Browsing by Author "Machado, Adriana"
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- Development of an organic culture medium for autotrophic production of chlorella vulgaris biomassPublication . Machado, Adriana; Pereira, Hugo; Costa, Margarida; Santos, Tamara; Carvalho, Bernardo; Soares, Maria; Quelhas, Pedro; Silva, Joana T.; Trovão, Mafalda; Barros, Ana; Varela, João; Vicente, António A.; Silva, JoanaMicroalgal biomass has gained increasing attention in the last decade for various biotechnological applications, including human nutrition. Certified organic products are currently a growing niche market in which the food industry has shown great interest. In this context, this work aimed at developing a certified organic culture medium for the production of autotrophic Chlorella vulgaris biomass. A preliminary assay in 2 L bubble column photobioreactors was performed in order to screen di erent commercial organic substrates (OS) at a normalized concentration of N (2 mmol L1). The highest growth performance was obtained using EcoMix4 and Bioscape which showed similar biomass concentrations compared to the synthetic culture medium (control). In order to meet the nutrient needs of Chlorella, both OS underwent elemental analyses to assess their nutrient composition. The laboratory findings allowed the development of a final organic culture medium using a proportion of Bioscape/EcoMix4 (1:1.2, m/m). This organic culture medium was later validated outdoors in 125 L flat panel and 10 m3 tubular flow through photobioreactors. The results obtained revealed that the developed organic medium led to similar microalgal growth performance and biochemical composition of produced biomass, as compared to the traditional synthetic medium. Overall, the formulated organic medium was e ective for the autotrophic production of organic C. vulgaris biomass.
- Growth performance, biochemical composition and sedimentation velocity of Tetraselmis sp. CTP4 under different salinities using low-cost lab- and pilot-scale systemsPublication . Trovão, Mafalda; Pereira, Hugo; Silva, Joana; Páramo, Jaime; Quelhas, Pedro; Santos, Tamara; Silva, Joana T.; Machado, Adriana; Gouveia, Luísa; Barreira, Luísa; Varela, JoãoBiomass harvesting is one of the most expensive steps of the whole microalgal production pipeline. Therefore, the present work aimed to understand the effect of salinity on the growth performance, biochemical composition and sedimentation velocity of Tetraselmis sp. CTP4, in order to establish an effective low-cost pilot-scale harvesting system for this strain. At lab scale, similar growth performance was obtained in cultures grown at salinities of 5, 10 and 20 g L-1 NaCl. In addition, identical settling velocities (2.4-3.6 cm h-1) were observed on all salinities under study, regardless of the growth stage. However, higher salinities (20 g L-1) promoted a significant increase in lipid contents in this strain compared to when this microalga was cultivated at 5 or 10 g L-1 NaCl. At pilot-scale, cultures were cultivated semi-continuously in 2.5-m3 tubular photobioreactors, fed every four days, and stored in a 1-m3 harvesting tank. Upon a 24-hour settling step, natural sedimentation of the microalgal cells resulted in the removal of 93% of the culture medium in the form of a clear liquid containing only vestigial amounts of biomass (0.07 ± 0.02 g L-1 dry weight; DW). The remaining culture was recovered as a highly concentrated culture (19.53 ± 4.83 g L-1 DW) and wet microalgal paste (272.7 ± 18.5 g L-1 DW). Overall, this method provided an effective recovery of 97% of the total biomass, decreasing significantly the harvesting costs.
- Heterotrophic cultivation of Chlorella vulgaris yellow mutant on sidestreams: Medium formulation and process scale-upPublication . Trovão, Mafalda; Barros, Ana; Machado, Adriana; Reis, Ana; Pedroso, Humberto; Espírito Santo, Gonçalo; Correia, Nádia; Costa, Monya; Ferreira, Sara; Varela, João; Cardoso, Helena; Silva, Joana; Pereira, Hugo; Freitas, FilomenaMicroalgal protein is a promising feedstock to complement and/or replace other protein sources. Besides requiring less land and water usage, microalgae production is a more sustainable process, especially if industrial sidestreams are used as nutrient sources. Additionally, the heterotrophic cultivation of microalgae, such as Chlorella vulgaris, enables the achievement of much higher biomass productivity and lower areal footprint than autotrophic cultivation. Chlorophyll-deficient strains of C. vulgaris, as the yellow strain 7Y, provide microalgal biomass with improved sensory properties. In line with this, a waste-based medium was formulated to cultivate this strain, aiming at maximum biomass productivity. In this context, several industrial sidestreams were screened, and two food wastes and corn molasses were selected for their high nitrogen and glucose concentrations, respectively. The waste-based medium formulated was compared to the inorganic optimised medium at laboratory scale in Erlenmeyer flasks and 7-L reactors. The results obtained in the 7-L fermenters revealed that both conditions achieved similar biomass productivities and growth rates of approximately 14 g L-1 d-1 and 0.8 d-1, respectively. The biomass and protein productivities were further enhanced by supplying a higher nitrogen concentration in the feeding solution when the process was scaled-up to 200-L reactors, reaching 22 and 6.3 g L-1 d-1, respectively, thus validating the developed industrial waste-based medium for the efficient cultivation of C. vulgaris under heterotrophic conditions.
- Industrial production of Phaeodactylum tricornutum for CO2 mitigation: biomass productivity and photosynthetic efficiency using photobioreactors of different volumesPublication . Quelhas, Pedro M.; Trovao, Mafalda; Silva, Joana T.; Machado, Adriana; Santos, Tamara; Pereira, Hugo; Varela, João; Simoes, Manuel; Silva, Joana L.The photosynthetic efficiency (PE) and potential of Phaeodactylum tricornutum for CO2 mitigation in industrial tubular photobioreactors (PBRs) of different volumes were evaluated. A preliminary assay was performed at lab-scale to optimize the salt concentration of the culture medium. Interestingly, salinity did not affect the growth of P. tricornutum at concentrations of 2.5, 5, 10, and 20 g L-1. Higher volumetric productivities were achieved in the 2.5-m(3) tubular PBR (0.235 g L-1 day(-1)), followed by 35- and 10-m(3) PBRs. Maximum areal productivities corresponded to 48.5, 45.0, and 12.8 g m(-2) day(-1) for the 35-, 10-, and 2.5-m(3) PBRs, respectively. PE was thus higher in the 35- and 10-m(3) PBRs (2.21 and 2.08%, respectively). The 10- and 35-m(3) PBR showed CO2 mitigation efficiencies of 60 and 41%, respectively, of the CO2 introduced into the PBR, corresponding to 2.3 and 2.5 g of fixed CO2 per g of biomass. Overall, cultivation of P. tricornutum couples high PE and areal productivity when the industrial PBRs were used, particularly PBRs of larger volumes. This improved PE performance with larger PBR volumes strongly suggests that large-scale cultivation of this diatom holds great potential for industrial CO2 mitigation.
- Isolation and characterization of novel chlorella vulgaris mutants with low chlorophyll and improved protein contents for food applicationsPublication . Lisa Schueler, Lisa M. Schueler; Morais, Etiele Greque de; Trovão, Mafalda; Machado, Adriana; Carvalho, Bernardo; Carneiro, Mariana; Maia, Inês; Soares, Maria; Duarte, Paulo; Barros, Ana; Pereira, Hugo; Silva, Joana; Varela, JoãoMicroalgae are widely used as food supplements due to their high protein content, essential fatty acids and amino acids as well as carotenoids. The addition of microalgal biomass to food products (e.g., baked confectioneries) is a common strategy to attract novel consumers. However, organoleptic factors such as color, taste and smell can be decisive for the acceptability of foods supplemented with microalgae. The aim of this work was to develop chlorophyll-deficient mutants of Chlorella vulgaris by chemically induced random mutagenesis to obtain biomass with different pigmentations for nutritional applications. Using this strategy, two C. vulgaris mutants with yellow (MT01) and white (MT02) color were successfully isolated, scaled up and characterized. The changes in color of MT01 and MT02 mutant strains were due to an 80 and 99% decrease in their chlorophyll contents, respectively, as compared to the original wild type (WT) strain. Under heterotrophic growth, MT01 showed a growth performance similar to that of the WT, reaching a concentration of 5.84 and 6.06 g L-1, respectively, whereas MT02 displayed slightly lower growth (4.59 g L-1). When grown under a light intensity of 100 μmol m-2 s-1, the pigment content in MT01 increased without compromising growth, while MT02 was not able to grow under this light intensity, a strong indication that it became light-sensitive. The yellow color of MT01 in the dark was mainly due to the presence of the xanthophyll lutein. On the other hand, phytoene was the only carotenoid detected in MT02, which is known to be colorless. Concomitantly, MT02 contained the highest protein content, reaching 48.7% of DW, a 60% increase as compared to the WT. MT01 exhibited a 30% increase when compared to that of the WT, reaching a protein content of 39.5% of DW. Taken together, the results strongly suggest that the partial abrogation of pigment biosynthesis is a factor that might promote higher protein contents in this species. Moreover, because of their higher protein and lower chlorophyll contents, the MT01 and MT02 strains are likely candidates to be feedstocks for the development of novel, innovative food supplements and foods.
- Oxyfluorfen: a novel metabolic inhibitor to select microalgal chlorophyll-deficient mutant strains for nutritional applicationsPublication . Trovão dos Santos, Mafalda; Cardoso, Lucas; Schüler, Lisa; Machado, Adriana; Santo, Gonçalo Espírito; Pedroso, Humberto; Reis, Ana; Barros, Ana; Correia, Nádia; Costa, Monya; Ferreira, Sara; Cardoso, Helena; Mateus, Marília; Silva, Joana; Pereira, Hugo; Freitas, Filomena; Varela, JoãoNowadays, there is an increasing demand for novel feedstocks and alternative protein sources to meet global needs. Because of their rich nutritional profiles and high protein contents, microalgae-based food products and supplements are being developed. Nonetheless, these products present organoleptic characteristics such as taste, smell and colour that are often considered unpleasant by human and animal consumers. To address this constraint, strain improvement approaches such as random mutagenesis have been used, which combined with the right selection strategy, lead to more appealing microalgal biomass. In this work, a novel selection strategy using oxyfluorfen, an inhibitor of the chlorophyll synthesis pathway, was applied for the first time to isolate chlorophyll-deficient strains of Scenedesmus rubescens and Chlorella vulgaris upon treatment with ethyl methanesulfonate (EMS). With this approach, one S. rubescens brownish (37Y01) mutant strain, as well as two C. vulgaris mutant strains, one yellow (31Y15) and one white (31W62), were obtained. S. rubescens 37Y01 displayed a reduced protein content of 19.1% dry weight (DW) compared to that of the wildtype, which presented a protein content of 25.0% DW. C. vulgaris wildtype and mutants exhibited higher protein contents, in the 42.844.3% DW range, compared to Scenedesmus rubescens (p < 0.05). The selective pressure of this inhibitor allowed the selection of S. rubescens and C. vulgaris mutants displaying 55% and 95% decrease in chlorophyll content, respectively. The reduced chlorophyll content greatly improves the sensory properties and consumer acceptance of established mutants, increasing the potential of both strains as feedstocks to develop novel food products.
- Random mutagenesis as a promising tool for microalgal strain improvement towards industrial productionPublication . Trovão Dos Santos, Mafalda; Schüler, Lisa M.; Machado, Adriana; Bombo, Gabriel; Navalho, Sofia; Barros, Ana; Pereira, Hugo; Silva, Joana; Freitas, Filomena; Varela, JoãoMicroalgae have become a promising novel and sustainable feedstock for meeting the rising demand for food and feed. However, microalgae-based products are currently hindered by high production costs. One major reason for this is that commonly cultivated wildtype strains do not possess the robustness and productivity required for successful industrial production. Several strain improvement technologies have been developed towards creating more stress tolerant and productive strains. While classical methods of forward genetics have been extensively used to determine gene function of randomly generated mutants, reverse genetics has been explored to generate specific mutations and target phenotypes. Site-directed mutagenesis can be accomplished by employing different gene editing tools, which enable the generation of tailor-made genotypes. Nevertheless, strategies promoting the selection of randomly generated mutants avoid the introduction of foreign genetic material. In this paper, we review different microalgal strain improvement approaches and their applications, with a primary focus on random mutagenesis. Current challenges hampering strain improvement, selection, and commercialization will be discussed. The combination of these approaches with high-throughput technologies, such as fluorescence-activated cell sorting, as tools to select the most promising mutants, will also be discussed.