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1.
The effects of wavelengths of light-emitting diode (LED), nitrate concentration, and salt concentration were evaluated for the two-phase culture of the microalgal species Phaeodactylum tricornutum, Dunaliella tertiolecta, and Isochrysis galbana on cell growth and lipid production. Blue LEDs produced the highest biomass of P. tricornutum at a nitrate concentration of 8 mg/L, reaching 0.97 g dcw/L with a specific growth rate (μ) of 0.047 h−1, followed by I. galbana with 0.79 g dcw/L and μ = 0.040 h−1 and D. tertiolecta with 0.55 g dcw/L and μ = 0.028 h−1. Of the three microalgae, P. tricornutum had the highest specific growth rate of μmax = 0.070 h−1 and lowest saturation constant of Ks = 4.18 mg/L, resulting in fast cell growth. The highest lipid production was obtained under green LED wavelength stress on day 14, reaching 60.6% (w/w) of the dry cell weight among the three microalgae. The main fatty acids produced by the three microalgae were myristic acid (C14:0), palmitic acid (C16:0), oleic acid (C18:1), and arachidic acid (C20:0), which comprised 72.68%–84.16% (w/w) of the total fatty acids content under three stresses.  相似文献   

2.
This study investigated the changes in lipid and starch contents, lipid fraction, and lipid profile in the nitrogen-starved Scenedesmus obtusus XJ-15 at different temperatures (17, 25, and 33 °C). The optimal temperature for both growth and lipid accumulation under nitrogen-sufficient condition was found to be 25 °C. However, under nitrogen deprivation, the total and neutral lipids increased with increasing temperature, and achieved the highest lipid content of 47.60 % of dry cell weight and the highest TAG content of 79.66 % of total lipid at 33 °C. In the meantime, the stored cellular starch content decreased with the increasing temperature. Thus, high temperature induced carbon flux from starch toward TAG accumulation in microalgae during nitrogen starvation. In addition, the decreased polar lipids may also serve for TAG synthesis under high temperature, and high temperature further reduced the degree of the fatty acid unsaturation and favored a better biodiesel production. These results suggested that high-temperature stress can be a good strategy for enhancing biofuel production in oleaginous microalgae during nitrogen deficiency.  相似文献   

3.
Diatoms are photosynthetic microalgae that fix a significant fraction of the world’s carbon. Because of their photosynthetic efficiency and high-lipid content, diatoms are priority candidates for biofuel production. Here, we report that sporulating Bacillus thuringiensis and other members of the Bacillus cereus group, when in co-culture with the marine diatom Phaeodactylum tricornutum, significantly increase diatom cell count. Bioassay-guided purification of the mother cell lysate of B. thuringiensis led to the identification of two diketopiperazines (DKPs) that stimulate both P. tricornutum growth and increase its lipid content. These findings may be exploited to enhance P. tricornutum growth and microalgae-based biofuel production. As increasing numbers of DKPs are isolated from marine microbes, the work gives potential clues to bacterial-produced growth factors for marine microalgae.  相似文献   

4.
Monoclonal antibodies (mAbs) represent actually the major class of biopharmaceuticals. They are produced recombinantly using living cells as biofactories. Among the different expression systems currently available, microalgae represent an emerging alternative which displays several biotechnological advantages. Indeed, microalgae are classified as generally recognized as safe organisms and can be grown easily in bioreactors with high growth rates similarly to CHO cells. Moreover, microalgae exhibit a phototrophic lifestyle involving low production costs as protein expression is fueled by photosynthesis. However, questions remain to be solved before any industrial production of algae-made biopharmaceuticals. Among them, protein heterogeneity as well as protein post-translational modifications need to be evaluated. Especially, N-glycosylation acquired by the secreted recombinant proteins is of major concern since most of the biopharmaceuticals including mAbs are N-glycosylated and it is well recognized that glycosylation represent one of their critical quality attribute. In this paper, we assess the quality of the first recombinant algae-made mAbs produced in the diatom, Phaeodactylum tricornutum. We are focusing on the characterization of their C- and N-terminal extremities, their signal peptide cleavage and their post-translational modifications including N-glycosylation macro- and microheterogeneity. This study brings understanding on diatom cellular biology, especially secretion and intracellular trafficking of proteins. Overall, it reinforces the positioning of P. tricornutum as an emerging host for the production of biopharmaceuticals and prove that P. tricornutum is suitable for producing recombinant proteins bearing high mannose-type N-glycans.  相似文献   

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Phaeodactylum tricornutum is a marine diatom in the class Bacillariophyceae and is important ecologically and industrially with regards to ocean primary production and lipid accumulation for biofuel production, respectively. Triacylglyceride (TAG) accumulation has been reported in P. tricornutum under different nutrient stresses, and our results show that lipid accumulation can occur with nitrate or phosphate depletion. However, greater lipid accumulation was observed when both nutrients were depleted as observed using a Nile Red assay and fatty acid methyl ester (FAME) profiles. Nitrate depletion had a greater effect on lipid accumulation than phosphate depletion. Lipid accumulation in P. tricornutum was arrested upon resupplementation with the depleted nutrient. Cells depleted of nitrogen showed a distinct shift from a lipid accumulation mode to cellular growth post-resupplementation with nitrate, as observed through increased cell numbers and consumption of accumulated lipid. Phosphate depletion caused lipid accumulation that was arrested upon phosphate resupplementation. The cessation of lipid accumulation was followed by lipid consumption without an increase in cell numbers. Cells depleted in both nitrate and phosphate displayed cell growth upon the addition of both nitrate and phosphate and had the largest observed lipid consumption upon resupplementation. These results indicate that phosphate resupplementation can shut down lipid accumulation but does not cause cells to shift into cellular growth, unlike nitrate resupplementation. These data suggest that nutrient resupplementation will arrest lipid accumulation and that switching between cellular growth and lipid accumulation can be regulated upon the availability of nitrogen and phosphorus.  相似文献   

7.
We tested the hypothesis that increased pH reduces the amount of structural lipids. To do this, we used three different diatoms (Phaeodactylum tricornutum CCAP strain, P. tricornutum TV strain and Amphiprora sp). We tested the effect of rapid increase from pH?7.5 to 10 by adding NaOH. The total lipid content was reduced by 13, 36 and 47 % in the P. tricornutum CCAP strain, TV strain and Amphiprora sp., respectively, 1 h after increasing the pH. The P. tricornutum CCAP strain was used for further testing the effect of pH on the lipid content during active growth. This strain was cultivated at pH?7.5 and 10, and the pH was regulated by the CO2 inflow. The growth rate was similar (0.3 day?1) in both pH treatments, but the lipid content in the pH?10 treatment was on average 28 % lower than in the pH?7.5 treatment. Our data support the hypothesis that structural lipids are reduced when pH increases to high levels. The results suggest that regulating the pH during algae cultivation could be used to refine the lipid composition in the harvested algal biomass.  相似文献   

8.
Microalgae contain lipid bodies (LBs) composed of triacylglycerols, which can be converted to biodiesel. Here we demonstrate a method to study the accumulation patterns of LBs in different microalgae strains and culture conditions utilizing laser scanning confocal microscopy (LSCM) with BODIPY 505/515 (4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene) staining, in parallel with Nile Red (9-diethylamino-5H-benzo-a-phenoxazine-5-one) fluorescence analysis of intracellular lipids in microplates. Phaeodactylum tricornutum and Tetraselmis suecica were selected as model organisms and monitored throughout the growth phases in standard and nitrogen-deficient growth conditions. Utilizing image quantification techniques, the number and morphology of LBs suggest that P. tricornutum accumulates lipids by merging with existing LBs, while T. suecica synthesizes new LBs. We observed that T. suecica accumulates a higher number of LBs and total volume of lipids per cell, while P. tricornutum accumulates only 1–2 LBs with a larger volume per LB. LSCM analysis complements Nile Red (NR) methods because LSCM provides three-dimensional images of lipid accumulation at a cellular level, while NR analysis can quickly monitor the total levels of intracellular lipids for phenotypic screening. Using NR analysis, we have observed that the optimal harvest date for P. tricornutum and T. suecica in standard cultivation conditions is 24 and 42 days, respectively. Comparison with nitrogen-deficient growth conditions is utilized as a model to confirm that LSCM and NR analysis can be used to study lipid storage and productivity for diverse growth conditions and various strains of microalgae.  相似文献   

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11.
Oils, carbohydrates, and fats generated by microalgae are being refined in an effort to produce biofuels. The research presented here examines two marine microalgae, Nannochloropsis salina (green alga) and Phaeodactylum tricornutum (diatom), when grown with 0 (no addition), 0.5, 1.0, 2.0, and 5.0 g L?1 NaHCO3 added to an f/2 medium during the growth phase (GP) and a nutrient induced (nitrate limitation) lipid formation phase (LP). We hypothesize that the addition of NaHCO3 is a sustainable and practical strategy to increase cellular density and concentrations of lipids in microalgae as well as the rate of lipid accumulation. In N. salina, final cell densities were significantly (p?<?0.05) higher in the NaHCO3-treated cells than the control while in P. tricornutum the cell densities were higher with >[NaHCO3] during the GP. During the LP, cell densities were generally higher in the NaHCO3-treated cells compared with controls. F V/F M (efficiency of photosystem II) patterns paralleled those for cell density with generally higher values with higher concentrations of NaHCO3 and significantly different values between controls and 5.0 g L?1 NaHCO3 at the end of the GP (p?<?0.05). F V/F M was variable between treatments in P. tricornutum (0.3–0.65) but less so in N. salina for (0.5–0.7) regardless of [NaHCO3]. The lipid index (measured with Nile red), used as a proxy for triacylglycerides (TAGs), was 10.2?±?6.5 and 4.4?±?2.9 (fluorescence units/OD cells ×1000) for N. salina and P. tricornutum, respectively, at the end of the GP. At the end of the LP, the lipid index was eight and four times higher than during the GP in the corresponding 5.0 g L?1 NaHCO3 treatments, revealing that N. salina was accumulating more lipid than P. tricornutum. Dry weights essentially doubled during LP compared with GP for N. salina; this was not the case for P. tricornutum. In general, the percentage of ash in dry weights was significantly higher in the LP relative to the corresponding GP treatments for P. tricornutum; this was not the case for N. salina. During the LP, there was also less soluble protein in N. salina compared to GP; differences were not significant in cells growing with 2.0 or 5.0 g L?1 NaHCO3. In P. tricornutum, faster growing cells had more soluble protein during the GP and LP; differences between treatments were significant. P. tricornutum generally accumulated significantly more crude protein than N. salina at higher [NaHCO3]; there was three times more crude protein in the highest NaHCO3 (5.0 g L?1) treatment compared with the controls. C:N ratios (mol:mol) were similar across treatments during GP: 7.03?±?0.12 and 10.16?±?0.41 for N. salina and P. tricornutum, respectively. Further, C:N ratios increased with increasing [NaHCO3] during LP. Species-specific fatty acid methyl ester (FAMEs) profiles were observed. While C16:0 was lower in P. tricornutum compared to N. salina, the diatom produced more C16:1 and C14 but not C18:3. Monounsaturated fatty acids (MUFA) significantly increased in N. salina in the LP compared to GP and in response to increasing [NaHCO3] (t tests; p?<?0.05). Saturated fatty acids (SFA) responded similarly but to a lesser degree. There were more polyunsaturated fatty acids (PUFA) in N. salina than MUFAs or SFAs. In P. tricornutum, there were generally more SFAs, MUFAs and PUFAs in P. tricornutum during LP than GP in the corresponding NaHCO3 treatments. These findings reveal the importance of considering NaHCO3 as a supplemental carbon source in the culturing marine phytoplankton in large-scale production for biofuels.  相似文献   

12.
We evaluated the salt tolerance of hybrids of pepper (Capsicum annuum L.) during germination. Treatments were applied at 0, 25, and 50 mM NaCl with preparations of supplemental extracts of the microalgae Dunaliella salina and Phaeodactylum tricornutum to determine the percentage germination rate as well as measured indicators of oxidative stress caused by the salt treatments during seed germination. We found that root growth was favorably influenced by the microalgae leading to increased germination rate. Tissues were analyzed in terms of superoxide radical production, lipid peroxidation, and activity of antioxidant enzymes viz. superoxide dismutase, catalase, and glutathione peroxidase. Our results suggest that application of microalgae extracts significantly reduced (p?<?0.05) superoxide radical production, as well as lower lipid peroxidation in comparison to plants without extracts of microalgae. The antioxidant enzymes increased in the presence of microalgae showing a significant difference (p?<?0.05). The results suggest differences in oxidative metabolism in response to the magnitude of salt stress and concentrations of microalgae help mitigate salt stress in plants during the germination process.  相似文献   

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14.
As one of the promising species of microalgae for biofuel production, Chlorella vulgaris CS-42 was cultivated phototrophically in two cylindrical photobioreactors with aeration of 5 % (v/v) CO2 or air for 13 days to evaluate the effects of CO2 supplementation on biomass, CO2 fixation performance, and biochemical content. Significant increases of specific growth rate and total carbon content in biomass resulting in a higher CO2 fixation rate were found with 5 % CO2. The maximum biomass concentration, carbohydrate and fatty acid contents with 5 % CO2 were significantly higher than those with air, while carbohydrate biosynthesis was most affected as compared to other biochemical components. Cytomic analysis revealed a rapid accumulation of neutral lipid in the late growth phase with more lipid bodies visualized by confocal laser scanning microscopy (CLSM), when nitrate consumption was accelerated with CO2 supplementation. Gas chromatography mass spectrometry (GC-MS) analysis indicated that 5 % CO2 favored the formation of C18:2, which led to a decrease in the degree of lipid unsaturation (DLU). These results proved that CO2 supplementation was one of the most efficient methods to significantly prompt the growth of microalgae and increase the C/N ratio in the medium, which in turn regulated the carbon metabolic flux to enhance neutral lipid and fatty acid production in C. vulgaris.  相似文献   

15.
The effect of overexpression of endogenous delta-6 fatty acid desaturase gene (ER<DELTA>6FAD) on eicosapentaenoic acid (EPA) production and total lipid content was investigated in Phaeodactylum tricornutum. All three randomly selected transformants exhibited significant increase (47.66%) in their EPA content, which reached up to 38.101 mg/g DW in algal strain D63. The total lipid content of all the three transformants was significantly higher (16.40–18.64%) than that of the wild-type strain. These findings suggested that the reaction catalyzed by ER<DELTA>6FAD is a limiting step for EPA biosynthesis, overexpression of endogenous ER<DELTA>6FAD gene is an effective method for improving EPA production in eukaryotic microalgae, and fatty acid metabolic pathway in microalgae can be genetically modified.  相似文献   

16.
The microalgae Phaeodactylum tricornutum (PT) is known for its high content of omega-3 fatty acids, which are known to attenuate inflammation. Additionally, this microalga contains other nutrients such as carbohydrates, vitamins, proteins, and carotenoids and therefore could be of interest for animal and human nutrition. Here, we investigated the effects of hexane, ethanolic, and aqueous extracts on lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells (PBMCs) and on the murine macrophage cell line RAW 264.7. Further, the cytotoxicity of the extracts was studied using the MTT assay. The production of pro-inflammatory cytokines was significantly inhibited by the ethanolic and aqueous P. tricornutum extracts but not by the hexane extract. Both at the mRNA and at the protein levels, the aqueous extract inhibited LPS-induced IL-6, IL-1β, TNFα, and COX-2 expression and release by up to 96% (mRNA) and 79% (protein) in a dose-dependent manner. Compared to the aqueous extract, the ethanolic extract was less effective in cytokine inhibition. The production of nitric oxide in RAW 264.7 cells was significantly reduced by all extracts. We showed that the anti-inflammatory effect of P. tricornutum is exerted through inhibition of nuclear factor-κB activation and dependent on the mitogen-activated protein (MAP) kinase pathway. Our data indicate anti-inflammatory effects of the aqueous P. tricornutum extract and provide a basis information on the safety and potential health benefits of P. tricornutum usage for future animal and human nutrition.  相似文献   

17.
Under nutrient deplete conditions, diatoms accumulate between 15% to 25% of their dry weight as lipids, primarily as triacylglycerols (TAGs). As in most eukaryotes, these organisms produce TAGs via the acyl‐CoA dependent Kennedy pathway. The last step in this pathway is catalyzed by diacylglycerol acyltransferase (DGAT) that acylates diacylglycerol (DAG) to produce TAG. To test our hypothesis that DGAT plays a major role in controlling the flux of carbon towards lipids, we overexpressed a specific type II DGAT gene, DGAT2D, in the model diatom Phaeodactylum tricornutum. The transformants had 50‐ to 100‐fold higher DGAT2D mRNA levels and the abundance of the enzyme increased 30‐ to 50‐fold. More important, these cells had a 2‐fold higher total lipid content and incorporated carbon into lipids more efficiently than the wild type (WT) while growing only 15% slower at light saturation. Based on a flux analysis using 13C as a tracer, we found that the increase in lipids was achieved via increased fluxes through pyruvate and acetyl‐CoA. Our results reveal overexpression of DAGT2D increases the flux of photosynthetically fixed carbon towards lipids, and leads to a higher lipid content than exponentially grown WT cells.  相似文献   

18.
Microalgae as sources for biodiesel production have been widely investigated. Microalgae biomass, lipid content and fatty acid profiles of microalgae are limiting factors for the cost-effective production of biodiesel. In this paper, the effects of high ferric ion concentrations on three species of microalgae (Tetraselmis subcordiformis, Nannochloropsis oculata and Pavlova viridis) were studied. The microalgae were cultured in different concentrations (1.2?×?10?2, 1.2?×?10?1, 1.2 and 12 mmol L?1) of ferric ion. The growth, lipid content and fatty acid profiles of the three microalgae were analysed. When algae were cultured in 1.2 mmol L?1 ferric ion for 10 days, the final cell density and specific growth rates of T. subcordiformis, N. oculata and P. viridis decreased significantly (p?<?0.05), and the total lipid contents of the microalgae, 33.72, 37.34 and 29.48 % (dry mass) in T. subcordiformis, N. oculata and P. viridis, respectively, were higher than those at other concentrations. The neutral lipid/total lipid ratios of the three microalgae species increased with increasing ferric ion concentration. Neutral lipids accounted for 50.75, 48.37 and 46.59 % of the total lipid in T. subcordiformis, N. oculata and P. viridis, respectively, when cultured in 12 mmol L?1 ferric ion. The proportions of saturated fatty acids in all three species cultured in 12 mmol L?1 ferric ion were significantly higher than those cultured in lower ferric ion concentrations. An optimum ferric ion concentration can improve the properties of T. subcordiformis, N. oculata and P. viridis as sources for biodiesel.  相似文献   

19.
Photosynthetic microalgae are of burgeoning interest in the generation of commercial bioproducts. Microalgae accumulate high lipid content under adverse conditions, which in turn compromise their growth and hinder their commercial potential. Hence, it is necessary to engineer microalgae to mitigate elevated lipid accumulation and biomass. In this study, we identified acetyl-CoA carboxylase (ACCase) in oleaginous microalga Phaeodactylum tricornutum (PtACC2) and expressed constitutively in the chloroplast to demonstrate the potential of chloroplast engineering. Molecular characterization of transplastomic microalgae revealed that PtACC2 was integrated, transcribed and expressed successfully, and localized in the chloroplast. Enzymatic activity of ACCase was elevated by 3.3-fold, and the relative neutral lipid content increased substantially by 1.77-fold, and lipid content reached up to 40.8% of dry weight. Accordingly, the number and size of oil bodies markedly increased. Fatty acid profiling showed that the content of monounsaturated fatty acids increased, while polyunsaturated fatty acids decreased. This method provides a valuable genetic engineering toolbox for microalgal bioreactors with industrial significance.  相似文献   

20.
E3 ubiquitin ligases determine the substrate specificity of ubiquitination. Plant U-box (PUB) E3 ligases, with a typical 70-amino acid U-box domain, participate in plant developmental processes and environmental responses. Thus far, 64 PUB proteins have been identified in Arabidopsis and 77 PUB proteins have been identified in Oryza. However, detailed studies on U-box genes in the model microalgae Chlamydomonas reinhardtii are lacking. Here, we present a comprehensive analysis of the genes encoding U-box family proteins in C. reinhardtii. Following BLASTP analysis, 30 full-length U-box genes were identified in the C. reinhardtii genome sequence. Bioinformatics analyses of CrPUB genes were performed to characterize the phylogenetic relationships, chromosomal locations and gene structures of each member. The 30 identified CrPUB proteins are clustered into 3 distinct subfamilies, and the genes for these proteins are unevenly distributed among 14 chromosomes. Furthermore, the quantitative real-time RT-PCR or semi-quantitative RT-PCR analysis of 30 CrPUB mRNA abundances under nitrogen starvation showed that 18 CrPUB genes were induced by N starvation and that 7 genes were repressed in the N-poor environment. We selected five CrPUB genes exhibiting marked changes in expression under N-free conditions for further analysis in RNAi experiments and examined the oil content of these gene-silenced transgenic strains. The silencing of CrPUB5 and CrPUB14, which are typically down-regulated under N starvation, induced 9.8%-45.0% and 14.4%-61.8% lipid accumulation, respectively. In contrast, the silencing of CrPUB11, CrPUB23 and CrPUB28, which are markedly up-regulated under N-free conditions, decreased the lipid content by 5.5%-27.8%, 8.1%-27.3% and 6.6%-27.9%, respectively. These results provide a useful reference for the identification and functional analysis of this gene family and fundamental information for microalgae lipid metabolism research.  相似文献   

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