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- Optimisation of biomass production and nutritional value of two marine diatoms (Bacillariophyceae), Skeletonema costatum and Chaetoceros calcitransPublication . Bastos, Carolina R. V.; Maia, Inês Beatriz; Pereira, Hugo; Navalho, João; Varela, JoãoS. costatum and C. calcitrans are two cosmopolitan high-value centric diatoms, with a rich nutritional profile. The following work optimised the culture medium of S. costatum and C. calcitrans cultures, respectively, in a stepwise process as follows: 2.4 mM and 1.2 mM of silicate, 4 mM of nitrate, 100 µM of phosphate, 20 and 80 µM iron, and 0.5 mL L−1 of micronutrients. The results that were obtained revealed an increase in biomass productivity with a 1.8- and 3.2-fold increase in biomass that was produced by S. costatum and C. calcitrans, respectively. The biochemical profile showed an increase in high-value PUFAs such as 2.6-fold and 2.3-fold increase in EPA for S. costatum and C. calcitrans, respectively, whilst a 2.6-fold increase in DHA was detected in S. costatum cultures. The present work provides the basic tools for the industrial cultivation of S. costatum and C. calcitrans with enhanced productivity as well as improved biomass quality, two factors which are highly relevant for a more effective application of these diatoms to aquaculture and nutraceutical production.
- Effects of LED lighting on Nannochloropsis oceanica grown in outdoor raceway pondsPublication . Carneiro, M.; Maia, Inês Beatriz; Cunha, P.; Guerra, I.; Magina, T.; Santos, Tamara; Schulze, Peter S.C.; Pereira, H.; Malcata, F. X.; Navalho, J.; Silva, J.; Otero, A.; Varela, JoãoGrowth in most microalgal mass cultivation systems is light-limited, particularly in raceway ponds (RWP) where the light path is higher. Artificial lighting can be a promising solution to diminishing dark zones and enhance microalgal productivity. Therefore, our goal was to prevent the cell shift from photosynthesis to a respiration-only stage by resorting to LED illumination. Nannochloropsis oceanica cultures were accordingly grown out-doors in a preliminary small-scaleexperiment, followed by pilot-scale trials. In the former, three 3.0-m(2) RWP were set up under three distinct conditions: 1) without LEDs (control); 2) LEDs turned on during the night; and 3) LEDs turned on for 24 h. In the pilot-scale trial, one of two 28.9-m(2) pilot-scale RWPs was coupled to the best LED setup - determined in the small-scale preliminary experiment - using the same light intensity (normal mode) and half of the intensity (economy mode), with the second RWP serving as a control. In the preliminary experiment, the use of LEDs for 24 h was deemed as not helpful during daytime, before the culture reached asymptotic to 0.5 g DW L-1 - when dark zones appeared during the day due to sunlight attenuation in the 0.1 m-deep cultures. Overall, use of LEDs increased biomass growth chiefly by increasing nighttime productivities - materialized in higher chlorophyll, protein, and carbohydrate productivities in LED-lit cultures. A higher impact of LED lighting was observed under lower sunlight irradiances. A preliminary economic analysis indicates that use of LEDs in RWPs outdoors should be considered for high-value metabolites only.
- Drying microalgae using an industrial solar dryer: a biomass quality assessmentPublication . Schmid, Benjamin; Navalho, Sofia; Schulze, Peter S.C.; Van De Walle, Simon; Van Royen, Geert; Schüler, Lisa M.; Maia, Inês Beatriz; Bastos, Carolina; Baune, Marie-Christin; Januschewski, Edwin; Coelho, Ana; Pereira, Hugo; Varela, João; Navalho, João; Rodrigues, Alexandre Miguel CavacoMicroalgae are considered a promising resource of proteins, lipids, carbohydrates, and other functional biomolecules for food and feed markets. Competitive drying solutions are required to meet future demands for high-quality algal biomass while ensuring proper preservation at reduced costs. Since often used drying methods, such as freeze or spray drying, are energy and time consuming, more sustainable processes remain to be developed. This study tested an indirect and hybrid solar dryer as an alternative to conventional freeze drying of industrially produced Tetraselmis chui and Nannochloropsis oceanica wet paste. The effects of the drying method on biomass quality parameters, including biochemical profiles, functional properties, and microbial safety, were assessed. No significant differences were found between the applied drying technologies for total proteins, carbohydrates, lipids, and fatty acid profiles. On the other hand, some pigments showed significant differences, displaying up to 44.5% higher contents in freeze-dried samples. Minor differences were also registered in the mineral profiles (<10%). Analyses of microbial safety and functional properties of the solar-dried biomass appear adequate for food and feed products. In conclusion, industrial solar drying is a sustainable technology with a high potential to preserve high-quality microalgal biomass for various markets at expected lower costs.
- Diel biochemical and photosynthetic monitorization of Skeletonema costatum and Phaeodactylum tricornutum grown in outdoor pilot-scale flat panel photobioreactorsPublication . Maia, Inês Beatriz; Carneiro, Mariana; Magina, Tânia; Malcata, F. Xavier; Otero, Ana; Navalho, João; Varela, João; Pereira, HugoDiatoms are currently considered valuable feedstocks for different biotechnological applications. To deepen the knowledge on the production of these microalgae, the diel pattern of batch growth, photosystem II performance, and accumulation of target metabolites of two commercially relevant diatoms, Phaeodactylum tricornutum and Skeletonema costatum, were followed outdoors in 100-L flat panel photobioreactors. S. costatum presented a higher light-to-biomass conversion resulting in higher growth than P. tricornutum. Both fluorescence data and principal component analysis pointed to temperature as a limiting factor for the growth of P. tricornutum. Higher protein and carbohydrate contents were found in P. tricornutum, whereas S. costatum fatty acids were charac-terized by a higher unsaturation degree. Higher productivities were found at 1 p.m. for protein, lipid, and ash in the case of S. costatum. Overall, S. costatum showed great potential for outdoor cultivation, revealing a broader temperature tolerance and increased biomass productivity than P. tricornutum.
- Chrysotila pseudoroscoffensis as a source of high-value polar lipids with antioxidant activity: A lipidomic approachPublication . Moreira, Ana S. P.; Gonçalves, Joana; Conde, Tiago A.; Couto, Daniela; Melo, Tânia; Maia, Inês Beatriz; Pereira, Hugo; Silva, Joana; Domingues, M. Rosário; Nunes, CláudiaMicroalgae are emerging as sustainable sources of a wide range of high-value compounds. However, the knowledge about microalgae polar lipids is still limited, despite their interest due to their chemical diversity and bioactivity. This study shows, for the first time, the polar lipidome of the haptophyte microalga Chrysotila pseudoroscoffensis unveiled by using hydrophilic interaction liquid chromatography-high resolution mass spectrometry (HILIC-HRMS and MS/MS) and gas chromatography-mass spectrometry (GC-MS). Freeze-dried C. pseudoroscoffensis biomass has a lipid content of 6.4 %, containing higher amounts of ash (45.5 %), proteins (11.6 %), and sugars (11.0 %). Uronic acids (53.8 mol%) are the sugars present in higher content, followed by glucose (13.7 mol%) and galactose (12.7 mol%). The polar lipid species identified by HILIC-MS/MS included betaine lipids, glycolipids, and phospholipids, some of them with recognised bioactive properties. Among the lipid classes found from C. pseudoroscoffensis, some are less reported in algae: betaine lipids diacylglycerylcarboxyhydroxymethylcholine (DGCC) and monoacylglycerylcarboxyhydroxymethylcholine (MGCC) (characteristic of haptophyte microalgae); acid glycolipid class glucuronosyldiacylglycerol (GlcADG) (mainly reported in plants with protective effects in phosphate-deprivation conditions); and phospholipid classes monomethylphosphatidylethanolamine (MMPE) and lysomonomethylphosphatidylethanolamine (MMLPE). As estimated by colorimetric assays, glycolipids and phospholipids accounted for 64 and 3 % of the total lipid extracts, respectively. Fatty acid profiling by GC-MS showed that total esterified fatty acids accounted for about 32 % of the total lipid extracts, of which 23 % were omega-3 polyunsaturated fatty acids (PUFA). Four lipid extract concentrations (12.5, 62.5, 125 and 250 mu g mL-1 in ethanol) were tested and displayed antioxidant capacity toward 2,2 & PRIME;-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals. A dose-dependent behaviour was observed with IC50 of 111.9 mu g mL(-1) for ABTS and IC35 of 234.8 mu g mL(-1) for DPPH assay. In conclusion, the lipid extracts of C. pseudoroscoffensis may be a source of high-value lipids for the development of novel microalgae-based products, namely nutraceuticals and cosmeceuticals.
- Fucoxanthin production from Tisochrysis lutea and Phaeodactylum tricornutum at industrial scalePublication . Pereira, Hugo; Sá, Marta; Maia, Inês Beatriz; Rodrigues, Alexandre; Teles, Iago; Wijffels, Rene H.; Navalho, João; Barbosa, MariaFucoxanthin is a xanthophyll carotenoid with high market value. Currently, seaweeds are the primary source for fucoxanthin industrial production. However, marine microalgae reach 5 to 10 times higher concentrations (2.24 to 26.6 mg g-1 DW) and are considered a promising feedstock. In this work, two marine microalgae were produced at industrial scale to evaluate biomass and fucoxanthin production: Phaeodactylum tricornutum for autumn/winter and Tisochrysis lutea for spring/summer. Both strains were grown in 15 m3 tubular flow-through photobioreactors; for 170 consecutive days of semi-continuous cultivation regime. The average volumetric biomass productivities of P. tricornutum and T. lutea were 0.11 and 0.09 g DW L-1 day-1. P. tricornutum reached higher maximum biomass concentration (2.87 g DW L-1) than T. lutea (1.47 g DW L-1). P. tricornutum fucoxanthin content ranged between 0.2 and 0.7% DW, while T. lutea between 0.2 and 0.6% DW. The fucoxanthin content was correlated with the irradiation (MJ m-2) and biomass concentration in the photobioreactor (g L-1). This is the first work in literature reporting a long-term industrial production of T. lutea. Overall, we showed possible scenarios for fucoxanthin production from microalgae, increasing the window to supply the industry with steady production throughout the year.
- In situ monitoring of chlorophyll a fluorescence in Nannochloropsis oceanica cultures to assess photochemical changes and the onset of lipid accumulation during nitrogen deprivationPublication . Carneiro, Mariana; Chini Zittelli, Graziella; Cicchi, Bernardo; Touloupakis, Eleftherios; Faraloni, Cecilia; Maia, Inês Beatriz; Pereira, Hugo; Santos, Tamára; Malcata, Francisco X.; Otero, Ana; Varela, João; Torzillo, GiuseppeIn situ chlorophyll a fluorescence measurements were applied to monitor changes in the photochemical variables of Nannochloropsis oceanica cultures under nitrogen-deplete and nitrogen-replete (control) conditions. In addition, growth, lipid, fatty acid, and pigment contents were also followed. In the control culture, growth was promoted along with pigment content, electron transport rate (ETR), and polyunsaturated fatty acids, while total lipid content and fatty acid saturation level diminished. Under nitrogen-deplete conditions, the culture showed a higher de-epoxidation state of the xanthophyll cycle pigments. Fast transients revealed a poor processing efficiency for electron transfer beyond Q(A), which was in line with the low ETR due to nitrogen depletion. Lipid content and the de-epoxidation state were the first biochemical variables triggered by the change in nutrient status, which coincided with a 20% drop in the in situ effective quantum yield of PSII (Delta F'/F-m'), and a raise in the V-j measurements. A good correlation was found between the changes in Delta F'/F-m' and lipid content (r = -0.96, p < 0.01). The results confirm the reliability and applicability of in situ fluorescence measurements to monitor lipid induction in N. oceanica.
- Carotenoid biosynthetic gene expression, pigment and n-3 fatty acid contents in carotenoid-rich Tetraselmis striata CTP4 strains under heat stress combined with high lightPublication . Schüler, Lisa Maylin; Bombo, Gabriel; Duarte, Paulo; Santos, Tamara; Maia, Inês Beatriz; Pinheiro, Filipa; Marques, José; Jacinto, Rita; Schulze, Peter S.C.; Pereira, Hugo; Barreira, Luísa; Varela, JoãoIn this study, two carotenoid-rich strains of the euryhaline microalga Tetraselmis striata CTP4 were isolated by random mutagenesis combined with selection via fluorescence activated cell sorting and growth on norflurazon. Both strains, ED5 and B11, showed an up to 1.5-fold increase in carotenoid contents as compared with the wildtype, independent of the growth conditions. More specifically, violaxanthin, beta-carotene and lutein contents reached as high as 1.63, 4.20 and 3.81 mg g-1 DW, respectively. Genes coding for phytoene synthase, phytoene desaturase, lycopene-beta-cyclase and epsilon-ring hydroxylase involved in carotenoid biosynthesis were found to be upregulated in ED5 and B11 cells as compared to the wildtype. Both strains showed higher contents of eicosapentaenoic acid as compared with those of the wildtype, reaching up to 4.41 and 2.88 mg g-1 DW, respectively. Overall, these results highlight the complexity of changes in carotenoid biosynthesis regulation that are required to improve pigment contents in microalgae.
- Microalgal systems for wastewater treatment: technological trends and challenges towards waste recoveryPublication . Morais, Etiele; Cristofoli, N.L.; Maia, Inês Beatriz; Magina, Tânia; Cerqueira, Paulo R.; Teixeira, Margarida Ribau; Varela, João; Barreira, Luísa; Gouveia, LuisaWastewater (WW) treatment using microalgae has become a growing trend due the economic and environmental benefits of the process. As microalgae need CO2, nitrogen, and phosphorus to grow, they remove these potential pollutants from wastewaters, making them able to replace energetically expensive treatment steps in conventional WW treatment. Unlike traditional sludge, biomass can be used to produce biofuels, biofertilizers, high value chemicals, and even next-generation growth media for “organically” grown microalgal biomass targeting zero-waste policies and contributing to a more sustainable circular bioeconomy. The main challenge in this technology is the techno-economic feasibility of the system. Alternatives such as the isolation of novel strains, the use of native consortia, and the design of new bioreactors have been studied to overcome this and aid the scale-up of microalgal systems. This review focuses on the treatment of urban, industrial, and agricultural wastewaters by microalgae and their ability to not only remove, but also promote the reuse, of those pollutants. Opportunities and future prospects are discussed, including the upgrading of the produced biomass into valuable compounds, mainly biofuels.
- Bioprospecting for industrially relevant exopolysaccharide-producing cyanobacteria under portuguese simulated climatePublication . Cruz, José Diogo; Delattre, Cédric; Felpeto, Aldo Barreiro; Pereira, Hugo; Pierre, Guillaume; Morais, João; Petit, Emmanuel; Silva, Joana; Azevedo, Joana; Elboutachfaiti, Redouan; Maia, Inês Beatriz; Dubessay, Pascal; Michaud, Philippe; Vasconcelos, VitorCyanobacterial exopolysaccharides (EPS) are potential candidates for the production of sustainable biopolymers. Although the bioactive and physicochemical properties of cyanobacterial-based EPS are attractive, their commercial exploitation is limited by the high production costs. Bioprospecting and characterizing novel EPS-producing strains for industrially relevant conditions is key to facilitate their implementation in various biotechnological applications and fields. In the present work, we selected twenty-five Portuguese cyanobacterial strains from a diverse taxonomic range (including some genera studied for the first time) to be grown in diel light and temperature, simulating the Portuguese climate conditions, and evaluated their growth performance and proximal composition of macronutrients. Synechocystis and Cyanobium genera, from marine and freshwater origin, were highlighted as fast-growing (0.1-0.2 g L-1 day(-1)) with distinct biomass composition. Synechocystis sp. LEGE 07367 and Chroococcales cyanobacterium LEGE 19970, showed a production of 0.3 and 0.4 g L-1 of released polysaccharides (RPS). These were found to be glucan-based polymers with high molecular weight and a low number of monosaccharides than usually reported for cyanobacterial EPS. In addition, the absence of known cyanotoxins in these two RPS producers was also confirmed. This work provides the initial steps for the development of cyanobacterial EPS bioprocesses under the Portuguese climate.