Improvement Of Lipid Production Research Articles

Comparative transcriptomic insights into key genes for biomass production and lipid synthesis in Chlorella sorokiniana mutated by argon and air atmospheric and room temperature plasma at different culture stages: Common mechanisms and unique differences

Metabolic mechanisms driving rapid lipid production in atmospheric and room temperature plasma-mutagenized Chlorella sorokiniana remain unclear, hindering improvement in lipid productivity through genetic modification. This study investigated two dominant strains, ArX13 and AirX12, mutated by argon and air ARTP, for growth, lipid production, and transcriptomic changes across various culture stages. Results showed that biomass and lipid contents peaked in the late stage, with ArX13 and AirX12 achieving final biomass of 0.71 g/L and 0.70 g/L, and lipid content of 58.57 % glipid gbiomass−1 and 46.90 % glipid gbiomass−1, 2.20 and 1.77 times higher than the control, respectively. The maximum biomass and lipid productivity occurred at the mid-stage, with ArX13 and AirX12 showing biomass productivity of 0.025 gbiomass L−1 day−1 and 0.024 gbiomass L−1 day−1, and lipid productivity of 17.79 mglipid L−1 day−1 and 11.40 mglipid L−1 day−1, respectively. Transcriptomic analysis revealed minimal changes of gene expression during early and late stages, but significant biomass and lipid synthesis during the middle stage. Key genes like PDHB, ATP6A, LSC1 and OGDH were identified in the PPI network throughout the culture cycle, with PDHB remarkably promoting lipid precursor production, up-regulated 8.37 and 4.99-fold in M_ArX13-M_X0 and M_AirX12-M_X0, respectively. In addition, high expression of photosynthesis-related genes (psaL, psaO, psaD, psaK, psbP) contributed the superior lipid production for the argon-based ARTP case, highlighting common and intrinsic mechanisms regulating microalgal lipid accumulation under different ARTP mutations. This study shows that the selection of appropriate ARTP gases can maximize the lipid production of microalgal, and the discovery of some key genes can provide a theoretical basis for the selection of target genes for microalgal research, which is of great significance for promoting the research and development of microalgal lipid engineering.

Improvement in lipid production and biodiesel quality of Pavlova sp. by monochromatic illumination

Improvement in lipid production and biodiesel quality of Pavlova sp. by monochromatic illumination

Nutrient removal and lipid production by the co-cultivation of Chlorella vulgaris and Scenedesmus dimorphus in landfill leachate diluted with recycled harvesting water

Applying microalgae for landfill leachate (LL) treatment is promising. However, LL usually needs to be diluted with much fresh water, aggravating water shortage. In this study, mono- and co-culturing microalgae (Chlorella vulgaris and Scenedesmus dimorphus) were used to treat LL diluted with recycled harvesting water, to investigate nutrient removal and lipid production. The results showed that microalgae in co-culture treatment had more biomass and stronger superoxide dismutase activity, which might be related to humic acids contained in recycled harvesting water, according to dissolved organic matters (DOMs) analysis. In addition, the lipid content and yield of co-cultured microalgae reached 27.60 % and 66.87 mg·L−1, respectively, higher than those of mono-culture, proving the potential of co-culture for the improvement of lipid production. This study provided a freshwater-saving dilution method for LL treatment with recycled harvesting water as well as a strategy for the increase of biomass and lipid accumulation by microalgae co-cultivation.

Lipid metabolism of the oleaginous yeast Lipomyces starkeyi.

The oleaginous yeast Lipomyces starkeyi is an excellent sustainable lipid producer, which can convert industrial wastes into lipids and accumulate triacylglycerols (TAG) by > 70% of its dry cell weight. Recent studies using omics technologies applied in L. starkeyi have aided in obtaining greater understanding of the important mechanisms of lipid metabolism in L. starkeyi. Therefore, the development of genetic engineering tools for L. starkeyi has led to accelerated efforts for a highly efficient production of lipids.This review focuses on the aspects of TAG and fatty acid synthesis pathways in L. starkeyi. We also present a quite effective strategy to obtain L. starkeyi mutants accumulating a larger amount of lipids and having a higher lipid production rate than the wild-type strain. The analysis of these mutants exhibiting high lipid production has led to the identification of important genes for achieving highly effective lipid production and thus advanced improvement in lipid production. Herein, our aim was to provide useful information to advance the development of L. starkeyi as a cost-effective TAG feedstock.Key Points•Oleaginous yeast Lipomyces starkeyi is an excellent sustainable lipid producer.•Efficient isolation of lipid-enriched L. starkeyi mutants depends on the low density of lipids.•Increased acyl-CoA synthesis pathway is important for improving lipid productivity.

A novel electroporation procedure for highly efficient transformation of Lipomyces starkeyi

Microbial lipids produced by oleaginous microorganisms as raw materials for the production of oleochemicals and biodiesel are sustainable while avoiding competition with food products. The oleaginous yeast Lipomyces starkeyi is an excellent lipid producer with a great industrial potential that is suitable as a valuable host to improve lipid production through genetic engineering modifications. However, genetic tools, including effective transformation methods, for L. starkeyi are insufficient for improvement of lipid production and analysis of lipid production mechanisms. We previously developed a polyethylene glycol (PEG)-mediated spheroplast transformation method that significantly improved the homologous recombination efficiency of L. starkeyi strain ∆lslig4. Although other transformation methods, including lithium acetate (LiAc)-mediated transformation and Agrobacterium tumefaciens-mediated transformation, have been reported, a more efficient and convenient transformation method for L. starkeyi is desired. In this study, we developed a novel electroporation transformation method that was first applied for integration of drug-resistance gene markers into the genome of L. starkeyi strain ∆lslig4 at the 18S ribosomal DNA locus of a multiple-copy gene, which yielded approximately 60 transformants/μg of DNA. Optimization of five parameters (i.e., cell growth phase, cell density, osmotic stabilizers, pretreatment agents, and electric conditions) enhanced the efficiency of transformation to approximately 1.5 × 104 transformants/μg of DNA. As compared with those of LiAc-mediated transformation and PEG-mediated spheroplast transformation, the efficiency of the proposed transformation method was increased by about 111- and 7-fold, respectively. Additionally, the transformation efficiency of our proposed electroporation method targeting a single-copy gene locus yielded 273 transformants/μg of DNA. To our knowledge, this is the first report of a successful electroporation method to accelerate analysis of lipid production by L. starkeyi.

Improvement in lipid production in Monoraphidium sp. QLY-1 by combining fulvic acid treatment and salinity stress

The effects of the combined treatment of fulvic acid (FA) and salinity stress on lipid production in Monoraphidium sp. QLY-1 at multiple levels was investigated in this study. The results indicated that the highest lipid content (59.53%) in QLY-1 was achieved by combining FA treatment and salinity stress. Compared with the control group and FA addition alone, the group treated with both FA and salinity stress had increased contents of reactive oxygen species (ROS), antioxidases, and nitric oxide (NO). Additionally, the addition of FA enhanced the expression levels of mitogen-activated protein kinases (MAPKs) and key genes related to lipid biosynthesis in QLY-1 under salinity stress. Collectively, biochemical analyses indicated that ROS, NO, MAPK, expression of lipid biosynthesis-related genes and antioxidant systems were involved in the lipid biosynthesis pathways of QLY-1 under the combined treatment of FA and salinity stress.

Improvement of lipid production in oleaginous yeast Rhodosporidium toruloides by ultraviolet mutagenesis.

The oleaginous yeast Rhodosporidium toruloides AS 2.1389 is viewed as desirable industrial microorganisms that can accumulate a high content of lipids for biodiesel production. In this study, we attempted to improve lipid accumulation in the yeast Rhodosporidium toruloides by UV irradiation mutagenesis and selection based on lithium chloride tolerance or ethanol-H2O2 tolerance. The biomass concentration, lipid yield and glucose consumption of mutant R. toruloides were determined. The transcription levels of lipid accumulation-related genes in the wild-type and mutant strains were also determined. The lithium chloride-tolerant strain R-ZL2 and the ethanol-H2O2-resistant strain R-ZY13 were generated by UV mutagenesis. The two mutant strains showed greater lipid productivity and lipid yield compared to the wild type. Transcriptional analysis revealed that IDP1, GPD1 and GND were expressed at significantly higher levels in the two high-lipid-producing mutants. In conclusion, lipid productivity and lipid yield in R. toruloides were successfully improved via UV mutagenesis and selection. We also identified some lipid accumulation-related genes for improving lipid productivity through genetic engineering.

Fungi (Mold)-Based Lipid Production.

There is an increasing need for the development of alternative energy sources with a focus on reducing greenhouse gas emissions and striving toward a sustainable economy. Bioethanol and biodiesel are currently the primary choices of alternative transportation fuels. At present, biodiesel is not competitive with conventional fuel due to its high price, and the only way to compete with conventional fuel is to improve the quality, reduce the costs, and coproduce value-added products. With the high demand for lipids in the energy sector and other industrial applications, microbial lipids accumulated from microorganisms, especially oleaginous fungi and yeasts have been the important topic of many recent research studies. This chapter summarizes the current status of knowledge and technology about lipid production by oleaginous fungi and yeasts for biofuel applications and other value-added products. The chapter focuses on several aspects such as the most promising oleaginous strains, strain development, improvement of lipid production, methods and protocols to cultivate oleaginous fungi, substrate utilization, fermentation process design, and downstream processing. The feasibility and challenges during the large-scale commercial production of microbial lipids as fuel sources are also discussed. It provides an overview of microbial lipid production biorefinery and also future development directions.

Concomitant production of chitosan and lipids from a newly isolated Mucor circinelloides ZSKP for biodiesel production

A newly-isolated oleaginous fungus Mucor circinelloides ZSKP concurrently yielded 21.4% lipids and 11.2% chitosan per gram of biomass. Parameters affecting the co-production were identified using Plackett-Burman design and were statistically optimized using Response Surface Methodology, which resulted in a 3-fold improvement in lipid production. The lipid profile showed a high content of unsaturated fatty acids including oleic (37%), linolenic (14%) and linoleic acids (19%), while palmitic acid was the major saturated fatty acid (21%). A comparative study to evaluate the efficacy of enzymatic (lipase) and chemical treatments for biodiesel production from fungal lipids and sunflower oil revealed enhanced production of biodiesel from fungal lipids. Synthesized biodiesel from M. circinelloides ZSKP satisfied standard specifications and had a higher cetane number (56), lower kinematic viscosity (4.6 mm2/s) and lower acid number (0.03) compared to sunflower oil. Results suggest Mucor circinelloides ZSKP is a promising candidate for implementation of the biorefinery concept.

Enhanced biological fixation of methane for microbial lipid production by recombinant Methylomicrobium buryatense

BackgroundDue to the success of shale gas development in the US, the production cost of natural gas has been reduced significantly, which in turn has made methane (CH4), the major component of natural gas, a potential alternative substrate for bioconversion processes compared with other high-price raw material sources or edible feedstocks. Therefore, exploring effective ways to use CH4 for the production of biofuels is attractive. Biological fixation of CH4 by methanotrophic bacteria capable of using CH4 as their sole carbon and energy source has obtained great attention for biofuel production from this resource.ResultsIn this study, a fast-growing and lipid-rich methanotroph, Methylomicrobium buryatense 5GB1 and its glycogen-knock-out mutant (AP18) were investigated for the production of lipids derived from intracellular membranes, which are key precursors for the production of green diesel. The effects of culture conditions on cell growth and lipid production were investigated in high cell density cultivation with continuous feeding of CH4 and O2. The highest dry cell weight observed was 21.4 g/L and the maximum lipid productivity observed was 45.4 mg/L/h obtained in batch cultures, which corresponds to a 2-fold enhancement in cell density and 3-fold improvement in lipid production, compared with previous reported data from cultures of 5GB1. A 90% enhancement of lipid content was achieved by limiting the biosynthesis of glycogen in strain AP18. Increased CH4/O2 uptake and CO2 evaluation rates were observed in AP18 cultures suggesting that more carbon substrate and energy are needed for AP18 growth while producing lipids. The lipid produced by M. buryatense was estimated to have a cetane number of 75, which is 50% higher than biofuel standards requested by US and EU.ConclusionsCell growth and lipid production were significantly influenced by culture conditions for both 5GB1 and AP18. Enhanced lipid production in terms of titer, productivity, and content was achieved under high cell density culture conditions by blocking glycogen accumulation as a carbon sink in the strain AP18. Differences observed in CH4/O2 gas uptake and CO2 evolution rates as well as cell growth and glycogen accumulation between 5GB1 and AP18 suggest changes in the metabolic network between these strains. This bioconversion process provides a promising opportunity to transform CH4 into biofuel molecules and encourages further investigation to elucidate the remarkable CH4 biofixation mechanism used by these bacteria.

Pelagibaca bermudensis promotes biofuel competence of Tetraselmis striata in a broad range of abiotic stressors: dynamics of quorum-sensing precursors and strategic improvement in lipid productivity

BackgroundAmelioration of biofuel feedstock of microalgae using sustainable means through synthetic ecology is a promising strategy. The co-cultivation model (Tetraselmis striata and Pelagibaca bermudensis) was evaluated for the robust biofuel production under varying stressors as well as with the selected two-stage cultivation modes. In addition, the role of metabolic exudates including the quorum-sensing precursors was assessed.ResultsThe co-cultivation model innovated in this study supported the biomass production of T. striata in a saline/marine medium at a broad range of pH, salinity, and temperature/light conditions, as well as nutrient limitation with a growth promotion of 1.2–3.6-fold. Hence, this developed model could contribute to abiotic stress mitigation of T. striata. The quorum-sensing precursor dynamics of the growth promoting bacteria P. bermudensis exhibited unique pattern under varying stressors as revealed through targeted metabolomics (using liquid chromatography–mass spectrometry, LC–MS). P. bermudensis and its metabolic exudates mutually promoted the growth of T. striata, which elevated the lipid productivity. Interestingly, hydroxy alkyl quinolones independently showed growth inhibition of T. striata on elevated concentration. Among two-stage cultivation modes (low pH, elevated salinity, and nitrate limitation), specifically, nitrate limitation induced a 1.5 times higher lipid content (30–31%) than control in both axenic and co-cultivated conditions.ConclusionPelagibaca bermudensis is established as a potential growth promoting native phycospheric bacteria for robust biomass generation of T. striata in varying environment, and two-stage cultivation using nitrate limitation strategically maximized the biofuel precursors for both axenic and co-cultivation conditions (T and T-PB, respectively). Optimum metabolic exudate of P. bermudensis which act as a growth substrate to T. striata surpasses the antagonistic effect of excessive hydroxy alkyl quinolones [HHQ, 4-hydroxy-2-alkylquinolines and PQS (pseudomonas quorum signal), 2-heptyl-3-hydroxy-4(1H)-quinolone].

Evaluation of lipid production from xylose and glucose/xylose mixed sugar in various oleaginous yeasts and improvement of lipid production by UV mutagenesis

Microbial lipids from oleaginous yeasts are recognized as promising alternative lipid sources. Although lipid productivity of oleaginous yeasts from glucose has been well-studied, knowledge of lipid productivity from xylose is limited. In this study, we aimed to evaluate and improve lipid productivity from xylose or glucose/xylose mixed sugar in various oleaginous yeasts. The xylose-based growth capability of 15 oleaginous strains belonging to Cryptococcus curvatus, Lipomyces starkeyi, Rhodosporidium toruloides, and Rhodotorula glutinis was tested. L. starkeyi strains showed relatively low growth capability. Ultraviolet mutagenesis combined with selection using cerulenin was applied for C. curvatus and R. toruloides strains. A mutant R. toruloides 8766 3-11C 2-53C exhibiting higher lipid productivity from xylose and glucose/xylose was obtained. The transcription levels of various lipid production-related genes were upregulated, whereas xylose metabolism-related genes were almost unchanged in the mutant. The results reveal the very diverse xylose-utilizing capability in the various oleaginous yeast species and strains. Promising target genes to improve lipid productivity from xylose through gene recombination were identified.

Improvement of lipid production from an oil-producing filamentous fungus, Penicillium brevicompactum NRC 829, through central composite statistical design

In the present study, 13 filamentous fungi were screened for their lipid production and an oleaginous fungus, Penicillium brevicompactum NRC 829, was found to be the highest lipid producer. Screening of various agro-industrial residues was performed and sunflower oil cake proved to be the best substrate for lipid production. A central composite design was employed to investigate the optimum concentrations of the most significant medium components required to improve the lipid production by P. brevicompactum. The results clearly revealed that the maximal lipid production of 8.014 ± 0.06 gL−1 (representing 57.6% lipid/dry biomass) was achieved by the fungus when grown for 6 days at 30 °C under static condition in a medium containing sunflower oil cake, NaNO3 and KCl at final concentrations of 8, 0.75 and 0.25 gL−1, respectively. Gas chromatography-mass spectrometry analysis of P. brevicompactum lipid indicated that linoleic acid (LA) (C18:2–6, 9) was the most abundant fatty acid, accounting for up to 62% of the total fatty acid profile, followed by palmitoleic acid (C16:1, 16%) and linolenic acid (C18:3, 8%). These results suggest that P. brevicompactum NRC 829 may have potential for commercial development for the production of LA by fermentation using cheap raw material.

Improvement of lipid production by the oleaginous yeast Rhodosporidium toruloides through UV mutagenesis.

Oleaginous yeasts are considered a promising alternative lipid source for biodiesel fuel production. In this study, we attempted to improve the lipid productivity of the oleaginous yeast Rhodosporidium toruloides through UV irradiation mutagenesis and selection based on ethanol and H2O2 tolerance or cerulenin, a fatty acid synthetase inhibitor. Glucose consumption, cell growth, and lipid production of mutants were evaluated. The transcription level of genes involved in lipid production was also evaluated in mutants. The ethanol and H2O2 tolerant strain 8766 2-31M and the cerulenin resistant strain 8766 3-11C were generated by UV mutagenesis. The 8766 2-31M mutant showed a higher lipid production rate, and the 8766 3-11C mutant produced a larger amount of lipid and had a higher lipid production rate than the wild type strain. Transcriptional analysis revealed that, similar to the wild type strain, the ACL1 and GND1 genes were expressed at significantly low levels, whereas IDP1 and ME1 were highly expressed. In conclusion, lipid productivity in the oleaginous yeast R. toruloides was successfully improved via UV mutagenesis and selection. The study also identified target genes for improving lipid productivity through gene recombination.

Exploration of sodium lignosulphonate's effects on lipid production by Rhodosporidium toruloides

Microbial oil is drawing increasing interest all over the world as an alternative non-food feedstock for biodiesel production. Rhodosporidium toruloides has been considered as a promising candidate to convert glycerol into microbial oils due to its good substrate adaptability, high intracellular lipid content, and the potential of co-production of some pigments. Interestingly, we have also found that sodium lignosulphonate (SL) could significantly enhance the cell growth and lipid accumulation of R. toruloides. In the present work, we investigated the possible mechanisms for the enhancement of lipid production by SL. It was found that SL could not be utilized by R. toruloides as a carbon source. No apparent changes on the morphology of cell membranes and the fatty acid profiles of microbial lipids were found when SL was added. Further experiments indicated that the volumetric oxygen transfer coefficient was increased by nearly 15% on average by adding 4g/L SL, which might be the main mechanism for the improvement of lipid production.

Application of magnesium sulfate and its nanoparticles for enhanced lipid production by mixotrophic cultivation of algae using biodiesel waste

CG (Crude glycerol) is one of the major wastes of biodiesel production process. It can be used as a substrate for lipid production by algae and the produced lipid can be recycled as a feedstock for biodiesel production. In order to avoid substrate inhibition, lipid production media are prepared by diluting the CG with distilled water. However, CG contains only a small amount of Mg (57.41 ± 18 ppm) and its concentration is further decreased to around 0.57 ppm during the dilution process. Apart from having a number of roles in algal physiology, Mg is the central atom of chlorophyll. Therefore, MgSO4 was evaluated as a Mg source to supplement the CG based media used for lipid production by Chlorella vulgaris. By supplementing the process with 1 g/L of MgSO4, nearly 185.29 ± 4.53% improvement in lipid production has been achieved. Further, application of MgSO4 nanoparticles was found to improve the lipid production by 118.23 ± 5.67%. Interestingly, unlike MgSO4, its nanoparticles were found to enhance the lipid production at the expense of only a small amount of glycerol. Thus, application of MgSO4 nanoparticles could be a potential strategy for enhanced lipid yield.