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Abstract

Large-scale production of energetic storage compounds by microalgae is hampered by competition and evolution. Both phenomena result in contamination and arise due to a mismatch between the desired productive microalgal strain and the constructed environment. The prevailing approach to solve this issue involves increasing the survival potential of the desired strain, for example by working in closed systems or at extreme conditions. We advocate adjusting the environment in such a way that lipid production, or any other desired characteristic, gives a competitive advantage. Competition and evolution become a value rather than a threat to processes in which the desired characteristic is ensured by a selective environment. Research and cultivation efforts will benefit from this approach as it harnesses the microalgal diversity in nature.

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... However, maintaining a large-scale axenic phototrophic system is expensive and cumbersome [48]. Instead, we propose to grow microalgae in an open system and to use selective pressure to enrich algae with a high storage compound production capacity, as described by Mooij et al. [49]. In their approach, Mooij et al. [49] subjected an open algal culture in a photobioreactor to a cyclical light/dark regimen. ...
... Instead, we propose to grow microalgae in an open system and to use selective pressure to enrich algae with a high storage compound production capacity, as described by Mooij et al. [49]. In their approach, Mooij et al. [49] subjected an open algal culture in a photobioreactor to a cyclical light/dark regimen. During the light period carbon dioxide was supplied, while nitrogen was supplied only during the dark period. ...
... A separation step after the anaerobic digester should prevent solids from entering the phototrophic system; otherwise, the turbidity in the tank would become too high and prevent light from reaching the algal biofilm. the polyglucose level in the biomass produced by Mooij et al. [49] was comparable with the highest reported values for pure cultures (57 AE 2% of volatile suspended solids). ...
Article
Microalgal biomass can be converted to biofuels to replace nonsustainable fossil fuels, but the widespread use of microalgal biofuels remains hampered by the high energetic and monetary costs related to carbon dioxide supply and downstream processing. Growing microalgae in mixed culture biofilms reduces energy demands for mixing, maintaining axenic conditions, and biomass concentration. Furthermore, maintaining a high pH improves carbon dioxide absorption rates and inorganic carbon solubility, thus overcoming the carbon limitation and increasing the volumetric productivity of the microalgal biomass. Digesting the microalgal biomass anaerobically at high pH results in biogas that is enriched in methane, while the dissolved carbon dioxide is recycled to the phototrophic reactor. All of the required haloalkaline conversions are known in nature.
... Production of bulk compounds, such as triacylglycerides, at an industrial scale is however troublesome in an axenic microalgal culture (Kazamia et al. 2012;Shurin et al. 2013;McBride et al. 2014). Previously, we described an ecology-based enrichment and cultivation method which allows for stable storage compound production under nonaxenic conditions (Mooij et al. 2013(Mooij et al. , 2015. The basis of this approach is uncoupling of carbon fixation in the light and nitrogen uptake in the dark by limiting the presence of an essential growth nutrient (such as nitrate) to the dark phase. ...
... Limiting the presence of nitrate to the dark period provides a competitive advantage for storage compound-producing microalgae. Microalgae that efficiently convert CO 2 into storage compounds in the light phase can take up the nitrate in the dark phase for biomass production, resulting in an enrichment culture consisting of efficient storage compound-producing microalgae (Mooij et al. 2013). ...
... Apparently, Chlamydomonas and Nitzschia outcompete Chlorella if carbon fixation and nitrate uptake are uncoupled. In previous work, Chlorella luteoviridis dominated the system throughout the experiment (Mooij et al. 2013). Operational differences between the previous and current work, such as the nitrogen source used and the solid retention time, could explain the disappearance of C. luteoviridis under the conditions applied in this experiment. ...
Article
Microalgae have the potential to supply a biobased society with essential feedstocks like sugar and lipids. Besides being productive, strains used for this purpose should grow fast, be resistant to predators, and have good harvestability properties. Diatoms, a class of siliceous algae, have these and other preferred characteristics. In this paper, we describe the enrichment of microalgae in sequencing batch reactors with and without supply of silicate. Both reactors were operated with a light–dark cycle. To maximize storage compound production, carbon fixation and nitrogen uptake were uncoupled by limiting the availability of nitrate to the dark phase. After ten cycles, a stable culture was established in both reactors. The diatom Nitzschia sp. dominated the silicate-rich reactor, and the green algae Chlamydomonas sp. dominated the silicate-depleted reactor. Over the remaining 27 cycles of the experiment, the microalgal community structure did not change, indicating a highly stable system. Although the dominant microalga was highly dependent on the presence of silicate, the performance of both microalgal enrichments was similar. Polymers of glucose were stored during the nitrogen-limited light period. On organic matter dry weight basis, the sugar content of the biomass increased during the light period from 17 ± 4 to 53 ± 4 % for the silicate-limited culture, and from 14 ± 4 to 43 ± 4 % (w w −1) for the silicate excess culture. These results show that storage compound production can be achieved under various conditions, as long as a selective environment is maintained.
... Selecting an appropriate species of microalgae is the first and most important step through the whole process of production of storage compounds (Mooij et al., 2013;Talebi et al., 2014). This selection is based on high storage capacity or high productivity. ...
... Cultivation of microalgae in pure culture will require an aseptic process condition resulting in a high investment and operating cost. Such preparations present serious barriers to large scale application (Mooij et al., 2013). In addition, even in specific environmental conditions such as high salinity for Dunaliella Salina cultivation, where contamination likelihood is low, strain evolution remains as a great threat, since mutated strains with higher growth rate are able to outcompete the desired strain leading to a drop in productivity (Mooij et al., 2015). ...
... As an alternative to pure culture, the application of a mixed-culture-based process would be a technically attractive and an economically feasible solution (Reis et al., 2003). Screening mixed culture based on ecological role of product, would be a good solution to overcome cultivation contamination risk (Mooij et al., 2013). Exploring microbial diversity and operation in various conditions are the main advantages of using mixed culture under selective environment (Johnson et al., 2009;Kleerebezem and van Loosdrecht, 2007). ...
... In these conditions the accumulation ability for lipids or polysaccharides is not a competitive advantage. Therefore, when microorganisms with high accumulation ability are cultivated in culture medium with high nutrient concentration, culture contaminations by unproductive cells (low accumulation ability, but higher grow rate) can rapidly replace the productive strains (Chubukov et al., 2017;Di Caprio et al., 2019;Mooij et al., 2015bMooij et al., , 2013. In this work the word "contaminant" is used to indicate every microbial cell, in the culture medium, with an accumulation ability lower than the target strain cultivated. ...
... These processes are usually based on the alternance of periods in which substrate (energy/C source) is available, named feast phase, and periods in which substrate is no longer available, named famine phase. The aim of these processes is pushing towards the survival of the "fattest" strains in place of the survival of the sole fastest strains (> μ max ) (Mooij et al., 2015b(Mooij et al., , 2013, otherwise obtained with culture medium fully replete in every nutrient. ...
... The general approach used is to alternate an energy replete condition with an energy starvation condition, to select those strains with more accumulated storage compounds (Di Caprio et al., 2019;Mooij et al., 2015b;Oliveira et al., 2017). This concept can be synthetized as the survival of the "fattest" (Mooij et al., 2015b(Mooij et al., , 2013 in place of the common survival of the "fastest" (Fig. 2). ...
Article
The microbial ability to accumulate biomolecules is fundamental for different biotechnological applications aiming at the production of biofuels, food and bioplastics. However, high accumulation is a selective advantage only under certain stressful conditions, such as nutrient depletion, characterized by lower growth rate. Conventional bioprocesses maintain an optimal and stable environment for large part of the cultivation, that doesn't reward cells for their accumulation ability, raising the risk of selection of contaminant strains with higher growth rate, but lower accumulation of products. Here in this work the physiological responses of different microorganisms (microalgae, bacteria, yeasts) under N-starvation and energy starvation are reviewed, with the aim to furnish relevant insights exploitable to develop tailored bioprocesses to select specific strains for their higher accumulation ability. Microorganism responses to starvation are reviewed focusing on cell cycle, biomass production and variations in biochemical composition. Then, the work describes different innovative bioprocess configurations exploiting uncoupled nutrient feeding strategies (feast-famine), tailored to maintain a selective pressure to reward the strains with higher accumulation ability in mixed microbial populations. Finally, the main models developed in recent studies to describe and predict microbial growth and intracellular accumulation upon N-starvation and feast-famine conditions have been reviewed.
... Ecological theory also suggests that maintaining crop diversity in both time and space appears to be a critical element in the creation of sustainable, pest-resistant agroecosystems [16,[22][23][24][25][26][27]. Both sets of principles can potentially be applied to the design and operation of industrial-scale algal cultivation platforms [28][29][30][31]. ...
... Hence, careful testing of candidate algal consortia must be tested in the pilot-scale for an extended period of time to evaluate their potential application in industrial algal biofuels production. Another approach that may be combined with the careful survey of endogenous algal species pool is to impose a selective environment that will favor the proliferation of commercially-desirable algal consortia [28][29][30]. ...
Article
Full-text available
BACKGROUND: Consideration of ecological principles has brought new ideas that can be implemented to achieve sustainable production of algal biomass at the commercial-scale. In particular, the key ideas of top-down control of algal pests and the potential advantages of diversifying algal crops have encouraged researchers to explore food-web interactions in algal biomass cultivation platforms, and to investigate the characteristics of algal strains that could be used to assemble designed, multi-species consortia that outperform algal monocultures. OBJECTIVE/METHODS: To explore the practical applications of top-down control of algal pests and algal crop diversification, this paper reviews literature on agricultural and aquatic systems with consideration of the implementations of these ecological principles in managing commercial-scale algal cultivation. RESULTS: Our review suggests that careful management of food-web structure in algal cultivation platforms will be needed to maximize crop protection, and that temporal and spatial diversification of algal crops also may benefit industrial algal biofuels production. Extensive domestication and genetic improvements of algal strains are currently underway worldwide, and we suggest that careful selection of endogenous algal community which proliferates under selective environmental condition has the potential to engineer algal communities of high commercial interest. CONCLUSIONS: Overall, our review suggests that the careful management of food-web structure and algal crop diversity, as well as experiences and insights from modern agriculture can be used to guide the design and operation of industrial-scale algal biomass production systems. We urge thorough experimental tests of these ideas in both laboratory and field settings.
... Furthermore, the potential for outdoors wastewater treatment (Ledda et al., 2015) and abatement performance of CO 2 and toluene contained in exhaust gas by marine microalgae using diluted centrate in seawater as nutrient source (Anbalagan et al., 2017) have been recently addressed in closed photobioreactors. On the other hand, this photosynthetic CO 2abatement process can be further optimized by implementing the novel nutrient supplementation strategy developed by Mooij et al. (2013) to promote the cyclic production of storage compounds in the algal biomass. In this context, the production of a biomass with a high content in the metabolites of interest will increase the economic sustainability of the process (Toledo-Cervantes et al., 2017a). ...
... However, it is important to highlight that this operation is typically performed batch-wise in two operation stages with different nutrient requirements: a preliminary biomass production stage is necessary before inducing carbohydrates accumulation in the second stage. In this sense, a novel strategy of feeding nitrogen only during the dark period was here applied as a strategy to induce the continuous accumulation of high-energy storage compounds (Mooij et al., 2013). It is worthy to highlight that this strategy has been implemented exclusively in stirred reactors operated in sequencing batch mode (Mooij et al., 2015). ...
Article
The long-term performance of a tubular photobioreactor interconnected to a gas absorption column for the abatement of CO2 from biogas and flue-gas was investigated. Additionally, a novel nitrogen feast-famine regime was implemented during the flue-gas feeding stage in order to promote the continuous storage of highly-energetic compounds. Results showed effective CO2 (~98%) and H2S (~99%) removals from synthetic biogas, supported by the high photosynthetic activity of microalgae which resulted in an alkaline pH (~10). In addition, CO2 removals of 99 and 91% were observed during the flue-gas operation depending on the nutrients source: mineral salt medium and digestate, respectively. A biomass productivity of ~8 g m⁻² d⁻¹ was obtained during both stages, with a complete nitrogen and carbon recovery from the cultivation broth. Moreover, the strategy of feeding nutrients during the dark period promoted the continuous accumulation of carbohydrates, their concentration increasing from 22% under normal nutrition up to 37% during the feast-famine cycle. This represents a productivity of ~3 g-carbohydrates m⁻² d⁻¹, which can be further valorized to contribute to the economic sustainability of the photosynthetic CO2 removal process.
... Sustainable large-scale microalgae cultivation will therefore most likely involve the use of marine microalgae (Vasudevan et al. 2012). Previously, we have described a cultivation strategy to enrich and sustain a community of storage compoundproducing freshwater algae based on the ecophysiological role of storage compounds (Mooij et al. 2013). This strategy enriches microalgae with high levels of internal storage compounds, which make the overall process economically more interesting (Borowitzka 1992). ...
... In the dark period, with ammonium present, part of these storage compounds was consumed and residual biomass was produced (Fig. 2). These findings are in line with the behaviour observed under freshwater conditions in previous work (Mooij et al. 2013). The yield of biomass on starch during the dark period was 0.62± 0.19 mg X mg −1 starch (or 0.56±0.17 ...
Article
Large-scale production of microalgal storage compounds will likely involve marine microalgae. Previously, we described a method to enrich microalgae with a high storage compound productivity from a natural inoculum. Here, this strategy was implemented under marine conditions in a sequencing batch reactor. The influence of the volume exchange ratio and the moment of ammonium addition in the day-night cycle on the storage compound productivity are described. Storage compound productivity was maximal if ammonium was supplied at the start of the dark period rather than the light period, irrespective of the volume exchange ratio. Increasing the volume exchange ratio from 33 to 50 % per cycle induced a decrease in storage compound production if ammonium was supplied in the light whereas the storage compound productivity was comparable when ammonium was supplied in the dark. The latter indicates a shift of cell division processes to the light period at increasing volume exchange ratio, although ammonium uptake completely occurred in the dark period.
... To increase competitiveness, the use of waste substrates must be combined with mixed culture biotechnology and engineering designs with reduced costs and easily maintained operational conditions (Gujjala et al., 2019;Kourmentza et al., 2017;Kumar et al., 2020;Yadav et al., 2020). This solution considers engineering the ecosystem rather than specific strains to maintain a characteristic or functionality in the culture (Kourmentza et al., 2017;Mooij et al., 2013). Mooij et al. (2013) showed how to obtain a stable, open system enriched in storing populations inspired by Darwin's Theory of evolution by natural selection. ...
... This solution considers engineering the ecosystem rather than specific strains to maintain a characteristic or functionality in the culture (Kourmentza et al., 2017;Mooij et al., 2013). Mooij et al. (2013) showed how to obtain a stable, open system enriched in storing populations inspired by Darwin's Theory of evolution by natural selection. To design better biotechnologies, they proposed that imposing the right selective environment can emphasize the ecological role of storage compounds, allowing for the robust enrichment of a complex culture with optimal storage capacity. ...
Article
Operational conditions select for preferent TAG or PHA storage from waste lipids. • Coupled C and N supply promotes TAG production. • Uncoupled C and N feedings along with limited C excess favours PHA over TAG storage. • Pathways analysis unravelled the experimentally slower synthesis of PHA than TAG. Editor: Daniel CW Tsang The lipid fraction of the effluents generated in several food-processing activities can be transformed into polyhydroxyalkanoates (PHAs) and triacylglycerides (TAGs), through open culture biotechnologies. Although competition between storing and non-storing populations in mixed microbial cultures (MMCs) has been widely studied, the right selective environment allowing for the robust enrichment of a community when different types of accumulators coexist is still not clear. In this research, comprehensive metabolic analyses of PHA and TAG synthesis and degradation, and concomitant respiration of external carbon, were used to understand and explain the changes observed in a laboratory-scale bioreactor fed with the lipid-rich fraction (mainly oleic acid) of a waste-water stream produced in the fish-canning industry. It was concluded that the mode of oxygen, carbon, and nitrogen supply determines the enrichment of the culture in specific populations, and hence the type of intracellular compounds preferentially accumulated. Coupled carbon and nitrogen feeding regime mainly selects for TAG producers whereas uncoupled feeding leads to PHA or TAG production function of the rate of carbon supply under specific aeration rates and feast and famine phases lengths.
... Microalgae are unicellular and photosynthetic microorganisms, ranging from 0.2 to 2 μm (picoplankton) up to filamentous forms with sizes of 100 μm or higher ( Figure 2) [5][6][7]. These are unicellular organisms that consist of both prokaryotic (Cyanophyceae) or eukaryotic (Chlorophyta) organisms and they can grow rapidly in aquatic environments such as fresh water, waste water, and the marine environment. ...
... Microalgae are unicellular and photosynthetic microorganisms, ranging from 0.2 to 2 µm (picoplankton) up to filamentous forms with sizes of 100 µm or higher ( Figure 2) [5][6][7]. These are unicellular organisms that consist of both prokaryotic (Cyanophyceae) or eukaryotic (Chlorophyta) organisms and they can grow rapidly in aquatic environments such as fresh water, waste water, and the marine environment. ...
Article
Full-text available
Abstract: Substantial progress has been made in algal technologies in past few decades. Initially, microalgae drew the attention of the scientific community as a renewable source of biofuels due to its high productivity over a short period of time and potential of significant lipid accumulation. As of now, a technological upsurge has elaborated its scope in phycoremediation of both organic and inorganic pollutants. The dual role of microalgae—i.e., phycoremediation coupled with energy production—is well established, however, commercially, algal biofuel production is not yet sustainable due to high energy inputs. Efforts are being made to make the algal biofuel economy through modification in the cultivation conditions, harvesting, and extraction of value added products. Recent studies have demonstrated algal biomass production with various types of wastewater and industrial effluents. Similarly, the recent advent of eco-friendly harvesting technologies—such as low-cost green coagulants, electrochemical harvesting, etc.—are energy efficient and economical. Contemporary improvement in efficient lipid extraction from biomass will make algal biodiesel economical. The absolute extraction of all the value added products from algal biomass, either whole cell or lipid extracted biomass, in a complete biorefinery approach will be more economical and eco-friendly.
... When this innate behaviour is amplified in designed processes, algae with high storage compound productivity will be selected. A way to enhance storage compound consumption in the dark is to limit the availability of an essential growth nutrient other than carbon to the dark period [28 ]. Such cultivation conditions will favour growth of algae strains with the capacity to produce storage compounds in the light, which are required for nutrient uptake and heterotrophic growth on internal storage compounds in the dark. ...
... Such cultivation conditions will favour growth of algae strains with the capacity to produce storage compounds in the light, which are required for nutrient uptake and heterotrophic growth on internal storage compounds in the dark. This ecological concept, which we recently introduced as 'Survival of the Fattest', allowed the enrichment of microalgae with a high starch productivity, reaching 57% of glucose polymers on organic dry weight after eight hours of light [28 ]. These values are comparable to the highest values reported in literature for pure cultures [29]. ...
... reducing the carbon flow towards polysaccharide and glycoprotein matrix of the cell wall). Besides metabolic engineering, also adaptive laboratory evolution to a selective pressure [48] combined with FACS (Fluorescence Activated Cell Sorting) [38,49,50] or RACS (Raman Activated Cell Sorting) [51] can lead to increased TAG productivities. ...
Article
Full-text available
Background Microalgal triglycerides (TAGs) hold great promise as sustainable feedstock for commodity industries. However, to determine research priorities and support business decisions, solid techno-economic studies are essential. Here, we present a techno-economic analysis of two-step TAG production (growth reactors are operated in continuous mode such that multiple batch-operated stress reactors are inoculated and harvested sequentially) for a 100-ha plant in southern Spain using vertically stacked tubular photobioreactors. The base case is established with outdoor pilot-scale data and based on current process technology. Results For the base case, production costs of 6.7 € per kg of biomass containing 24% TAG (w/w) were found. Several scenarios with reduced production costs were then presented based on the latest biological and technological advances. For instance, much effort should focus on increasing the photosynthetic efficiency during the stress and growth phases, as this is the most influential parameter on production costs (30 and 14% cost reduction from base case). Next, biological and technological solutions should be implemented for a reduction in cooling requirements (10 and 4.5% cost reduction from base case when active cooling is avoided and cooling setpoint is increased, respectively). When implementing all the suggested improvements, production costs can be decreased to 3.3 € per kg of biomass containing 60% TAG (w/w) within the next 8 years. Conclusions With our techno-economic analysis, we indicated a roadmap for a substantial cost reduction. However, microalgal TAGs are not yet cost efficient when compared to their present market value. Cost-competiveness strictly relies on the valorization of the whole biomass components and on cheaper PBR designs (e.g. plastic film flat panels). In particular, further research should focus on the development and commercialization of PBRs where active cooling is avoided and stable operating temperatures are maintained by the water basin in which the reactor is placed. Electronic supplementary material The online version of this article (doi:10.1186/s13068-017-0873-2) contains supplementary material, which is available to authorized users.
... However, this new combination is still under investigation and a hybrid system can be developed: While microalgae remove the nutrient, they serve as a substrate for anaerobic digestion to produce methane and to recover nutrients. In the literature, there are limited studies on this (Alcántara et al. 2013;Mahdy et al. 2014;Mooij et al. 2013;Posadas et al. 2013). These studies showed that algal biomass is a potential co-substrate for biogas production. ...
... Furthermore, higher lipid accumulation in Pi-Vitamin medium may also lead to conclude that the supplementation provides a selective environment in which species with higher lipid production capacity are enriched. There are other studies with the aim of selective environment for lipid-producing algae (Hassanpour et al., 2015;Mooij et al., 2013). ...
Article
The present study investigated the interaction between starch and lipid accumulation was in a green microalgae enrichment culture. The objective was to optimize the lipid content by manipulation of the medium in regular batch culture. Two medium designs were evaluated: first a high ortho-P concentration with vitamin supplement (Pi-vitamins supplemented medium), second normal growth medium (control). Both media contained a low amount of nitrogen which was consumed during batch growth in three days. The batch experiments were prolonged continued for another four days with the absence of soluble nitrogen in the medium. When the mixed microalgal culture was incubated with in the Pi-vitamin supplemented medium, the lipid and starch content of the culture increased within the first three days to 102.0 ± 5.2 mg.l⁻¹ (12.7 ± 0.6% of DW) and 31.7 ± 1.6 mg.l⁻¹ (4.0 ± 0.2% of DW), respectively. On the last day of the experiment, the lipid and starch content in Pi-vitamin medium increased to 663.1 ± 32.5 (33.4 ± 1.6% of DW) and 127.5 ± 5.2 (6.4 ± 0.3% of DW) mg.l⁻¹. and However, the lipid and starch content in the control process, reached to 334.7 ± 16.4 (20.1 ± 1.0% of DW) and 94.3 ± 4.6 (5.7 ± 0.3% of DW), respectively. The high Pi-vitamin medium induced storing lipid formation clearly while the starch formation was not affected. The lipid contents reported here are among the high reported in the literature, note that already under full growth conditions significant lipid levels occurred in the algal enrichment culture. The high lipid productivity of the reported mixed microalgae culture provides an efficient route for efficient algal biodiesel production. This article is protected by copyright. All rights reserved
... Recently, Mooij et al. (2015) have proposed to apply selection pressures for microalgae culture in order to select species of interest, and to avoid contaminations. In particular, periodic nutrient stress can be carried out for the selection of microalgae with a high storage capacity (Masci et al., 2008;Mooij et al., 2013). ...
Article
Microalgae have recently attracted attention for their potential to produce high added compounds, proteins, and even biofuels. Our paper seeks to develop a control strategy for light-limited continuous culture imposing a stress for which microalgae have to adapt. This operating mode - called photoinhibistat - consists, for a culture with a constant dilution rate, in varying the incident light in order to regulate the light at the bottom of the reactor, inducing a light stress. Based on a simple model of light-limited growth, we analyze the dynamics of the photoinhibistat in monoculture and in competition. It appears that the photoinhibistat can be used to select, from the initial microalgae population, the strain with the highest resistance to photoinhibition.
... Thus, mixed cultures are considered as an attractive research area compared to traditional pure culture. Mooij et al. introduced the concept of ''survival of the fattest", a strategy for enrichment of species with a high storage compound productivity in mixed microalgae culture [18]. Later on, Hassanpour et al. applied a gravimetric enrichment method for screening carbohydrate and lipid accumulating species in a mixed microalgae culture [19]. ...
Article
Mixed microalgae cultures are considered as an attractive research area compared to traditional pure culture to dominate cultivation contamination risk and enhance economic feasibility of large-scale biofuel production. However, pre-treatment and bioethanol production from mixed microalgae culture has not been reported yet. Therefore, this study was aimed to evaluate the effect of different pre-treatment strategies including acidic, alkaline, and enzymatic hydrolysis on the sugar extraction from mixed microalgae. Besides, the effects of MgSO4 and CaCl2 as lewis acids in acidic pre-treatment on reducing sugar yield were studied.
... Open pond systems offer advantages compared to closed photobioreactor systems in terms of energy input and ease of operation. Recent work by Mooij et al. (2013) showed that one of the main disadvantages of open pond systems, contamination by less productive species, can be overcome by creating a selective environment. The temperature for the culture was assumed to be optimal and the energy-input of the microalgal cultivation was assumed to be covered by renewable resources, such as wind-or tidal power. ...
Article
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The disruption of nutrient cycles caused by human activities such as agriculture and burning fossil fuels is impacting ecosystem services on global and local scales. The increasing concentration of carbon dioxide in the atmosphere contributes to rising global temperatures and ocean acidification, whereas the accumulation of nutrients in water systems is leading to degradation of water quality and biodiversity. City populations play a major role in carbon dioxide and nutrient emissions as ‘end consumers’ of resources. The current challenge towards more resource-efficient cities is to transform urban metabolism from linear to cyclical. Discharged nutrients and carbon dioxide can be used as input for algae, which fixate carbon very efficiently into energetic storage compounds as starch or lipids. However, cities often lack the space to implement large-scale algae production. This article evaluates the potential of reusing nutrients and carbon dioxide to produce algae, food and biofuel on water nearby coastal and delta cities. First, nutrients and carbon dioxide discharge is estimated and two scenarios are developed. From the cities nutrient production, the potential algal yield is evaluated and translated into feed, food and oil yields. Two delta cities are chosen as case studies: Rotterdam and Metro Manila. The conclusion of this article is that Floating Production can help cities increasing their resilience in the field of food and energy. Floating Production can also contribute to a solution for global land shortage. The combination of food and energy production with floating urban development provides a climate-proof urban expansion in delta and coastal areas.
... Therefore, the ad-hoc control of microalgae biomass composition during the photosynthetic biogas upgrading can enhance the economic sustainability of the process by producing a biomass feedstock with a tailored valorization potential. In this context, Kleerebezem and co-workers reported a significant increase in the carbohydrate content of microalgae when nutrients were supplied during the dark period, which promoted the enrichment of microalgae capable of growing based on the intracellular polyglucose accumulated during the illuminated period in excess of CO 2 [16]. ...
Article
The performance of a pilot high rate algal pond (HRAP) interconnected with a biogas absorption column during the simultaneous upgrading of biogas and treatment of digestate was evaluated under two innovative biogas and nutrient supply strategies. Process operation with biogas supply during the night at a liquid recirculation/biogas ratio of 0.5 to prevent N2 and O2 stripping resulted in a biomethane complying with most international regulations for injection into natural gas grids (99.1 ± 1% CH4, 0.5 ± 0.2% CO2, 0.6 ± 0.5% N2 and 0.07 ± 0.08% O2). The potential of this technology to remove methyl mercaptan (MeSH), toluene and hexane from biogas (typically present at trace levels) was assessed, for the first time, with removal efficiencies under steady-state correlating with pollutant hydrophobicity (7 ± 7% for hexane, 66 ± 4% for MeSH and 98 ± 1% for toluene). Finally, the supply of digestate during the dark period shifted both microalgae population structure and biomass composition in the HRAP without a significant impact on biomethane quality. Overall, the removal of nitrogen and phosphorous from digestate in the HRAP was almost complete (96–99%) regardless of the nutrient supply strategy.
... Particularly, no study on ANN modeling of enzymatic hydrolysis of pure or mixed microalgae culture has yet been reported. Application of mixed culture of microalgae will decline the risk of system contamination, and thus operation in different conditions and process feasibility will be increased (Hassanpour et al., 2015;Mooij et al., 2013). ...
Article
The efficient production of reducing sugars is an extremely important requirement in the utilization of microalgae as a feedstock in bioethanol production. In this study, for the first time, the time course of reducing sugar production during starch hydrolysis of mixed microalgal biomass under different operational conditions was modeled by two different methods; a-Michaelis-Menten kinetic model and b-artificial neural network (ANN) method. The results from both models revealed that predicted values are in good agreement with the experimental results. Also, sensitivity analysis indicated that the kinetic model results are less sensitive to Km and Ki than to . The applied ANN was a feed-forward back propagation network with Levenberg-Marquardt algorithm. It was found that the order of relative importance of the input variables on reducing sugar concentration predicted by ANN model was as follows: pH>substrate concentration>temperature>hydrolysis time. Subsequently, the results indicated that the maximum reducing sugar yield (96.3%) was achieved by adding enzymes with the sequence of first cellulases, at 50 °C, pH 4.5, second α -amylase, at 70 °C, and pH 6 with a substrate concentration of 50 g/L. These findings may be useful for improving the enzymatic hydrolysis of mixed microalgae for bioethanol production.
... This has been supported by the observation that polycultures are more stable and productive tan monocultures under some scenarios of growth (Shurin et al., 2013). Furthermore, it has been shown that it is possible to use an environmental selective pressure to force consortia of microalgae to exhibit a certain train, such as increased lipid productivity (Mooij et al., 2013). These methods are attractive because they bypass chemical use and harness the dynamics of natural communities (Smith and Mcbride, 2015;Smith et al., 2010). ...
Article
Full-text available
Microalgal cultivation that takes advantage of solar energy is one of the most cost-effective systems for the biotechnological production of biofuels, and a range of high value products, including pharmaceuticals, fertilizers and feed. However, one of the main constraints for the cultivation of microalgae is the potential contamination with biological pollutants, such as bacteria, fungi, zooplankton or other undesirable microalgae. In closed bioreactors the control of contamination requires the sterilization of the media, containers, and all materials, which increases the cost of production, whereas in open pond systems severely limits the number of species that can be cultivated. Here, we report the metabolic engineering of Chlamydomonas reinhardtii to use phosphite as its sole phosphorus source by expressing the ptxD gene from Pseudomonas stutzeri WM88, which encodes a phosphite oxidoreductase able to oxidize phosphite into phosphate using NAD as a cofactor. Engineered C. reinhardtii lines are capable of becoming the dominant species in a mixed culture when fertilized with phosphite as a sole phosphorus source. Our results represent a new platform for the production of microalgae, potentially useful for both closed photobioreactors and open pond systems without the need for using sterile conditions nor antibiotics or herbicides to prevent contamination with biological pollutants. This article is protected by copyright. All rights reserved.
... Microalgae are considered as a lowcost novel expression system for the biosynthesis of biopolymers (Daniell et al., 2009;Hempel et al., 2011). It is known that microalgae under selective environment like absence of nutrients and varied photoperiod produce storage compounds which are natural polymers (Mooij et al., 2013). Microalgae excrete extracellular polymeric substance (EPS) into its environment comprising of polysaccharides, proteins, lipids, uronic and nucleic acids under controlled conditions. ...
Article
Microalgae due to its metabolic versatility have received a focal attention in the biorefinery and bioeconomy context. Microalgae products have broad and promising application potential in the domain of renewable fuels/energy, nutraceutical, pharmaceuticals and cosmetics. Biorefining of microalgal biomass in a circular loop with an aim to maximize resource recovery is being considered as one of the sustainable option that will have both economical and environmental viability. The expansive scope of microalgae cultivation with self-sustainability approach was discussed in this communication in the framework of blue-bioeconomy. Microalgae based primary products, cultivation strategies, valorization of microalgae biomass for secondary products and integrated biorefinery models for the production of multi-based products were discussed. The need and prospect of self-sustainable models in closed loop format was also elaborated.
... Such trait combinations are normally mutually exclusive. Approaches include genetic engineering (Zeng et al., 2011), directed evolution via successive rounds of mutagenesis and selection (Johnson et al., 2016;Lewin et al., 2016), and selection on the existing levels of genetic variation in a population (Mooij et al., 2013;Shurin et al., 2016). ...
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Drawing insights from multiple disciplines is essential for finding integrative solutions that are required to tackle complex environmental problems. Human activities are causing unprecedented influence on global ecosystems, culminating in the loss of species and fundamental changes in the selective environments of organisms across the tree of life. Our collective understanding about biological evolution can help identify and mitigate many of the environmental problems in the Anthropocene. To this end, we propose a stronger integration of environmental sciences with evolutionary biology.
... In some phytoplankton-dominated lakes, cyanobacteria are harvested for food (e.g. Spirulina or Arthrospira; Habib, 2008), and phytoplankton-dominated lakes may provide a genetic resource for the synthesis of valuable biochemicals (Mooij, Stouten, Tamis, van Loosdrecht, & Kleerebezem, 2013;Muys et al., 2019). ...
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• Shallow lakes can shift between stable states as a result of anthropogenic or natural drivers. Four common stable states differ in dominant groups of primary producers: submerged, floating, or emergent macrophytes or phytoplankton. Shifts in primary producer dominance affect key supporting, provisioning, regulating, and cultural ecosystem services supplied by lakes. However, links between states and services are often neglected or unknown in lake management, resulting in conflicts and additional costs. • Here, we identify major shallow lake ecosystem services and their links to Sustainable Development Goals (SDGs), compare service provisioning among the four ecosystem states and discuss potential trade‐offs. • We identified 39 ecosystem services potentially provided by shallow lakes. Submerged macrophytes facilitate most of the supporting (86%) and cultural (63%) services, emergent macrophytes facilitate most regulating services (60%), and both emergent and floating macrophytes facilitate most provisioning services (63%). Phytoplankton dominance supports fewer ecosystem services, and contributes most to provisioning services (42%). • The shallow lake ecosystem services we identified could be linked to 10 different SDGs, notably zero hunger (SDG 2), clean water and sanitation (SDG 6), sustainable cities and communities (SDG 11), and climate action (SDG13). • We highlighted several trade‐offs (1) among ecosystem services, (2) within ecosystem services, and (3) between ecosystem services across ecosystems. These trade‐offs can have significant ecological and economic consequences that may be prevented by early identification in water quality management. • In conclusion, common stable states in shallow lakes provide a different and diverse set of ecosystem services with numerous links to the majority of SDGs. Conserving and restoring ecosystem states should account for potential trade‐offs between ecosystem services and preserving the natural value of shallow lakes.
... When the cells reached ≥ 3.0 g/L concentration, nitrogen starvation was implemented in an N-free BG-11 medium for 2 days. Nitrogen-starved microalgae can assimilate ammonia faster, and only the microalga assimilating ammonia the fastest survives in the mixed algal culture (Mooij et al., 2015). They were harvested by centrifugation at 1590 × g for 15 min (RC 5B plus, Thermo Fisher, USA). ...
Article
Microalgal treatment of undiluted raw piggery wastewater is challenging due to ammonia toxicity and a deep dark color hampering photosynthesis. To overcome these problems, (1) a microalga (Coelastrella sp.) was isolated from an ammonia-rich environment, (2) the wastewater treatment was divided into two steps: a heterotrophic process followed by a mixotrophic process, and (3) a narrower transparent photobioreactor was employed with higher light intensity in the mixotrophic process. Coelastrella sp. removed 99% of ammonia, 92% of chemical oxygen demand (COD), and 100% of phosphorus during the 4-day process. Acetate in the wastewater relieved the ammonia stress on microalgae and promoted algal lipid and triacylglycerol productivity. Oxidative stability and low-temperature fluidity of triacylglycerols in lipids were improved by means of an altered fatty acid profile. Aside from the overall microalgal treatment performance, the proposed processing of piggery wastewater yielded a material suitable for possible production of algal biodiesel of better quality.
... Although many microbes have been enriched in a wide range of conditions, all of these enrichments are based on imposing a specific selective pressure in either a continuous or batch system. Recent work has demonstrated that through imposing dynamic process conditions an additional wealth of microbial diversity is revealed, compared to traditional chemostat or batch cultivation (6)(7)(8)(9). Essentially all natural environments are subject to ever changing conditions, from scarce nutrient availability to its sudden abundance and more regular day-night cycles leading to fluctuations in temperature and substrate supply. An abundant portion of microbes evolved under dynamic conditions and as such developed different functional strategies to exploit a competitive edge. ...
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Determining the functional development and dominant competitive strategy in microbial community enrichments is complicated by the extensive measurement campaigns required for off-line system analysis. This study demonstrates that detailed system characterization of aerobic pulse fed enrichments can be established using on-line measurements combined with automated data analysis. By incorporating the physicochemical processes in on-line data processing with a Particle Filter and kinetic process model, an accurate reconstruction of the dominant biological rates can be made. We hereby can differentiate between storage compound production and biomass growth in sequencing batch bioreactors. The method proposed allows for close monitoring of changes in functional behavior of long-running enrichment cultures, without the need for off-line samples, therewith enabling the identification of new insights in process dynamics with a minimal experimental effort. Even though a specific example application of the method proposed is described here, the approach can readily be extended to a wide range of dynamic experimental systems that can be characterized based on on-line measurements.
... . Indeed, data from this experimentation are in the low range, and this is in agreement with the low lipid content of the algal biomass. Indeed, the algal BMP is expected to be higher for those microalgae that are naturally richer in lipid or that are grown under environmental conditions that con induce lipid production (among others Li et al. 2008;Mooij et al. 2013). ...
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This paper discusses the possibility of including the culturing of microalgae within a conventional wastewater treatment sequence by growing them on the blackwater (BW) from biosolid dewatering to produce biomass to feed the anaerobic digester. Two photobioreactors were used: a 12 L plexiglas column for in-door, lab-scale tests and a 85 L plexiglas column for outdoor culturing. Microalgae (Chlorella sp and Scenedesmus sp) could easily grow on the tested blackwater. The average specific growth rate in indoor and outdoor batch tests was satisfactory, ranging between 0.14-0.16 d-1. During a continuous test performed under outdoor conditions from May to November, in which the off-gas from the combined heat and power unit was used as the CO2 source, an average biomass production of 50 mgTSS L-1 d-1 was obtained. However, statistical analyses confirmed that microalgal growth was affected by environmental conditions (temperature and season) and that it was negatively correlated with the occurrence of nitrification. Finally, the biochemical methane potential of the algal biomass was slightly higher than that from waste sludge (208 mLCH4 gVS-1 vs. 190 mLCH4 gVS-1).
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Microalgae are a diverse group of single-cell photosynthetic organisms that include cyanobacteria and a wide range of eukaryotic algae. A number of microalgae contain high-value compounds such as oils, colorants, and polysaccharides, which are used by the food additive, oil, and cosmetic industries, among others. They offer the potential for rapid growth under photoautotrophic conditions, and they can grow in a wide range of habitats. More recently, the development of genetic tools means that a number of species can be transformed and hence used as cell factories for the production of high-value chemicals or recombinant proteins. In this article, we review exploitation use of microalgae with a special emphasis on genetic engineering approaches to develop cell factories, and the use of synthetic ecology approaches to maximize productivity. We discuss the success stories in these areas, the hurdles that need to be overcome, and the potential for expanding the industry in general.
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Although no species lives in isolation in nature, efforts to grow organisms for use in biotechnology have generally focused on a single-species approach, particularly where a product is required at high purity. In such scenarios, preventing the establishment of contaminants requires considerable effort that is economically justified. However, for some applications in biotechnology where the focus is on lower-margin biofuel production, axenic culture is not necessary, provided yields of the desired strain are unaffected by contaminants. In this article, we review what is known about interspecific interactions of natural algal communities, the dynamics of which are likely to parallel contamination in industrial systems. Furthermore, we discuss the opportunities to improve both yields and the stability of cultures by growing algae in multi-species consortia.
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This paper presents a continuous-flow cell screening device to isolate and separate microalgae cells (Chlamydomonas reinhardtii) based on lipid content using high frequency (50 MHz) dielectrophoresis. This device enables screening of microalgae due to the balance between lateral DEP forces relative to hydrodynamic forces. Positive DEP force along with amplitude-modulated electric field exerted on the cells flowing over the planar interdigitated electrodes, manipulated low-lipid cell trajectories in a zigzag pattern. Theoretical modelling confirmed cell trajectories during sorting. Separation quantification and sensitivity analysis were conducted with time-course experiments and collected samples were analysed by flow cytometry. Experimental testing with nitrogen starved dw15-1 (high-lipid, HL) and pgd1 mutant (low-lipid, LL) strains were carried out at different time periods, and clear separation of the two populations was achieved. Experimental results demonstrated that three populations were produced during nitrogen starvation: HL, LL and low-chlorophyll (LC) populations. Presence of the LC population can affect the binary separation performance. The continuous-flow micro-separator can separate 74% of the HL and 75% of the LL out of the starting sample using a 50 MHz, 30 voltages peak-to-peak AC electric field at Day 6 of the nitrogen starvation. The separation occurred between LL (low-lipid: 86.1% at Outlet # 1) and LC (88.8% at Outlet # 2) at Day 9 of the nitrogen starvation. This device can be used for onsite monitoring; therefore, it has the potential to reduce biofuel production costs.
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Modern society is fueled by fossil energy produced millions of years ago by photosynthetic organisms. Cultivating contemporary photosynthetic producers to generate energy and capture carbon from the atmosphere is one potential approach to sustaining society without disrupting the climate. Algae, photosynthetic aquatic microorganisms, are the fastest growing primary producers in the world and can therefore produce more energy with less land, water, and nutrients than terrestrial plant crops. We review recent progress and challenges in developing bioenergy technology based on algae. A variety of high-value products in addition to biofuels can be harvested from algal biomass, and these may be key to developing algal biotechnology and realizing the commercial potential of these organisms. Aspects of algal biology that differentiate them from plants demand an integrative approach based on genetics, cell biology, ecology, and evolution. We call for a systems approach to research on algal biotechnology rooted in understanding their biology, from the level of genes to ecosystems, and integrating perspectives from physical, chemical, and social sciences to solve one of the most critical outstanding technological problems.
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The high frequency dielectrophoresis (>20 MHz) response of microalgae cells with different lipid content was monitored over time. Chlamydomonas reinhardtii was cultured in regular medium and under nitrogen-depleted conditions in order to produce populations of cells with low and high lipid content, respectively. The electrical conductivity (EC) of the culture media was also monitored over the same time. The upper crossover frequency (UCOF) decreased for high-lipid cells over time. The single-shell model predicts that the upper crossover frequency is dictated primarily by the dielectric properties of the cytoplasm. The high frequency DEP response of the high-lipid cells' cytoplasm was changed by lipid accumulation. DEP response of the low-lipid cells also varied with the conductivity of the culture media due to nutrient consumption. Relative lipid content was estimated with BODIPY 505/515 dye by calculating the area-weighted intensity average of fluorescent images. Finally, microalgae cells were successfully separated based on lipid content at 41 MHz and DEP media conductivity 106 ± 1 μS/cm. This article is protected by copyright. All rights reserved.
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The use of microalgae as a production system has gained huge interest in recent years. Recent research has concentrated on single aspects, such as the microalgal cells or the photobioreactors. The design of sustainable, effective and economic processes for microalgal products requires the integration of microalgal biology including strain selection and genetic engineering, process and reactor design and integration into environmental mass and energy fluxes. This involves tools of biotechnology and process engineering. Several attempts of such integrated processes have been developed. This review groups the integration aspects into several degrees of integration with respect to the classical upstream, bioreaction, and downstream steps. The integration levels consider metabolic, process, environmental and social integration. At the end, the question remains: How far can microalgal biotechnology help to solve the problems of feed, food and fuel supply and contribute to a better society? Overall, this review gives an overview of the different stages of integration in microalgal processes. It is a guide towards conceptual positioning and planning of microalgal plants and may be used as a decision guide for politicians, which could help support microalgal technologies.This article is protected by copyright. All rights reserved
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The enzymatic hydrolysis of microalgae has already been studied for bioethanol production. However, the enzymatic hydrolysis of microalgae under various fluid hydrodynamic conditions has not been simulated yet. Accordingly, the present study investigated the effect of various stirrer speed values, impeller types, and the presence of baffles on glucose extraction from the mixed microalgae using cellulase. The Michaelis-Menten kinetic constants of enzymatic hydrolysis were calculated in the AQUASIM and were used for simulation in COMSOL. The simulated values agreed with the experimental results and revealed that the low stirring speed reduced glucose extraction through undesirable enzyme distribution and the formation of regions with a high concentration of hydrolysis products to inhibit the enzyme's operation. The higher uniformity of glucose concentration at higher stirring speeds compared to lower stirring speeds ensured the higher efficiency of the mixing process in this reactor. The study findings suggested that a sufficient understanding of the mixing effects in the enzymatic hydrolysis could improve the economic feasibility of the process. Furthermore, compared to the unbaffled reactor, the proper mixing in the baffled reactor increased the enzymatic hydrolysis rate. It was found that altering the impeller type leads to a negligible change in the enzymatic hydrolysis rate.
Article
Microalgal treatment systems could advance nutrient recovery from wastewater by achieving effluent nitrogen (N) and phosphorus (P) levels below the current limit of technology, but their successful implementation requires an understanding of how design decisions influence nutrient uptake over daily (i.e., diel) cycles. This work demonstrates the ability to influence microalgal N:P recovery ratio via solids residence time (SRT) while maintaining complete nutrient removal across day/night cycles through carbon storage and mobilization. Experiments were conducted with two microalgal species, Scenedesmus obliquus and Chlamydomonas reinhardtii, in photobioreactors (PBRs) operated as cyclostats (chemostats subjected to simulated natural light cycles) with retention times of 6–22 days (S. obliquus) and 7–13 days (C. reinhardtii). Nutrient loading and all other factors were fixed across all experiments. Elevated SRTs (>8 days) achieved limiting nutrient concentrations (either N or P) below the detection limit throughout the diel cycle. N:P mass ratio in algal biomass was linearly correlated with SRT, varying from 9.9:1 to 5.0:1 (S. obliquus) and 4.7:1 to 4.3:1 (C. reinhardtii). Carbohydrate content of biomass increased in high irradiance and decreased in low irradiance and darkness across all experiments, whereas lipid dynamics were minimal over 24-h cycles. Across all nutrient-limited cultures, specific (i.e., protein-normalized) dynamic carbohydrate generally decreased with increasing SRT. Nighttime consumption of stored carbohydrate fueled uptake of nutrients, enabling complete nutrient limitation throughout the night. Dynamic carbohydrate consumption for nutrient assimilation was consistent with dark protein synthesis but less than that of heterotrophic growth, underscoring the need for algal process models to decouple growth from nutrient uptake in periods of low/no light. The ability to tailor microalgal N:P uptake ratio and target an optimal energy storage metabolism with traditional engineering process controls (such as SRT) may enable advanced nutrient recovery facilities to target continuous and reliable dual limitation of nitrogen and phosphorus.
Article
A two-stage cultivation strategy was applied to mixed microalgae, which were first cultured in complete nutrient medium then switched to different nutrient-free mediums in order to assess the impact of nutrient starvation on intracellular biochemical components of mixed microalgae. The effects of nitrogen, sulfur and phosphorus starvation on cell counts, chlorophyll, carotenoid, protein, starch and lipid content of the mixed microalgae are compared in this study. The obtained results revealed that starch, as a dominant storage compound, was the highest in the nitrogen-free medium up to 49% of dry weight (DW). Protein and chlorophyll content declined slightly from 512 to 472.96 mg.l⁻¹ and 29.43 to 26.58 mg.l⁻¹ only in the case of nitrogen starvation. Nitrogen starvation showed the best result as it ceased cell division immediately whereby in the case of sulfur and phosphorus starvation, cell division was not interrupted as microalgae are a pool of phosphorus and store sulfur. Calculation of starch and lipid energy content on the basis of electron equivalent and J/mg dry weight showed the higher energy content of lipid in compare with that of starch for all nutrients starvation. Nitrogen deprivation condition represents the superlative energy value of storage compounds content with the value of 17.61 J/mg dry weight. The finding proves the potential of attractive and economically feasible mixed microalgae cultivation for high percentages of storage compounds production under nitrogen starvation
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Microalgae offer a multiplicity of applications for the production of bio-sourced compounds such as proteins, pigments, sugars and oils. However, the energy spent for algae culture and lipid extraction hinder the energetic viability of the process for the production of biofuel derived from algae oils. Among possible improvements, pulsed electric fields (PEF) may be used as a pre-treatment to extract valuable compounds from microalgae and making the process less energy demanding.This project started with a collaboration between the team of bio-micro-systems Biomis, laboratory SATIE, with the team of bio-process engineering laboratory LGPM to study in situ the effects of PEF on microalgae.First, a energetic study is performed in a micro-system specially built for this project to characterize in situ, the effect of various treatment parameters (pulse duration / electric field) on Chlamydomonas reinhardtii cells with high lipid content.Among the outputs of this study, an energetic optimization of PEF conditions shows that a high level of permeability and low energy consumption are obtained when using short pulses of 5 µs. Associated with an electric field of 4.5 kV/cm, the pores are reversible (80% of the cells) during few seconds, and with a field of 7 kV/cm or higher, the permeabilization is irreversible. Afterwards, this PEFpre-treatment is associated with solvent mixing (hexane) to evaluate if lipid extraction is improved.
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Suspended growth, mixed community phototrophic wastewater treatment systems (including high-rate algal ponds and photobioreactors) have the potential to achieve biological nitrogen and phosphorus recovery with effluent nutrient concentrations below the current limit-of-technology. In order to achieve reliable and predictive performance, it is necessary to establish a thorough understanding of how design and operational decisions influence the complex community structure governing nutrient recovery in these systems. Solids residence time (SRT), a critical operational parameter governing growth rate, was leveraged as a selective pressure to shape microbial community structure in laboratory-scale photobioreactors fed secondary effluent from a local wastewater treatment plant. In order to decouple the effects of SRT and hydraulic retention time (HRT), nutrient loading was fixed across all experimental conditions and the effect of changing SRT on microbial community structure, diversity, and stability, as well as its impact on nutrient recovery, was characterized. Reactors were operated at distinct SRTs (5, 10, and 15 days) with diurnal lighting over long-term operation (>6 SRTs), and in-depth examination of the eukaryotic and bacterial community structure was performed using amplicon-based sequencing of the 18S and 16S rRNA genes, respectively. In order to better represent the microalgal community structure, this study leveraged improved 18S rRNA gene primers that have been shown to provide a more accurate representation of the wastewater process-relevant algal community members. Long-term operation resulted in distinct eukaryotic communities across SRTs, independent of the relative abundance of Operational Taxonomic Units (OTUs) in the inoculum. The longest SRT (15 days, SRT 15) resulted in a more stable algal community along with stable bacterial nitrification, while the shortest SRT (5 days, SRT 5) resulted in a less stable, more dynamic community. Although SRT was not strongly associated with overall bacterial diversity, the eukaryotic community of SRT 15 was significantly less diverse and less even than SRT 5, with a few dominant OTUs making up a majority of the eukaryotic community structure in the former. Overall, although longer SRTs promote stable bacterial nitrification, short SRTs promote higher eukaryotic diversity, increased functional stability, and better total N removal via biomass assimilation. These results indicate that SRT may be a key factor in not only controlling microalgal community membership, but community diversity and functional stability as well. Ultimately, the efficacy and reliability of NH4⁺ removal may be in tension with TN removal in mixed phototrophic systems given that lower SRTs may achieve better total N removal (via biomass assimilation) through increased eukaryotic diversity, biomass productivity, and functional stability.
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The capacity of microalgae to advance the limit of technology of nutrient recovery and accumulate storage carbon make them promising candidates for wastewater treatment. However, the extent to which these capabilities are influenced by microbial community composition remains poorly understood. To address this knowledge gap, 3 mixed phototrophic communities sourced from distinct latitudes within the continental United States (28° N, Tampa, FL; 36° N, Durham, NC; and 40° N, Urbana, IL) were operated in sequencing batch reactors (8 day solids residence time, SRT) subjected to identical diel light cycles with media addition at the start of the nighttime period. Despite persistent differences in community structure as determined via 18S rRNA (V4 and V8-V9 hypervariable regions) and 16S rRNA (V1-V3) gene amplicon sequencing, reactors achieved similar and stable nutrient recovery after 2 months (8 SRTs) of operation. Intrinsic carbohydrate and lipid storage capacity and maximum specific carbon storage rates differed significantly across communities despite consistent levels of observed carbon storage across reactors. This work supports the assertion that distinct algal communities cultivated under a common selective environment can achieve consistent performance while maintaining independent community structures and intrinsic carbon storage capabilities, providing further motivation for the development of engineered phototrophic processes for wastewater management.
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A paradigm shift is underway in wastewater treatment as the industry heads toward ~3% of global electricity consumption and contributes ~1.6% of greenhouse gas emissions. Although incremental improvements to energy efficiency and renewable energy recovery are underway, studies considering wastewater for carbon capture and utilization are few. This Review summarizes alternative wastewater treatment pathways capable of simultaneous CO2 capture and utilization, and demonstrates the environmental and economic benefits of microbial electrochemical and phototrophic processes. Preliminary estimates demonstrate that re-envisioning wastewater treatment may entirely offset the industry’s greenhouse gas footprint and make it a globally significant contributor of negative carbon emissions.
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The prospect of using constructed communities of microalgae in algal cultivation was confirmed in this study. Three different algal communities, constructed of diatoms (Diatom), green algae (Green) and cyanobacteria (Cyano), each mixed with a natural community of microalgae were cultivated in batch and semi‐continuous mode and fed CO2 or cement flue gas (12‐15 % CO2). Diatom had the highest growth rate but Green had the highest yield. Changes in the community composition occurred throughout the experiment. Green algae were the most competitive group, while filamentous cyanobacteria were outcompeted. Euglenoids, recruited from scarce species in the natural community became a large part of the biomass in semi‐steady state in all communities. High temporal and yield stability were demonstrated in all communities during semi‐steady state. Valuable products (lipids, proteins and carbohydrates) comprised of 61.5 ±5 % of ash free biomass and were similar for the three communities with lipids ranging 14–26 % of dry mass (DM), proteins (15‐28 % DM) and carbohydrates (9‐23 % DM). Our results indicate that culture functions (stability, biomass quality) were maintained while dynamic changes occurred in community composition. We propose that a multi‐species community approach can aid sustainability in microalgal cultivation, through complementary use of resources and higher culture stability. This article is protected by copyright. All rights reserved
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Use of diverse algal polycultures could enhance the viability of mass algal cultivation by increasing the productivity and stability of production. However, there are multiple approaches for selecting productive polycultures, ranging from a synthetic ecology approach using carefully selected laboratory strains, to collection and cultivation of naturally occurring polycultures. In this study, we compared the performance of a systematically assembled and a naturally occurring polyculture under two conditions: 1) pilot-scale, semi-continuous culture conditions and 2) grazing and resource stress conditions. The naturally occurring polyculture had 25% higher mean biomass productivity and 19% higher N-removal efficiency than the assembled polyculture under semi-continuous culture. Differences became even more pronounced under grazing and resource stress conditions; mean biomass productivity was 60% higher in the naturally occurring polyculture when exposed to stress. Lipid and protein content (% dry mass) were significantly higher in the assembled polyculture, while carbohydrate content was higher in the naturally occurring polyculture. The naturally occurring polyculture maintained higher diversity than the assembled polyculture throughout the experiment, suggesting a greater degree of niche complementarity as a potential mechanism for differences in biomass yields and nutrient uptake. Our results show that systematically engineered algal communities may not always lead to greater productivity than algal communities found in nature per se. We suggest that future studies using polycultures assembled from lab strains should test them against those occurring naturally; collectively, this would both serve as a benchmark for progress in synthetic ecology and would test whether there are generalities in the performance of natural vs. assembled polycultures.
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A high ammonia concentration and chemical oxygen demand (COD) in piggery wastewater force it to be diluted before conventional microalgal treatment to reduce ammonia toxicity. Incomplete treatment of ammonia and COD in piggery wastewater may cause eutrophication, resulting in algal blooms. This study tried to treat raw piggery wastewater without dilution, using three strains of microalgae (Chlorella sorokiniana, Coelastrella sp. and Acutodesmus nygaardii) that outcompeted other algae under heterotrophic, mixotrophic, and autotrophic conditions, respectively, through adaptive evolution at high ammonia concentration. The three stepwise processes were designed to remove (1) small particles, COD, and phosphorus in the 1st heterotrophic C. sorokiniana cultivation, (2) ammonia and COD in the 2nd mixotrophic Coelastrella sp. cultivation, and (3) the remaining ammonia in the 3rd photoautotrophic A. nygaardii cultivation. To enhance ammonia uptake rate, each algal species were inoculated after 2-day nitrogen starvation. When the N-starved three species were inoculated at each step sequentially at 7 g/L for 2 days, the final phosphorus, COD, and ammonia removal efficiencies were 100% (16.4 to 0 mg/L), 92% (6820 to 545 mg/L), 90% (850 to 81 mg/L) and turbidity (99%) after total 6 days.
Article
In this work, the algal biomass productivity and its lipid content were explored using a database containing 4670 instances extracted from the experimental results reported in 102 published articles. First, the influences of critical factors such as microalgae species, cultivation conditions, light intensity, CO2 amount, nutrient concentrations, reactor type, stress conditions, cell disruption methods, and lipid extraction solvents on the biomass and lipid production were reviewed. Then, the database was analyzed using machine learning techniques; decision trees were utilized to determine the combination of variables leading to high biomass and lipid content while association rule mining was used to find the specific conditions leading to very high biomass and lipid levels. Decision tree analysis discovered 11 different combinations of variables leading to high biomass productivity and 13 combinations for high lipid content; whereas, association rule mining analysis helped to identify the levels of specific factors for very high biomass and lipid production. It was then concluded that machine learning methods can help to determine the best conditions for optimum biomass growth and lipid yield for microalgae to manufacture renewable biofuels, and this can guide the planning of new experimental works.
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Beta-carotene, a pigment found in plants, is mainly produced by microalgae. Nevertheless, this production has only been investigated in pure cultures. Beta-carotene production through mixed culture eliminates the costly procedure of sterilization and contamination prevention needed for pure cultures. In this study, for the first time, the growth, beta-carotene, and chlorophyll production of a mixed culture of microalgae from Caspian Sea was investiagted under different stress conditions. At the condition of tripled light intensity and nitrogen starvation, beta-carotene content increased from 18.03 to 43.8 and 46.5 mol beta-carotene g ⁻¹ protein, respectively. However, the salinity of 4 mol L ⁻¹ caused the beta-carotene content to fall to zero. The blank sample reached a constant value of 23 mol beta-carotene g ⁻¹ protein. The comparable results with the specific monoculture species exhibit the high potential of a mixed culture of microalgae for beta-carotene production without need of the high sterilization cost. Nevertheless, more research is needed for where it can be a good substitute for pure culture.
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One of the most energy-consuming processes in conventional microalgal biofuel production is harvesting cells from dilute media. Cell harvesting by centrifugation or membrane filtration requires equal or more energy than is captured by photosynthesis, resulting in a negative net energy and CO2 balance. As a cost6 effective alternative to this approach, we investigated the possibility of using the inherent motility and behavioral responses of the green microalgae Euglena gracilis to light stimuli, to promote cell aggregation. Irradiation of cells with light stimuli of different wavelengths and intensities revealed that E. gracilis cells are specifically attracted to green light. The cell aggregation rate for cultures irradiated with green light for 24 h was 8.7 fold and the cell collection rate reached 70%, which is comparable to the efficiency of centrifugal separation. Utilization of green light for cell aggregation does not compete with the light absorption by chlorophylls in photosystems I and II (PSI and PSII). Therefore, the findings in the present study offer the use of green light in solar radiation, which was originally wasted energy in photosynthesis, as the energy source for one of the most energy-intensive downstream processes in microalgal biofuel production.
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Although open outdoor pond systems are the most economically viable option for mass cultivation of algae as a biofuel source, such systems face a number of limitations. Open ponds experience environmental fluctuations (i.e., light levels, nutrient ratios, and temperature), invasion pressure by undesired algal species, pathogen infections, and herbivory by invading zooplankton, all of which may negatively influence the system's overall harvestable yield. Using ecological principles to address the limitations of open-pond cultivation is a promising direction in algal biofuel research. This review surveys the growing body of work on these topics and offers a mechanistic framework for optimizing algal biofuel production while minimizing the negative effects of invasion, infection, and herbivory. High levels of productivity (in terms of biomass and lipids) are crucial for viable biofuel production and can be achieved by increasing algal diversity and assembling communities based on species' eco-physiological traits. Herbivory can be significantly reduced by choosing algal species resistant to grazing or by introducing biotic controls on herbivores. Diverse assemblages of algal species can be constructed to fill in the available ecological niche space, leading not only to high productivity but also reduced invasibility by undesired strains and potentially reduced susceptibility to algal diseases. Optimization of the mass cultivation of algae requires an interdisciplinary approach that includes using ecological principles for designing productive, resistant, and resilient algal communities.
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Background Microalgae are a promising platform for producing neutral lipids, to be used in the application for biofuels or commodities in the feed and food industry. A very promising candidate is the oleaginous green microalga Scenedesmus obliquus, because it accumulates up to 45% w/w triacylglycerol (TAG) under nitrogen starvation. Under these conditions, starch is accumulated as well. Starch can amount up to 38% w/w under nitrogen starvation, which is a substantial part of the total carbon captured. When aiming for optimized TAG production, blocking the formation of starch could potentially increase carbon allocation towards TAG. In an attempt to increase TAG content, productivity and yield, starchless mutants of this high potential strain were generated using UV mutagenesis. Previous studies in Chlamydomonas reinhardtii have shown that blocking the starch synthesis yields higher TAG contents, although these TAG contents do not surpass those of oleaginous microalgae yet. So far no starchless mutants in oleaginous green microalgae have been isolated that result in higher TAG productivities. Results Five starchless mutants have been isolated successfully from over 3,500 mutants. The effect of the mutation on biomass and total fatty acid (TFA) and TAG productivity under nitrogen-replete and nitrogen-depleted conditions was studied. All five starchless mutants showed a decreased or completely absent starch content. In parallel, an increased TAG accumulation rate was observed for the starchless mutants and no substantial decrease in biomass productivity was perceived. The most promising mutant showed an increase in TFA productivity of 41% at 4 days after nitrogen depletion, reached a TAG content of 49.4% (% of dry weight) and had no substantial change in biomass productivity compared to the wild type. Conclusions The improved S. obliquus TAG production strains are the first starchless mutants in an oleaginous green microalga that show enhanced TAG content under photoautotrophic conditions. These results can pave the way towards a more feasible microalgae-driven TAG production platform.
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Microalgae-derived lipids in the form of triacylglycerols (TAGs) are considered an alternative resource for the production of biofuels and food commodities. Large scale production of microalgal TAGs is currently uneconomical. The cost price could be reduced by improving the areal and volumetric TAG productivity. The economic value could be increased by enhancing the TAG quality. To improve these characteristics, the impact of light intensity, and the combined impact of pH and temperature on TAG accumulation were studied for Scenedesmus obliquus UTEX 393 under nitrogen starved conditions. The maximum TAG content was independent of light intensity, but varied between 18% and 40% of dry weight for different combinations of pH and temperature. The highest yield of fatty acids on light (0.263g/mol photon) was achieved at the lowest light intensity, pH 7 and 27.5°C.
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Microalgae are among the most diverse organisms on the planet, and as a result of symbioses and evolutionary selection, the configuration of core metabolic networks is highly varied across distinct algal classes. The differences in photosynthesis, carbon fixation and processing, carbon storage, and the compartmentation of cellular and metabolic processes are substantial and likely to transcend into the efficiency of various steps involved in biofuel molecule production. By highlighting these differences, we hope to provide a framework for comparative analyses to determine the efficiency of the different arrangements or processes. This sets the stage for optimization on the based on information derived from evolutionary selection to diverse algal classes and to synthetic systems.
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Polyhydroxyalkanoates (PHAs) are microbial storage polymers that attract interest as bioplastics. PHAs can be produced with open mixed cultures if a suitable enrichment step based on the ecological role of PHA is used. An acetate-fed sequencing batch reactor operated with 1 day biomass residence time and with feast-famine cycles of 12 h was used to enrich a mixed culture of PHA producers. In subsequent fed-batch experiments under growth limiting conditions, the enriched mixed culture produced PHA up to a cellular content of 89 wt % within 7.6 h (average rate of 1.2 g/g/h). The PHA produced from acetate was the homopolymer polyhydroxybutyrate. The culture was dominated by a Gammaproteobacterium that showed little similarity on 16S rRNA level with known bacteria (<90% sequence similarity). The mixed culture process for PHA production does not require aseptic conditions. Waste streams rather than pure substrates could be used as raw materials.
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Microalgae are a promising alternative source of lipid for biodiesel production. One of the most important decisions is the choice of species to use. High lipid productivity is a key desirable characteristic of a species for biodiesel production. This paper reviews information available in the literature on microalgal growth rates, lipid content and lipid productivities for 55 species of microalgae, including 17 Chlorophyta, 11 Bacillariophyta and five Cyanobacteria as well as other taxa. The data available in the literature are far from complete and rigorous comparison across experiments carried out under different conditions is not possible. However, the collated information provides a framework for decision-making and a starting point for further investigation of species selection. Shortcomings in the current dataset are highlighted. The importance of lipid productivity as a selection parameter over lipid content and growth rate individually is demonstrated.
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The interaction of whole cell metabolism with the distribution of excitation energy between photosystem 2 (PS2) and photosystem 1 (PS1), the light state transition, was investigated in vivo in the green alga Selenastrumminutum. Nitrogen limited cells of S. minutum were presented with a pulse of either NH4+ or NO3− in the light. As shown previously, CO2 fixation is inhibited and high rates of N assimilation ensue [(1986) Plant Physiol. 81, 273–279]. NH4+ assimilation has a much higher requirement ratio for ATP/NADPH than either CO2 or NO3− assimilation and thus drastically increases the demand for ATP relative to reducing power. Room temperature chlorophyll a fluorescence kinetic measurements showed that a reversible non-photochemical quenching of PS2 fluorescence accompanied the assimilation of NH4+ but not the assimilation of NO3− or CO2. 77K fluorescence emission spectra taken from samples removed at regular intervals during NH+4assimilation showed that the non-photochemical quenching of PS2 was accompanied by a complementary increase in the fluorescence yield of PS1, characteristic of a transition to state 2. Our data suggests that S. minutum responds to the increased demand for ATP/NADPH during NH4 assimilation by inducing the light state transition to direct more excitation energy to PS1 at the expense of PS2 to increase the production of ATP by cyclic electron transport.
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In her News Feature “Biotech’s green gold”, Emily Waltz details the ‘hype’ being propagated around emerging microalgal biofuel technologies, which often exceeds the physical and thermodynamic constraints that ultimately define their economic viability. Our calculations counter such excessive claims and demonstrate that 22 MJ m−2 d−1 solar radiation supports practical yield maxima of ∼60 to 100 kl oil ha−1 y−1 (∼6,600 to 10,800 gal ac−1 y−1) and an absolute theoretical ceiling of ∼94 to 155 kl oil ha−1 y−1, assuming a maximum photosynthetic conversion efficiency of 10%. To evaluate claims and provide an accurate analysis of the potential of microalgal biofuel systems, we have conducted industrial feasibility studies and sensitivity analyses based on peer-reviewed data and industrial expertise. Given that microalgal biofuel research is still young and its development still in flux, we anticipate that the stringent assessment of the technology's economic potential presented below will assist R&D investment and policy development in the area going forward.