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Unlocking the potential of microalgae as sustainable bioresources from up to downstream processing: A critical review
... Customized for C. reinhardtii, the MoClo toolkit consists of 119 publicly available genetic components. These include promoters, terminators, UTRs, tags, antibiotic resistance genes, reporters, and introns, strategically arranged for optimal modularity [81]. This toolkit is positioned to streamline the formulating codon-optimized gene expression cassettes, aiming for increased gene expression in Chlamydomonas by leveraging standardized synthetic biology tools [82]. ...
... Requires additional processing and infrastructure, increasing production complexity. [81,386] Green extraction methods Future research will focus on developing environmentally friendly and sustainable methods for extracting lipids and other valuable compounds from microalgae. Green extraction methods aim to reduce the ecological footprint of the industry and minimize the use of hazardous solvents. ...
Microalgal biofuels have emerged as a promising avenue for meeting the growing demands for clean and efficient energy. However, the integration of microalgae into the biofuel industry is still in the early stages, primarily due to low productivity and high production costs. To address these challenges, researchers are actively exploring innovative methods to enhance biomass, concurrently increasing lipid and carbohydrate content. This review paper discusses the unique attributes of microalgae that make them attractive candidates for biofuel production. Advancements in cultivation techniques, such as photobioreactor design, co-cultivation strategies (microalgae-microalgae, microalgae-bacteria, and microalgae-fungi), and the optimization of nutrient conditions (carbon, nitrogen, and phosphorus) as well as environmental factors (salinity, light, and temperature) were explored to enhance biomass and lipid productivity. Furthermore, genetic engineering tools (genetic elements, gene interference , genome editing, and genome reconstruction) and omics technologies (genomics, transcriptomics, and proteomics) were discussed to gain a deeper understanding of microalgal lipid synthesis metabolism. The application of these techniques in microalgae facilitates enhanced lipid productivity, improved stress tolerance, optimized carbon sequestration and utilization, and reduced harvesting and processing costs. The study also delves into the decision-making process related to software selection, with the overarching goal of improving performance, profitability, and sustainability while mitigating risks, operational costs, and environmental impacts. Additionally, this review highlights future perspectives on large-scale microalgal biofuel production and its industry.
... It is the primary requirement (Purohit and Chaturvedi (2018) Their metabolic energy by a long list of photosynthetic processes shows the enormous importance of light supply for their growth (OR* 400-700 nm) Temperature It changes the rate of chemical reactions (Diankristanti et al. 2024) It affects the physiology and biochemistry of microalgae The growth rate increased at an optimum range of 25-35 °C Beyond the range, structural integrity turns down Nutrients 7-10% of algal biomass is comprised of nitrogen, making it an essential nutrient (Vickram et al. 2023) Higher concentrations increase biomass growth Phosphorus is a second essential nutrient for algae, and its higher concentrations increase biomass a significant influence on the rate of growth, efficiency of photosynthetic and biobased carbon capture, biomass composition, and downstream processing costs, in addition to strain selection (Cheah et al. 2015). The ideal algal species would have a high sinking capacity, as well as a high tolerance for CO 2 concentrations, temperature, nutrients, toxic pollutants, and pH effects. ...
... The open pond system is widely used for large-scale cultivation of microalgae due to its minimal cost and ease of operation and maintenance. Open raceways and circular ponds are popular, particularly in industrial applications to produce significant products at low costs (Diankristanti et al. 2024). Typically, these ponds are 0.25 m wide and cover 0.2-0.5 hectares (Zhao and Su 2014), offering a high surface area-to-volume ratio for effective CO 2 sequestration. ...
Anthropogenic CO2 emissions are the prime cause of global warming and climate change, promoting researchers to develop suitable technologies to reduce carbon footprints. Among various CO2 sequestration technologies, microalgal-based methods are found to be promising due to their easier operation, environmental benefits, and simpler equipment requirements. Microalgae-based carbon capture and storage (CCS) technology is essential for addressing challenges related to the use of industrial-emitted flue gases. This review focuses on the literature concerning the microalgal application for CO2 sequestration. It highlights the primary physiochemical parameters that affect microalgal-based CO2 biofixation, including light exposure, microalgal strain, temperature, inoculum size, pH levels, mass transfer, CO2 concentration, flow rate, cultivation system, and mixing mechanisms. Moreover, the inhibition effect of different flue gas components including NOx, SOx, and Hg on growth kinetics is discussed to enhance the capacity of microalgal-based CO2 biofixation, along with deliberated challenges and prospects for future development. Overall, the review indicated microalgal-based flue gas CO2 fixation rates range from 80 mg L⁻¹ day⁻¹ to over 578 mg L⁻¹ day⁻¹, primarily influenced by physiochemical parameters and flue gas composition. This article summarizes the mechanisms and stages of microalgal-based CO2 sequestration and provides a comprehensive review based on international interest in this green technology.
... In this context, the United Nations (UNs) established 17 Sustainable Development Goals (SDGs) targeting their implementation by 2030. Microalgae can assist in fulfilling this objective by providing a wide portfolio of bioproducts with promising applications in the biofuel, food, and nutraceutical industries [1]. A microalgae-based biorefinery concept can contribute to a circular economy scenario, highlighting the significance of these microorganisms in creating a greener future, leveraging their potential as main resources for the production of thirdgeneration biofuels [2]. ...
... Mixotrophic growth emerges as a combination of these two cultivation modes, benefiting from the distinct energy sources [22,23]. Key cultivation parameters such as carbon and nitrogen sources, C/N ratio, and light intensity critically affect microalgal growth performance and biochemical composition and their interactions are crucial in determining the outcome of the cultivation [1]. Given the increasing number of studies on isolating new microalgal strains and the vast diversity of known species, adopting a universal cultivation approach is not feasible. ...
... Consequently, the appropriate selection of the technology and its proper design for Chlorella cultivation also determines the sustainability of biomass production. Open cultivation systems such as open ponds or raceway ponds can be applied for large-scale biomass production, as they demand less construction and operational cost than closed cultivation systems (Diankristanti et al. 2024) However, the open cultivation system is typically more susceptible to predators and contamination of other organisms, which may decrease the overall biomass production (Tan et al. 2020). In contrast, closed cultivation systems such as photobioreactors require high investment and operational costs, but they can significantly reduce predation and contamination (Posten 2009). ...
... Generally, extraction processes are categorized into physical, chemical, and enzymatic techniques. Among the physical extraction methods is the use of bead milling which has high cell disruption efficiency (Diankristanti et al. 2024). Low temperature may also be employed to preserve the structure of targeted compounds during the extraction process. ...
Chlorella and its metabolites have been known for decades to affect skin and beauty positively. With its increasing use in cosmeceuticals, there is a growing interest in elucidating its potential, particularly the bioactive compounds extracted from Chlorella biomass. Chlorella has numerous advantages as the key ingredient in cosmeceutical products as it can be easily cultivated, is highly adaptable in various environmental conditions, and contains high-value metabolites. The bioactive compounds include polysaccharides, chlorophyll, carotenoids, lutein, and peptides, demonstrating antioxidant, anti-aging, moisturizing, brightening, and UV-protection properties that are highly desirable in cosmeceutical products. This review highlights the biodiversity of Chlorella bioactive compounds, the application of Chlorella and its bioactive compounds for commercial cosmeceutical products, along with the production and extraction systems that have not been discussed elsewhere. Further optimization on the extraction and purification of bioactive compounds from microalgae biomass and the use of genetic engineering and omics of Chlorella will have the potential to improve the purity and yield of bioactive compounds.
... Improving the sustainability of microalgal PLA production involves addressing several key stages of the process, from cultivation to final processing. Scaling up microalgae cultivation poses unique challenges, including high nutrient demands, complex downstream processing, and limited compatibility with concentrated CO 2 sources (Diankristanti et al., 2024). Enhancing this stage could involve selecting or genetically engineering algal strains with higher biomass productivity, increased tolerance to environmental stresses, and greater lactic acid yield potential. ...
Polylactic acid (PLA), a bioplastic widely used in sustainable packaging, offers notable environmental advantages over petro-plastics. However, the environmental footprint of PLA production depends heavily on biomass feedstock. This study provides a comparative life cycle assessment (LCA) of PLA production from cane-sugar and microalgal biomass, analyzing contributions across production stages and performing sensitivity analysis to identify key environmental factors. The study also highlights the recommendations for optimizing processes to reduce environmental burdens. Conducted per ISO 14044:2006 standards, the analysis revealed distinct trade-offs between the feedstocks. Cane-sugar based PLA demonstrated lower global warming potential (GWP) (406 kg CO2 eq/tonne PLA) and energy consumption due to established agricultural practices but required higher land (959 m2 × a crop eq/tonne PLA) and water demand (183 m3 eq/tonne PLA). Conversely, microalgae-based production significantly reduced land use (75 m2 × a crop eq/tonne PLA) due to utilizing non-arable lands but incurred elevated impacts such as freshwater eutrophication (11.93 kg P eq/tonne PLA) due to high energy and nutrient demands. Sensitivity analysis identified fermentable sugar content in microalgae as critical for reducing GWP, emphasizing the importance of strain selection and optimized cultivation. High sensitivity ratios and sensitivity coefficients for energy inputs across both feedstocks highlighted their significant impact on environmental performance. While cane-sugar based PLA currently offers lower impacts, microalgae is a promising alternative for regions with limited arable land, contingent on improvements in energy efficiency and nutrient recycling. These findings contribute to the broader effort of optimizing PLA production systems for enhanced sustainability and reduced environmental footprints.
... These systems are designed to withstand the harsh marine environment while maximizing light exposure, which is critical for algae growth [46]. Photobioreactors offer several advantages over traditional open pond systems, including higher biomass productivity, reduced contamination risks, and better control over growth conditions [47]. Recent studies have demonstrated that floating photobioreactors can achieve high levels of carbon capture and biofuel production, making them a promising option for offshore integration [48]. ...
Marine algae have shown great promise for both capturing carbon and producing bio-fuels, but their use in offshore oil and gas operations is still largely unexplored. This review paper looks into how marine algae could play a dual role: acting as a carbon sink and providing a renewable energy source. Known for their fast growth and ability to absorb carbon dioxide, algae offer a potential way to reduce the carbon footprint of offshore activities. The paper explores current research on algae-based bio-fuels and assesses how feasible it would be to integrate these into offshore platforms. It also considers how marine algae might be used to capture and store CO₂ emissions, contributing to overall emission reduction efforts. Key issues discussed include the practical challenges of growing algae in marine settings, the economic viability of algae-derived bio-fuels, and the environmental effects of scaling up these technologies. By reviewing existing knowledge and highlighting areas that need more research, this paper aims to provide a detailed look at how marine algae could help make offshore oil and gas operations more sustainable.
... The full biotechnological exploitation of non-conventional microalgae, however, requires improvements in biomass yield and downstream processing to retrieve target metabolites [202,203]. ...
From sea shores to the abysses of the deep ocean, marine ecosystems have provided humanity with valuable medicinal resources. The use of marine organisms is discussed in ancient pharmacopoeias of different times and geographic regions and is still deeply rooted in traditional medicine. Thanks to present-day, large-scale bioprospecting and rigorous screening for bioactive metabolites, the ocean is coming back as an untapped resource of natural compounds with therapeutic potential. This renewed interest in marine drugs is propelled by a burgeoning research field investigating the molecular mechanisms by which newly identified compounds intervene in the pathophysiology of human diseases. Of great clinical relevance are molecules endowed with anti-inflammatory and immunomodulatory properties with emerging applications in the management of chronic inflammatory disorders, autoimmune diseases, and cancer. Here, we review the historical development of marine pharmacology in the Eastern and Western worlds and describe the status of marine drug discovery. Finally, we discuss the importance of conducting sustainable exploitation of marine resources through biotechnology.
This chapter delves into the processing techniques for bio-based products in the Global South, emphasizing the intersection of traditional methods and modern advancements. The analysis begins with defining bio-based products and their importance in the context of sustainable development and environmental conservation. It explores the historical background and common traditional methods, particularly focusing on fermentation techniques prevalent in West Africa. The chapter then transitions to modern processing techniques, highlighting their potential to enhance efficiency and product quality while addressing challenges such as economic constraints and technical difficulties. The socioeconomic impacts of these processing activities are examined, noting their influence on small-scale farmers, gender dynamics, and the broader community. The discussion also encompasses the environmental benefits of adopting sustainable practices. The chapter concludes by identifying opportunities for innovation and growth in the bio-based products sector, advocating for policies and investments that support sustainable processing practices. Through this comprehensive analysis, the chapter provides insights into the challenges and opportunities within the processing of bio-based products, offering a pathway toward sustainable development in the Global South.
Bio-based plastics, primarily polyhydroxyalkanoates (PHAs), offer a hopeful alternative to petroleum-derived plastics. Third-generation (3G; microalgae/cyanobacteria) biomass has gained significant importance due to its rapid biomass productivity and metabolic versatility. Microalgae can produce PHAs by utilizing CO2 and wastewater, establishing them as highly promising and eco-friendly systems for bioplastic production. This comprehensive review presents comprehensive insights into microalgae-PHA production, from optimization of physicochemical and cultural conditions to effective PHA purification processes. The critical review also examines the latest advancements in cultivation strategies, metabolic engineering, and bioreactor developments, which may lead to more sustainable and progressive microalgal-based bioplastic accumulation. The effectiveness of algae biomass generation for PHA accumulation through integrated wastewater treatment has been addressed. This review examines the role of mathematical modeling and emerging artificial intelligence in advancing algae-based PHA production processes. Finally, the review concludes with a discussion of the economic and social challenges, life cycle analysis, and prospects for research and development of advanced microalgal-derived bioplastics production and predictions of potential solutions for economically feasible and sustainable microalgae-based PHA production at the industrial scale.
The production of high-value products from microalgae, one of the preferred emerging biorefineries’ feedstocks, relies on the crucial step of biomass fractionation. In this work, the fractionation of Chlorella vulgaris and Scenedesmus obliquus biomass was tested for protein extraction using a wide range of physical, chemical, and enzymatic treatment combinations, including ultrasound, cell homogenizer, cellulase, and alcalase combinations in aqueous and alkali extraction conditions. The impact of these processes on biomass carbohydrates was also evaluated. Alkaline-assisted ultrasound treatments using alcalase presented the highest protein extraction yield, reaching 90 g/100 g protein on C. vulgaris, closely followed by the same treatment in aqueous conditions (85 g/100 g protein). The same aqueous treatment achieved the best performance on S. obliquus, reaching 82 g/100 g protein. All treatments on both microalgae partially solubilized the polysaccharide fraction with all alkaline treatments solubilizing over 50 g/100 g sugars for all conditions. Overall, all the treatments applied were effective methods for biomass fractionation, although they showed low selectivity regarding the individual extraction of protein or carbohydrates.
We investigated two non-ionising mutagens in the form of ultraviolet radiation (UV) and ethyl methanosulfonate (EMS) and an ionising mutagen (X-ray) as methods to increase fucoxanthin content in the model diatom Phaeodactylum tricornutum. We implemented an ultra-high throughput method using fluorescence-activated cell sorting (FACS) and live culture spectral deconvolution for isolation and screening of potential pigment mutants, and assessed phenotype stability by measuring pigment content over 6 months using high-performance liquid chromatography (HPLC) to investigate the viability of long-term mutants. Both UV and EMS resulted in significantly higher fucoxanthin within the 6 month period after treatment, likely as a result of phenotype instability. A maximum fucoxanthin content of 135 ± 10% wild-type found in the EMS strain, a 35% increase. We found mutants generated using all methods underwent reversion to the wild-type phenotype within a 6 month time period. X-ray treatments produced a consistently unstable phenotype even at the maximum treatment of 1000 Grays, while a UV mutant and an EMS mutant reverted to wild-type after 4 months and 6 months, respectively, despite showing previously higher fucoxanthin than wild-type. This work provides new insights into key areas of microalgal biotechnology, by (i) demonstrating the use of an ionising mutagen (X-ray) on a biotechnologically relevant microalga, and by (ii) introducing temporal analysis of mutants which has substantial implications for strain creation and utility for industrial applications.
Background
Microalgae are emerging hosts for the sustainable production of lutein, a high-value carotenoid; however, to be commercially competitive with existing systems, their capacity for lutein sequestration must be augmented. Previous attempts to boost microalgal lutein production have focussed on upregulating carotenoid biosynthetic enzymes, in part due to a lack of metabolic engineering targets for expanding lutein storage.
Results
Here, we isolated a lutein hyper-producing mutant of the model green microalga Chlamydomonas reinhardtii and characterized the metabolic mechanisms driving its enhanced lutein accumulation using label-free quantitative proteomics. Norflurazon- and high light-resistant C. reinhardtii mutants were screened to yield four mutant lines that produced significantly more lutein per cell compared to the CC-125 parental strain. Mutant 5 (Mut-5) exhibited a 5.4-fold increase in lutein content per cell, which to our knowledge is the highest fold increase of lutein in C. reinhardtii resulting from mutagenesis or metabolic engineering so far. Comparative proteomics of Mut-5 against its parental strain CC-125 revealed an increased abundance of light-harvesting complex-like proteins involved in photoprotection, among differences in pigment biosynthesis, central carbon metabolism, and translation. Further characterization of Mut-5 under varying light conditions revealed constitutive overexpression of the photoprotective proteins light-harvesting complex stress-related 1 (LHCSR1) and LHCSR3 and PSII subunit S regardless of light intensity, and increased accrual of total chlorophyll and carotenoids as light intensity increased. Although the photosynthetic efficiency of Mut-5 was comparatively lower than CC-125, the amplitude of non-photochemical quenching responses of Mut-5 was 4.5-fold higher than in CC-125 at low irradiance.
Conclusions
We used C. reinhardtii as a model green alga and identified light-harvesting complex-like proteins (among others) as potential metabolic engineering targets to enhance lutein accumulation in microalgae. These have the added value of imparting resistance to high light, although partially compromising photosynthetic efficiency. Further genetic characterization and engineering of Mut-5 could lead to the discovery of unknown players in photoprotective mechanisms and the development of a potent microalgal lutein production system.
Algae-based biofuel developed over the past decade has become a viable substitute for petroleum-based energy sources. Due to their high lipid accumulation rates and low carbon dioxide emissions, microalgal species are considered highly valuable feedstock for biofuel generation. This review article presented the importance of biofuel and the flaws that need to be overcome to ensure algae-based biofuels are effective for future-ready bioenergy sources. Besides, several issues related to the optimization and engineering strategies to be implemented for microalgae-based biofuel derivatives and their production were evaluated. In addition, the fundamental studies on the microalgae technology, experimental cultivation, and engineering processes involved in the development are all measures that are commendably used in the pre-treatment processes. The review article also provides a comprehensive overview of the latest findings about various algae species cultivation and biomass production. It concludes with the most recent data on environmental consequences, their relevance to global efforts to create microalgae-based biomass as effective biofuels, and the most significant threats and future possibilities.
Microalgae and cyanobacteria are a precious source for the production of biofuels/bioenergy, biomaterials and valuable biochemicals. Beyond photosynthetic CO2 conversion, microalgal systems can involve the valorisation of waste streams and the implementation of green chemistry, industrial symbiosis, and circular bioeconomy approaches. However, their sustainability is uncertain, thus their large-scale application is hindered. The numerous life cycle assessments (LCAs) performed so far are mostly based on data extrapolated from lab-scale experiments or the literature, leading to qualitative and controversial results. This paper reviews primary data-based LCA studies on microalgal pilot to industrial-scale plants. Sixteen studies satisfied the selection criteria, despite they used primary data almost exclusively for cultivation and harvesting. The outlined current status (methodology, inventory, energy performance and environmental impacts) highlighted the lack of uniformity in the applied methods and the presentation of results, as well as some lack of transparency. Nevertheless, the review concluded that electricity consumption and infrastructure are major hotspots. Therefore, the use of renewable energy for supplying the process and of sunlight for biomass photosynthesis should be preferred. The upstream processes produce large impacts. Thus, a suitable reactor, geographic location, and harvesting method should be selected. Biofuels are not competitive in most cases, but some promising multi-product biorefinery scenarios have been presented. To improve the environmental profile of microalgal high-value compounds (e.g., astaxanthin or biostimulants), co-product valorisation, waste stream utilization, renewable energy deployment, and compound productivity should be enhanced. More efforts on LCA of large-scale plants are required, especially looking at integrated biorefinery concepts, to take a crucial step towards the implementation of sustainable commercial systems.
Currently dairy processing by-products, such as whey, still propose a significant threat to the environment if unproperly disposed. Microalgal bioconversion of such lactose containing substrates can be used for production of valuable microalgae-derived bio-products as well as for significant reduction of environmental risks. Moreover, it could significantly reduce microalgae biomass production costs, being a significant obstacle in commercialization of many microalgae species. This review summarizes current knowledge on the use of lactose containing substrates, e.g. whey, for the production of value-added products by microalgae, including information on producer cultures, fermentation methods and cultivation conditions, bioprocess productivity and ability of microalgal cultures to produce β-galactosidases. It can be stated, that despite several limitations lactose-containing substrates can be successfully used for both—the production of microalgal biomass and removal of high amounts of excess nutrients from the cultivation media. Moreover, co-cultivation of microalgae and other microorganisms can further increase the removal of nutrients and the production of biomass. Further investigations on lactose metabolism by microalgae, selection of suitable strains and optimisation of the cultivation process is required in order to enable large-scale microalgae production on these substrates.
Deep generative models, which can approximate complex data distribution from large datasets, are widely used in biological dataset analysis. In particular, they can identify and unravel hidden traits encoded within a complicated nucleotide sequence, allowing us to design genetic parts with accuracy. Here, we provide a deep-learning based generic framework to design and evaluate synthetic promoters for cyanobacteria using generative models, which was in turn validated with cell-free transcription assay. We developed a deep generative model and a predictive model using a variational autoencoder and convolutional neural network, respectively. Using native promoter sequences of the model unicellular cyanobacterium Synechocystis sp. PCC 6803 as a training dataset, we generated 10 000 synthetic promoter sequences and predicted their strengths. By position weight matrix and k-mer analyses, we confirmed that our model captured a valid feature of cyanobacteria promoters from the dataset. Furthermore, critical subregion identification analysis consistently revealed the importance of the -10 box sequence motif in cyanobacteria promoters. Moreover, we validated that the generated promoter sequence can efficiently drive transcription via cell-free transcription assay. This approach, combining in silico and in vitro studies, will provide a foundation for the rapid design and validation of synthetic promoters, especially for non-model organisms.
Purpose
Microalgae-derived biofuels are considered a low-carbon alternative to fossil fuels. Nevertheless, as with all biofuels, there is still uncertainty around their sustainability. Most life cycle assessments (LCA) of microalgae biofuels so far used lab-based, scaled-up lab experimental data or data from the scientific literature. This article, provides evidence and analysis, undertaking an LCA using real-world data from an industrial facility that uses a combination of photobioreactor and fermenter systems.
Methods
The current well-to-wheel LCA study aimed to compare the environmental impacts of microalgae biodiesel production—under different energy regimes—and with petroleum-derived diesel. The functional unit was considered as “combustion of 1 MJ (Lower Heating Value) of algal biodiesel in an internal combustion engine (as B100)”. This LCA study considers the environmental and energy impacts from the construction of the facility, as well as those impacts from the operation of the facility. The foreground LCI data was collected from a real-world one-hectare microalgae production pilot facility. ReCiPe, IPCC AR5 (GWP100 and GWP20) and Global Temperature Potential (GTP) were implemented to assess the life cycle environmental impacts.
Results and discussion
The assessment shows that when infrastructure is included, microalgae-derived biofuels are not yet favourable over petroleum-derived fuels on GWP100, and this becomes worse over shorter timescales. In terms of climate change (GWP100), whilst 1 MJ (LHV) of fossil-derived diesel would emit 8.84 × 10⁻² kg CO2eq, 1 MJ of microalgae-derived biodiesel from a solar photovoltaic powered facility would emit 1.48 × 10⁻¹ kg CO2eq. To be equal to petroleum-derived diesel in terms of GWP100, or perform better, productivity of the microalgae production system needs to be improved as the most effective solution. The results also showed that electricity and infrastructure were major sources of environmental impacts, as well as the yeast used within the fermenter. Moreover, it takes 0.99 MJ of direct energy per 1 MJ of microalgae biofuel produced, similar to the fossil fuel industry for 1 MJ of diesel.
Conclusions
Using infrastructure and operational models, the study shows that the facility does not compare well with petroleum-derived diesel unless productivity can be increased. Productivity improvements, be it through improvements to microalgae strains or improved photobioreactor designs, should be a priority to ensure microalgae become a sustainable fuel feedstock. Electricity use should be reduced as well, again, through improved cultivation system designs. In terms of the current system, the high impacts of yeast should be addressed, either through co-locating yeast production or through ensuring specific sources with lower impacts. Extracting lipids will effectively waste some high-value products, whilst the waste can be expected to be a mixture of unextracted lipids, polysaccharides or fibre, some proteins and minerals. It is also shown that harmonisations of the assessments are needed for future studies and real-world operation facilities to conclusively decide if microalgae should be used as fuel or if they would be better used for other products, such as feed or high-value products.
Here, an unprecedented biorefinery approach has been designed to recover high-added value bioproducts starting from the culture ofPorphyridium cruentum. This unicellular marine red alga can secrete and accumulate high-value compounds that can find applications in a wide variety of industrial fields. 300 ± 67 mg/L of exopolysaccharides were obtained from cell culture medium; phycoerythrin was efficiently extracted (40% of total extract) and isolated by single chromatography, with a purity grade that allowed the crystal structure determination at 1.60 Å; a twofold increase in β-carotene yield was obtained from the residual biomass; the final residual biomass was found to be enriched in saturated fatty acids. Thus, for the first time, a complete exploitation ofP. cruentumculture was set up.
Microalgal-based biofuel replaces fossil fuel to meet the current global energy demand. The mutant strains, UV-2 and 5’FDU-1, generated using random mutagenesis through UV light exposure (60 s) and 5’fluorodeoxyuridine (5’FDU-0.25 mM) enhanced the lipid production in marine microalgae, Chlorella vulgaris, with suitable fatty acid profile for biodiesel production, under laboratory conditions. Augmentation in the lipid content in UV-2 (26%) and 5’FDU-1 (23%) mutants was observed in outdoor conditions. Decreased growth rate observed in the mutants under indoor conditions was reversed under outdoor environment compared to wild type. Transcriptome-based gene expression analysis of wild type and mutants explained the variation in growth rate and lipid production. Acetyl-CoA carboxylase and other important fatty acid biosynthesis were significantly upregulated in mutant strains under outdoor conditions, correlating with the increased lipid production in outdoor mutants. Significant upregulation observed in genes related to photosystems in the mutant samples under both indoor and outdoor conditions corroborated with the increased biomass observed in mutant samples. Transcriptome study also provided pointers to the expression levels of key genes involved in carotenoid biosynthesis. Impact of physiological parameters like temperature played vital role in regulating genes related to growth, carotenoid biosynthesis, etc. Variation of gene expression levels of carotenoid biosynthesis genes and transcription factors with respect to indoor and outdoor conditions provided crucial pointers for further downstream studies. The current study evaluating the role of mutation substantiated with transcriptome data could be a precursor towards exploring specific targets for augmenting biomass, lipid leading to enhanced biodiesel production.
Continuously escalating energy demand and depletion of conventional fossil fuels has challenged and urged our globe to search for alternative resources which are economically viable, renewable and sustainable. Microalgae has emerged as an excellent resource in this regard but the excessive cost of nutrients is a vital restriction for producing economically viable algal fuels. In this study, waste banana peels were used as the feedstock for cultivation medium for microalgae after proper pre-treatment and hydrolysis. Various pre-treatment techniques were executed and the best conditions (40 min, autoclaving) furnishing the higher glucose concentration were chosen for further investigation. Rationale for the enzyme hydrolysis of the pre-treated peels was created through central composite design (CCD) using response surface methodology (RSM) to attain higher concentration of glucose to optimize the hydrolysis parameters. Temperature (°C), time (h) and the rotation frequency or agitation speed (revolutions per min, RPM) of the incubator were the three chosen parameters. 29.84 ± 0.57 g L⁻¹ glucose was obtained from the banana peels. The peel extract after hydrolysis was used as the medium for microalgal growth, and the growth kinetics of the two microalgal isolates (Chlorella sorokiniana KMBM_I and Chlorella sorokiniana KMBM_K) cultivated in the waste peel hydrolysate were evaluated. Chlorella sorokiniana KMBM_K and Chlorella sorokiniana KMBM_I displayed a biomass yield of respectively 1.72 and 1.53 g L⁻¹ with a lipid content of 22.83% and 22.17% respectively. This study demonstrated that this organic peel waste can act as an effective, eco-friendly, efficient and natural substrate for cost effective microalgal cultivation.
In this paper, microalgae are considered to produce bioenergy through firing. Emitted CO2 would then be captured and stored. This “negative emissions” process, called Microalgae-based Bioenergy with Carbon Capture and Storage (MBECCS), is quantified for the first time, and compared to “classical” Bioenergy with Carbon Capture and Storage (BECCS) using plants. Our goal is to remove 10 Gt per year of CO2 from the atmosphere using MBECCS only. The needed area for cultivation is 4 times less than with BECCS. Because of the high water consumption, marine microalgae should be used. On the other hand, nutrients taken from world wastewater are insufficient, and remain an issue. Planktonic and biofilm microalgae are considered: the latter offer a better areal productivity and require less energy since they are in a concentrated form.
For both planktonic and biofilm microalgae, the process produces net heat. Concerning electrical efficiency, we show that it is better for biofilms, as expected. Moreover, for low productivities, MBECCS consumes electricity, comparably to Direct Air Capture, another energy-consuming negative emissions process. Finally, for higher productivities, we show that MBECCS produces net electricity, comparably to other renewable sources such as wind turbines and to BECCS.
Growth 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 outdoors 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 ≈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.
Chromochloris zofingiensis has obtained particular interest as a promising candidate for natural astaxanthin production. In this study, we established a two-stage heterotrophic cultivation process, by using which both the growth of C. zofingiensis and astaxanthin accumulation are substantially enhanced. Specifically, the ultrahigh biomass concentration of 221.3 g L⁻¹ was achieved under the optimum culture conditions in 7.5 L fermenter during 12 days. When scaled-up in the 500 L fermentor, the biomass yield reached 182.3 g L⁻¹ in 9 days, while the astaxanthin content was 0.068% of DW. To further promote astaxanthin accumulation, gibberellic Acid-3 (GA3) was screened from a variety of phytohormones and was combined with increased C/N ratio and NaCl concentration for induction. When C. zofingiensis was grown with the two-stage cultivation strategy, the astaxanthin yield reached 0.318 g L⁻¹, of which the biomass yield was 235.4 g L⁻¹ and astaxanthin content was 0.144% of DW. The content of the total fatty acids increased from 23 to 42% of DW simultaneously. Such an astaxanthin yield was 5.4-fold higher than the reported highest record and surpassed the level of Haematococcus pluvialis. This study demonstrated that heterotrophic cultivation of C. zofingiensis is competitive for industrial astaxanthin production.
Algae-derived protein has immense potential to provide high-quality protein foods for the expanding human population. To meet its potential, a broad range of scientific tools are required to identify optimal algal strains from the hundreds of thousands available and identify ideal growing conditions for strains that produce high-quality protein with functional benefits. A research pipeline that includes proteomics can provide a deeper interpretation of microalgal composition and biochemistry in the pursuit of these goals. To date, proteomic investigations have largely focused on pathways that involve lipid production in selected microalgae species. Herein, we report the current state of microalgal proteome measurement and discuss promising approaches for the development of protein-containing food products derived from algae.
Some mixotrophic microalgae appear to exceed the sum of photoautotrophy and heterotrophy in terms of biomass production. This paper mainly reviews the carbon and energy metabolism of microalgae to reveal the synergistic mechanisms of the mixotrophic mode from multiple aspects. It explains the shortcomings of photoautotrophic and heterotrophic growth, highlighting that the mixotrophic mode is not simply the sum of photoautotrophy and heterotrophy. Specifically, microalgae in mixotrophic mode can be divided into separate parts of photoautotrophic and heterotrophic cultures, and the synergistic parts of photoautotrophic culture enhance aerobic respiration and heterotrophic culture enhance the Calvin cycle. Additionally, this review argues that current deficiencies in mixotrophic culture can be improved by uncovering the synergistic mechanism of the mixotrophic mode, aiming to increase biomass growth and improve quality. This approach will enable the full utilization of advantagesin various fields, and provide research directions for future microalgal culture.
Chaetoceros, the most abundant genus of marine planktonic diatoms, can be used in mariculture. An effective genetic transformation system with a short transformation period was established in Chaetoceros muelleri by electroporation in our previous study. In this study, a sequence-specific clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 vector applicable for C. muelleri was constructed, and the expressions of sgRNA, resistance gene, and Cas9 gene were driven by the endogenous promoters U6, acetyl-CoA acetyltransferase, and fucoxanthin chlorophyll a/c binding protein, respectively, in the vector. Nitrate reductase (NR) and urease (URE) genes were edited in C. muelleri, and the NR knockout and NR/URE double-knockout lines displayed the strict auxotrophic phenotype. In addition, the DNA double-strand break was repaired by homologous recombination when a donor DNA was introduced. CRISPR/Cas9 technology was successfully applied to C. muelleri with an editing efficiency of up to 86%, providing a molecular tool for the study of basic biology in C. muelleri and its synthetic biology applications.
In recent years, the digital transformation of bioprocesses, which focuses on interconnectivity, online monitoring, process automation, artificial intelligence (AI) and machine learning (ML), and real-time data acquisition, has gained considerable attention. AI can systematically analyze and forecast high-dimensional data obtained from the operating dynamics of bioprocess, allowing for precise control and synchronization of the process to improve performance and efficiency. Data-driven bioprocessing is a promising technology for tackling emerging challenges in bioprocesses, such as resource availability, parameter dimensionality, nonlinearity, risk mitigation, and complex metabolisms. This special issue entitled “Machine Learning for Smart Bioprocesses (MLSB-2022)” was conceptualized to incorporate some of the recent advances in applications of emerging tools such as ML and AI in bioprocesses. This VSI: MLSB-2022 contains 23 manuscripts, and summarizes the major findings that can serve as a valuable resource for researchers to learn major advances in applications of ML and AI in bioprocesses.
Addition of phytohormones (diethyl aminoethyl hexanoate and indole acetic acid) at 10⁻⁵ M significantly increased the growth of a green microalga, Chlorella pyrenoidosa up to 62.25% achieving cell concentrations of 5.13╳10⁷ cells ml⁻¹. Yielding of high-value added byproducts (pigments, carbohydrate, proteins, and fatty acids) was more in presence of phytohormones comparing to the control. Further transcriptomics analysis demonstrated phytohormones upregulated numerous genes involved in DNA replication and repair pathways, and energy metabolisms (glycolysis, citrate cycle, and oxidative phosphorylation). Moreover, genes in purine metabolism, and porphyrin and chlorophyll metabolism were also more expressed by up to 6.49 times with phytohormones. These pathways can supply more cellular signaling molecules and antioxidant contents. This study carried new insights on the key driving factors in phytohormone enhanced microalgal biomass production processes, which can help to find more feasible application solutions of phytohormones in microalgal biofactories.
Technoeconomic analysis and life-cycle assessment are critical to guiding and prioritizing bench-scale experiments and to evaluating economic and environmental performance of biofuel or biochemical production processes at scale. Traditionally, commercial process simulation tools have been used to develop detailed models for these purposes. However, developing and running such models can be costly and computationally intensive, which limits the degree to which they can be shared and reproduced in the broader research community. This study evaluates the potential of an automated machine learning approach to develop surrogate models based on conventional process simulation models. The analysis focuses on several high-value biofuels and bioproducts for which pathways of production from biomass feedstocks have been well-established. The results demonstrate that surrogate models can be an accurate and effective tool for approximating the cost, mass and energy balance outputs of more complex process simulations at a fraction of the computational expense.
Carbon dioxide is the major greenhouse gas and regards as the critical issue of global warming and climate changes. The inconspicuous microalgae are responsible for 40% of carbon fixation among all photosynthetic plants along with a higher photosynthetic efficiency and convert the carbon into lipids, protein, pigments, and bioactive compounds. Genetic approach and metabolic engineering are applied to accelerate the growth rate and biomass of microalgae, hence achieve the mission of carbon neutrality. Meanwhile, CRISPR/Cas9 is efficiently to enhance the productivity of high-value compounds in microalgae for it is easier operation, more affordable and is able to regulate multiple genes simultaneously. The genetic engineering strategies provide the multidisciplinary concept to evolute and increase the CO2 fixation rate through Calvin–Benson–Bassham cycle. Therefore, the technologies, bioinformatics tools, systematic engineering approaches for carbon neutrality and circular economy are summarized and leading one step closer to the decarbonization society in this review.
Microalgae have been identified as a promising option for concomitant bioremediation of wastewaters and biodiesel generation. However, the nutritional composition of wastewater differs between locations, and some wastewater contains low nitrogen levels that inhibit microalgae growth, resulting in poor total nutrient removal and biomass output. Furthermore, the high cost of harvesting microalgae biomass is a barrier to commercial microalgae farming. This study aims to employ a novel microalga, Scenedesmus sp. DDVG I for bioremediation of low-nitrogen domestic wastewater. The nutritional balance of the wastewater was achieved with the addition of urea at two different levels and then was used for the cultivation of microalgae. The results showed that the optimized nutritional condition included total nitrogen (TN) of 250 mg/L and total phosphorus (TP) of 5 mg/L (N250/P5). The culture cultivated in this optimal medium improved chemical oxygen demand (COD), TN, and TP removal by as much as 89.5 %, 99.98 %, and 99.1 %, respectively. Scenedesmus sp. DDVG I growth was likewise at its peak, with a value of 5.1 g/L, but with a low harvesting efficiency (0.02%). Subsequently, Scenedesmus sp. DDVG I was co-cultured with a novel cyanobacterium, Limnothrix sp. DDVG II under the optimal nutritional condition. The results revealed a 99 % increase in harvesting efficiency with effective wastewater treatment. The co-culture biodiesel also complied with the standard specifications. Further, analysis of the essential amino acid content and in vitro digestibility of the biomass proved its usefulness as a feed supplement. Overall, this study demonstrated a promising future for a cleaner environment and sustainable bio-economy of microalgae.
The culture method using sodium acetate and glucose, widely used as organic carbon sources in the mixotrophy of Haematococcus pluvialis, was compared with its autotrophy. In the 12-day culture, mixotrophy using sodium acetate and glucose increased by 40.4% and 77.1%, respectively, compared to autotrophy, but the mechanisms for the increasing biomass were different. The analysis of the mechanism was divided into autotrophic and heterotrophic metabolism. The mixotrophy with glucose increased the biomass by directly supplying the substrate and ATP to the TCA cycle while inhibiting photosynthesis. Gene expressions related to glycolysis and carbon fixation pathway were confirmed in autotrophy and mixotrophy with glucose and acetate. The metabolism predicted in the mixotrophy with acetate and glucose was proposed via autotrophic and heterotrophic metabolism analysis. The mechanism of Haematococcus pluvialis under mixotrophic conditions with high CO2 concentration was confirmed through this study.
Excessive carbon dioxide (CO2) emissions into the atmosphere have become a
dire threat to the human race and environmental sustainability. The ultimate
goal of net zero emissions requires combined efforts on CO2 sequestration
(natural sinks, biomass fixation, engineered approaches) and reduction in CO2
emissions while delivering economic growth (CO2 valorization for a circular
carbon bioeconomy, CCE). We discuss microalgae-based CO2 biosequestration,
including flue gas cultivation, biotechnological approaches for enhanced CO2
biosequestration, technological innovations for microalgal cultivation, and CO2
valorization/biofuel productions. We highlight challenges to current practices
and future perspectiveswith the goal of contributing to environmental sustainability,
net zero emissions, and the CCE.
For commercial scale algal biorefining, harvesting cost is a major bottleneck. Thus, a cost-effective, less-energy intensive, and efficient harvesting method is being investigated. Among several harvesting methods, magnetic flocculation offers the benefits of modest operation, energy savings and quick separation. This study aims to develop novel magnetite-(Fe3O4) nanoparticles (MNPs) of 20 nm average size and their high reusability potential to reduce the harvesting cost of microalgae biomass. The MNPs were synthesized and characterized using FTIR, Zeta analyzer, and SEM before performing on Chlorella sorokiniana Kh12 and Tu5. For maximum harvesting efficiency >99%, the optimal culture pH, MNPs concentration, and agitation speed were 3, 200 mg/L, and 450 rpm, respectively. Subsequently, MNPs were recovered via alkaline treatment and reused up to 5 cycles as they retained their reactivity and harvesting efficiency. The studied MNPs-based harvesting method could be adopted at a commercial scale for cost-effective algae biorefinery in the future.
Microalgae are a promising resource for biofuel production, although the lack of effective harvesting techniques limits their industrial use. In this context, flotation, and in particular dissolved air flotation (DAF), is an interesting separation technique that could drastically reduce harvesting costs and make biofuel-production systems more economically viable. But because of the repulsive interaction between cells and bubbles in water, the efficiency of this technique can be limited. To solve this problem, we propose here an original DAF process where bubbles are functionalized with a bio-sourced polymer able to specifically bind to the surface of cells, chitosan. In a first part, we modify chitosan by adding hydrophobic groups on its backbone to obtain an amphiphilic molecule, PO-chitosan, able to assemble at the surface of bubbles. Then, using a recently developed technique based on atomic force microscopy (AFM) combined with microfluidics, we probe the interactions between PO-chitosan coated bubbles and cells at the molecular scale; results show an enhanced adhesion of functionalized bubbles to cells (from 3.5 to 12.8 nN) that is pH-dependent. Separation efficiencies obtained in flotation experiments with functionalized bubbles are in line with AFM data, and a microalgae separation efficiency of approximately 60% could be reached in a single step. In addition, we also found that PO-chitosan could be used efficiently as a flocculant (nearly 100% of cells removed), and in this case AFM experiments revealed that the flocculation mechanism is based on hydrophobic interactions between cells and PO-chitosan. Altogether, this comprehensive study shows the interest of PO-chitosan to harvest cells in flotation or flocculation/flotation processes.
Microalgae are photosynthetic microbes that can synthesize compounds of therapeutic potential with wide applications in the food, bioprocessing and pharmaceutical sector. Recent research advances have therefore, focused on finding suitable economic substrates for the sustainable cultivation of microalgae. Among such substrates, food derived waste specifically from the starch, meat, dairy, brewery, oil and fruit and vegetable processing industries has gained popularity but poses numerous challenges. Pretreatment, dilution of waste water supernatants, mixing of different food waste streams, utilizing two-stage cultivation and other biorefinery approaches have been intensively explored for multi-fold improvement in microalgal biomass recovery from food waste. This review discusses the advances and challenges associated with cultivation of microalgae on food waste. The review suggests that there is a need to standardize different waste substrates in terms of general composition, genetically engineered microalgal strains, tackling process scalability issues, controlling wastewater toxicity and establishing a waste transportation chain.
The mixotrophic cultivation approach, consisting of heterotrophy and photoautotrophy, has been suggested as a promising strategy to reach a high growth rate and biomass productivity. However, the underlying growth-boosting mechanisms remain elusive. In this study, Chromochloris zofingiensis, a promising high-value-added astaxanthin producer, was selected to elucidate how mixotrophy benefits cellular growth. Results showed that mixotrophy yielded the highest biomass, which was significantly higher than the sum of photoautotrophic and heterotrophic cultivation, indicating a significant growth-boosting effect. The chlorophyll fluorescence and transcriptomics results confirmed that the highest biomass production under mixotrophy was attributed to a higher photosynthesis performance, decreased light damage and photorespiration, as well as significantly upregulated glucose metabolism compared to photoautotrophy. Compared to heterotrophy, mixotrophy consumed less glucose and had a lower glucose metabolic efficiency, but led to higher biomass yield. Taken together, we propose a synergistic growth-promoting mechanism between photosynthesis and glucose metabolism under mixotrophy, in which cells not only directly benefit from extra energy from exogenous organic carbon metabolism but also the reutilization of the byproduct CO2 for photosynthesis. In return, O2 evolved from photosynthesis was consumed by glucose metabolism as well. Furthermore, the coexistence of photosynthesis and glucose metabolism contributes to higher energy usage efficiency and biomass yield, correspondingly diminishing the waste and decreasing the damage. This study provides insights into the synergistic mechanism under mixotrophy and helps to better select growth strategies for commercially valuable algal species.
Techno-economic analysis (TEA) has been widely utilized to evaluate the commercial viability of different microalgae cultivation systems, including open raceway ponds, photobioreactors, and algal turf scrubbers. A disparity in the overall findings of the studies has been observed due to differences in the boundary conditions and selection of TEA parameters. The present review aims to provide a critical evaluation of scientific literature on TEA of microalgae cultivation systems, published in the last decade, to provide a roadmap for future research. The microalgae cultivation systems and their design, dynamics, and associated expenses were thoroughly reviewed. Emphasis was given to highlight the factors influencing the system economics. Moving from local boundaries, the effect of geospatial variability on economic performance and the scalability aspects of cultivation systems was accentuated. Data based on assumptions, simulations, and hypothetical plant layouts was observed to result in biased findings. Henceforth, step-wise process scale-up from lab to target capacity, careful inclusion of influence of environmental conditions on associated expenses, and appropriate selection of TEA parameters is recommended.
In this study, Auxenochlorella protothecoides (AP-CK) was selected due to its reported high growth potential in sterilized black and odorous water (SBOW). In order to improve the resource utilization level of microalgae for wastewater treatment, AP-CK was mutated using ¹²C⁶⁺ heavy-ion beam irradiation, and a high lipid-containing mutant (AP-34#) was isolated and further evaluated to treat original black and odorous water (OBOW). Compared with the wild type, the maximum removal rates of COD, NH4⁺-N and TP of the mutant increased by 8.12 ± 0.33%, 10.43 ± 0.54% and 11.97 ± 0.16%, respectively, while maximum dissolved oxygen content increased from 0 to 4.36 ± 0.25 mg/L. Besides, the mutant lipid yield increased by 115.87 ± 3.22% over the wild type in OBOW. The fatty acid profile of AP-34# grown in SBOW and OBOW showed higher proportion of saturated fatty acids (C16:0 and C18:0) and valuable polyunsaturated fatty acids (mainly C20:5n3 and C22:6n3) which are more suitable for biodiesel production and value-added products, respectively. This work provides a new perspective on improving the characteristics of microalgae and an innovative approach for resource-based microalgae wastewater treatment through bioremediation of black and odorous water.
Microalgae produced lipids, pigments, lutein, and valuable chemicals under stressful conditions, making them as attractive feedstock for biorefineries. However, high cell density culture of microalgae in harsh condition using seawater is still a challenge. In this study, we used aminobutyric acid (GABA) from an in vitro biotransformation of monosodium glutamate (MSG) to alleviate salinity stress in Chlorella sorokiniana (Cs). Initially, Chlorophyll a/b content was suppressed under the stress of 20 g/L sodium chloride (NaCl), leading to an extreme decline in protein and pigment accumulation. While the biomass and protein of Cs cultured in 10 g/L NaCl reached 3.44 g/L and 1.14 g/L with 5 mM GABA, with 56.3% and 171% increase compared to control. The optimal conditions were 10 mM GABA and 20 g/L NaCl in modified BG11 medium, with a biomass, protein, lutein and β-carotene content of 4.56 g/L, 2.07 g/L, 24 mg/L and 27.48 mg/L, respectively. Finally, protein from Cs was used as a prebiotic for cultivating Escherichia coli Nissle 1917 (EcN) and Lactobacillus rhamnosus ZY (LGG). We provided an effective and sustainable approach for the simultaneous production of biomass and high-end products by microalgae using GABA and seawater for the first time.
This study determined that 60% is the most appropriate concentration of CO2 for the domestication of microalgae to obtain strains with improved flue gas CO2-adapting ability. The effect of long-term high CO2 stress (6–99% concentrations) on microalgal gene mutations was first clarified with genomic and transcriptomic analyses. The most beneficial long fragment indel/SV gene mutations in microalgae were obtained under 60% CO2. However, > 60% CO2 domestication caused genotoxicity of the microalgae cells via the following mechanisms: (1) it was not conducive to forming more stable long fragment indel/SV gene mutations, thus preventing further gene mutation; (2) gene mutations did not generate successful linkage to the regulation in transcription and translation; and (3) inhibition of the mismatch repair damaged the specialized ability of genetic variation, leading to the disrepair of harmful gene mismatches and fewer beneficial mutations. These novel results revealed that higher concentrations of CO2 for microalgal domestication did not necessarily result in microalgae that were tolerant to CO2 owing to the genotoxicity of long-term high CO2 stress. This conclusion informs futures efforts of domestication in the microalgae industry.
Microalgae are in the focus of research and industry because of their potential as high-value substance producers. Especially diatoms and within this group Phaeodactylum tricornutum are of high interest as biofactory, for e.g. synthesizing high amounts of fucoxanthin or as important source for ω-3 fatty acids. As model organism P. tricornutum can be used for genetic engineering. Here we showed the potential of P. tricornutum as efficient biofactory under mixotrophic conditions, combining the advantages of hetero- and phototrophic growth. We expressed the hexose uptake protein (HUP1) from the Chlorophyte Chlorella kessleri under the control of the light-inducible promoter fcpA or the nitrate reductase promoter. Only in case of fcpA significant overexpression could be demonstrated. With an in situ localization study, we demonstrated that the N-terminal sequence of the heterologous HUP1 protein is sufficient for insertion in the ER, whereas only the full-length protein is correctly targeted to the plasma membrane. The heterologous expression of HUP1 enables the cells to take up glucose, leading to increased biomass, especially when grown under low light intensities. The results demonstrate the capability of P. tricornutum to grow more efficiently using glucose and shed light on the targeting pathway of plasma membrane transporters.
Contaminating microzooplankton have a great influence on the growth of microalgae. To illustrate how harmful contaminants affect the growth of Chlorella sorokiniana and to investigate the microzooplankton community composition, the succession of microzooplankton contaminating pilot-scale C. sorokiniana cultures was surveyed in four different seasons. Nineteen species of microzooplankton (including a cyst) were detected in the C. sorokiniana cultures, with the flagellate Poterioochromonas malhamensis and the amoeba Vannella sp. being the most harmful species to C. sorokiniana due to their high abundance and ability to graze C. sorokiniana, resulting in a decrease of 38% to 59% of C. sorokiniana production. Once the P. malhamensis abundance reached more than 1.0 × 10⁴ ind. mL⁻¹ or the Vannella sp. abundance exceeded 2.0 × 10³ ind. mL⁻¹, the C. sorokiniana population declined dramatically. More species and a greater abundance of microzooplankton were found in summer with the lowest algal biomass by the end of the cultivation. The occurrence of microzooplankton in the C. sorokiniana culture system showed a certain regularity. Network analysis indicated that the two microzooplankton species of P. malhamensis and Vannella sp. had positive correlations with the growth of C. sorokiniana and the occurrence of the other contaminating microzooplankton. Overall, this study provided novel insights into the microzooplankton community and their co-occurrence networks during C. sorokiniana cultivation, which will help provide reliable theoretical support for the effective control of contaminating microzooplankton.
Sunlight-driven microalgal conversion of CO2 is a promising carbon neutral strategy for the sustainable production of various value-added products. However, its real-world application is challenging due to the irregularity of sunlight which reduces photosynthetic efficiency. Here, we report that microdroplet-based screening of photosynthesis-augmented microalgae mutant under fluctuating light accelerates mass production of CO2-derived algal biofuel in natural sunlight. Random insertional Chlamydomonas reinhardtii mutants were cultivated in single-layered microdroplet photobioreactors to be effectively applied to light conditions mimicking natural sunlight without self-shading. High-density microdroplets containing fast-growing mutants were efficiently selected at the single-droplet level by a centrifugation-assisted droplet sorting platform for high-throughput screening. Consequently, we isolated a mutant exhibiting cell growth 1.85-fold that of the wild-type even under continuous and rapid alteration in light intensity, which can serve as a cell stressor. After whole-genome resequencing, we found disruption in the SNF2 gene encoding ATP-dependent chromatin remodeling complexes, which reveal its importance for the regulation of chlorophyll biosynthesis and accumulation of reactive oxygen species under fluctuating light. After a 1.6 ton-scale field culture utilizing natural sunlight and industrial CO2, the mutant showed increased biomass productivity, CO2 fixation rate, and calorific value, a crucial parameter of fuel performance, by 34.38%, 45.07%, and 16.82%, respectively. Our results indicate that improving photosynthesis of microalgae in fluctuating light environments elucidates the mechanisms responsible for enhancing the sunlight utilization, allowing industrial-scale biological CO2 conversion.
Although microalgae biofuel has many advantages, its production is faced with difficulties regarding algal cell harvesting and low lipid content. In this experiment, mixtures of seawater and domestic sewage were prepared for the cultivation of freshwater microalgae. The results showed that the proportion of seawater had an important effect on the precipitation and biofuel production of the microalgae. The increasing proportion of seawater in the mixed wastewater improved the sedimentation performance and lipid content of microalgae, but the growth of microalgae was greatly inhibited under 80% seawater stress. Therefore, the highest algal lipid productivity of 10.74 mg L⁻¹ d⁻¹ was achieved in the wastewater containing 60% seawater, which was 2.15-fold higher than that of the control. Furthermore, the fatty acid saturation and the C16-C18 fatty acid proportion of the microalgae cultured in wastewater containing 60% seawater were relatively ideal, which was beneficial for improving the quality of biodiesel. Therefore, wastewater containing 60% seawater was the most favourable for the cultivation of biofuel-producing microalgae. The mechanism by which seawater improved the precipitation performance of freshwater microalgae was also investigated. The addition of seawater to the sewage increased the cell diameter of the microalgae, reduced the zeta potential of microalgal colloids, and promoted the synthesis of extracellular polymeric substances by the microalgal cells. Thus, the precipitation performance of the microalgae cells was effectively improved. The pigment contents and antioxidant enzyme activities of the microalgae were also determined, and it was found that the glutathione peroxidase activity of microalgae increased under certain seawater stress, indicating that glutathione peroxidase played an indispensable role in the removal of reactive oxygen species produced by seawater stress.
Chlamydomonas reinhardtii is a model microalga that has a higher growth rate and produces high levels of lutein and lipids, but biomass production is limited. Carbonic anhydrase (CA) converts atmospheric CO2 to bicarbonate which is crucial for carbon‐concentrating mechanism (CCM) in microalgae and boosts cell density. Therefore, C. reinhardtii harboring the heterologous CA from Mesorhizobium loti (MlCA) and Sulfurihydrogenibium yellowstonense (SyCA) were explored to increase CO2 capture and utilization (CCU) through different culture devices. Genetically modified C. reinhardtii was able to grow from mixotrophic to autotrophic conditions. Subsequently, biomass, lutein, and lipid were maximized to OD680 of 4.56, 21.32 mg/L and 672 mg/L using photo-bioreactor (PBR) with 5% CO2. Moreover, CO2 assimilation rate was 2.748 g-CO2/g-DCW and 2.792 g-CO2/g-DCW under mixotrophic and autotrophic conditions, respectively. The biomass accumulation correlated with CA activity. In addition, the transcript levels of major genes in metabolic pathways of lutein and lipid were dramatically increased.
Microalgae express high protein levels and can be produced in contained cultivation systems with low water requirements and complete fertilizer use. The production potential is 22–44 tons of protein per hectare per year although the current production scale is small. Techno economic analyses have shown good potential for scale-up and cost reduction. Large-scale production of microalgae in the post-fossil era will rely on the capture of carbon dioxide from the air, or sugars from crops. Microalgal amino acid composition matches well with requirements for food and feed, which, in combination with novel biomass pre-treatment steps, will guarantee high-quality microalgal protein. For a broadening of the microalgae species available as single-cell protein, novel food approval is required.
The global trend is shifting toward circular economy systems. It is a sustainable environmental approach that sustains economic growth from the use of resources while minimizing environmental impacts. The multiple industrial use of microalgal biomass has received great attention due to its high content of essential nutrients and elements. Nevertheless, low biomass productivity, unbalanced carbon to nitrogen (C/N) ratio, resistant cellular constituents, and the high cost of microalgal harvesting represent the major obstacles for valorization algal biomass. In recent years, microalgae biomass has been candidate as a potential feedstock for different bioenergy generation with simultaneous treating wastewater and CO2 capture. An overview of the appealing features and needed advancements are urgently essential for microalgae-derived bioenergy generation. The present review provides a timely outlook and evaluation of biomethane production from microalgal biomass and related challenges. Moreover, the biogas recovery potential from microalgal biomass through different pretreatment and synergism anaerobic co-digestion (AcoD) with another biowastes are evaluated. In addition, the removal of micropollutants and heavy metals by microalgal cells via adsorption and bioaccumulation in their biomass are discussed. Herein, a comprehensive review is presented about a successive high-throughput for anaerobic digestion (AD) of the microalgal biomass in order to achieve for sustainable energy source. Lastly, the valorization of the digestate from AD of microalgae for agricultural reuse is highlighted.