Search Results (15)

A novel urea complexation technology was developed based on the agglomeration phenomenon induced by ambient-temperature agitation of a ternary system consisting of urea, water and fatty acid ethyl esters (EEs). The agglomeration phenomenon can be regarded as an intuitive indicator to judge the occurrence of urea complexation. Using docosahexaenoic acid (DHA) containing EE (DHA-EE) from Crypthecodinium cohnii oil as the substrate, key variables including agglomeration time, urea/DHA-EE ratio, water/DHA-EE ratio, and temperature were investigated. The urea complexation predominantly occurred within 15 min following agglomerate formation. Temperature in the range of 0–40°C exerted no significant effect on the yield of the non-urea-complexed fraction or its DHA content, enabling the operation to be conducted at room temperature without heating or cooling. Under unoptimized conditions, the proposed method effectively increased the DHA content of EE from Crypthecodinium cohnii oil from 40.73% to 89.87%. For EE from Schizochytrium sp. oil, the contents of DHA and docosapentaenoic acid were improved from 47.17% and 13.93% to 69.30% and 20.29%, respectively. Meanwhile, the contents of eicosapentaenoic acid and DHA in two EE form fish oils were enhanced from 18.26% and 11.76% to 34.86% and 22.96%, and from 13.30% and 57.24% to 15.66% and 68.68%, respectively. The present study provided a novel technical pathway for the efficient enrichment of polyunsaturated fatty acids. Full article

In this review dedicated to the great protistologist Edouard Chatton (1883–1947), I wanted to highlight the originality and remarkable diversity of some dinoflagellate protists through the lens of cell biology. Their fossilized traces date back to more than 538 million years (Phanerozoic eon). However, they may be much older because acritarchs from the (Meso) Proterozoic era (1500 million years ago) could be their most primitive ancestors. Here, I described several representative examples of the various lifestyles of free-living (the autotrophic thecate Prorocentrum micans Ehrenberg and the heterotrophic athecate Noctiluca scintillans McCartney and other “pseudo-noctilucidae”, as well as the thecate Crypthecodinium cohnii Biecheler) and of parasitic dinoflagellates (the mixotroph Syndinium Chatton). Then, I compared the different dinoflagellate mitotic systems and reported observations on the eyespot (ocelloid), an organelle that is present in the binucleated Glenodinium foliaceum Stein and in some Warnowiidae dinoflagellates and can be considered an evolutionary marker. The diversity and innovations observed in mitosis, meiosis, reproduction, sexuality, cell cycle, locomotion, and nutrition allow us to affirm that dinoflagellates are among the most innovative unicells in the Kingdom Protista. Full article

Crypthecodinium cohnii is a marine microalga that can accumulate high amounts of polyunsaturated fatty acids (PUFAs) and thus replace conventional routes of fish oil production. They are associated with the destruction of marine resources and multiple downstream/purification complications. The major drawbacks of using C. cohnii for industrial-scale production are associated with low PUFA productivity. One of the means of increasing the PUFA synthesis rate is to maintain the medium component concentrations at optimal values throughout cultivation, thus increasing PUFA production efficiency, which can result in the successful transfer of the process to pilot and/or industrial scale. The goal of the present research was to develop techniques for increasing the efficiency of PUFA production via C. cohnii cultivation. Multiple experiments were carried out to test and fine-tune the cultivation medium composition and oxygen transfer factors. The biomass yields from individual components, yeast extract, sea salts, and glucose amounted to 5.5, 0.65, and 0.61 g·g⁻¹, respectively. C. cohnii cell susceptibility to mechanical damage was experimentally evaluated. Power inputs of <276.5 W/m³ did not seem to promote cell destruction when Pitched-blade impellers were used. The obtained cultivation conditions were shown to be efficient in terms of increasing the biomass productivity and the omega-3 fatty acid content in C. cohnii. By using the applied methods, the maximal biomass productivity reached 8.0 g·L⁻¹·day⁻¹, while the highest obtained biomass concentration reached 110 g·L⁻¹. A steady increase in the concentration of PUFAs during cultivation was observed from the FTIR data. Full article

The heterotrophic Crypthecodinium cohnii is a major model for dinoflagellate cell biology, and a major industrial producer of docosahexaenoic acid, a key nutraceutical and added pharmaceutical compound. Despite these factors, the family Crypthecodiniaceae is not fully described, which is partly attributable to their degenerative thecal plates, as well as the lack of ribotype-referred morphological description in many taxons. We report here significant genetic distances and phylogenetic cladding that support inter-specific variations within the Crypthecodiniaceae. We describe Crypthecodinium croucheri sp. nov. Kwok, Law and Wong, that have different genome sizes, ribotypes, and amplification fragment length polymorphism profiles when compared to the C. cohnii. The interspecific ribotypes were supported by distinctive truncation-insertion at the ITS regions that were conserved at intraspecific level. The long genetic distances between Crypthecodiniaceae and other dinoflagellate orders support the separation of the group, which includes related taxons with high oil content and degenerative thecal plates, to be ratified to the order level. The current study provides the basis for future specific demarcation-differentiation, which is an important facet in food safety, biosecurity, sustainable agriculture feeds, and biotechnology licensing of new oleaginous models. Full article

Wastes generated during the cultivation of marine microalga Crypthecodinium cohnii and after the lipid extraction process, were energetically valorized into biogas production through anaerobic digestion (AD). The tested wastes were extracted microalgae (Ae) with hexane (AeH) using supercritical extraction methods (AeS) and the supernatant obtained after culture medium centrifugation (M). The digestion of the algae biomass in the admixture with the supernatant medium (AeH+M+I and AeS+M+I) provided a higher methane content and a higher methane yield (582 and 440 L CH₄/kg VS) than the substrates Ae and M, individually digested (155 and 96 L CH₄/kg VS, respectively). Flow cytometry monitoring processes during AD indicated that the yield of the accumulated biogas was influenced by the operating conditions. The mixture of AeH+M+I was the only assay with a proportion of cells with less damaged membranes after AD, providing the highest methane yield and productivity (582 L CH₄/kg VS and 31 L CH₄/kg VS.d, respectively) and the highest energetic potential of 5.8 KWh/kg VS of all the substrates. From the results, AD integration to lipid production by C. cohnii to recover energy from the generated wastes enhanced the sustainability of the entire process and promoted the practice of zero waste. Full article

Chrypthecodinium cohnii lipids have been almost exclusively used as a source of Docosahexaenoic acid (DHA). Such an approach wastes the remaining microalgal lipid fraction. The present work presents a novel process to produce C. cohnii biomass, using low-cost industrial by-products (raw glycerol and corn steep liquor), in a 7L-bioreactor, under fed-batch regime. At the end of the fermentation, the biomass concentration reached 9.2 g/L and the lipid content and lipid average productivity attained 28.0% (w/w dry cell weight) and 13.6 mg/L h, respectively. Afterwards the microalgal biomass underwent a saponification reaction to produce fatty acid (FA) soaps, which were further converted into FA ethyl ester (FA EE). C. cohnii FA EE mixture was then fractionated, using the urea complexation method at different temperatures, in order to obtain a polyunsaturated fatty acid ethyl ester (PUFA EE) rich fraction, that could be used for food/pharmaceutical/cosmetic purposes, and a saturated fatty acid ethyl ester (SAT EE) rich fraction, which could be used as biodiesel. The temperature that promoted the best separation between PUFA and SAT EE, was −18 °C, resulting in a liquid fraction with 91.6% (w/w) DHA, and a solid phase with 88.2% of SAT and monounsaturated fatty acid ethyl ester (MONOUNSAT), which could be used for biodiesel purposes after a hydrogenation step. Full article

Prior to the commissioning of a new industrial biorefinery it is deemed necessary to evaluate if the new project will be beneficial or detrimental to climate change, one of the main drivers for the sustainable development goals (SDG) of the United Nations. In particular, how SDG 7, Clean and Efficient Energy, SDG 3, Good Health and Well Being, SDG 9, Industry Innovation and Infrastructure, and SDG 12, Responsible Production and Consumption, would engage in a new biorefinery design, beneficial to climate change, i.e., fostering SDG 13, Climate Action. This study uses life cycle assessment methodology (LCA) to delve in detail into the Global Warming Impact category, project scenario GHG savings, using a conventional and a dynamic emission flux approach until 2060 (30-year lifetime). Water, heat and electricity circularity are in place by using a water recirculation process and a combined heat and power unit (CHP). A new historical approach to derive low and higher-end commodity prices (chemicals, electricity, heat, jet/maritime fuel, DHA, N-fertilizer) is used for the calculation of the economic indicators: Return of investment (ROI) and inflation-adjusted return (IAR), based upon the consumer price index (CPI). Main conclusions are: supercritical fluid extraction is the hotspot of energy consumption; C. cohnii bio-oil without DHA has higher sulfur concentration than crude oil based jet fuel requiring desulfurization, however the sulfur levels are compatible with maritime fuels; starting its operation in 2030, by 2100 an overall GHG savings of 73% (conventional LCA approach) or 85% (dynamic LCA approach) is projected; economic feasibility for oil productivity and content of 0.14 g/L/h and 27% (w/w) oil content, respectively (of which 31% is DHA), occurs for DHA-cost 100 times higher than reference fish oil based DHA; however future genetic engineering achieving 0.4 g/L/h and 70% (w/w) oil content (of which 31% is DHA), reduces the threshold to 20 times higher cost than reference fish oil based DHA; N-fertilizer, district heating and jet fuel may have similar values then their fossil counterparts. Full article

Docosahexaenoic acid (DHA) is one of the most important long-chain polyunsaturated fatty acids (LC-PUFAs), with numerous health benefits. Crypthecodinium cohnii, a marine heterotrophic dinoflagellate, is successfully used for the industrial production of DHA because it can accumulate DHA at high concentrations within the cells. Glycerol is an interesting renewable substrate for DHA production since it is a by-product of biodiesel production and other industries, and is globally generated in large quantities. The DHA production potential from glycerol, ethanol and glucose is compared by combining fermentation experiments with the pathway-scale kinetic modeling and constraint-based stoichiometric modeling of C. cohnii metabolism. Glycerol has the slowest biomass growth rate among the tested substrates. This is partially compensated by the highest PUFAs fraction, where DHA is dominant. Mathematical modeling reveals that glycerol has the best experimentally observed carbon transformation rate into biomass, reaching the closest values to the theoretical upper limit. In addition to our observations, the published experimental evidence indicates that crude glycerol is readily consumed by C. cohnii, making glycerol an attractive substrate for DHA production. Full article

Crypthecodinium cohnii is a marine heterotrophic dinoflagellate that can accumulate high amounts of omega-3 polyunsaturated fatty acids (PUFAs), and thus has the potential to replace conventional PUFAs production with eco-friendlier technology. So far, C. cohnii cultivation has been mainly carried out with the use of yeast extract (YE) as a nitrogen source. In the present study, alternative carbon and nitrogen sources were studied: the extraction ethanol (EE), remaining after lipid extraction, as a carbon source, and dinoflagellate extract (DE) from recycled algae biomass C. cohnii as a source of carbon, nitrogen, and vitamins. In mediums with glucose and DE, the highest specific biomass growth rate reached a maximum of 1.012 h⁻¹, while the biomass yield from substrate reached 0.601 g·g⁻¹. EE as the carbon source, in comparison to pure ethanol, showed good results in terms of stimulating the biomass growth rate (an 18.5% increase in specific biomass growth rate was observed). DE supplement to the EE-based mediums promoted both the biomass growth (the specific growth rate reached 0.701 h⁻¹) and yield from the substrate (0.234 g·g⁻¹). The FTIR spectroscopy data showed that mediums supplemented with EE or DE promoted the accumulation of PUFAs/docosahexaenoic acid (DHA), when compared to mediums containing glucose and commercial YE. Full article

Crude glycerol, a biodiesel industry byproduct, and corn steep liquor (CSL) derived from a starch industry, were used as carbon and nitrogen sources, respectively, for lipid production, using the heterotrophic microalga C. cohnii grown in a bench bioreactor, in a batch culture. The maximum biomass concentration, lipid content and lipid productivity attained were 5.34 g/L, 24.6% (w/w Dry Cell Weight-DCW) and 0.016 g L⁻¹ h⁻¹, respectively. Flow cytometry analysis was used to evaluate the impact of these substrates on the microalgae cells. A high proportion of intact cells with enzymatic (esterases) activity (>50%) was present throughout the cultivation time course. These results indicate that crude glycerol and CSL can be used in the medium formulation for DHA and lipid production using this microalga, which reduce the process costs in an expected maximum of 84%. Full article

The core objective of this work was to take advantage of the unexploited wheat straw biomass, currently considered as a broadly available waste stream from the Greek agricultural sector, towards the integrated valorization of sugar streams for the microbial production of polyunsaturated omega-3 fatty acids (PUFAs). The OxiOrganosolv pretreatment process was applied using acetone and ethanol as organic solvents without any additional catalyst. The results proved that both cellulose-rich solid pulp and hemicellulosic oligosaccharides-rich aqueous liquid fraction after pretreatment can be efficiently hydrolyzed enzymatically, thus resulting in high yields of fermentable monosaccharides. The latter were supplied as carbon sources to the heterotrophic microalga Crypthecodinium cohnii for the production of PUFAs, more specifically docosahexaenoic acid (DHA). The solid fractions consisted mainly of hexose sugars and led to higher DHA productivity than their pentose-rich liquid counterparts, which can be attributed to the different carbon source and C/N ratio in the two streams. The best performance was obtained with the solid pulp pretreated with ethanol at 160 °C for 120 min and an O₂ pressure of 16 bar. The total fatty acids content reached 70.3 wt% of dried cell biomass, of which 32.2% was DHA. The total DHA produced was 7.1 mg per g of untreated wheat straw biomass. Full article

In the last decades, several plant-based materials were used for the substitution of fish meal and oil in aquaculture. The present study evaluated the fish quality and the sensory differences of rainbow trout (Oncorhynchus mykiss) and pike-perch (Sander lucioperca) from three different feeding groups, which were fed a commercially available industrial (standard) diet, a control diet, and a special microorganism-based feed mix. This feed mainly consisted of a mix made of Rhodotorula glutinis, Crypthecodinium cohnii, and Arthrospira sp. and had 50% less fish meal and fish oil compared to typical control diets. At the beginning, the pike-perch population was six months old, and the rainbow trout population was 15 months old. The feeding study duration was 16 weeks and every four weeks the growth performance and several morphometric parameters were recorded. Afterwards, sensory evaluation took place to identify possible trends. Sensory evaluation revealed that the rainbow trout groups did not show any significant differences to the standard and control fish fillets with regard to odor, texture, and taste. The effects on rainbow trout growth performances and carcass parameters were similar to the standard group. The feed mix was not optimal for pike-perch farming, which was also reflected by significantly adversely affected growth performance and carcass parameters. The sensorial evaluation showed an opposite trend: here, only small differences in the fillets from the feed mix and standard/control diet were observed. Full article

Exopolysaccharide from Crypthecodinium cohnii (EPCP1-2) is a marine exopolysaccharide that evidences a variety of biological activities. We isolated a neutral polysaccharide from the fermentation liquid of Crypthecodinium cohnii (CP). In this study, a polysaccharide that is derived from Crypthecodinium cohnii were analyzed and its anti-inflammatory effect was evaluated on protein expression of toll-like receptor 4 and nuclear factor κB pathways in macrophages. The structural characteristics of EPCP1-2 were characterized by GC (gas chromatography) and GC-MS (gas Chromatography-Mass Spectrometer) analyses. The molecular weight was about 82.5 kDa. The main chain of EPCP1-2 consisted of (1→6)-linked mannopyranosyl, (1→6)-linked glucopyranosyl, branched-chain consisted of (1→3,6)-linked galactopyranosyl and terminal consisted of t-l-Rhapyranosyl. The in vitro anti-inflammatory activity was representated through assay of proliferation rate, pro-inflammatory factor (NO) and expressions of proteins on RAW 264.7, the macrophage cell line. The results revealed that EPCP1-2 exhibited significant anti-inflammatory activity by regulating the expression of toll-like receptor 4, mitogen-activated protein kinases, and Nuclear Factor-κB protein. Full article

Extracellular polysaccharides (EPS) produced by marine microalgae have the potential to be used as antioxidants, antiviral agents, immunomodulators, and anti-inflammatory agents. Although the marine microalga Crypthecodinium cohnii releases EPS during the process of docosahexaenoic acid (DHA) production, the yield of EPS remains relatively low. To improve the EPS production, a novel mutagenesis of C. cohnii was conducted by atmospheric and room temperature plasma (ARTP). Of the 12 mutants obtained, 10 mutants exhibited significantly enhanced EPS yield on biomass as compared with the wild type strain. Among them, mutant M7 was the best as it could produce an EPS volumetric yield of 1.02 g/L, EPS yield on biomass of 0.39 g/g and EPS yield on glucose of 94 mg/g, which were 33.85%, 85.35% and 57.17% higher than that of the wild type strain, respectively. Results of the present study indicated that mutagenesis of the marine microalga C. cohnii by ARTP was highly effective leading to the high-yield production of EPS. Full article