da produtividade molar volumétrica (PrM) de 24,5 para 81,2 molH2.m-3.d-1. A maior produtividade molar específica (PrME) obtida foi de 8,71 molH2.kgSVT-1.d-1 para a COVAS de 18,0 gDQO.L-1.d-1. A diminuição do tempo de ciclo resultou na diminuição do consumo de sacarose e no aumento da PrM. Foi verificado também que a diminuição do tC de 4h para 3h contribuiu para o aumento da PrME. O aumento da concentração afluente resultou na diminuição do consumo de sacarose apenas na faixa de 2h, no aumento do RMCRC,n e da PrM em todas as faixas de tC, e no aumento da PrME nas faixas de 4h e 3h. A estratégia de alimentação mostrou ser um parâmetro operacional de grande importância, sendo o aumento do tempo de enchimento responsável pelo aumento do consumo de sacarose, da PrM, da PrME e do RMCRC,n para todas as COAVS investigadas. Em todas as condições, houve o predomínio do ácido acético seguido pelo etanol, ácido butírico e propiônico. / This study investigated the feasibility of an anaerobic sequencing batch biofilm reactor (AnSBBR) with agitation on biohydrogen production treating synthetic wastewater from sucrose, the performance of the bioreactor was evaluated according the combined influence of fill time, cycle period, influent concentration and applied organic loading rate (COAVS) . The bioreactor, with working volume of 5,6L, was divided in 3 parts: 1,5L of fed volume per cycle, 2,0L of residual medium and 2,1L of inert support and biomass. The reactor was operated under six operating conditions with different COAVS ranging from 9,0 to 27,0 gCOD.L-1.d-1, obtained by the combination of different influent concentrations (3500 e 5400 mgCOD.L-1), cycle periods (4, 3 e 2h) and fill time corresponding to 50% of cycle period. The results showed that increasing COAVS resulted in lesser sucrose removal from 99% to 86% and improved yield per removed loading rate (RMCRC,n) of 1,02 molH2.molSUC-1 in COAVS of 9,0 gCOD.L-1.d-1 to maximum value of 1,48 molH2.molSUC-1 in COAVS of 18,0 gCOD.L-1.d-1 decreasing after that. Increasing COAVS improved molar productivity (PrM) from 24,5 to 81,2 molH2.m-3.d-1. The higher specific molar productivity (PrME) obtained was 8,71 molH2.kgTVS-1.d-1 in COAVS of 18,0 gCOD.L-1.d-1. Decreasing cycle period resulted in less sucrose consumption and increased PrM. It was observed that decreasing cycle period of 4h to 3h improved PrME. Increasing influent concentration resulted in less sucrose degradation only on range of 2h, in an increase of RMCRC,n and in an increase of PrM in all ranges of tC, and increased PrME on ranges of 4h and 3h. In all operational conditions, the main intermediate metabolic was acetic acid followed by ethanol, butyric and propionic acids. The feeding strategy had a great effective on hydrogen production, longer fill times resulted in better sucrose removal, PrM, PrME and RMCRC,n for all COAVS investigated.