Microscopic green algae produce a number of beneficial substances. The commercially used ones include mainly pigments, especially secondary carotenoids. While primary carotenoids are part of the photosynthetic apparatus, secondary carotenoids are produced only under certain specific conditions (e.g. high exposure to light, nitrogen deficiency). Secondary carotenoids have antioxidant properties that protect the cell from adverse effects. The strains currently in commercial use and the strains that are tested for potential use come almost exclusively from algal collections, so this thesis focuses on strains isolated from nature. Two strains of aeroterrestrial green unicellular algae, which showed potential of producing carotenoids, were found and isolated. These strains were identified as Tetracystis pulchra (clade Dunaliellinia) and Tetracystis sp., a strain related to Tetracystis tetraspora SAG 98.80, belonging to the clade Stephanosphaerinia. The culture conditions for T. pulchra were subsequently optimized and then the culture was subjected to a series of experiments, examining the effect of stress factors on carotenoid formation and accumulation. Specifically, the effect of nitrogen starvation, light intensity, temperature and UVA radiation was tested. It has been shown that especially the light...

Microalgae are considered one of the most promising feedstocks for biofuel production for the future. The most efficient way to produce vast amounts of algal biomass is the use of closed tubular photobioreactors (PBR). The heat requirement for a given system is a major concern since the best algae growth rates are obtained between 25-30 °C, depending on the specific strain. A procedure to determine temperature influence on algal growth rates was developed for a lab-scale PBR system using the species Chlorella. A maximum growth rate of 1.44 doublings per day at 29 °C (optimal temperature) was determined. In addition, a dynamic mathematical model was developed to simulate heating and cooling energy requirements of tubular PBRs for any desired location. Operating the model with hourly weather data as input, heating and cooling loads can be calculated early in the planning stage of a project. Furthermore, the model makes it possible to compare the operation inside a greenhouse to the outdoor operations, and consequently provides fundamental information for an economic feasibility study. The best configuration for a specific location can be evaluated easily. The model was exemplary tested for a hypothetical 100,000 l photobioreactor located in San Luis Obispo, California, U.S.A. Average algae productivity rates of 23% and 67% for outdoor and indoor PBR operations, respectively, were obtained. Actual energy loads (heating and cooling) needed to maintain the PBR at optimal temperature were determined and compared. Sensitivity analyses had been performed for abrupt temperature and solar radiation steps, PBR row distances, ground reflectivities, and ventilation rates of the greenhouse. An optimal row distance of 0.75 m was determined for the specific PBR. The least amount of energy was needed for a ground reflectivity of 20%. The ventilation rate had no major influence on the productivity rate of the system. Results demonstrated the importance of a simulation model as well as the economic impact of a sophisticated heat management system. Energy savings due to an optimized heat management system will eventually increase proficiency of the systems, which will support a new sustainable industry and future developmental potential.

Tesis por compendio / [ES] En el contexto actual de escasez de recursos que sufre el planeta (biomasa, agua y energía), la tecnología basada en los cultivos de microalgas para el tratamiento de aguas residuales aparece como una tecnología muy interesante que permite no sólo la eliminación de los nutrientes (N y P) presentes en el agua, sino también la recuperación de estos nutrientes en la producción de una biomasa algal de alto valor con diversas aplicaciones: generación de biogás, producción de biocombustibles y biofertilizantes, elaboración de fármacos y cosméticos, etc. Estudios previos han demostrado que el efluente de un reactor anaerobio de membranas (AnMBR) resulta ser un medio de cultivo óptimo para el crecimiento de las microalgas. No obstante, la mayoría de los estudios existentes se han llevado a cabo a escala laboratorio en condiciones controladas de luz, temperatura, pH, carga de nutrientes, etc., y normalmente siempre en experimentos batch. Este trabajo consiste en el estudio y modelación matemática del proceso de cultivo de microalgas en una planta piloto de fotobiorreactores de membrana (MPBR) operando en continuo y en condiciones outdoor para el tratamiento del efluente de un sistema AnMBR que trata agua residual urbana real. Durante la fase de experimentación de los cultivos de microalgas se han llevado a cabo diversos experimentos en la planta MPBR donde se han evaluado diversos factores que afectan al crecimiento de las microalgas: temperatura, luz solar, tiempo de retención celular (TRC), carga de nutrientes o tiempo de retención hidráulico (TRH), sistema de recirculación del cultivo y el volumen en zona oscura. Los resultados obtenidos muestran la enorme importancia de las condiciones ambientales (luz solar y temperatura) en el rendimiento de los cultivos de microalgas. La temperatura óptima del cultivo de microalgas con predominancia del género Scenedesmus sp. resultó estar en torno a los 25ºC, mientras que temperaturas por debajo de 20ºC y por encima de 25ºC afectaron negativamente a la productividad de biomasa. La operación del sistema de fotobiorreactores (FBR) sin membranas para TRH 8 días y en condiciones ambientales favorables consiguió reducir la concentración de nutrientes por debajo de los límites de vertido que marca la Directiva 98/15/CE (10 mg N·L-1 y 1 mg P·L-1) alcanzando valores de eliminación de 75,2% de N y 77,9% de P. La operación del sistema MPBR permitió desacoplar el TRC del TRH en la operación de los FBR, lo que resultó en una mejora general del rendimiento de los cultivos de microalgas y permitió obtener un efluente libre de sólidos con alto potencial de reutilización. Los sistemas de recirculación del cultivo de microalgas comparados en el estudio (bombeo mecánico vs sistema airlift) no afectaron significativamente al rendimiento del cultivo. Por otro lado, reduciendo el volumen en zona oscura de un 27,2% al 13,6% en el sistema MPBR se consiguió un incremento del 40% en la productividad de biomasa. Mediante el uso de los datos obtenidos en planta piloto se ha desarrollado un modelo matemático de crecimiento de microalgas que permite simular de manera muy precisa (R2 = 0,9954) el comportamiento de los cultivos de microalgas en un sistema MPBR. Este modelo utiliza la notación y terminología de los modelos ASM, y consta de un total de 14 componentes (10 solubles y 4 suspendidos), 11 procesos gobernados por la cinética y los equilibrios ácido-base que determinan el pH del medio. Además, el modelo considera los efectos la luz y la temperatura en el crecimiento. Como novedad interesante respecto a otros modelos matemáticos de crecimiento de microalgas ya publicados, este modelo contempla, en condiciones de ausencia de P en el medio de cultivo, el crecimiento de las microalgas a partir del polifosfato almacenado internamente. El modelo desarrollado en este trabajo pretende ser una herramienta para facilitar la implementación futura de la tecnología de cultivos de microalgas en una EDAR a escala industrial. / [CAT] En el context actual d'escassetat de recursos que sofreix el planeta (biomassa, agua i energia), la tecnologia basada en els cultius de microalgues per al tractament d'aigües residuals apareix com una tecnologia molt interessant que permet no només l'eliminació dels nutrients (N i P) presents a l'aigua, sinó també la recuperació d'aquests nutrients amb la producció d'una biomassa algal d'alt valor amb diverses aplicacions: generació de biogàs, producció de biocombustibles i biofertilitzants, elaboració de fàrmacs i cosmètics, etc. Estudis previs han demostrat que l'efluent d'un reactor anaerobi de membranes (AnMBR) resulta ser un mitjà de cultiu òptim per al creixement de les microalgues. Tot i això, la majoria dels estudis existents s'han dut a terme a escala laboratori en condicions controlades de llum, temperatura, pH, càrrega de nutrients, etc., i normalment sempre en experiments batch. Aquest treball consisteix en l'estudi i la modelació matemàtica del procés de cultiu de microalgues en una planta pilot de fotobioreactors de membrana (MPBR) operant en continu i en condicions outdoor per al tractament de l'efluent d'un sistema AnMBR que tracta aigua residual urbana real. Durant la fase d'experimentació dels cultius de microalgues s'han dut a terme diversos experiments a la planta MPBR on s'han avaluat diversos factors que afecten al creixement de les microalgues: temperatura, llum solar, temps de retenció cel·lular (TRC), càrrega de nutrients o temps de retenció hidràulic (TRH), sistema de recirculació del cultiu i el volum en zona obscura. Els resultats obtinguts mostren l'enorme importància de les condicions ambientals (llum solar i temperatura) en el rendiment dels cultius de microalgues. La temperatura òptima del cultiu de microalgues amb predominança del gènere Scenedesmus sp. va resultar estar entorn als 25ºC, mentre que temperatures per sota de 20ºC i per sobre de 25ºC van afectar negativament a la productivitat de biomassa. L'operació del sistema de fotobioreactors (FBR) sense membranes per a TRH 8 dies i en condicions ambientals favorables va aconseguir reduir la concentració de nutrients per sota dels límits d'abocament que marca la Directiva 98/15/CE (10 mg N·L-1 i 1 mg (P·L-1) assolint valors d'eliminació de 75,2% de N i 77,9% de P. L'operació del sistema MPBR va permetre desacoblar el TRC del TRH en l'operació dels FBR, la qual cosa va resultar en una millora general del rendiment dels cultius de microalgues i va permetre obtenir un efluent lliure de sòlids amb un alt potencial de reutilització. Els sistemes de recirculació del cultiu de microalgues comparats en aquest estudi (bombeig mecànic vs sistema airlift) no van afectar significativament al rendiment del cultiu. D'altra banda, reduint el volum en zona obscura del 27,2% al 13,6% al sistema MPBR es va aconseguir un increment del 40% en la productivitat de biomassa. Mitjançant l'ús de les dades obtingudes a la planta pilot s'ha desenvolupat un model matemàtic de creixement de microalgues que permet simular de manera molt precisa (R2 = 0,9954) el comportament dels cultius de microalgues en un sistema MPBR. Aquest model utilitza la notació i la terminologia dels models ASM, i consta d'un total de 14 components (10 solubles i 4 suspesos), 11 processos governats per la cinètica i els equilibris àcid-base que determinen el pH del medi. A més, el model considera els efectes de la llum i la temperatura en el creixement. Com a novetat interessant respecte d'altres models matemàtics de creixement de microalgues ja publicats, aquest model contempla, en condicions d'absència de P en el mitjà de cultiu, el creixement de les microalgues a partir del polifosfat emmagatzemat internament. El model desenvolupat en aquest treball pretén ser una eina per facilitar la implementació futura de la tecnologia de cultius de microalgues a una EDAR a escala industrial. / [EN] In the actual context of resource scarcity along the world (biomass, water and energy), microalgae-based technology for wastewater treatment appears as a promising technology that allows not only nutrient removal (N and P) from wastewater, but also the recovery of these nutrients for the production of high-value algal biomass which has different applications: biogas generation, biofuel and biofertilizer production, pharmaceuticals and cosmetics manufacturing, etc. Previous studies have proved that the effluent from an anaerobic membrane bioreactor (AnMBR) could be a suitable growth medium for microalgae cultivation. However, most of the existing studies have been carried out at bench scale under controlled conditions of light, temperature, pH, nutrient load, etc., when working in batch mode. The present work consists of the study and mathematical modelling of an outdoor pilot-scale membrane photobioreactor (MPBR) for microalgae cultivation under continuous operation for treating the effluent of an AnMBR system fed with real municipal wastewater. During the experimental phase of microalgae cultivation, different experiments were carried out in the MPBR plant to evaluate the main factors that affect microalgae growth: temperature, solar light irradiance, biomass retention time (BRT), nutrient load or hydraulic retention time (HRT), the algae culture recirculation system and the non-photic volume. The results obtained show the significant effect of the environmental conditions (solar light and temperature) on the microalgae cultivation performance. Optimum temperature for the microalgae cultures with a predominance of the genus Scenedesmus sp. resulted to be around 25ºC, while temperatures below 20ºC and above 25ºC negatively affected biomass productivity. During the operation of the photobioreactors (PBRs) system without membranes at HRT of 8 days and under favourable environmental conditions, it was possible to comply with effluent nutrient discharge limits established by Directive 98/15/CE (10 mg N·L-1 and 1 mg P·L-1) and to achieve nutrient removal efficiencies of 75.2% of N and 77.9% of P. The MPBR plant allowed decoupling BRT and TRH in the PBRs operation, which resulted in a general improvement of the microalgae cultivation performance and allowed to obtain a solid-free effluent with high potential for reuse applications. The microalgae culture recirculation systems compared in the study (mechanical pumping vs airlift system) did not significantly affect the culture performance. Moreover, reducing the non-photic volume fraction in the MPBR system from 27.2% to 13.6% resulted in an increase of 40% in biomass productivity. A mathematical model of microalgal growth was developed by making use of the data obtained in the pilot plant. This model was able to reproduce accurately (R2 = 0.9954) the overall microalgae cultivation performance in an MPBR system. This model uses the notation and terminology of the ASM models, and it considers a total of 14 components (10 soluble and 4 suspended), 11 processes governed by kinetics and acid-base equilibria to calculate the pH of the medium. In addition, the model considers the effects of solar light and temperature on microalgae growth. As an interesting novelty with respect to other published mathematical models of microalgae growth, this model contemplates the possibility of using the stored polyphosphate for growing in the absence of P in the culture medium. The model developed in this work is intended to be a tool to promote the future implementation of microalgae cultivation technology on full-scale WWTP. / This research was supported by the Spanish Ministry of Economy and Competitiveness (MINECO, Projects CTM2011-28595-C02-01/02, CTM2014-54980-C2-1-R and CTM2014-54980-C2-2-R) jointly with the European Regional Development Fund (ERDF) and Generalitat Valenciana (GVA-ACOMP2013/203), which are gratefully acknowledged. The authors also like to acknowledge the support received from Generalitat Valenciana via one VALi+d post-doctoral grant (APOSTD/2014/049). / Viruela Navarro, A. (2023). Modelación matemática del proceso de crecimiento de microalgas en el tratamiento de aguas residuales Aplicación a un fotobiorreactor de membranas (MPBR) [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/195826 / Compendio

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Fundação Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro / A aplicabilidade de um método selecionado de medição indireta de vitelogenina (Vtg) em plasma sanguíneo de peixe, baseado na quantificação de fosfato álcali-lábil (alkali-labile phosphate-ALP) para acessar estrogenicidade em água, foi investigada na presente tese. O método foi originalmente desenvolvido para a espécie de peixe Carassius carassius (Carpa cruciana) e aplicado pela primeira vez na espécie Oreochromis niloticus (Tilápia do Nilo) no presente estudo. Com o objetivo de acessar a sensibilidade do método, em uma primeira etapa da investigação foram realizados estudos laboratoriais com soluções estoques de 17-ethinylestradiol (EE2), 17-estradiol (E2), e estrona (E1). Os efeitos destes hormônios foram investigados com base tanto na concentração quanto na carga, utilizando-se para tanto, unidades experimentais com volumes distintos (2 L e 130 L). Após a validação do método de ALP, a estrogenicidade foi avaliada nas seguintes águas contaminadas: (i) afluente e efluente de uma grande estação de tratamento de esgotos convencional (ETE) e de uma estação descentralizada de tratamento de esgoto de pequeno porte (Ecossistema Engenheirado-DEE); (ii) água superficial (SW) e água subterrânea (GW) coletadas em uma área de brejo contaminada com gasolina; (iii) água de uma lagoa urbana (LRF) da cidade do Rio de Janeiro, com alta densidade populacional e descarte clandestino de esgoto. Na segunda etapa foram analisados em microalgas os efeitos (outros que não disrupção endócrina) causados pelos hormônios EE2, E2 e E1. Os hormônios foram testados individualmente e em misturas, em culturas individuais e combinada (S+) das espécies de microalgas unicelulares P. subcapitata e D. subspicatus. Com base nos níveis de ALP para a espécie de peixe e no EC50 para as espécies de algas, os resultados mostraram que o EE2 e o E2 causaram disrupção endócrina superior e foram mais tóxicos do que o E1 para peixes e microalgas respectivamente. Quando em misturas (E+) de concentrações equivalentes (EE2:E2:E1), os estrogênios resultaram em efeito aditivo para as espécies O. niloticus e P. subcapitata, e menos que aditivo para D. subspicatus e cultivo misto de algas (S+). Culturas contendo ambas as espécies de algas (S+) por um longo período de exposição (96 h) resultaram na atenuação dos efeitos tóxicos causados pela exposição, tanto individual (EE2, E2 ou E1), quanto na mistura (E+) dos estrogênios, medidos em termos de EC50 (T0h 0,07; 0,09; 0,18; e 0,06 g mL-1; e T96h 1,29; 1,87; 5,58; e 4,61 g mL-1, respectivamente). O DEE apresentou uma maior eficiência na remoção dos disrutores endócrinos do que a ETE convencional. Foi detectada estrogenicidade em amostras da LRF, e de água SW e GW em área brejosa contaminada com gasolina. Os resultados dos ensaios sugerem que as interações (efeitos aditivos ou menos que aditivo) causadas pela mistura dos estrogênios assim como, as interações entre as espécies de algas afetaram o resultado final dos ensaios ecotoxicológicos. Um fator raramente abordado em estudos ecotoxicológicos que foi destacado na presente tese refere-se à importância de considerar não somente a concentração e a dosagem, mas também a carga aplicada e o volume das unidades experimentais. Devido à boa sensibilidade do O. niloticus quando exposto às concentrações relativamente baixas dos estrogênios, a combinação do método de ALP com os biomarcadores auxiliares (particularmente MN) pode ser um protocolo adequado para a detecção de estogenicidade e genotoxicidade respectivamente em diferentes ambiente aquáticos contaminados, como parte de um programa de monitoramento ambiental / The applicability of one selected method for indirect measurement of vitellogenin (Vtg) in fish plasma based on the quantification of alkali-labile phosphates (ALP method) to assess estrogenicity in water was investigated. The ALP method applied in this investigation was originally developed with Carassius carassius (Crucian carps). This thesis describes the first attempt of using this method with Oreochromis niloticus (Nile tilapia). In a first part of the investigation, laboratory studies were conducted with water spiked with 17-ethinylestradiol (EE2), 17-estradiol (E2), and estrone (E1) in order to assess the method sensitivity. The effects of these estrogens were investigated on the basis of both load and concentration, using experimental units with two different volumes (2 L and 130 L). After validation of the method, the estrogenicity of the following contaminated waters was assessed: (i) affluent and effluent of one large conventional municipal wastewater treatment plant (WWTP) and one small decentralized wastewater treatment plant (Decentralized Engineered Ecosystem-DEE); (ii) surface water (SW) and groundwater (GW) obtained from a gasoline-contaminated marshland; (iii) samples from a urban lagoon (LRF) located in Rio de Janeiro city with high density population and clandestine sewage discharge. An additional goal of the thesis was to assess the effect (other than endocrine disruption) caused by EE2, E2 and E1 to microalgae. Assays with single and mixed estrogens and single and combined cultures (S+) of the green microalgae P. subcapitata and D. subspicatus were carried out. The results have shown that EE2 and E2 were more estrogenic and toxic than E1 to the fishes and to the microalgae respectively. Mixed solutions of estrogens (E+) in equal proportions (EE2:E2:E1) resulted in additive effect on O. niloticus and P. subcapitata and less-than-additive effect on D. subspicatus and S+ measured as ALP (for fish) and EC50 (for microalgae). Combined cultivation of both algae species and longer exposure time (96 h) resulted in attenuation of the toxic effects caused by single (EE2, E2 or E1) and mixed (E+) estrogens according to EC50 (T0h 0.07, 0.09, 0.18, and 0.06 g mL-1; and T96h 1.29, 1.87, 5.58, and 4.61 g mL-1, respectively). The decentralized engineered ecosystem was more efficient than the conventional WWTP regarding estrogenicity removal from the final effluent. Estrogenicity was detected in some samples of the LRF and the SW and GW of the gasoline-contaminated marshland. Therefore, the investigations suggested that interactions (additive and less-than additive effect) take place when different estrogens are present in the water environment and interactions also occur between algae species, which affect the final toxicity. Additionally, the study highlighted the importance of taking into account not only concentration and dose regime but also the mass load and therefore, the volume used in the experimental units, which are rarely addressed in ecotoxicity assays. Considering the good sensitivity of O. niloticus exposed to relatively low concentrations of estrogens, the combination of the ALP method with auxiliary biomarkers (particularly MN) can be a suitable protocol for estrogenicity and genotoxicity detection in different contaminated waters as part of water environmental monitoring programs

Fundação Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro / A aplicabilidade de um método selecionado de medição indireta de vitelogenina (Vtg) em plasma sanguíneo de peixe, baseado na quantificação de fosfato álcali-lábil (alkali-labile phosphate-ALP) para acessar estrogenicidade em água, foi investigada na presente tese. O método foi originalmente desenvolvido para a espécie de peixe Carassius carassius (Carpa cruciana) e aplicado pela primeira vez na espécie Oreochromis niloticus (Tilápia do Nilo) no presente estudo. Com o objetivo de acessar a sensibilidade do método, em uma primeira etapa da investigação foram realizados estudos laboratoriais com soluções estoques de 17-ethinylestradiol (EE2), 17-estradiol (E2), e estrona (E1). Os efeitos destes hormônios foram investigados com base tanto na concentração quanto na carga, utilizando-se para tanto, unidades experimentais com volumes distintos (2 L e 130 L). Após a validação do método de ALP, a estrogenicidade foi avaliada nas seguintes águas contaminadas: (i) afluente e efluente de uma grande estação de tratamento de esgotos convencional (ETE) e de uma estação descentralizada de tratamento de esgoto de pequeno porte (Ecossistema Engenheirado-DEE); (ii) água superficial (SW) e água subterrânea (GW) coletadas em uma área de brejo contaminada com gasolina; (iii) água de uma lagoa urbana (LRF) da cidade do Rio de Janeiro, com alta densidade populacional e descarte clandestino de esgoto. Na segunda etapa foram analisados em microalgas os efeitos (outros que não disrupção endócrina) causados pelos hormônios EE2, E2 e E1. Os hormônios foram testados individualmente e em misturas, em culturas individuais e combinada (S+) das espécies de microalgas unicelulares P. subcapitata e D. subspicatus. Com base nos níveis de ALP para a espécie de peixe e no EC50 para as espécies de algas, os resultados mostraram que o EE2 e o E2 causaram disrupção endócrina superior e foram mais tóxicos do que o E1 para peixes e microalgas respectivamente. Quando em misturas (E+) de concentrações equivalentes (EE2:E2:E1), os estrogênios resultaram em efeito aditivo para as espécies O. niloticus e P. subcapitata, e menos que aditivo para D. subspicatus e cultivo misto de algas (S+). Culturas contendo ambas as espécies de algas (S+) por um longo período de exposição (96 h) resultaram na atenuação dos efeitos tóxicos causados pela exposição, tanto individual (EE2, E2 ou E1), quanto na mistura (E+) dos estrogênios, medidos em termos de EC50 (T0h 0,07; 0,09; 0,18; e 0,06 g mL-1; e T96h 1,29; 1,87; 5,58; e 4,61 g mL-1, respectivamente). O DEE apresentou uma maior eficiência na remoção dos disrutores endócrinos do que a ETE convencional. Foi detectada estrogenicidade em amostras da LRF, e de água SW e GW em área brejosa contaminada com gasolina. Os resultados dos ensaios sugerem que as interações (efeitos aditivos ou menos que aditivo) causadas pela mistura dos estrogênios assim como, as interações entre as espécies de algas afetaram o resultado final dos ensaios ecotoxicológicos. Um fator raramente abordado em estudos ecotoxicológicos que foi destacado na presente tese refere-se à importância de considerar não somente a concentração e a dosagem, mas também a carga aplicada e o volume das unidades experimentais. Devido à boa sensibilidade do O. niloticus quando exposto às concentrações relativamente baixas dos estrogênios, a combinação do método de ALP com os biomarcadores auxiliares (particularmente MN) pode ser um protocolo adequado para a detecção de estogenicidade e genotoxicidade respectivamente em diferentes ambiente aquáticos contaminados, como parte de um programa de monitoramento ambiental / The applicability of one selected method for indirect measurement of vitellogenin (Vtg) in fish plasma based on the quantification of alkali-labile phosphates (ALP method) to assess estrogenicity in water was investigated. The ALP method applied in this investigation was originally developed with Carassius carassius (Crucian carps). This thesis describes the first attempt of using this method with Oreochromis niloticus (Nile tilapia). In a first part of the investigation, laboratory studies were conducted with water spiked with 17-ethinylestradiol (EE2), 17-estradiol (E2), and estrone (E1) in order to assess the method sensitivity. The effects of these estrogens were investigated on the basis of both load and concentration, using experimental units with two different volumes (2 L and 130 L). After validation of the method, the estrogenicity of the following contaminated waters was assessed: (i) affluent and effluent of one large conventional municipal wastewater treatment plant (WWTP) and one small decentralized wastewater treatment plant (Decentralized Engineered Ecosystem-DEE); (ii) surface water (SW) and groundwater (GW) obtained from a gasoline-contaminated marshland; (iii) samples from a urban lagoon (LRF) located in Rio de Janeiro city with high density population and clandestine sewage discharge. An additional goal of the thesis was to assess the effect (other than endocrine disruption) caused by EE2, E2 and E1 to microalgae. Assays with single and mixed estrogens and single and combined cultures (S+) of the green microalgae P. subcapitata and D. subspicatus were carried out. The results have shown that EE2 and E2 were more estrogenic and toxic than E1 to the fishes and to the microalgae respectively. Mixed solutions of estrogens (E+) in equal proportions (EE2:E2:E1) resulted in additive effect on O. niloticus and P. subcapitata and less-than-additive effect on D. subspicatus and S+ measured as ALP (for fish) and EC50 (for microalgae). Combined cultivation of both algae species and longer exposure time (96 h) resulted in attenuation of the toxic effects caused by single (EE2, E2 or E1) and mixed (E+) estrogens according to EC50 (T0h 0.07, 0.09, 0.18, and 0.06 g mL-1; and T96h 1.29, 1.87, 5.58, and 4.61 g mL-1, respectively). The decentralized engineered ecosystem was more efficient than the conventional WWTP regarding estrogenicity removal from the final effluent. Estrogenicity was detected in some samples of the LRF and the SW and GW of the gasoline-contaminated marshland. Therefore, the investigations suggested that interactions (additive and less-than additive effect) take place when different estrogens are present in the water environment and interactions also occur between algae species, which affect the final toxicity. Additionally, the study highlighted the importance of taking into account not only concentration and dose regime but also the mass load and therefore, the volume used in the experimental units, which are rarely addressed in ecotoxicity assays. Considering the good sensitivity of O. niloticus exposed to relatively low concentrations of estrogens, the combination of the ALP method with auxiliary biomarkers (particularly MN) can be a suitable protocol for estrogenicity and genotoxicity detection in different contaminated waters as part of water environmental monitoring programs

Tesis por compendio / [ES] Las microalgas son organismos unicelulares fotosintéticos microscópicos que se encuentran en gran variedad de ambientes y son muy eficientes en la transformación de energía solar en biomasa. Los estudios realizados hasta el momento hacen referencia a posibles beneficios de la incorporación de microalgas en la dieta, por la mejora del sistema cardiovascular, las propiedades adelgazantes y energizantes, capacidad antioxidante, o la reducción del colesterol y los triglicéridos. La forma más habitual de consumir las microalgas es como suplemento dietético en forma de tabletas, cápsulas o polvo. La incorporación de biomasa de microalgas en productos tradicionales se ha enfrentado al reto de la aparición de colores verdes fuertes, así como su consistencia pulverulenta que puede afectar la textura y percepción del producto. Todos estos aspectos constituyen las principales áreas de mejora para conseguir un mayor grado de aceptación de productos con microalgas, y son la base del reto de este proyecto. El objetivo de la presente tesis doctoral fue el desarrollo de nuevos productos alimentarios incorporando las propiedades nutricionales de la biomasa de microalgas, incrementando o mejorando con ello, las propiedades nutricionales del alimento original. Para conseguir este objetivo se evaluaron a nivel fisicoquímico, reológico y textural, la incorporación de diferentes especies de microalgas (Arthrospira platensis (Spirulina), Chlorella vulgaris, Dunaliella salina y Nannochloropsis gaditana) en distintas matrices alimentarias (productos horneados, emulsiones y extrusionados). Por otra parte, se evaluó y caracterizó la incorporación de las microalgas utilizando diferentes tecnologías como la impresión 3D o la extrusión. Además de cómo afecta la incorporación de las microalgas a los productos obtenidos, se evaluaron los aspectos nutricionales de su incorporación, en cuanto al aporte de minerales y su biodisponibilidad. Las propiedades reológicas de las masas y emulsiones enriquecidas con microalgas (Spirulina, Chlorella y Dunaliella) indicaron que su comportamiento viscoelástico fue modificado y mejorado, mostrando características aptas para este tipo de productos. La adición de microalgas (Spirulina y Chlorella) a las masas utilizadas para la impresión 3D de galletas y snacks, permitió una mejor extrusión o impresión de éstas, obteniendo muestras impresas en 3D de forma cilíndrica, más precisas en cuanto a sus dimensiones con respecto a la estructura cilíndrica diseñada. Además, las muestras impresas presentaron mayor estabilidad y resistencia, antes y después del proceso de horneado comparadas con la muestra control. Para los productos horneados, tanto para las rosquilletas como los snacks impresos en 3D, la adición de microalgas (Spirulina y Chlorella) permitió mayor estabilidad en términos de textura. Ligeros cambios en los parámetros fisicoquímicos y de expansión se produjeron por la adición de Spirulina y Chlorella en los productos extrusionados. Además, los extrusionados enriquecidos con Nannochloropsis, mostraron parámetros similares a los de la muestra de control. Todos los productos presentaron colores luminosos y apariencias innovadoras y atractivas. En cuanto a los minerales, se observó un aumento de P, K, Ca, Na, Mg, Fe y Se con la adición de Spirulina y Chlorella, junto con el aumento de la concentración adicionada de microalgas. Siguiendo la normativa sobre etiquetado nutricional de los alimentos, el enriquecimiento con microalgas en rosquilletas se puede clasificar como un alimento "rico en hierro (Fe)". De igual forma, las rosquilletas y galletas enriquecidas con microalgas pueden considerarse un alimento "alto en selenio (Se)". Además, la incorporación de Spirulina y Chlorella en las formulaciones de galletas, permitió una mayor bioaccesibilidad del contenido de P, K, Ca, Mg, Fe, Zn y Se para la absorción en el cuerpo comparado con las muestras control. / [CAT] Les microalgues són organismes unicellulars fotosintètics microscòpics. Són molt eficients a l'hora de transformar l'energia solar en biomassa. Els estudis realitzats fins ara fan referència a possibles beneficis de la incorporació de microalgues en la dieta per produir una millora del sistema cardiovascular, per presentar propietats per aprimar i donar energia, per mostrar capacitat antioxidant o per afavorir una reducció del colesterol i els triglicèrids. La forma més habitual de consumir microalgues és com a suplement dietètic en forma de tauleta, càpsula o en pols. La incorporació de biomassa de microalgues en productes tradicionals s'ha afrontat al repte de l'aparició d'un color verd fosc i d'una consistència polsosa que pot afectar a la textura i, per tant, a la percepció del producte. Aquests aspectes constituïxen les principals àrees de millora per aconseguir un major grau d'acceptació de productes amb microalgues i són la base del repte d'aquest projecte. L'objectiu d'aquesta tesi doctoral és el desenvolupament de nous productes alimentaris que incorporen les propietats nutricionals de la biomassa de microalgues, de manera que s'incrementen o es milloren les propietats nutricionals de l'aliment original. Per aconseguir aquest objectiu s'avaluaren a escala fisicoquímica, reològica i de textura la incorporació de diferents espècies de microalgues (Arthrospira platensis (Spirulina), Chlorella vulgaris, Dunaliella salina i Nannochloropsis gaditana) en diferents matrius alimentàries (productes fornejats, emulsions i extrudits). D'altra banda, s'avaluà i caracteritzà la incorporació de les microalgues utilitzant diferents tecnologies com la impressió en 3D o l'extrusió. A banda de valorar com afecta la incorporació de microalgues als productes elaborats, s'avaluaren els aspectes nutricionals, pel que fa a l'aportació i biodisponibilitat de minerals. Les propietats reològiques de les masses i emulsions enriquides amb microalgues (Spirulina, Chlorella i Dunaliella) indicaren que el seu comportament viscoelàstic fou modificat i millorat, de tal manera que mostrà característiques aptes per aquest tipus de productes. L'addició de microalgues (Spirulina i Chlorella) en les masses utilitzades per a la impressió 3D de galetes i snacks permeté una millor impressió, ja que s'obtingueren mostres impreses de forma cilíndrica amb unes dimensions més precises respecte a l'estructura cilíndrica dissenyada. A més, les mostres impreses presentaren una major estabilitat i resistència abans i després del procés de fornejat en comparació amb la mostra control. Respecte als productes fornejats, l'addició de microalgues (Spirulina i Chlorella) a les rosquilletes i els snacks impresos en 3D permeté una major estabilitat en termes de textura. Lleugers canvis als paràmetres fisicoquímics i d'expansió es produïren per l'addició d'Spirulina i Chlorella en els productes extrudits. A més, els extrudits que foren enriquits amb Nannochloropsis mostraren paràmetres similars als de la mostra control. Tots els productes presentaren colors lluminosos i aparences innovadores i atractives. Pel que fa als minerals, s'observà un augment de P, K, Ca, Na, Mg, Fe i Se quan s'afegí Spirulina i Chlorella, directament relacionat amb l'augment de la concentració de microalgues. Seguint la normativa sobre etiquetatge nutricional dels aliments, l'enriquiment amb microalgues en rosquilletes ens permet classificar-les com a aliment "ric en ferro (Fe)". De la mateixa manera, les rosquilletes i galetes enriquides amb microalgues poden considerar-se un aliment "alt en seleni (Se)". A més a més, la incorporació de Spirulina i Chlorella en les formulacions de galetes, permeté una major bioaccessibiltat del contingut de P, K, Ca, Mg, Fe, Zn i Se comparat amb les mostres control. / [EN] Microalgae are microscopic unicellular and photosynthetic organisms that can be found in a wide variety of environments. These microorganisms are very efficient when transforming solar energy into biomass, due to their cellular structure, which is completely submerged in an aqueous medium, forming an adequate surface for the exchange of nutrients and gases. Microalgae compounds are now known to exhibit cardioprotective, immunomodulatory, anti-proliferative, anti-inflammatory, cognitive, neurobehavioral and antimicrobial properties, amongst others. Researchers have shown possible benefits of the incorporation of microalgae in the diet so far. The most common way to consume microalgae is as a dietary supplement in the form of tablets, capsules or powder. The incorporation of microalgae biomass in traditional products has faced the challenge of the appearance of strong green colours, as well as its powdery consistency that can affect the texture and perception of the product. All these aspects constitute the main areas for improvement the development of microalgae-based products, and they are the challenges faced of this project. The main objective of this PhD Thesis was the development of novel food products incorporating the nutritional properties of microalgae biomass, thereby increasing or improving the nutritional properties of the original food matrix. To achieve this goal, the effect of the addition of different species of microalgae (Arthrospira platensis (Spirulina), Chlorella vulgaris, Dunaliella salina and Nannochloropsis gaditana) on the physicochemical, rheological and textural properties of different food matrices (baked products, emulsions and extrudates) was evaluated. Furthermore, the effect of the incorporation of microalgae using different technologies such as 3D printing or extrusion to obtain food products was studied. In addition, how the incorporation of microalgae affects the nutritional aspects of the food products in terms of the contribution of minerals and their bioavailability was also evaluated. The rheological properties of doughs, batters and emulsions enriched with microalgae (Spirulina, Chlorella and Dunaliella) indicated that their viscoelastic behaviour was modified and improved, showing characteristics suitable for this type of products. The addition of microalgae (Spirulina and Chlorella) to the doughs and batters used for the 3D printing of cookies and snacks, allowed a better extrusion or printing behaviour. This allowed obtaining cylindrical 3D printed samples, more precise in terms of their dimensions with respect to the designed cylindrical structure. In addition, the 3D microalgae-printed sample structures presented greater stability and resistance, before and after the baking process compared to the control sample. For baked products, both for breadsticks and 3D printed snacks, the addition of microalgae (Spirulina and Chlorella) allowed greater stability in terms of texture. Slight changes in the physicochemical and expansion parameters were produced by the addition of Spirulina and Chlorella in the extruded products. In addition, the extrudates enriched with Nannochloropsis, showed similar parameters to those of the control sample. Microalgae-enriched obtained products showed bright colours with appealing appearances. Regarding minerals, an increase in P, K, Ca, Na, Mg, Fe and Se was observed with the addition of Spirulina and Chlorella, along the increase of concentration of microalgae addition. Following the regulations on nutrition labelling for food stuffs, breadstick enrichment with microalgae are a food "high in iron (Fe)" In the same way, breadsticks and cookies enriched with microalgae can be considered a "high in selenium (Se)" food. Going a step further, Spirulina and Chlorella vulgaris incorporation in cookie formulations allowed for greater bioaccessibility of P, K, Ca, Mg, Fe, Zn, and Se content for absorption in the body than control cookies. / Uribe Wandurraga, ZN. (2020). Microalgae as novel ingredients for the formulation of food products [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/158743 / Compendio

<p dir="ltr">With the global push for sustainable energy solutions, microalgae have gained attention as a promising biofuel source, with new methods like virus-assisted lipid extraction making algae-based biofuels more cost-effective. This thesis investigated the interaction between <i>Chlorella</i> algae and <i>Paramecium bursaria chlorella virus</i>-1 (PBCV-1) using variations of the SID (Susceptible-Infected-Dead) model to improve algae biofuel production. By leveraging epidemiological modeling, the thesis aimed to understand viral infection dynamics in algae and optimize the viral infection for efficient and effective lipid extraction for biofuels. Two distinct experimental setups were conducted to evaluate how environmental conditions impact infection outcomes. Mathematical models, including SID and SEID (Susceptible-Exposed-Infected-Dead) versions, were calibrated using Bayesian inference and nonlinear optimization to estimate critical parameters such as infection and mortality rates. Key findings reveal that the Susceptible-Infected-Dead model with variable initial infection (SIDVII), which included the initial concentration of infected algae as a model parameter, offered the most reliable fit by balancing biological relevance and statistical performance. In contrast, the SEID-based models, while able to capture a latent infection phase, posed risks of overfitting with too many parameters. The study emphasized the complexity of applying modeling tools to characterize viral infections at scale, suggesting that future research should address potential trade-offs between lipid productivity and infection dynamics. The findings offer potential insights not only for biofuel production but also for broader ecological management strategies, including the targeted control of harmful algal blooms.</p>