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Abstract
Algae have long been recognized as potential feedstock to produce oils. In recent years, the use of algae, in particular, Chlorella, for heterotrophic oil production has gained increasing interest due to its fast growth, ultrahigh cell density, and superior oil productivity. The current technology for heterotrophic production of algal oils, however, is still far from economically viable because of its high production cost. The opportunities that lie ahead for improving the production economics of heterotrophic algal oils will be the advances in exploration of low-cost carbon alternatives, advanced culture systems, and genetic engineering of algal strains for improvement as well as the biorefinery-based integrated production of oils and coproducts. Breakthroughs and innovations in these areas are sought to expand heterotrophic production of algae from high-value products to cheap commodity products of oils.
... However, we are far from a consensus about the most economic production mode. While some authors claimed heterotrophic production as the most cost-viable method [12,13], there is also a movement that strongly encouraged the use of photosynthetic microalgae as the best option due to economic advantages [2,14]. Albeit apparently none of these opinions were based on an impartial comparison arising from a techno-economic assessment. ...
... Glucose is used as the substrate, at a cost of 0.44 €⋅kg − 1 , and associated mineral nutrients [23,24]. This is the preferred organic source [14], with a growth yield for heterotrophic Chlorella spp. of 0.35 g biomass⋅g sugar − 1 [23]. ...
... Nineteen fermenters, with a working volume of 250.7 m 3 each, are then required for this base case. As a reference, large-scale production of heterotrophic microalgae was already done in the '90s, in fermenters with capacities up to 150 m 3 [14]. Nowadays, Solazyme, Roquette, and Corbion have fermentation tanks with even greater capacities. ...
Heterotrophic or autotrophic? This is the continuous question the industry faces when microalgae production is the endeavor. Surprisingly, nowadays specialists have not reached a consensus on which is the most economical option. The current work analyses costs for heterotrophic and autotrophic cultivation of microalgae at an industrial scale. Heterotrophic cultivation of microalgae results in a production cost of 4.00 €·kg⁻¹ of dry weight as a centrifuged paste. This is within the range of autotrophic costs, but still above the production cost in some photobioreactors. The study also identifies the current limitations on the technology and studies the effect on the cost of overcoming these. Once achieved, the advances in the process could result in a heterotrophic production cost reduced to 1.08 €·kg⁻¹. Autotrophic cultivation seems competitive with heterotrophic production. It is time to leap forward in the autotrophic production scale to achieve the critical reduction in production cost.
... Among them, N limitation has been broadly reported as an effective approach for overproduction of storage lipids in oleaginous microalgal species [12]. In heterotrophic cultures, organic carbon sources are used, and carbon/nitrogen (C/N) ratio controls the switch between protein and lipid synthesis; thus, the C/N ratio of culture media is one of the most critical nutritional factors affecting lipid content [13]. It was reported that the initial C/N ratio must be greater than 20 for maximum lipid production by oleaginous microorganisms [14]. ...
... When the glucose concentration was more than 60 g/L, biomass no longer increased but decreased. The decrease in biomass was likely due to high glucose inhibition, which is a common phenomenon in heterotrophic batch cultures [13]. The highest glucose-to-biomass conversion rate was 43.4%, which was obtained at a glucose concentration of 20 g/L. ...
... When the glucose concentration exceeded 40 g/L, the lipid content and lipid productivity no longer increased (Fig. 1g, h). In photoautotrophic culture, stress conditions commonly increase lipid content but decrease the biomass, whereas heterotrophic culture simultaneously accumulates lipids and biomass [13]. It was verified in T. bernardii cultured heterotrophically in both NaNO 3 and urea as a nitrogen source. ...
Background
Heterotrophic cultivation of microalgae has been proposed as a viable alternative method for novel high-value biomolecules, enriched biomass, and biofuel production because of their allowance of high cell density levels, as well as simple production technology. Tetradesmus bernardii , a newly isolated high-yielding oleaginous microalga under photoautotrophic conditions, is able to grow heterotrophically, meaning that it can consume organic carbon sources in dark condition. We investigated the effect of different carbon/nitrogen (C/N) ratios on the growth and lipid accumulation of T. bernardii in heterotrophic batch culture under two nitrogen sources (NaNO 3 and CO(NH 2 ) 2 ). In addition, we conducted time-resolved transcriptome analysis to reveal the metabolic mechanism of T. bernardii in heterotrophic culture.
Results
T. bernardii can accumulate high biomass concentrations in heterotrophic batch culture where the highest biomass of 46.09 g/L was achieved at 100 g/L glucose concentration. The rate of glucose to biomass exceeded 55% when the glucose concentration was less than 80 g/L, and the C/N ratio was 44 at urea treatment. The culture was beneficial to lipid accumulation at a C/N ratio between 110 and 130. NaNO 3 used as a nitrogen source enhanced the lipid content more than urea, and the highest lipid content was 45% of dry weight. We performed RNA-seq to analyze the time-resolved transcriptome of T. bernardii . As the nitrogen was consumed in the medium, nitrogen metabolism-related genes were significantly up-regulated to speed up the N metabolic cycle. As chloroplasts were destroyed in the dark, the metabolism of cells was transferred from chloroplasts to cytoplasm. However, storage of carbohydrate in chloroplast remained active, mainly the synthesis of starch, and the precursor of starch synthesis in heterotrophic culture may largely come from the absorption of organic carbon source (glucose). With regard to lipid metabolism, the related genes of fatty acid synthesis in low nitrogen concentration increased gradually with the extension of cultivation time.
Conclusion
T. bernardii exhibited rapid growth and high lipid accumulation in heterotrophic culture. It may be a potential candidate for biomass and biofuel production. Transcriptome analysis showed that multilevel regulation ensured the conversion from carbon to the synthesis of carbohydrate and lipid.
... Among the nutrients, carbon is the most prominent element of algae and accounts for approximately 50% of the biomass. Algae can grow photoautotrophically by utilizing carbon dioxide or heterotrophically with organic carbon such as sugar to achieve high cell density (Liu et al. 2011(Liu et al. , 2014aSirisansaneeyakul et al. 2011;Breuer et al. 2012). Nitrogen and phosphorus represent the additional important nutrients and their availability impacts algal growth and lipid production considerably (Breuer et al. 2012;Liu et al. 2013;Sun et al. 2015Sun et al. , 2018. ...
... Many algal strains are able to utilize glucose for growth and lipid production (Liu et al. 2014a), as they have a hexose/H + symport system to transport glucose into cytosol for metabolism (Tanner 2000). We firstly tested the effect of glucose on growth and lipid production of M. inermum JL1 under light ( Fig. 4a-d). ...
... The heterotrophic lipid productivity (0.68 g L −1 day −1 ) for M. inermum JL1 is considerably greater than the photoautotrophic one. Furthermore, the welldeveloped fermentation technologies, e.g., fed-batch cultivation, continuous cultivation, and continuous cultivation with cell recycling (Liu et al. 2014a), can be applied to M. inermum JL1 for achieving much higher yield and productivity. However, glucose is used as the carbon source for growth, adding cost to heterotrophic production. ...
Molecular characterization based on the internal transcribed spacer 2 (ITS2) sequence identified Micractinium inermum JL1, a local isolate of green alga capable of growing robustly under photoautotrophic, mixotrophic, and heterotrophic culture modes. Physiochemical analyses of the alga revealed great variations in growth, lipid content, oleic acid abundance, and productivities of lipids and triacylglycerol (TAG) under various culture conditions including nitrogen concentrations, phosphorus concentrations, light intensities, salinity levels, and glucose concentrations with or without light illumination. The carbon shunt from starch and to a lesser extent from membrane lipids likely contributed to the TAG accumulation. Noteworthily, salt supplementation promoted TAG content and productivity by ~ 88% and ~ 55%, respectively. The alga could accumulate ~ 52% lipids per dry weight and achieved a high lipid productivity of 0.68 g L⁻¹ day⁻¹. C18:1 percentage, a biodiesel quality indicator, reached ~ 50%, and correlated well with TAG contents in a positive manner, demonstrating the feasibility of using C18:1 percentage for TAG quantification thereby avoiding time- and labor-intensive analysis. These results together indicate the potential of M. inermum JL1 as a lipid producer for future exploration.
... Based on the obtained results, this species only grew well on glucose at 0.02 and 0.05 M concentrations. This finding was not surprising, as glucose is the most commonly used carbon source for the heterotrophic culture of microalgae (Park et al. 2002;Liang et al. 2009;Li et al. 2013;Liu et al. 2014;Gim et al. 2016). A study by Gim et al. (2016) showed that the supplementation of glucose led to a significant improvement in the growth of I. galbana, Nannochloropsis oculata and Dunaliella salina compared to other studied carbon sources. ...
... This activated system transports glucose and H + with a stoichiometry of 1:1 (Komor et al. 1973) into the cytosol and invests one molecule of ATP per molecule of sugar transported (Tanner 2000). The oxidative assimilation of glucose begins with the phosphorylation of hexose to form glucose-6-phosphate, which is an important intermediate product for storage, cell synthesis and respiration (Perez-Garcia et al. 2011;Liu et al. 2014), yielding the reducing equivalents (FADH 2 and NADPH) from the ATP synthesis. Hence, all of the produced ATP is used for cell growth and lipid accumulation (Gim et al. 2016). ...
... The low maximum cell density at 0.1 M (19.8 g L −1 ) recorded for all carbon sources was most likely due to an excess of organic substrate, which inhibited the growth of I. maritima (Perez-Garcia et al. 2011;Liu et al. 2014). Ogawa and Aiba (1981) suggested that the initial concentration of glucose should be limited to 10 and 1 g L −1 to promote the cellular growth of Chlorella vulgaris and Scenedesmus acutus, respectively. ...
The growth and biochemical composition of photoautotrophic and heterotrophic Isochrysis maritima in 50 L of Walne’s medium were compared. Heterotrophic I. maritima fed with 0.02 M glucose had a 4.6-fold higher maximum cell density (38.17 ± 0.23 × 10⁶ cells mL⁻¹) than photoautotrophic cells (8.29 ± 0.70 × 10⁶ cells mL⁻¹). The carbohydrate content was slightly higher in heterotrophic cells at all growth stages (mid-exponential, 40.8%; early stationary, 48.3%; and late stationary, 47.6%), but there was no significant effect on the protein content under either trophic condition. The total saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs) were higher under heterotrophic conditions than those under photoautotrophic conditions. However, because omega-3 PUFAs are the most essential element in feed nutrition, low results for eicosapentaenoic acid (EPA) (0.28 ± 0.06%) and docosahexaenoic acid (DHA) (3.22 ± 0.26%) in the heterotrophic cells compared to the photoautotrophic cells (EPA: 0.44 ± 0.11%; DHA: 8.58 ± 0.73%) plus a low omega-3/6 PUFAs ratio (heterotrophic: 0.16–0.47; photoautotrophic: 2.60–2.88) and high value of (SFA + MUFA)/PUFA (heterotrophic: 5.50–6.81; photoautotrophic: 2.64–3.60) showed that this species is not suitable for aquaculture feed when cultivated under heterotrophic conditions.
... Up to now, many species of microalgae, such as species of Chlorella, Euglena, and Scenedesmus, have been cultivated on an industrial scale to achieve high cell densities using fermenters (Jin et al., 2020). Generally, the biomass of these microalgae achieved by chemoheterotrophic cultivation is 10 times more than that achieved by autotrophic cultivation (Liu et al., 2014). For instance, a maximum biomass of 32.5 g L −1 for Ochromonas danica has been achieved by heterotrophic fermentation (Lin et al., 2014). ...
... The present study provides a promising fermentation technology for acquiring abundant P. malhamensis cells, and the maximum biomass achieved (more than 3 × 10 8 cells mL −1 ) in this study exceeds that reported in other studies (Lewitus & Caron, 1991;Rottberger et al., 2013) by more than an order of magnitude. In recent years, fermentation of microalgae has attracted increasing interest due to the ultrahigh cell densities produced, fast growth, and low occurrence of contaminants (Liu et al., 2014). Our results provide useful insights into the optimal culture conditions, in terms of nutrients and environmental factors, for high-cell-density fermentation of P. malhamensis. ...
The chrysophyte Poterioochromonas malhamensis has potential for controlling algal blooms through rapid grazing of toxic Microcystis cells and efficient degradation of microcystin. However, this method has not been used in practice because a high-cell-density method for cultivating P. malhamensis has not yet been established and the actual effect of the chrysophyte in controlling Microcystis blooms in the field is still unknown. To achieve the application of this method, high-cell-density heterotrophic cultivation of P. malhamensis was established through optimizing the carbon/glucose concentration, C:N ratio, temperature, pH, and dissolved oxygen concentration. Under optimized conditions, the cell concentration of P. malhamensis reached more than 3 × 10⁸ cells mL⁻¹, which exceeds that reported in other studies by more than an order of magnitude. The ability of the chemoheterotrophic P. malhamensis to graze unicellular Microcystis cells was comparable to that of autotrophic and phagotrophic P. malhamensis. A controlled field experiment showed that chemoheterotrophic P. malhamensis could live in the aquatic environment with a Microcystis bloom and decrease the Microcystis biomass on the surface of the water by promoting the sedimentation of colonial Microcystis cells. This study offers an opportunity to drive the development of methods to control Microcystis blooms using predatory P. malhamensis.
... When the glucose concentration was more than 60 g/L, biomass no longer increased but decreased. The decrease in biomass was likely due to high glucose inhibition, which is a common phenomenon in heterotrophic batch cultures [13]. The highest glucose-to-biomass conversion rate was 43.4%, which was obtained at a glucose concentration of 20 g/L. ...
... When the glucose concentration exceeded 40 g/L, the lipid content and lipid productivity no longer increased (Fig. 1g, h). In photoautotrophic culture, stress conditions commonly increase lipid content but decrease the biomass, whereas heterotrophic culture simultaneously accumulate lipids and biomass [13]. It was veri ed in T. bernardii cultured heterotrophically both NaNO 3 or urea as a nitrogen source. ...
Background: Heterotrophic cultivation of microalgae has been proposed as a viable alternative method for novel high-value biomolecules, enriched biomass and biofuel production because of their allowance of high cell density levels, as well as simple production technology. Tetradesmus bernardii, a newly isolated high-yielding oleaginous microalga under photoautotrophic conditions, is able to grow heterotrophically, meaning that it can consume organic carbon sources in dark condition. We investigated the effect of different carbon/nitrogen (C/N) ratios on the growth and lipid accumulation of T. bernardii in heterotrophic batch culture under two nitrogen sources (NaNO3, CO(NH2)2). In addition, we conducted time-resolved transcriptome analysis to reveal the metabolic mechanism of T. bernardii in heterotrophic culture.
Results: T. bernardii can accumulated high biomass concentrations in heterotrophic batch culture which the highest biomass of 46.09 g/L was achieved at 100 g/L glucose concentration. The rate of glucose to biomass was exceed 55% when the glucose concentration was less than 80 g/L, and the C/N ratio was 44 at urea treatment. The culture was beneficial to lipid accumulation at a C/N ratio between 110 and 130. NaNO3 used as a nitrogen source enhanced the lipid content more than urea, and the highest lipid content was 45% of dry weight. We performed RNA-seq to analyze the time-resolved transcriptome of T. bernardii. As the nitrogen was consumed in the medium, nitrogen metabolism related genes were significantly up-regulated to speed up the N metabolic cycle. As chloroplasts were destroyed in the dark, the metabolism of cells was transferred from chloroplasts to cytoplasm. However, storage of carbohydrate in chloroplast remained active, mainly the synthesis of starch, and the precursor of starch synthesis in heterotrophic culture may largely came from the absorption of organic carbon source (glucose). With regard to lipid metabolism, the related genes of fatty acid synthesis in low nitrogen concentration increased gradually with the extension of cultivation time.
Conclusion: T. bernardii exhibited rapid growth and high lipid accumulation in heterotrophic culture. It may be a potential candidate for biomass and biofuel production. Transcriptome analysis showed that multilevel regulation ensured the conversion from carbon to the synthesis of carbohydrate and lipid.
... Industrially, they are mostly used in biofuel refineries, as food additives and as health-promoting supplements (Chisti 2007;Koller et al. 2014;Mata et al. 2010). Microalgae can produce a wide range and large amounts of lipids, i.e. contents from 4.8 to 60% of dry weight (DW) depending on the algal strain and culture conditions (Liu et al. 2014b;Sajjadi et al. 2018). Among all the range of lipids, neutral lipids are the most interesting ones, in particular, triacylglycerol (TAG) which may account for 80% of total neutral lipids (Morales-Sánchez et al. 2014). ...
... These carbon sources may be used at different concentrations which were found to impact the lipid distribution. Indeed, when sugar concentration increases, the proportion of neutral lipids and TAGs tends to increase and, in contrast, the content of membrane lipids (phospholipids and glycolipids) tends to decrease (Liu et al. 2014b). Moreover, saturation of the fatty acids is directly dependent on the amount of excess sugars: as the concentration of the sugars increases, the fatty acids become more saturated (Wood et al. 1999). ...
Microalgae constitute a phylogenetically diverse group of eukaryotic photosynthetic microorganisms that thrive in various environments. They are the subject of increasing interest from both the scientific community and the industrial world due, on the one hand, to their ecological significance at the global scale and, on the other hand, to their large potential as sources of economically important products, from biofuels to new pharmaceutical drugs. Some species (such as Chlorella sp., Haematococcus pluvialis or Dunaliella salina) are already cultivated on large scales, mainly in the photoautotrophic mode. Due to the production cost of microalgal biomass, relatively high-value compounds are targeted for economically viable processes. Among them, fatty acids and pigments are common products for which relevant species and cultivation processes are studied. Antibacterial substances appear as promising but less studied targets. This chapter makes a review about the production of fatty acids, pigments and antibacterial/antifungal compounds, with a special emphasis on their production in the heterotrophic mode. Compared to phototrophic growth, heterotrophic growth may result in higher biomass productivity by avoiding light limitations. In addition, it decreases area footprint and installation costs. The heterotrophic metabolism of some model microalgae is also described as well as the productivities that could be encountered in heterotrophy.
... In fermentation media, the carbon/nitrogen (C/N) ratio controls the switch between protein and lipid synthesis; therefore, the C/N ratio of culture media is one of the most critical nutritional factors affecting the total amount of produced lipids. Moreover, it has been observed that the initial C/N ratio should be higher than 20 for maximum lipid accumulation by oleaginous organisms (23,24). ...
In this study, 18 standard amino acids were tested as a single nitrogen source on biomass, total lipid, total fatty acid (TFA) production, and yield of γ-linolenic acid (GLA) in Rhizomucor pusillus AUMC 11616.A and Mucor circinelloides AUMC 6696.A isolated from unusual habitats. Grown for 4 days at 28 • C, shaking at 150 rpm, the maximum fungal biomass for AUMC 6696.A was 14.6 ± 0.2 g/L with arginine and 13.68 ± 0.1 g/L with asparagine, when these amino acids were used as single nitrogen sources, while AUMC 11616.A maximum biomass was 10.73 ± 0.8 g/L with glycine and 9.44 ± 0.6 g/L with valine. These were significantly higher than the ammonium nitrate control (p < 0.05). The highest levels of TFA were achieved with glycine for AUMC 11616.A, 26.2 ± 0.8% w/w of cell dry weight, and glutamic acid for AUMC 6696.A, 23.1 ± 1.3%. The highest GLA yield was seen with proline for AUMC 11616.A, 13.4 ± 0.6% w/w of TFA, and tryptophan for AUMC 6696.A, 12.8 ± 0.3%, which were 38% and 25% higher than the ammonium tartrate control. The effects of environmental factors such as temperature, pH, fermentation time, and agitation speed on biomass, total lipids, TFA, and GLA concentration of the target strains have also been investigated. Our results demonstrated that nitrogen assimilation through amino acid metabolism, as well as the use of glucose as a carbon source and abiotic factors, are integral to increasing the oleaginicity of tested strains. Few studies have addressed the role of amino acids in fermentation media, and this study sheds light on R. pusillus and M. circinelloides as promising candidates for the potential applications of amino acids as nitrogen sources in the production of lipids.
... p = 0.12), which again showed more variation in non-sterile RAS water. The reverse results obtained with cell density (cells ml −1 ), where values for C. vulgaris were higher (Fig. 3), can be explained with the cell size of these species [33,34], where the larger species, T. obliquus, achieves a higher dry weight despite a lower cell count. These values are comparatively high [10,29,35]. ...
Microalgae biotechnology is of increasing importance and a central application concerns the treatment of wastewater. Here, its implementation in a recirculating aquaculture system (RAS) to lower the discharge of wastewater is studied. To better cope with external variations in culture conditions, a co-cultivation of two species of microalgae, Chlorella vulgaris and Tetradesmus obliquus, was used to obtain a more reliable and robust culture and was compared to monocultures. This approach was tested using RAS water both under sterile and non-sterile conditions at laboratory scale and then compared to a co-culture at pilot-scale in an open thin-layer photobioreactor. Performance of cultures was tested in terms of microalgae growth and nutrient removal efficiency. Furthermore, to better understand the interaction between environmental variables and each microalgae species, their relative frequencies in co-cultures as well as the presence of protozoa and bacteria were monitored. All growth experiments were carried out successfully and, unlike in a previous study, no crashes were observed. However, shifts in species frequency in co-cultures indicated that the two species were differentially affected by cultivation conditions. Despite nutrient limitation, the pilot-scale cultivation had a high productivity (13.3 g m⁻² d⁻¹) and final dry weight (11.1 g l⁻¹) after 29 days and demonstrated its suitability for RAS water treatment.
... This is in accordance with the general apprehension that polyunsaturated fatty acids (PUFA) synthesis is favored at low temperatures [20]. As reported previously, it is possible that the desaturases produced by the microalga cells exhibit a higher activity at low temperatures due to an increased availability of oxygen [21]. This is also apparent in the present results, with an exception at the temperature of 15 C. By carrying out an ANOVA analysis coupled with a Tukey test, it is clear that between the temperature range of 17e23 C, the difference of the DHA content (%TFA) of the cells is not significant (p ¼ 0.32), securing the conclusion that the DHA is indeed favored at lower temperatures. ...
Dark fermentation is an anaerobic digestion process of biowaste, used to produce hydrogen as a fuel, which however releases high amounts of polluting volatile fatty acids in the environment. In order for the process to become more competitive, the acids stream can be utilized through conversion to high added-value docosahexaenoic acid by the microalga Crypthecodinium cohnii. Docosahexaenoic acid is one of the two main omega-3 fatty acids, necessary for human nutrition. The purpose of this work was to optimize the production of omega-3 fatty acids by the cells, utilizing the organic content of a dark fermentation effluent. For that purpose, the effect of different fermentation conditions was examined, such as incubation temperature, nitrogen source and concentration, the addition of chemical modulators, as well as the feeding composition. The volatile fatty acid content of the effluent was totally depleted in a fed-batch culture of the microalga, while the cells accumulated DHA in a percentage of 35.6% of total lipids, when fed with yeast extract or 34.2% when fed with ammonium sulfate. Taking into consideration the economic feasibility of the culture conditions proposed it was concluded that the use of yeast extract could be substituted by the much economic ammonium sulfate.
... microscope observations were carried out. C. vulgaris size was around 5 µm diameter (in accordance with Liu et al., 2014) and T. obliquus between 5 and 12 µm length (in accordance with Belcher and Swale, 1977). Protozoa size ranged from 20 to 300 µm length (from ciliates and flagellates around 20 µm to 300 µm of rotifers). ...
... microscope observations were carried out. C. vulgaris size was around 5 µm diameter (in accordance with Liu et al., 2014) and T. obliquus between 5 and 12 µm length (in accordance with Belcher and Swale, 1977). Protozoa size ranged from 20 to 300 µm length (from ciliates and flagellates around 20 µm to 300 µm of rotifers). ...
The ongoing and increasing worldwide demand for fish has caused a steady increase in aquaculture production during the last decades. This emphasizes the importance of farming systems with a low ecological footprint, like recirculating aquaculture systems (RAS), which are an alternative to traditional open systems. Furthermore, implementing microalgae treatments in RAS, sustainable water management and low discharge of concentrated wastewater could be achieved, allowing its reuse in the system. The influence of three factors on microalgae treatment efficiency in RAS water were studied: i) microalgae species (Chlorella vulgaris, Tetradesmus obliquus), ii) water pre-treatment (sterile filtration), and iii) sampling location within the RAS (e.g. from fish tank, after UV-disinfection, etc.). To this end, fully factorial, replicated cultivations were carried out in 100-ml flasks, and nutrient removal, microalgae growth, and density of bacteria and protozoa were measured for up to 18 days. Results show that both species are able to grow in RAS water and effectively remove nutrients in it, yet their performance depended greatly on water quality. In sterile RAS water, growth and nutrient removal efficiency of C. vulgaris surpassed that of T. obliquus. In non-sterile RAS water, the pattern reversed because of grazing protozoa. The location of sampling within the RAS had no discernible effect on microalgae growth or nutrient removal efficiency. The results confirm that a microalgae-based technology to treat and valorise RAS water is technically feasible, yet caution that inferences made can be reversed depending on the choice of the species and the pretreatment of the RAS water prior to cultivation.
... The main advantages of SCO production are high diversity, capability of accumulating large amounts of oil within the cells of cultivated microorganisms, fast growth of biomass in comparison with plants and animals and reduced costs of production [50]- [52]. In 2002 the first industrially produced SCOs were recognized as being safe by the US Food and Drug Administration and since then they are included in baby formulas for newborns, which in general has promoted increase in production volumes of these oils all over the world [9], [33], [45], [50]- [54]. ...
Aquaculture is currently the fastest growing food-producing sector in the world. The growth of this industry has been rapid for the last 25 years, however, aquaculture still relies heavily on feed input from wild capture fisheries. Landings in wild capture fisheries have been stagnant for the last two decades; therefore, new alternatives for conventional fish meal and fish oil need to be found. In this review, various alternatives are described and their advantages and disadvantages are evaluated. Single cell oils (SCO) and single cell proteins (SCP) produced by microorganisms are recognized as the alternative with the most potential for replacing fish meal and fish oil in aquacultures. However, production costs of SCOs and SCPs are still higher than production costs of Omega-3 rich oils from other sources (wild capture, plant derived oils and genetically modified plants); therefore, currently used substrates need to be replaced with cheaper agriculture and industrial biomass residues applicable for microbial fermentation. In order to evaluate various biodegradable residues and find the most suitable ones for SCO and SCP production, methods analysing resource availability are reviewed.
... microscope observations were carried out. C. vulgaris size was around 5 µm diameter (in accordance with Liu et al., 2014) and T. obliquus between 5 and 12 µm length (in accordance with Belcher and Swale, 1977). Protozoa size ranged from 20 to 300 µm length (from ciliates and flagellates around 20 µm to 300 µm of rotifers). ...
The ongoing and increasing worldwide demand for fish has caused a steady increase in aquaculture production during the last decades. This development is expected to continue and therefore emphasizes the importance of farming systems with a low ecological footprint. Recirculating aquaculture systems (RAS) feature a sustainable water management and produce a low discharge of concentrated wastewater. Cultivation of microalgae has been suggested as a means to treat this wastewater.
The influence of three factors on treatment efficiency was studied: i) microalgae species (Chlorella vulgaris, Acutodesmus obliquus), ii) wastewater pre-treatment (sterile filtration), and iii) sampling location within the RAS (e.g. from fish tank, after UV-disinfection, etc.). To this end, fully factorial, replicated cultivations were carried out on a laboratory scale and nutrient removal, microalgae growth, and density of bacteria and protozoa were measured for up to 18 days.
When cultivated in sterile wastewater, growth and nutrient removal efficiency of C. vulgaris surpassed that of A. obliquus. In non-sterile wastewater, however, the pattern reversed because increasing numbers of grazing protozoa caused cultures of C. vulgaris to crash within ten days, while A. obliquus was less affected and maintained a high nutrient removal efficiency, which even tended to increase. The susceptibility of C. vulgaris towards protozoa was further supported by a negative correlation between cell density and numbers of protozoa, a relationship that was not present in A. obliquus. The location of sampling within the RAS had not discernible effect on microalgae growth or nutrient removal efficiency.
The results indicate that a microalgae-based technology to treat aquaculture wastewater could be successfully implemented. However, because an effective wastewater sterilization is not feasible on an industrial scale, our results also point to the importance of choosing the right microalgae species and caution that its neglection can cause results to reverse with corresponding consequences.
... This poor productivity is not a reflection of any inherent limitation in algal biology, but of inability to provide sufficient light to self-shaded algal cultures. Microalgae grown heterotrophically on organic carbon sources obtained from higher plants have a much greater (e.g., 10-fold greater) biomass productivity relative to photoautotrophically grown cultures (Liu et al., 2014). Therefore, heterotrophic production of algae may have a greater potential to provide PUFA-rich biomass for use in aquaculture to replace fish oils. ...
... The lipid content of Isochrysis strains showed a slight difference in response to the initial N concentration and was promoted slightly by lower N concentrations (Fig. 3c), differing a lot from other algae such as Chlorella and Nannochloropsis strains in which the lipid content was influenced considerably by N limitation/starvation [18,19]. This may be explained by the fact that Isochrysis strains utilize the recycled membrane lipids rather than the de novo synthesized fatty acyls for neutral lipid synthesis upon N stress [20]. Similarly, the lipid productivity of Isochrysis was in a N concentration-dependent manner, which reached the maximum at 100-200 mg L −1 N (Fig. 3d). ...
Isochrysis is a genus of marine algae without cell wall and capable of accumulating lipids. In this study, the lipid production potential of Isochrysis was assessed by comparing 15 Isochrysis strains with respect to their growth rate, lipid production, and fatty acid profiles. Three best strains were selected (lipid productivity, 103.0~121.7 mg L−1 day−1) and their lipid-producing capacities were further examined under different controlled parameters, e.g., growth phase, medium nutrient, and light intensity in laboratory cultures. Furthermore, the three Isochrysis strains were monitored in outdoor panel photobioreactors with various initial cell densities and optical paths, and the strain CS177 demonstrated the superior potential for outdoor cultivation. A two-stage semi-continuous strategy for CS177 was subsequently developed, where high productivities of biomass (1.1 g L−1 day−1) and lipid (0.35 g L−1 day−1) were achieved. This is a comprehensive study to evaluate the lipid-producing capability of Isochrysis strains under both indoor and outdoor conditions. Results of the present work lay a solid foundation for the physiological and biochemical responses of Isochrysis to various conditions, shedding light on the future utilization of this cell wall-lacking marine alga for biofuel production.
... [6] As a consequence, the production of omega-3 by microorganisms has increased worldwide. [76,78,[80][81][82] The dimension and potential of the DHA food supplement business may be illustrated by the sale of Martek Biosciences in 2011 to DSM, for USD 1.1 billion, reflecting the oil price sales -DHA-rich oil being considered the "cornerstone" of the company, which produced around 2000 t/year, and a revenue of USD 317 million in 2010. [6] Alongside David Kyle, Bill Barclay, head of Omega-Tech Inc., found that there was a large group of marine eukaryotic organisms, known as Thraustochytrids, capable of producing high concentrations of DHA. ...
In recent years, foods that contain omega-3 lipids have emerged as important promoters of human health. These lipids are essential for the functional development of the brain and retina, and reduction of the risk of cardiovascular and Alzheimer's diseases. The global market for omega-3 production, particularly docosahexaenoic acid (DHA), saw a large expansion in the last decade due to the increasing use of this lipid as an important component of infant food formulae and supplements. The production of omega-3 lipids from fish and vegetable oil sources has some drawbacks, such as complex purification procedures, unwanted contamination by marine pollutants, reduction or even extinction of several species of fish, and aspects related to sustainability. A promising alternative system for the production of omega-3 lipids is from microbial metabolism of yeast, fungi, or microalgae. The aim of this review is to discuss the various omega-3 sources in the context of the global demand and market potential for these bioactive compounds. To summarize, it is clear that fish and vegetable oil sources will not be sufficient to meet the future needs of the world population. The biotechnological production of single-cell oil comes as a sustainable alternative capable of supplementing the global demand for omega-3, causing less environmental impact.
... C. protothecoides also demonstrated a good potential for removing nutrient of N and P (Fig. 2E and 4B) and for wastewater treatment (Sforza et al., 2014). C. protothecoides has been explored for lipid production, to our best knowledge, under heterotrophic growth conditions using sugars , glucose in particular, as the solo carbon and energy source (Liu et al., 2014). Although heterotrophic C. protothecoides is able to achieve high cell density and thus high biomass and lipid productivities , it has intrinsic drawbacks such as low sugar-to-biomass conversion (commonly below 0.5) and relatively high production cost, and thus is less favorable for the production of low-cost commodity oils. ...
Biorefinery approaches offer the potential to improve the economics of the microalgae industry by producing multiple products from a single source of biomass. Chromochloris zofingiensis shows great promise for biorefinery due to high biomass productivity and a diverse range of products including secondary carotenoids, predominantly astaxanthin; lipids such as TAGs; carbohydrates including starch; and proteins and essential amino acids. Whilst this species has been demonstrated to accumulate multiple products, the development of an integrated downstream process to obtain these is lacking. The objective of this review paper is to assess the research that has taken place and to identify the steps that must be taken to establish a biorefinery approach for C. zofingiensis. In particular, the reasons why C. zofingiensis is a promising species to target for biorefinery are discussed in terms of cellular structure, potential products, and means to accumulate desirable components via the alteration of culture conditions. Future advances and the challenges that lie ahead for successful biorefinery of this species are also reviewed along with potential solutions to address them.
The discharge of palm oil mill effluent (POME) without effective treatment is highly toxic to the environment and receiving waters due to its high chemical oxygen demand , biochemical oxygen demand and increased levels of suspended solids. The ponding system is regarded as a conventional method for the treatment of POME but requires long retention time and extensive treatment areas. Other processes, such as physicochemical treatment, have also been applied as tertiary steps to ensure lower toxicity of the POME discharge. More recently, phycoremediation, a new biological treatment method using microalgae, has been reported as having great potential in being a highly efficient method of reducing the environmental impact of POME. This paper discusses the feasibility of the above treatment methods of POME, including the potential of phycoremediation, with a focus on their application in South East Asia.
Biofuels can be derived from waste biomass feedstocks, such as municipal, agricultural, forestry and industrial waste. There are several advantages in switching to microalgae for biofuel production. Microalgae has a rapid growth rate, so is more productive, so requires smaller areas for cultivation per unit of biomass produced. Microalgae can absorb “waste” CO2, does not compete with food crops (for land and freshwater), and can be cultivated in wastewater, doubling as a wastewater treatment. This paper gives an overview of microalgae cultivation, focusing on the early energy-intensive stages: growth, harvesting and drying. The harvesting and drying steps constitute a significant economic bottleneck, due to their high energy costs. This review also covers microalgal cultivation and its integration with wastewater treatment, carbon and energy sources, and the utilization of microalgal biofuel co-products from thermochemical conversion, as this route is the most likely to mitigate the techno-economic downsides of microalgal biofuel production.
No matter the scale, stirred tank bioreactors are the most commonly used systems in biotechnological production processes. Single-use and reusable systems are supplied by several manufacturers. The type, size, and number of impellers used in these systems have a significant influence on the characteristics and designs of bioreactors. Depending on the desired application, classic shaft-driven systems, bearing-mounted drives, or stirring elements that levitate freely in the vessel may be employed. In systems with drive shafts, process hygiene requirements also affect the type of seal used. For sensitive processes with high hygienic requirements, magnetic-driven stirring systems, which have been the focus of much research in recent years, are recommended. This review provides the reader with an overview of the most common agitation and seal types implemented in stirred bioreactor systems, highlights their advantages and disadvantages, and explains their possible fields of application. Special attention is paid to the development of magnetically driven agitators, which are widely used in reusable systems and are also becoming more and more important in their single-use counterparts.
Key Points
• Basic design of the most frequently used bioreactor type: the stirred tank bioreactor
• Differences in most common seal types in stirred systems and fields of application
• Comprehensive overview of commercially available bioreactor seal types
• Increased use of magnetically driven agitation systems in single-use bioreactors
This thesis summarizes the results of a doctoral research executed in the State
University of Campinas and in the Technical University of Delft as part of the PhD Dual Degree Program between the two universities. The research project was designed in partnership with Petrobras S. A. (Brazilian Petroleum Corporation), which provided most of the financial support as well as technical cooperation, with the goal of evaluating the potential of heterotrophic microalgae for biofuels production.
In order to shift the world to a more sustainable future, it is necessary to phase out the use of fossil fuels and focus on the development of low-carbon alternatives. However, this transition is slow so there still exists a large dependence on fossil-derived power and therefore CO2 release. Owing to the potential for assimilating and utilizing CO2 to generate carbon neutral products such as biodiesel, the application of microalgae technology to capture CO2 from flue gases has gained significant attention over the past decade. Microalgae offer a more sustainable source of biomass energy over conventional fuel crops since they grow more quickly and do not adversely affect food supply. This review focuses on the technical feasibility of combined carbon fixation and microalgae cultivation for carbon reuse. We appraise a range of different C metabolisms and the impact of flue gas compounds on microalgae. Fixation of flue gas CO2 is found to be dependent on the selected microalgae strain and on flue gas compounds/concentrations. Additionally, we assess current pilot-scale demonstrations of microalgae technology for CO2 capture as well as discuss its future prospects. Practical implementation of this technology at an industrial-scale still requires significant research, necessitating multidiscipline R&D to demonstrate its viability for CO2 capture from flue gases at the commercial level.
Chlorella is one of the best-studied green microalgal genera because of its wide use as a model system and its utilization in biotechnology. Since the description of the type species Chlorella vulgaris Beij., more than a hundred species have been established in the literature. However, the taxonomic description and identification of these small (<15 μm) spherical or elliptical coccoid cells is difficult due to the lack of characteristic morphologic features. In addition to molecular investigations, biochemical criteria are employed to distinguish between the numerous “Chlorella” species, of which the sterol composition seems to be a reliable chemotaxonomic marker within several groups of these morphologically similar algae. In this study, the distribution of ergosterol was examined in 20 species of the “true” genus Chlorella and more distant “Chlorella” species using HPLC. Ergosterol in concentrations up to 4.5 μg · mg−1 dry weight (dwt) was detected in nine species, which are all related representatives of the Chlorellaceae. More distant relatives within the Trebouxiophyceae or representatives of the Chlorophyceae did not contain ergosterol. The results coincide with the latest molecular investigations of the genus Chlorella and further promote the potential of ergosterol as chemical marker to differentiate between members of the Chlorellaceae and other “Chlorella-like” species.
Algae have recently gained attention as a potential source for biodiesel; however, much is still unknown about the biological
triggers that cause the production of triacylglycerols. We used RNA-Seq as a tool for discovering genes responsible for triacylglycerol
(TAG) production in Chlamydomonas and for the regulatory components that activate the pathway. Three genes encoding acyltransferases, DGAT1, DGTT1, and PDAT1, are induced by nitrogen starvation and are likely to have a role in TAG accumulation based on their patterns of expression.
DGAT1 and DGTT1 also show increased mRNA abundance in other TAG-accumulating conditions (minus sulfur, minus phosphorus, minus zinc, and
minus iron). Insertional mutants, pdat1-1 and pdat1-2, accumulate 25% less TAG compared with the parent strain, CC-4425, which demonstrates the relevance of the trans-acylation
pathway in Chlamydomonas. The biochemical functions of DGTT1 and PDAT1 were validated by rescue of oleic acid sensitivity and restoration of TAG accumulation in a yeast strain lacking all acyltransferase
activity. Time course analyses suggest than a SQUAMOSA promoter-binding protein domain transcription factor, whose mRNA increases precede that of lipid biosynthesis genes like
DGAT1, is a candidate regulator of the nitrogen deficiency responses. An insertional mutant, nrr1-1, accumulates only 50% of the TAG compared with the parental strain in nitrogen-starvation conditions and is unaffected by
other nutrient stresses, suggesting the specificity of this regulator for nitrogen-deprivation conditions.
The green alga Chlorella zofingiensis can accumulate high level of oleic acid (OA, C18:1△(9)) rich oils in response to stress conditions. To understand the regulation of biosynthesis of fatty acid in particular OA at the molecular level, we cloned and characterized the stearoyl acyl carrier protein (ACP) desaturase (SAD) responsible for OA formation through desaturation of stearic acid (C18:0) from C. zofingiensis. Southern blot indicated that the C. zofingiensis genome contained a single copy of SAD, from which the deduced amino acid sequence shared high identity to the corresponding homologs from other microalgae and higher plants. The desaturation activity of SAD was demonstrated in vitro using C18:0-ACP as a substrate. Stress conditions such as high light (HL), nitrogen deficiency (N(-)), or combination of HL and N(-) (HL + N(-)) drastically up-regulated the transcripts of biotin carboxylase (BC, a subunit of ACCase) and SAD, and therefore induced considerably the cellular accumulation of total fatty acids including OA. Glucose (50 mM) gave rise to the similar up-regulation of the two genes and induction of fatty acid accumulation. The accumulation of intracellular reactive oxygen species was found to be associated with the up-regulation of genes. This is the first report of characterization of Chlorella-derived SAD and the results may contribute to understanding of the mechanisms involved in fatty acid/lipid biosynthesis in microalgae.
A multimethod approach was used to characterize unicellular green algae that were traditionally assigned to the genus Chlorella Beijerinck and to resolve their phylogenetic relationships within the Chlorophyta. Biochemical, physiological, and ultrastructural characters, together with molecular data such as DNA base composition and DNA hybridization values, were compared with a molecular phylogeny based on complete 18S rRNA sequences. Our results show that Chlorella taxa are dispersed over two classes of chlorophytes, the Trebouxiophyceae and the Chlorophyceae. We propose that only four species should be kept in the genus Chlorella (Chlorophyta, Trebouxiophyceae): C. vulgaris Beijerinck, C. lobophora Andreyeva, C. sorokiniana Shih. et Krauss, and C. kessleri Fott et Nováková. Common characteristics of these taxa are glucosamine as a dominant cell wall component and the presence of a double thylakoid bisecting the pyrenoid matrix. Norspermine, norspermidine, and secondary carotenoids are never produced. Other “Chlorella” species belong to different taxa within the Trebouxiophyceae (“C.” protothecoides=Auxenochlorella protothecoides[Krüger] Kalina et Punčochářová, “C.” ellipsoidea, “C.” mirabilis, “C.” saccharophila, and “C.” luteoviridis) and Chlorophyceae (“C.” zofingiensis and “C.” homosphaera=Mychonastes homosphaera Kalina et Punčochářová). The latter taxa can easily be recognized by the production of secondary carotenoids under nitrogen-deficient conditions.
The species of the genus Chlorella exhibit considerable biochemical and physiological differences. Therefore, it is important to select for and utilize in research or biotechnology correctly identified strains of the species having the most favorable properties for the respective project. We examined the Chlorella strains of the University of Texas collection at Austin, Texas, according to species-specific chemotaxonomic characters and assigned 58 strains to 10 well-established species (only 17 of these strains were correctly named before).
Twelve salts were tested for their ability to coagulate microalgae cells in cultures of Chlorella minutissima. The final aim was to develop an easy and efficient approach for harvesting microalgae biomass in dense cultures. Aluminum,
ferric, and zinc salts coagulated C. minutissima cultures, while optimum concentration was 0.75 and 0.5gL−1 for sulfate and chloride salts, respectively. Aluminum salts were most efficient, but caused some cell lysis, which may render
this approach inappropriate in some cases. Ferric and zinc salts were ranked second and third, respectively, according to
their culture cell-coagulation efficiency. Ferric salts caused a change in the color of the cells, mainly at concentrations
higher than 1gL−1. Zinc salts were less harmful for the microalgal cells, but an additional problem was observed with cell aggregates adhering
to the walls of the glass test tubes. Selection of the appropriate coagulant is related to the purpose of the coagulation
process.
KeywordsAluminum salts-Ferric salts-Flocculation-Microalgae-Zinc salts
An isolate of Isochrysis galbana rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) has been grown as a chemostat culture at 20° C and pH 8.00 controlled by CO2 injection. From a low dilution rate (D) of 0.0024 h−1 to 0.0377 h−1, close to maximum growth, a decrease in EPA content from 5.21% dry weight (d.w.) to 2.80% d.w. was observed, although the percentage of EPA in the total fatty acids increased. Lipids were fractionated, EPA being the major fatty acid found in the glycolipid fraction, whereas in the neutral lipid fraction were mainly palmitic and palmitoleic acids. At the same time, the biomass concentration also decreased from 1015 mg·l−1 to 202 mg·l−1 over the range of Ds mentioned. Nonetheless, EPA productivity had a maximum value of 15.26 mg·l−1·day−1 at D=0.0208 h−1.
This is a well-rounded handbook of fermentation and biochemical engineering presenting techniques for the commercial production of chemicals and pharmaceuticals via fermentation. Emphasis is given to unit operations fermentation, separation, purification, and recovery. Principles, process design, and equipment are detailed. Environment aspects are covered. The practical aspects of development, design, and operation are stressed. Theory is included to provide the necessary insight for a particular operation. Problems addressed are the collection of pilot data, choice of scale-up parameters, selection of the right piece of equipment, pinpointing of likely trouble spots, and methods of troubleshooting. The text, written from a practical and operating viewpoint, will assist development, design, engineering and production personnel in the fermentation industry. Contributors were selected based on their industrial background and orientation. The book is illustrated with numerous figures, photographs and schematic diagrams. © 1996 Noyes Publications Published by Elsevier Inc. All rights reserved.
This work presents the experimental results obtained in the hetero- and mixotrophic culture of Chlorella vulgaris, a microalga belonging to the Chlorophyceae class. Objective of the study was evaluating the capability of the microalga to accumulate lipids, prospectively usable either in biodiesel production or in the extraction of nutraceuticals. Our study shows that C. vulgaris can increase its lipid content from similar to 15% (balanced-, autotrophically-grown biomass) to more than 50%, and highlighted the difference between the type (i.e., polar or nonpolar) of the lipids accumulated upon nitrogen and phosphorus starvation. An apparent interaction between diel rythms and glucose uptake was also suggested, together with its implications on productivity.
Among many challenges faced in the commercial cultivation of microalgae, low-cost water and nutrients availability is crucial. Our study aimed at testing and optimizing two agro-industrial co-products, dry-grind ethanol thin stillage (TS) and soy whey (SW), as nutrient feedstock for mixotrophic/heterotrophic microalgal cultivation. Heterotrophic growth of Chlorella vulgaris was first optimized in a Bioscreen turbidimeter and 250 mL Erlenmeyer flasks, then scaled up to a 6-L stirred bioreactor. Intracellular oil was extracted from dried microalgal biomass by ultrasonication and solvent extraction treatments for yield comparison, and fatty acid (FA) profile. Biomass yields (dry basis) from TS, SW and modified basal medium (MBM) after 4 days of incubation at mixotrophic conditions in the bioreactor were 9.8, 6.3 and 8.0 g.L− 1 with oil content at 43, 11, and 27% (w/w) respectively. FA profile of oil samples was found to vary and depend on growth media characteristics. C. vulgaris when grown on TS and MBM produced oil richer in linoleic and linolenic acids, respectively. This research highlights the potential of two agro-industrial co-products as microalgal growth media with consequent production of high-value microalgal oil and biomass.
In this study, initially, the carbon and nitrogen source preferences of Chlorella saccharophila were examined in terms of biomass productivities under heterotrophic growth conditions. It was shown that C. saccharophila could actively utilize glucose and glycerol as carbon sources and bacteriological peptone as a nitrogen source. Secondly, the concentrations of nitrogen and carbon sources that were found to significantly influence the biomass productivity of heterotrophic C. saccharophila were further optimized by using Box–Behnken experimental design. Lastly, in a scale-up attempt, the medium consisting of 20 g/L glucose and 1 g/L bacteriological peptone was used in a 3 L stirred tank bioreactor in which the final biomass concentration obtained was 7.7 fold higher than that of obtained under autotrophic conditions. Also, lipid content in heterotrophic cells of C. saccharophila was increased about 3 times compared to that of autotrophic cells. The principal fatty acids in heterotrophic C. saccharophila were oleic acid (C18:1) and linoleic acid (C18:2) constituting 34.4% and 30.1% of the total fatty acid contents, respectively. Moreover, a simple and rapid method determining the neutral lipid accumulation in C. saccharophila with spectrofluorimetry was developed and used easily for monitoring lipid accumulation in a stirred tank bioreactor.
Biodiesel is an alternative diesel fuel that is produced from vegetable oils and animal fats. It consists of the monoalkyl esters formed by a catalyzed reaction of the triglycerides in the oil or fat with a simple monohydric alcohol. The reaction conditions generally involve a trade-off between reaction time and temperature as reaction completeness is the most critical fuel quality parameter. Much of the process complexity originates from contaminants in the feedstock, such as water and free fatty acids, or impurities in the final product, such as methanol, free glycerol, and soap. Processes have been developed to produce biodiesel from high free fatty acid feedstocks, such as recycled restaurant grease, animal fats, and soapstock.
The heterotrophic growth of microalgae has been proposed as a potential mode of culture to achieve high cell densities on a large scale. In this paper, the heterotrophic growth of the green microalga, Chlamydomonas reinhardtii,on acetate as sole energy and carbon source in batch and single-stage continuous cultures is reported. No inhibition by acetate was observed at low concentrations (≤ 0·4 g/litre), in which case, cell growth was well described by the Monod model. However, above this acetate concentration, growth was progressively inhibited. Inhibition was observed as a decrease in both the specific growth rate and the observed growth yield. The former was well modelled using the Haldane Model. A model based on constant qm was proposed to describe the latter phenomenon. Whereas it predicted the trend of falling observed cell growth yield with initial acetate concentration in batch cultures, there was significant lack of fit between simulated and experimental data. In continuous cultures, a steady state cell concentration of 0·4 g/litre and maximum cell productivity of 0·02 g/litre/h was obtained for an acetate feed concentration of 0·85 g/litre. This compares well with values achieved by photosynthetic mass culture systems.
In this study different methods were applied for lipids extraction from the dry biomass of Chlorella pyrenoidosa. The survey was carried under different conditions seeking comparative assessment of extraction methods. The method using chloroform:methanol (2:1 v/v) showed the highest lipid extraction followed by methanol, chloroform, ethanol, and hexane. Afterward, we also assessed the relative influence of the solvent extractor selectivity on the overall FAMEs (Fatty Acids Methyl Esters) yield. The application of the transesterification process on the several lipidic extracts was compared with direct transesterification process from dry biomass. In the extraction using chloroform:methanol system a larger amount of lipids was obtained but the conversion to FAMEs using transesterification process was the lowest from lipids. However, despite the amount of extracted lipids with methanol being smaller, its conversion to FAMEs was higher from lipids. In addition, the extraction with methanol followed by transesterification process also resulted in a higher FAMEs yield from biomass than direct transesterification process using methanol.
Microbial lipid accumulation to provide alternative oil resources is an exciting research area, obtaining increasing attentions recently for the biodiesel production due to its high production efficiency and less demand of agricultural land. The aim of this study is to optimize the lipid accumulation of Chlorella vulgaris by using various carbon sources in heterotrophic and mixotrophic cultures. Different cultivation factors were studied on their influences to the cell growth and oil accumulation.Our results revealed that C. vulgaris could grow on autotrophic, mixotrophic and heterotrophic modes; and the mixotrophic cultivation especially could produce more cell biomass than the autotrophic or heterotrophic cultures individually or combined. The substrate concentration significantly influenced the final cell yield of the mixotrophic cultivations while the cell lipid content remained relatively constant. Glycerol was inhibitive to the cell growth while the microalgae strain could actively utilize acetate as the carbon source. This provides a promising niche in reducing the overall cost of biofuel production since this substrate can be obtained from some waste processes such as anaerobic digestion.
In this paper we present and analyse single-cell oil (SCO) production through the cultivation of the cyanobacterium Aphanothece microscopicaNägeli in fish processing wastewater at different temperature levels. The growth data were fitted into four mathematical models (Logistic, Gompertz, modified Gompertz and Baranyi), to define a mathematical expression for describing cell growth. Using statistical criteria, the Logistic model was considered the most adequate one to describe biomass growth. A negative relationship between the percentage of lipid and high growth temperature was evidenced. Mathematical modelling indicates the possibility of obtaining 0.26, 0.36 and 0.44gbiomass/Lreactorday at temperatures of 10, 20 and 30°C, respectively. This results in lipid productivity of 0.04, 0.05 and 0.03glipid/Lreactorday at 10, 20 and 30°C, respectively.
Biodiesel is an alternative to petroleum-derived diesel fuel composed of alkyl esters of vegetable oils, animal fats or other feedstocks such as used cooking oils. The fatty acid profile of biodiesel corresponds to that of its feedstock. Most common feedstocks possess fatty acid profiles consisting mainly of five C16 and C18 fatty acids, namely, palmitic (hexadecanoic), stearic (octadecanoic), oleic (9(Z)-octadecenoic), linoleic (9(Z),12(Z)-octadecadienoic) and linolenic (9(Z),12(Z),15(Z)-octadecatrienoic) acids, with the exception of a few oils such as coconut oil, which contains high amounts of saturated acids in the C12–C16 range or others. While in many respects biodiesel possesses advantages or is competitive with petroleum-derived diesel fuel, virtually all biodiesel fuels, typically the methyl esters, produced from these oils have performance problems such as poor low-temperature properties or insufficient oxidative stability. Considerable research has focused on solving or alleviating these problems and five approaches have been developed. Besides the approach of using additives, changing the fatty ester composition by either varying the alcohol or the fatty acid profile of the oil have been studied. Changing the fatty acid profile can be achieved by physical means, genetic modification of the feedstock or use of alternative feedstocks with different fatty acid profiles. In some cases approaches may overlap. This article briefly summarizes the five approaches currently used with an emphasis on those dealing with changing the fatty ester composition of a biodiesel fuel.
The marine PrasinophyteTetraselmis may be cultured under both mixotrophic (photoheterotrophic) and heterotrophic conditions. The growth rate was slightly lower, and pigment levels and lipid composition were radically affected on heterotrophic culture in 1 L fermenters. Total chlorophyll levels of dark grown cultures were less than 1% of those observed in mixotrophically grown cells, the chlorophylla : b ratio also decreased as did the carotenoid content. In addition, the total amounts of lipids including polyunsaturated fatty-acids were also lower in heterotrophically cultured cells: 6.4 mg g−1 (dried alga) and 0.35 mg g−1 (dried alga); as compared to 37.1 mg g−1 (dried alga) and 18.5 mg g−1 (dried alga), for cells grown in the light. However, gross morphology and final yield (>16 g l−1) were relatively unaffected. The algae produced were spray-dried and tested for their suitability as an aquaculture feed.
Spirulina sp. was able to grow photoautotrophically (in the light), heterotrophically (on glucose) and mixotrophically (simultaneously in the light and on glucose). The highest specific growth rate was reached in mixotrophic culture (0.055h−1, above I=30Wm−2, CGl=0.5gl−1). The specific growth rate of the alga on 2.5gl−1 glucose was markedly increased with increasing light intensity up to 30Wm−2. In photoautotrophic culture at a light intensity above 50Wm−2 photoinhibition occurred.Unstructured kinetic models to describe microalgal culture system including photoautotrophic, heterotrophic and mixotrophic specific growth rate as a function of light intensity and glucose concentration were proposed. The models demonstrated that the light intensity for autotrophic culture was 30–50Wm−2, glucose concentration for heterotrophic growth was >0.5gl−1 and for mixotrophic growth of Spirulina sp. in batch culture using 1l photobioreactor was >30Wm−2 and glucose concentration >0.5gl−1.
Micro-algae are widely used as a traditional feed for the early developmental stages of molluscs and crustaceans. A large spectrum of algae are cultivated; these have been selected because they have a suitable chemical and nutritional profile, are of the correct size and grow well photoautotrophically. A large number of potential feed organisms have been screened for their nutritional suitability and growth characteristics, with the above criteria expanded to include the capacity for heterotrophic growth. A process for the heterotrophic cultivation of Tetraselmis suecica CSL161, has been developed and scaled up to an industrial-level process. The biomass produced, after concentration, is stabilized by drying to give a free-flowing powder. Growth trials with the bivalves Crassostrea gigas, Crassostrea virginica, Tapes philippinarum, Mercenaria mercenaria and Ostrea edulis have demonstrated that dried Tetraselmis CSL161, sold as Celsys Algal 161, can directly replace live Tetraselmis in bivalve feed and may provide up to 70% replacement for hatchery-grown micro-algae.
Statistically based experimental designs were applied to the optimization of medium components and environmental factors for eicosapentaenoic acid (EPA) production by the diatom Nitzschia laevis in heterotrophic conditions. First, the Plackett-Burman design was used to evaluate the effects of variables including medium components and environmental factors on cell growth and EPA production. Among these variables, NaCl, CaCl2, PI metal solution, pH, and temperature were identified to have the significant effects (with confidence level > 90 %). Subsequently, the concentrations of NaCl, CaCl2, PI metal solution as well as the values of pH and temperature were optimized using central composite design. The cell growth and EPA production were found to respectively correlate to NaCl, CaCl2, pH, and temperature that could be represented by second-order polynomial models. The optimal values of the four parameters were determined by response surface and numerical analyses as 8 g/L NaCl, 0.10 g/L CaCl2, pH 8.5 and 19.8°C for cell dry weight (DW), and 14 g/L NaCl, 0.10 g/L CaCl2, pH 8.5 and 18°C for EPA production, respectively. The subsequent verification experiments confirmed the validity of the models. This optimization strategy led to a DW of 9 g/L, an EPA yield of 280 mg/L and an EPA productivity of 28 mg/L/d, respectively, which were considerably higher than those obtained in the previous studies. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 75: 159–169, 2001.
Three different techniques of harvesting microalgae from four pilot scale (125 m 2) and two demonstration ponds (1200 m 2) have been investigated. The processes included centrifugation, chemical flocculation followed by flotation and continuous filtration with a fine-weave belt filter. None of these processes was completely satisfactory. Centrifugation gave good recovery and a thickened slurry, but required high capital investment and energy inputs. Dissolved air flotation was more economical, but, if the recovered algae were to be incorporated into animal feed, the use of flocculants such as alum could have undesirable effects on the growth rate of the animals. This problem could be overcome by the use of non-toxic flocculants. The continuous filtration process had significant advantages in energy efficiency, economics and chemical-free operation. The only drawback of this process was that the efficiency depended on the size and morphology of the algae.
The diatom Nitzschia laevis Hust. is a potential producer of eicosapentaenoic acid (EPA). To elucidate its cellular response to salt stress, the effects of salinity on EPA production, lipid composition, and fatty acid distribution in the lipid pool were investigated. The highest contents of total fatty acids, EPA, and polar lipids were all obtained at NaCl of 20 g · L−1, under which 71.3% of total EPA existed in polar lipid fractions. In N. laevis, high salt concentration might induce the decrease in neutral lipids (NLs), whereas the production of polar lipids, including phospholipids (PLs) and glycolipids (GLs), was enhanced. The degree of fatty acid unsaturation of both neutral and polar lipid fractions increased sharply when NaCl concentration increased from 10 to 20 g · L−1 but decreased at NaCl concentration of 30 g · L−1. The amount of total free sterols was increased with the increase in salt concentration. All these changes in lipid and fatty acids suggested a decrease in membrane permeability and fluidity under high salt concentration, which could help the alga acclimate to the salinity stress.
In the development of high yield of eicosapentaenoic acid (EPA) by the diatom Nitzschia laevis, substrate limitation and toxic metabolite inhibition were the two principal obstacles. In the present study, a perfusion culture was developed to eliminate substrate (glucose) limitation and by-product inhibition simultaneously. Using an exponential feed strategy and manipulating glucose supply rate and by-product removal rate, the optimal glucose concentration in the feed medium was determined to be 50 gl−1. With this feed glucose concentration (S0=50 gl−1), the perfusion culture resulted to a cell dry weight concentration of 40 gl−1. Such a high cell density led to an EPA yield of 1112 mgl−1, which is the highest ever reported in microalgal cultures. In addition, the perfusion culture (at S0=50 gl−1) greatly reduced the amount of wastewater (broth) during cultures. Considering the simple set-up and operation, the proposed perfusion culture is potentially useful for large scale EPA production by N. laevis.
Growth Dl Euglena gracilis Z Pringsheim under pho- il
toheterotrophic conditions in a nitrogen-deprived medium
resulted in progressive loss of chloroplastic material until
total bleaching of the cells occurred. Biochemical analysis (:
and ultrastructural observation of the first stages of the
starvation process demonstrated an early lag Phase (from
O to 9 h) in which cells increased in size, followed by a
period of cell division, apparently supported by the mo
bilization of some chloroplastic proteins such as the pho-
tosynthetic CO2fixing enzyme ribulose-l,5-bisPhosPhate ~
carboxylase/ oxygenase. The degradation of the enzyme
started alter 9 h of starvation and was preceded by a
transient concentration this protein in pyrenoidal struc-
tures.Protein nitrogen and photosynthetic pigments as well as
number of chloroplasts per cell decreased during pro
liferation through mere distribution among daughterc ells.
However, alter 24 h, when cell division had almost ceased,
there was a slow but steady decline of phphotosynthetic pig
ments.This his was paralleled by observable ultrastructural
changes including progressive loss chloroPlast structure
and accumulation of paramylon granules and lipid glob
ules in the cytoplasm.These findings reinforcet he role of
chloroplastic materials as a nitrogen source during
starvation of E. gracilis in a carbon-rich medium. The "
excessof ribulose-l,5-bisphosphate carboxylasej / xygenase
acts as a first reservoir that, once exhausted is superseded
by the generalized disassembly of the photosynthetic
structures, if the adverse environment persists more than
24 h.
Partitioning of the carbon (C) fixed during photosynthesis between neutral lipids (NL) and carbohydrates was investigated in Isochrysis sp. (Haptophyceae) in relation to its nitrogen (N) status. Using batch and nitrate-limited continuous cultures, we studied the response of these energy reserve pools to both conditions of N starvation and limitation. During N starvation, NL and carbohydrate quotas increased but their specific growth rates (specific rates of variation, μCAR and μNL) decreased. When cells were successively deprived and then resupplied with NO3, both carbohydrates and neutral lipids were inversely related to the N quota (N:C). These negative relationships were not identical during N impoverishment and replenishment, indicating a hysteresis phenomenon between N and C reserve mobilizations. Cells acclimated to increasing degrees of N limitation in steady-state chemostat cultures showed decreasing NL quota and increasing carbohydrate quota. N starvation led to a visible but only transient increase of NL productivity. In continuous cultures, the highest NL productivity was obtained for the highest experimented dilution rate (D = 1.0 d−1; i.e., for non N-limited growth conditions), whereas the highest carbohydrate productivity was obtained at D = 0.67 d−1. We used these results to discuss the nitrogen conditions that optimize NL productivities in the context of biofuel production.
The dispersed air flotation (DiAF) process was utilized to separate algal cells (Chlorella sp.) from water. Two types of collector, cationic N-cetyl-N,N,N-trimethylammonium bromide (CTAB) and anionic sodium dodecylsulfate (SDS), were used. It was observed that 20% of cell removal was achieved in the presence of 40 mg/L of SDS, and ca. 86% of the cells were removed at 40 mg/L of CTAB. Upon the addition of 10 mg/L of chitosan, over 90% of the cells were removed when SDS (20 mg/L) was used as the collector. Air flow rate affected cell flotation slightly. Optimum pH values for cell flotation were from 4.0 to 5.0. Flotation efficiency decreased at high ionic strength. The electrostatic interaction between collector and cell surface plays a critical role in the separation processes.
The commercial culture of microalgae is now over 30 years old with the main microalgal species grown being Chlorella and Spirulina for health food, Dunaliella salina for β-carotene, Haematococcus pluvialis for astaxanthin and several species for aquaculture. The culture systems currently used to grow these algae are generally fairly unsophisticated. For example, Dunaliella salina is cultured in large (up to approx. 250 ha) shallow open-air ponds with no artificial mixing. Similarly, Chlorella and Spirulina also are grown outdoors in either paddle-wheel mixed ponds or circular ponds with a rotating mixing arm of up to about 1 ha in area per pond. The production of microalgae for aquaculture is generally on a much smaller scale, and in many cases is carried out indoors in 20–40 1 carboys or in large plastic bags of up to approximately 1000 1 in volume. More recently, a helical tubular photobioreactor system, the BIOCOIL™, has been developed which allows these algae to be grown reliably outdoors at high cell densities in semi-continuous culture. Other closed photobioreactors such as fiat panels are also being developed. The main problem facing the commercialisation of new microalgae and microalgal products is the need for closed culture systems and the fact that these are very capital intensive. The high cost of microalgal culture systems relates to the need for light and the relatively slow growth rate of the algae. Although this problem has been avoided in some instances by growing the algae heterotrophically, not all algae or algal products can be produced this way.
The diatom Achnanthes brevipes C.A. Ag. was cultured in the presence of limiting concentrations of nitrogen (N) or inorganic phosphate (Pi). Growth, in terms of final yield, was more affected by N limitation than Pi limitation; N limitation had a greater effect also on protein and chlorophyll content. Carbohydrate concentrations increased under both nutrient starvation treatments, but N or Pi limitation had different effects. Total (intracellular plus extracellular) sugar content increased when cells were exposed to both types of nutrient limitation, but the extracellular polysaccharide fraction increased only in the presence of Pi starvation. Analyses were performed to identify the metabolic changes occurring in cells exposed to low phosphate because this was the main condition that affected carbohydrate extrusion. Activities of several enzymes involved in carbohydrate metabolism showed that under Pi limitation there was no activation of alternative reactions that were found to result in Pi liberation, instead of its consumption, in some higher plants and in the green alga Selenastrum minutum Naeg. Collins. Results showed that activities of pyruvate kinase, phosphorylating NAD-dependent 3-phosphate-glyceraldehyde dehydrogenase, and 3-phospho-glycerate kinase were inhibited under Pi-limited conditions compared with control cells, indicating limited glucose catabolism. Activity of uridine diphosphate glucose pyrophosphorylase, a key enzyme for the biosynthesis of the storage compound crysolaminarin, was also partly inhibited in Pi-stressed cells. Our findings suggest that carbohydrate catabolism in A. brevipes is limited under Pi deficiency, whereas extracellular extrusion of carbohydrate is favored.
Cell walls of forty Chlorella strains covering all species of the Algal Collection of Göttingen (C. fusca var. vacuolata, C. kessleri, C. luteoviridis, C. minutissima, C. protothecoides, C. saccharophila, C. sorokiniana, C. vulgaris, and C. zofingiensis) were compared. The nine species were divided into two groups according to the major sugar in the rigid wall. The first group had a glucose-mannose-rigid wall and included C. fusca var. vacuolata, C. luteoviridis, C. minutissima, C. protothecoides, C. saccharophila, and C. zofingiensis. The second group, with a glucosamine-rigid wall, included C. kessleri, C. sorokiniana, and C. vulgaris. Chlorella strains of the nine species were further classified by constituent sugars, ruthenium red stainability, and anisotropy of the cell walls.
BACKGROUND: As a potential source of biomass, Jerusalem artichoke has been studied for bioethanol production; however, thus far it has not been investigated for the production of other liquid biofuels, such as biodiesel. This work aims to develop a novel approach for biodiesel production from Jerusalem artichoke tuber using heterotrophic microalgae.
RESULTS: In this study, Chlorella protothecoides utilized hydrolysate of Jerusalem artichoke tuber as carbon source and accumulated lipid in vivo, with lipid content as high as 44% by dry mass, and a carbon source to lipid conversion ratio of about 25% in a 4-day scale cultivation. The lipids were extracted and then converted into biodiesel by transesterification. Cetane acid methyl ester, linoleic acid methyl ester and oleic acid methyl ester were the dominating components of the biodiesel produced. Unsaturated fatty acids methyl ester constituted over 82% of the total biodiesel content.
CONCLUSION: This work suggests the feasibility of an alternative method of producing biodiesel from Jerusalem artichoke tuber using microalgae cultivation, and a cost reduction of carbon source feed in algal oil production can be expected. Copyright
Microalgae are a great source of many highly valuable products such as polyunsaturated fatty acids, astaxanthin and bioactive compounds. Large-scale production of these products, however, has been hindered by an inability to obtain high cell densities and productivities in conventional photoautotrophic systems. High cell density processes suitable for heterotrophic cultures of microalgae may provide an alternative means for the large-scale production of algal products of high value. This paper reviews recent studies on the formation of algal products in various cultivation systems, with emphasis on the use of heterotrophic techniques. The potential employment of heterotrophic high cell density strategies for commercial production is discussed.
This study investigates the capability of the oleaginous yeast Cryptococcus curvatus O3 to synthesize microbial lipids using glucose as its sole carbon source. Both glucose concentration and varying nitrogen sources have a significant effect on cell growth and microbial lipid accumulation in batch and fed-batch cultures. When cultivated in a shaking flask at 30 °C with glucose as sole carbon source, the cellular biomass and lipid content reached 51.8 kg m−3 and 651 g kg−1, respectively. The fed-batch culture in a 30 × 10−3 m3 stirred-tank fermentor run for 185 h produced a cellular biomass, lipid content, and lipid productivity rate of up to 104.1 kg m−3, 827 g kg−1, and 0.47 kg m−3 h−1, respectively. These data indicate that C. curvatus O3 can be used as an ideal oleaginous yeast for microbial lipid production. Gas chromatography analysis of the synthesized microbial lipids revealed that the major constituents are long-chain fatty acids, such as palmitic acid, stearic acid, oleic acid, and linoleic acid. The results suggest that the microbial lipids produced by C. curvatus O3 can be used to produce biodiesel.Highlights► In this manuscript, we use the fed-batch ferementation method to improve the lipid production. ► In a 30 × 10−3 m3 stirred-tank fermentor run for 185 h produced a cellular biomass, lipid content, and lipid productivity rate of up to 104.1 kg m−3, 827 g kg−1, and 0.47 kg m−3 h−1, respectively. ► These data indicate that C. curvatus O3 can be used as an ideal oleaginous yeast for microbial lipid production.
Due to increasing oil prices and climate change concerns, biodiesel has gained attention as an alternative energy source. Biodiesel derived from microalgae is a potentially renewable and carbon–neutral alternative to petroleum fuels. One of the most important decisions in obtaining oil from microalgae is the choice of algal species to use. Eight microalgae from a total of 33 isolated cultures were selected based on their morphology and ease of cultivation. Five cultures were isolated from river and identified as strains of Scenedesmus obliquus YSR01, Nitzschia cf. pusilla YSR02, Chlorella ellipsoidea YSR03, S. obliquus YSR04, and S. obliquus YSR05, and three were isolated from wastewater and identified as S. obliquus YSW06, Micractinium pusillum YSW07, and Ourococcus multisporus YSW08, based on LSU rDNA (D1-D2) and ITS sequence analyses. S. obliquus YSR01 reached a growth rate of 1.68 ± 0.28 day−1 at 680nm and a biomass concentration of 1.57 ± 0.67 g dwt L−1, with a high lipid content of 58 ± 1.5%. Under similar environmental conditions, M. pusillum reached a growth rate of 2.3 ± 0.55 day−1 and a biomass concentration of 2.28 ± 0.16 g dwt L−1, with a relatively low lipid content of 24 ± 0.5% w/w. The fatty acid compositions of the studied species were mainly myristic, palmitic, palmitoleic, oleic, linoleic, g-linolenic, and linolenic acids. Our results suggest that S. obliquus YSR01 can be a possible candidate species for producing oils for biodiesel, based on its high lipid and oleic acid contents.
Heterotrophic growth of Chlamydomonas reinhardtii using acetate as a carbon source and nitrate, ammonium and urea as nitrogen sources in batch cultrues was investigated. These nitrogen sources supported good growth of C. reinhardtii, the resulting cell concentrations for nitrate, ammonium chloride and urea were 0.48, 0.44 and 0.61 g l−1, respectively. The specific growth rate μx of the urea culture was the highest (0.071 h−1), followed by that of the nitrate culture (0.062 h−1), and the specific growth rate μx of the ammonium culture was the lowest (0.058 h−1). Urea is therefore considered to be the best nitrogen source for the growth of the alga. Based on these results fed-batch cultures were performed to reach the maximum cell concentration of 1.1482 g l−1, about 1.9-fold that obtained in batch culture. Finally, a group of kinetic models for describing cell growth, pH variation and acetate consumption were proposed and a satisfactory fit between the experimental results and predicted values was demonstrated. The effects of dilution rate and acetate feed concentration on cell growth were analyzed with those with these models.
Optimization of cellular productivity of an industrial microalgae fermentation was investigated. The fermentation was carried out at Coors Biotech Products Company, Fort Collins, Colorado. A mathematical model was developed based on the data collected from pilot plant test runs at different operating conditions. Pontryagin's maximum principle was used for determining the optimal feed policy. A feedback control algorithm was also studied for maximizing the cellular productivity. During continuous operation, the optimum dilution rate was determined by an adaptive optimization scheme based on the steepest descent technique and a recursive least squares estimation of model parameters. A direct search algorithm was also applied to determine the optimum feed rate. Comparison of the theoretical results of the different optimization schemes revealed that the direct search algorithm was preferable because of its simplicity. The experimental results of real time application of the feedback algorithm agreed fairly well with those of the theoretical analyses. © 1994 John Wiley & Sons, Inc.
Diatoms are the world's largest contributors to biosilicification and are one of the predominant contributors to global carbon fixation. Silicon is a major limiting nutrient for diatom growth and hence is a controlling factor in primary productivity. Because our understanding of the cellular metabolism of silicon is limited, we are not fully knowledgeable about intracellular factors that may affect diatom productivity in the oceans. The goal of this review is to present an overview of silicon metabolism in diatoms and to identify areas for future research.
Ab α-shape tubular photobioreactor was designed and constructed based on knowledge of algal growth physiology using sunlight.
The algal culture is lifted 5 m by air to a receiver tank. From the receiver tank, the culture flows down parallel polyvinyl-chloride
tubes of 25 m length and 2.5 cm internal diameter, placed at an angle of 25 ° with the horizontal to reach another set of
air riser tubes. Again the culture is lifted 5 m to another receiver tank, then flows down parallel tubes connected to the
base of the first set of riser tubes. Thus, the bioreactor system looks like the symbol α. As there is no change in the direction
of the liquid flow, high liquid flow rate and Reynolds Number can be achieved at relatively low air flow rate in the riser
tubes. Due to the high area-volume ratio of the bioreactor, and equable photosynthetically available radiance and culture
temperature, biomass density of exceeding 10 g dry weight L-1 and daily output rate of 72 g dry weight m-2 land d-1 were achieved.
The fatty acid composition of the alga Chlorella saccharophila was investigated under different growth conditions. Using glucose as the sole carbon source, heterotrophically-grown Chlorella saccharophila produced a greater proportion of the polyunsaturated fatty acids (C18: 2 and C18: 3) than photosynthetic cultures, with linoleic acid (C18: 2) predominating. An unexpected discovery was the observation that at the lowest glucose concentration (2.5 gl–1) the lipid content of the algae increased to between 36–47% of the cell weight, depending on the temperature. At glucose concentrations of 5 g l–1 or more, the lipid content fell to 10–12% of the cell, although total fatty acid yield was higher due to higher biomass concentrations. Aeration of heterotrophic cultures promoted the production of unsaturated fatty acids compared to non-aerated cultures.
An isolate of Isochrysis galbana rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) has been grown as a chemostat culture at 20 C and pH 8.00 controlled by CO2 injection. From a low dilution rate (D) of 0.0024 h–1 to 0.0377 h–1, close to maximum growth, a decrease in EPA content from 5.21% dry weight (d.w.) to 2.80% d.w. was observed, although the percentage of EPA in the total fatty acids increased. Lipids were fractionated, EPA being the major fatty acid found in the glycolipid fraction, whereas in the neutral lipid fraction were mainly palmitic and palmitoleic acids. At the same time, the biomass concentration also decreased from 1015 mgl–1 to 202 mgl–1 over the range of Ds mentioned. Nonetheless, EPA productivity had a maximum value of 15.26 mgl–1day–1 at D=0.0208 h–1.