Feedstock For Production Of Lipids Research Articles

Assessing oleaginous yeasts for their potentials on microbial lipid production from sugarcane bagasse and the effects of physical changes on lipid production

We assess oleaginous yeast strains from five genera to identify the strain suitable to use sugarcane bagasse as a feedstock for lipid production. Upon initial assessment in glucose or xylose media, five potential strains were chosen for further assessment in sugarcane bagasse hydrolysate. All strains exhibited improved growth and lipid production due to a buffering capacity of the hydrolysate, with R. glutinis showing the best lipid production. The yeast growth and lipid production were similar over the pH range between 5.5 and 7.5. It performed best at 30 °C. Using baffled flask at 200 rpm resulted in the highest cell dry weight of 40 g/L and 20 g/L of lipid, while using 100 rpm or normal flasks resulted in a much lower growth and lipid production. The results from this study suggest that sugarcane bagasse could be a raw material for microbial lipid production, providing the control criteria are suitable.

Harnessing pongamia shell hydrolysate for triacylglycerol agglomeration by novel oleaginous yeast Rhodotorula pacifica INDKK

BackgroundTo meet the present transportation demands and solve food versus fuel issue, microbial lipid-derived biofuels are gaining attention worldwide. This study is focussed on high-throughput screening of oleaginous yeast by microwave-aided Nile red spectrofluorimetry and exploring pongamia shell hydrolysate (PSH) as a feedstock for lipid production using novel oleaginous yeast Rhodotorula pacifica INDKK.ResultsA new oleaginous yeast R. pacifica INDKK was identified and selected for microbial lipid production. R. pacifica INDKK produced maximum 12.8 ± 0.66 g/L of dry cell weight and 6.78 ± 0.4 g/L of lipid titre after 120 h of growth, showed high tolerance to pre-treatment-derived inhibitors such as 5-hydroxymethyl furfural (5-HMF), (2 g/L), furfural (0.5 g/L) and acetic acid (0.5 g/L), and ability to assimilate C3, C5 and C6 sugars. Interestingly, R. pacifica INDKK showed higher lipid accumulation when grown in alkali-treated saccharified PSH (AS-PSH) (0.058 ± 0.006 g/L/h) as compared to acid-treated detoxified PSH (AD-PSH) (0.037 ± 0.006 g/L/h) and YNB medium (0.055 ± 0.003 g/L/h). The major fatty acid constituents are oleic, palmitic, linoleic and linolenic acids with an estimated cetane number (CN) of about 56.7, indicating the good quality of fuel.ConclusionThese results suggested that PSH and R. pacifica INDKK could be considered as potential feedstock for sustainable biodiesel production.

Production of microbial lipids utilizing volatile fatty acids derived from wastepaper: A biorefinery approach for biodiesel production

Volatile fatty acids (VFAs) derived from organic wastes are being considered as low-cost feedstock for microbial lipid production as a valuable alternative to plant derived oils/biodiesel. In this study, VFAs were produced from anaerobic open culture fermentation of wastepaper and subsequently, used as a feedstock for lipid production by Cryptococcus curvatus. Total VFAs, yield and productivity achieved from waste office paper (WOP) and waste newspaper (WNP) were 17.3 and 10.2 g/L, 0.17 and 0.10 g/g TS, and 0.86 and 0.51 g/L/day, respectively. Biomass, lipid content and productivity achieved utilizing VFAs derived from WOP and WNP were 4.3 and 2.9 g/L, 41.2 and 27.7% DCW, and 0.037 and 0.033 g/L/h, respectively. The dominance of fatty acids such as oleic, palmitic, stearic and linoleic acid in the lipids suggests a high level of similarity with plant/vegetable oils used for biodiesel production. Therefore, VFAs derived from wastepaper could be potentially used as feedstock to produce microbial lipids towards cost-effective production of biodiesel.

Production of biodiesel feedstock - microbial lipid from slaughterhouse wastewater

Biodiesel is an alternative diesel fuel, which can be synthesised from renewable biological sources. Lipid production using carbon source in wastewater is an emerging process as it purely depends on waste source. In the present study, the ability of Yarrowia lipolytica to accumulate lipids using slaughterhouse wastewater as substrate was investigated. Using raw wastewater as substrate, maximum lipid content (0.43 g/L) and biomass (1.2 g/g) were obtained. Various pre-treatment methods like acid, alkaline, heat, activated carbon and sawdust treatment were performed and two-fold increase in C/N ratio was observed in combined pre-treatment of sawdust with KOH. Using pretreated wastewater, lipid accumulation was enhanced to 32% with lipid content of 0.64 g/L. Results of this study conclude that the pre-treated slaughterhouse wastewater can be employed as a better feedstock for lipid production using Yarrowia lipolytica.

PRODUCTION OF BIODIESEL FEEDSTOCK - MICROBIAL LIPID FROM SLAUGHTERHOUSE WASTEWATER

Biodiesel is an alternative diesel fuel, which can be synthesised from renewable biological sources. Lipid production using carbon source in wastewater is an emerging process as it purely depends on waste source. In the present study, the ability of Yarrowia lipolytica to accumulate lipids using slaughterhouse wastewater as substrate was investigated. Using raw wastewater as substrate, maximum lipid content (0.43 g/L) and biomass (1.2 g/g) were obtained. Various pre-treatment methods like acid, alkaline, heat, activated carbon and sawdust treatment were performed and two-fold increase in C/N ratio was observed in combined pre-treatment of sawdust with KOH. Using pretreated wastewater, lipid accumulation was enhanced to 32% with lipid content of 0.64 g/L. Results of this study conclude that the pre-treated slaughterhouse wastewater can be employed as a better feedstock for lipid production using Yarrowia lipolytica.

Sustainable Production of Microbial Lipids from Lignocellulosic Biomass Using Oleaginous Yeast Cultures

Microbial lipids derived from oleaginous yeast could be a promising resource for biodiesel and other oleochemical materials. The objective of this study was to develop an efficient bioconversion process from lignocellulosic biomass to microbial lipids using three types of robust oleaginous yeast: T. oleaginosus, L. starkeyi, and C. albidus. Sorghum stalks and switchgrass were utilized as feed-stocks for lipid production. Among oleaginous yeast strains, T. oleaginous showed better performance for lipid production using sorghum stalk hydrolysates. Lipid titers of 13.1 g·L-1 were achieved by T. oleaginosus, using sorghum stalk hydrolysates with lipid content of 60% (wt·wt-1) and high lipid yield of 0.29 g·g-1, which was substantially higher than the value reported in literature. Assessment of overall lipid yield revealed a total of 14.3 g and 13.3 g lipids were produced by T. oleaginosus from 100 g of raw sorghum stalks and switchgrass, respectively. This study revealed that minimization of sugar loss during pretreatment and selection of appropriate yeast strains would be key factors to develop an efficient bioconversion process and improve the industrial feasibility in a lignocellulose-based biorefinery.

Mechanism of Nutrient Deprivation Induced Triacylglyceride Accumulation in Alga Indicated by Fluorescence Hyperspectral Imaging

Microalgae have been identified as a promising renewable feedstock for production of lipids for feeds and fuels. For a wide variety of algal species, depletion of major nutrients from the growth medium results in dramatically increased biosynthesis of triacylglycerides, in some cases exceeding 70% of the dry weight of the biomass. In recent work, we explored this process in a number of promising algal production strains (Chlorophyceae and Eustigmophyceae) using hyperspectral confocal fluorescence imaging with multivariate spectral unmixing. By coupling intensity modulation of the excitation laser and attenuation of the dominant chlorophyll fluorescence emission, we identified a distinct spectral component composed of chlorophyll fluorescence and resonance Raman emission for lipid-solubilized carotenoids allowing label-free projections of lipid bodies in addition to plastids. Furthermore, cells exposed to the lipophilic dye, Nile Red, exhibited distinct spectral components corresponding to the polar and neutral lipid fractions. Combining these capabilities for analysis of nutrient deprivation time courses revealed increases in the lipid fraction of the cellular volume of ∼500%, as well as a correlated decrease in the plastid fraction of the total cellular volume. Additionally, diffraction limited tubules connecting the lipids bodies to the plastids and the presence of unbound chlorophyll in the lipid bodies provided evidence for direct shuttling of lipids from the chloroplast to the lipid bodies, in opposition to the expected ER to lipid body transfer dictated by the Kennedy pathway. The results suggest that nutrient deprivation-based lipid accumulation in these species is dominated by plastid membrane recycling, independent of acyl-CoA. Finally, an additional spectral component exhibiting high spectral overlap with Nile Red and varying inversely with the chlorophyll emission was detected during senescence, potentially resulting in false positives for lipid enrichment in low spectral resolution assays.

Label-free measurement of algal triacylglyceride production using fluorescence hyperspectral imaging

Microalgae have been identified as a promising renewable feedstock for production of lipids for feeds and fuels. Current methods for identifying algae strains and growth conditions that support high lipid production require a variety of fluorescent chemical indicators, such as Nile Red and more recently, Bodipy. Despite notable successes using these approaches, chemical indicators exhibit several drawbacks, including non-uniform staining, low lipid specificity, cellular toxicity, and variable permeability based on cell-type, limiting their applicability for high-throughput bioprospecting. In this work, we used in vivo hyperspectral confocal fluorescence microscopy of a variety of potential microalgae production strains (Nannochloropsis sp., Dunaliella salina, Neochloris oleoabundans, and Chlamydomonas reinhardtii) to identify a label-free method for localizing lipid bodies and quantifying the lipid yield on a single-cell basis. By analyzing endogenous fluorescence from chlorophyll and resonance Raman emission from lipid-solubilized carotenoids we deconvolved pure component emission spectra and generated diffraction limited projections of the lipid bodies and chloroplast organelles, respectively. Applying this imaging method to nutrient depletion time-courses from lab-scale and outdoor cultivation systems revealed an additional autofluorescence spectral component that became more prominent over time, and varied inversely with the chlorophyll intensity, indicative of physiological compromise of the algal cell. This signal could result in false-positives for conventional measurements of lipid accumulation (via spectral overlap with Nile Red), however, the additional spectral feature was found to be useful for classification of lipid enrichment and culture crash conditions in the outdoor cultivation system. Under nutrient deprivation, increases in the lipid fraction of the cellular volume of ~500% were observed, as well as a correlated decrease in the chloroplast fraction of the total cellular volume. The results suggest that a membrane recycling mechanism dominates for nutrient deprivation-based lipid accumulation in the microalgae tested.

Nannochloropsis plastid and mitochondrial phylogenomes reveal organelle diversification mechanism and intragenus phylotyping strategy in microalgae

BackgroundMicroalgae are promising feedstock for production of lipids, sugars, bioactive compounds and in particular biofuels, yet development of sensitive and reliable phylotyping strategies for microalgae has been hindered by the paucity of phylogenetically closely-related finished genomes.ResultsUsing the oleaginous eustigmatophyte Nannochloropsis as a model, we assessed current intragenus phylotyping strategies by producing the complete plastid (pt) and mitochondrial (mt) genomes of seven strains from six Nannochloropsis species. Genes on the pt and mt genomes have been highly conserved in content, size and order, strongly negatively selected and evolving at a rate 33% and 66% of nuclear genomes respectively. Pt genome diversification was driven by asymmetric evolution of two inverted repeats (IRa and IRb): psbV and clpC in IRb are highly conserved whereas their counterparts in IRa exhibit three lineage-associated types of structural polymorphism via duplication or disruption of whole or partial genes. In the mt genomes, however, a single evolution hotspot varies in copy-number of a 3.5 Kb-long, cox1-harboring repeat. The organelle markers (e.g., cox1, cox2, psbA, rbcL and rrn16_mt) and nuclear markers (e.g., ITS2 and 18S) that are widely used for phylogenetic analysis obtained a divergent phylogeny for the seven strains, largely due to low SNP density. A new strategy for intragenus phylotyping of microalgae was thus proposed that includes (i) twelve sequence markers that are of higher sensitivity than ITS2 for interspecies phylogenetic analysis, (ii) multi-locus sequence typing based on rps11_mt-nad4, rps3_mt and cox2-rrn16_mt for intraspecies phylogenetic reconstruction and (iii) several SSR loci for identification of strains within a given species.ConclusionThis first comprehensive dataset of organelle genomes for a microalgal genus enabled exhaustive assessment and searches of all candidate phylogenetic markers on the organelle genomes. A new strategy for intragenus phylotyping of microalgae was proposed which might be generally applicable to other microalgal genera and should serve as a valuable tool in the expanding algal biotechnology industry.

Microbial lipid production by Rhodosporidium toruloides under sulfate-limited conditions

Novel biochemical approaches remain to be developed to improve microbial lipid technology. This study demonstrated that sulfate limitation was effective to promote accumulating substantial amounts of intracellular lipid by the oleaginous yeast Rhodosporidium toruloides Y4. When it was cultivated using a medium with an initial carbon-to-sulfur (C/S) molar ratio of 46,750, cellular lipid content reached up to 58.3%. The time courses of cell growth, lipid accumulation and nutrient depletion were analyzed and discussed in terms of lipid biosynthesis. Moreover, lipid accumulation under sulfate-limited conditions was effective regardless of the presence of a high concentration of nitrogen sources. Thus, lipid contents almost held constant at near 57% in the media with an initial C/S molar ratio of 11,380 although the carbon-to-nitrogen molar ratio ranged from 28.3 to 5.7. Taken together, our results established the sulfate-limitation approach to control lipid biosynthesis, which should be valuable to explore nitrogen-rich raw materials as the feedstock for lipid production.