京都大学 / 新制・課程博士 / 博士(農学) / 甲第24677号 / 農博第2560号 / 新制||農||1100(附属図書館) / 学位論文||R5||N5458(農学部図書室) / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 吉田 天士, 教授 澤山 茂樹, 准教授 神川 龍馬 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The project aimed to carry out the techno-economic modeling of the heterotrophic microalgae bioreactors application in the poultry and swine abattoirs and processing wastewater treatment and possible exploitation of bioproducts from biomass generated in the process. The project focus was directed to the techno-economic modeling of bioprocesses following: (i) agroindustrial wastewater treatment and integral biomass production (ii) obtaining of bulk oil and lipid extracted algal (LEA) as feedstock for biodiesel conversion process and protein production (iii) obtaining of microalgal carotenoid-rich oleoresin solubilized in soybean oil. The results indicated that the agroindustrial wastewater treatment costs and production of microalgal biomass were USD 2.66 per cubic meter and USD 30 per ton of dried biomass respectively. The obtaining of bulk oil and LEA had a production cost of approx. USD 386.5 and 70.4 per ton. Finally, the oleoresin produced had an estimated production cost in USD 146.9 per kilogram. The results evidenced that the techno-economic modeling of the heterotrophic microalgae bioreactors application are an alternative to substantially minimize the production costs, giving economic sustainability to the agroindustry of poultry and swine processing. / O projeto teve por objetivo realizar a modelagem técnico-econômica da aplicação de biorreatores heterotróficos microalgais no tratamento de efluentes provenientes do abate e processamento de aves e suínos e possível exploração dos bioprodutos provenientes da biomassa gerada no processo. O foco do projeto foi direcionado à modelagem técnicoeconômica dos seguintes bioprocessos: (i) tratamento do efluente agroindustrial e produção de biomassa integral (ii) obtenção de óleo a granel e farelo microalgal desengordurado, como matéria-prima para o processo de conversão de biodiesel microalgal e produção de proteína (iii) obtenção de oleorresina de carotenóides mistos solubilizada em óleo de soja a partir de microalgas. Os resultados indicaram que os custos de tratamento do efluente agroindustrial e produção de biomassa microalgal foram US$ 2,66 por m³ de efluente e US$ 30 por tonelada de biomassa seca respectivamente. A obtenção de óleo a granel e farelo de microalgas obtiveram um custo de produção de US$ 386,5 e 70,4 por tonelada de óleo e farelo, respectivamente. Por fim, a oleorresina produzida teve um custo de produção estimado em US$ 146,9 por quilograma. Os resultados evidenciaram que a modelagem técnico-econômica da aplicação do biorreator heterotrófico microalgal, são uma alternativa para minimizar substancialmente os custos de produção, dando sustentabilidade econômica às agroindústrias de processamento de aves e suínos.

High costs for aeration, greenhouse-gas emissions and excess sludge disposal have entailed a paradigm shift in the wastewater treatment. Microalgal-bacterial-based wastewater treatments have gained increasing attention because of their potential in energy demand reduction and biomass resource recovery. In particular, photosynthetic oxygenation is combined with bacterial activity to treat wastewater avoiding external artificial aeration. To optimize the technology in order to become more competitive than activated sludge, an in-depth investigation about the treatment performance and the microbiology interactions under real operational condition is needed. This work focused on the study of wastewater-borne microalgal-bacterial consortia treating real municipal wastewater. The main objectives were to: (i) Understand the removal mechanisms and the influence of operational conditions to optimize the process; (ii) Analyze the microbial community. At first, a photo-sequencing batch reactor (PSBR), called Pilot, was started up and continuously monitored for two years to analyze the evolution of the treatment performance and of the biomass composition. At the same time, other two lab-scale PSBRs were installed to evaluate if microalgal inoculation is essential to start up a consortium. Samples of these consortia were collected over a period of one year and analyzed through microscopic observations, flow cytometry and metagenomics, to investigate the microbial structure and diversity.A second part of the research focused on the optimization of the Pilot to explore its limit in view of the scale-up of the system. In addition, respirometry was adapted to test microalgal-bacterial consortia to estimate the removal kinetic parameters for future modelling. To conclude, the research project addressed many aspects and lay the foundation to apply a methodological research approach to scale-up this promising technology.

With the growing interest in converting a wide variety of biomass-based feedstocks to biofuels, there is a need to develop effective procedures for the design and optimization of multi-feedstock biorefineries. The unifying goal of this work is the development of systematic methodologies and procedures for designing flexible multifeedstock biorefineries. This work addresses four problems that constitute building blocks towards achieving the unifying goal of the dissertation. The first problem addresses the design and techno-economic analysis of an integrated system for the production of biodiesel from algal oil. With the sequestration of carbon dioxide from power plant flue gases, algae growth and processing has the potential to reduce greenhouse gas emissions. Algae are a non-food oil feedstock source and various pathways and technologies for obtaining algal oil were investigated. Detailed economic and sensitivity analysis reveal specific scenarios that lead to profitability of algal oil as an alternative feedstock. In the second problem, a new safety metric is introduced and utilized in process design and selection. A case study was solved to assess the potential of producing biodiesel from sewage sludge. The entire process was evaluated based on multiple criteria including cost, technology and safety. The third problem is concerned with incorporating flexibility in the design phase of the development of multi-feedstock biofuel production processes. A mathematical formulation is developed for determining the optimal flexible design for a biorefinery that is to accommodate the use of multiple feedstocks. Various objective functions may be utilized for the flexible plant depending on the purpose of the flexibility analysis and a case study is presented to demonstrate one such objective function. Finally, the development of a systematic procedure for incorporating flexibility and heat integration in the design phase of a flexible feedstock production process is introduced for the fourth problem. A mathematical formulation is developed for use in determining the heat exchange network design. By incorporating the feedstock scenarios under investigation, a mixed integer linear program is generated and a flexible heat exchange network scheme can be developed. The solution provides for a network that can accommodate the heating and cooling demands of the various scenarios while meeting minimum utility targets.

Ce travail étudie l’obtention de phospholipides structurés enrichis en DHA et en acide caprylique (PC DHA-C8) par voie enzymatique. Deux voies de synthèse sont étudiées, l’acidolyse et l’estérification. Suite à un criblage enzymatique, la lipase retenue pour les deux voies de synthèse est TL-IM. Une optimisation des paramètres de la réaction d’acidolyse a été réalisée entre l’acide caprylique (C8:0) et la phosphatidylcholine de tournesol (PC) par le biais d’un plan d’expériences. Les conditions optimales déterminées sont une température de 38°C, une activité de l’eau de 0,7, une quantité d’enzyme de 15% de la masse en substrat ainsi qu’un rapport molaire C8:0/PC de 18. Ces conditions ont ensuite été utilisées pour l’acidolyse de phospholipides microalgaux riches en DHA issus de la microalgue Tisochrysis lutea afin d’obtenir de la PC DHA-C8. Les résultats n’ont pas été concluants. L’autre voie de synthèse étudiée est l’estérification par des lipases de la GPC, de l’acide caprylique et du DHA en milieu fondu. Cette réaction a été optimisée par la technique du pas par pas. Les paramètres étudiés sont la température, la quantité d’enzyme, le rapport molaire GPC/C8:0/DHA et l’application d’un vide. Pour l’obtention de PC DHA-C8, il faut fixer chacun de ces paramètres respectivement de la sorte : 45°C, 20% d’enzyme, un rapport molaire de 1/3/15 et un vide de 100 mbar. La production de PC DHA-C8, bien qu’optimisée ne dépasse pas 2% de rendement. Cependant, durant cette expérience, il a été constaté une forte production de LPC DHA, atteignant 16% sans optimisation des paramètres de synthèse. / The enzymatic synthesis of structured phsopholipids enriched in DHA and caprylic acid (PC DHA-C8) is studied. Two different ways are studied, acidolysis and esterification. An enzymatic screening led to the choice of the immobilized lipase from Thermomyces lanuginosa (TL-IM) for the 2 reactions. Parameters of the acidolysis reaction between carpylic acid (C8:0) and sunflower phosphatidylcholine (PC) were optimized by means of an experimental design. The optimum conditions determined are a temperature of 38°C, an aw of 0.7, an amount of enzyme of 15% of the mass of substrate and a molar ratio of C8:0/PC of 18. These conditions were applied to the acidolysis of microalgal phospholipids from T. lutea, rich in DHA, in order to produce PC DHA-C8. The other studied reaction is the lipase catalyzed esterification of GPC with C8:0 and DHA in a solvent-free medium This reaction has been optimized by studying each factor independently. The parameters studied are the temperature, the amount of lipase, the molar ratio GPC/C8:0/DHA and the use of reduced pressure. In order to obtain PC DHA-C8, each of theses parameters are respectively set at: 45°C, 20% of enzyme, a molar ratio of 1/3/15 and a pressure of 100 mbar. The production of PC DHA-C8, although optimized, does not exceed a yield of 2%. However, during this experiment, a high production of LPC DHA is observed, up to 16% without optimization of the synthesis parameter.

Freshwater green algae within species complex Haematococcus pluvialis are best natural producers of secondary carotenoid astaxanthin famous for its strong antioxidant activity. Despite worldwide distribution, great biotechnological potential and extensive literature dealing with different approaches to the cultivation of these organisms, there still remains surprising gaps in knowledge of their complex life cycle and behaviour. First uncertainty concerns asexual cell division of zoospores, hypothetically limited to five cell doublings (Lee & Ding 1994). No such genetic limitation was observed in this thesis, number of cell divisions is thus influenced mainly by cultivation conditions. There is still considerable uncertainty regarding to the asexual reproduction, as well. Ambiguity might be caused by scarce observation of the whole process of gametogenesis and conjugation of gametes. Published studies differed in some particular points. Unfortunately, we did not succeed in induction of gametogenesis despite broad scale of tested cultivation conditions applied to two different species within the H. pluvialis complex, namely H. pluvialis and H. rubicundus. This failure might be caused by heterothalism of the studied strains. Haematococcus pluvialis inhabits mainly ephemeral pools. Its ability to...