Toruloides Y4 Research Articles

Catalytic Hydrodeoxygenation of Methyl Stearate and Microbial Lipids to Diesel-Range Alkanes over Pd/HPA-SiO2 Catalysts

Catalytic hydrotreating of renewable oils is a promising way to get diesel alkanes, with the use of microbial lipids further facilitating scalable production of green diesel. The core issue of triglyceride hydrodeoxygenation (HDO) research is still to develop efficient catalysts under mild reaction conditions. Herein, we demonstrate that the heteropolyacid (HPA)-modified SiO2 supported Pd catalysts showed high activity for the HDO reaction of methyl stearate and soybean oil to diesel alkanes (C15∼C18, >90 wt % at 200 °C). Characterizations show that the Brønsted acid sites introduced by grafting heteropolyacid on SiO2 and metal sites of small Pd particles contribute to the high HDO activity of this bifunctional catalyst. Additionally, lipids produced by Rhodosporidium toruloides Y4 from glucose or corn stover were also transformed to about 75 wt % of diesel-range alkane products. This integration of microbial lipid production with chemical catalysis should facilitate the engineering of green diesel production from lignocellulosic biomass.

Bio-refining corn stover into microbial lipid and advanced energy material using ionic liquid-based organic electrolyte

Full utilization of the entire components of lignocellulosic biomass is one of the challenges in modern biorefinery via integrated approaches. This study pioneers an integrated approach to convert carbohydrates and lignin from corn stover into microbial lipids and advanced energy carbon nanofibers for supercapacitors using ionic liquid-based organic electrolyte ([Emim][OAc]/DMSO (XILs = 0.2, XILs is the molar fraction of ILs in the electrolyte). The pretreated corn stover was delignified by 60 wt%, and its saccharification was enhanced from 0.11 g.g−1 total reducing sugar (TRS) and 0.08 g.g−1 glucose to 0.68 g.g−1 TRS and 0.43 g.g−1 glucose yields within 12 h by enzymatic hydrolysis. The hydrolysates were used as carbon sources for cultivating R. toruloides Y4 for lipid production, yielding 13.8 g/L biomass and 51.0 % lipid. The lignin after enzymatic saccharification was integrated with extracted lignin to form a lignin organic electrolyte solution, which was directly prepared into electrospun nanofibers combined with polyacrylonitrile that served as electrode materials for supercapacitors with good performance.

Recycling biodiesel-derived glycerol by the oleaginous yeast Rhodosporidium toruloides Y4 through the two-stage lipid production process

Direct utilization of crude glycerol, a major byproduct in biodiesel industry, becomes imperative, because its production has outpaced the demand recently. We demonstrated that the oleaginous yeast Rhodosporidium toruloides Y4 had a great capacity to convert glycerol into lipids with high yield using the two-stage production process. Significantly higher cell mass and lipid yield were observed when the media were made with synthetic crude glycerol than pure glycerol. The process achieved a lipid yield of 0.22gg−1 glycerol, which was comparable with the lipid yield using glucose as the substrate. Lipid samples showed similar fatty acid compositional profiles to those of vegetable oils, suggesting that such microbial lipids were potential feedstock for biodiesel production. Our data provided an attractive route to integrate biodiesel production with microbial lipid technology for better resource efficiency and economical viability.

Microbial biodiesel production by direct methanolysis of oleaginous biomass

Biodiesel is usually produced by the transesterification of vegetable oils and animal fats with methanol, catalyzed by strong acids or bases. This study introduces a novel biodiesel production method that features direct base-catalyzed methanolysis of the cellular biomass of oleaginous yeast Rhodosporidium toruloides Y4. NaOH was used as catalyst for transesterification reactions and the variables affecting the esterification level including catalyst concentration, reaction temperature, reaction time, solvent loading (methanol) and moisture content were investigated using the oleaginous yeast biomass. The most suitable pretreatment condition was found to be 4gL(-1) NaOH and 1:20 (w/v) dried biomass to methanol ratio for 10h at 50°C and under ambient pressure. Under these conditions, the fatty acid methyl ester (FAME) yield was 97.7%. Therefore, the novel method of direct base-catalyzed methanolysis of R. toruloides is a much simpler, less tedious and time-consuming, process than the conventional processes with higher FAME (biodiesel) conversion yield.

The integrated production of microbial lipids and bio-SiO2 from rice husks by an organic electrolytes pretreatment technology

In this study, a full dissolution behavior of rice husks (RHs) in ionic liquids-based organic electrolytes was achieved, and physicochemical effect of the dissolution pretreatment on the structures of RHs was elucidated. The physicochemical changes led to an enhanced subsequent enzymatic saccharification of RHs, and a total reducing sugars (TRSs) yield of 0.70gg(-1), and a glucose yield of 0.43gg(-1) were obtained. The hydrolysates could be used as carbon sources for the cultivation of Rhodosporidium toruloides Y4 for the production of microbial lipids with a satisfactory productivity of cell biomass (13.3gL(-1)) and lipid content (32.5%) after 100h cultivation. Further pyrolysis of the residuals after the enzymatic hydrolysis at 600°C for 3h resulted in new uniform, spherical silica powder materials with particle size around 150nm, and surface area of 179.3m(2)g(-1).

Phase‐switching Homogeneous Catalysis for Clean Production of Biodiesel and Glycerol from Soybean and Microbial Lipids

AbstractWith the increasing concerns into the utilization of lipids as renewable resources for production of biofuels, chemicals, and materials, the efficient extraction and conversion of lipid is one of most important issues for the foundation of cost competitive lipid‐economy. In this study, we found that by using a range of “switchable solvents” from 1,8‐diazabicyclo[5.4.0]undec‐7‐ene to methanol, CO2, a recognized greenhouse gas, can be used as a trigger to catalyze the methanolysis of soybean and microbial lipids, and facilitate the separation of fatty acid methyl esters (FAMEs) and glycerol. It was found that the FAMEs can be decanted from the system in a 95.2 % conversion efficiency. The produced glycerol can be extracted from the FAMEs completely by the “switchable solvents”, and can be separated with high purity after recycling the DBU by an easy extraction process. Our primary investigation of the in situ extraction and conversion of microbial lipids from oleaginous yeast (Rhodosporidium toruloides Y4) shows that, although a 95.4 % extraction yield was achieved, the methanolysis efficiency was only 21.9 %. Our study presents an efficient and facile process for the integrated clean production of biodiesel and glycerol from soybean and microbial lipids, and the catalyst DBU can be recycled by an easy extraction process.

Enhancing the value of nitrogen from rapeseed meal for microbial oil production

Rapeseed meal, a major byproduct of biodiesel production, has been used as a low-cost raw material for the production of a generic microbial feedstock through a consolidated bioconversion process. Various strategies were tested for the production of a novel fermentation medium, rich in free amino nitrogen (FAN): commercial enzymes (CEs) (2.7mgg−1 dry meal), liquid state fungal pre-treatment (LSF) using Aspergillus oryzae (4.6mgg−1), liquid state fungal pre-treatment followed by fungal autolysis (LSFA) (9.13mgg−1), liquid state pre-treatment using fungal enzymatic broth (EB) (2.1mgg−1), but the best strategy was a solid state fungal pre-treatment followed by fungal autolysis (34.5mgg−1).The bioavailability of the nitrogen sources in the novel medium was confirmed in fed-batch bioreactor studies, in which 82.3g dry cell L−1 of the oleaginous yeast Rhodosporidium toruloides Y4 was obtained with a lipid content of 48%. The dry cell weight obtained was higher than that obtained using conventional yeast extract, due to a higher total nitrogen content in the novel biomedium. The fatty acids obtained from the microbial oil were similar to those derived from rapeseed oil.

Enzymatic hydrolysates of corn stover pretreated by a N-methylpyrrolidone–ionic liquid solution for microbial lipid production

In this study, we reported a novel N-methylpyrrolidone (NMP)–1-ethyl-3-methyl imidazolium acetate (EmimAc) mixed solvent (XILs = 0.2, mole fraction of ionic liquids (ILs) in the mixed solution) that can dissolve up to 10 wt% corn stover at 140 °C in 60 min. Physiochemical analysis showed major differences in terms of cellulose crystallinity, compositional distribution and surface morphology between raw corn stover samples and the regenerated materials, indicating that dissolution in the NMP–EmimAc system followed by regeneration with anti-solvents could be used as an effective pretreatment technology. Enzymatic saccharification of the regenerated corn stover afforded an 82.9% total reducing sugars yield and a 60.8% glucose yield within 24 h. The hydrolysates were used, without detoxification, as the carbon sources for the cultivation of Rhodosporidium Toruloides Y4 for lipid production. Glucose and xylose in the hydrolysates were both consumed in our process. In a word, our study reported a novel dissolution pretreatment technology for biomass utilization with outstanding advantages compared with traditional pure ionic liquid processes, such as (I) minimal use of expensive ILs in NMP, and commercially available EmimAc and NMP can be used directly without any further dry purification; (II) high cellulose and hemicellulose regeneration; (III) high enzymatic hydrolysis efficiency of the pretreated sample with full conversion of carbohydrates in less than 24 h; (IV) air-dried corn stover can be used directly (water content = ∼5%); (V) the hydrolysates can be used for further bioconversion and on inhibitory effect was observed. We believe that all of these advantages determine its potential for practical applications.

Energy efficiency evaluation of lipid production by oleaginous yeast Rhodosporidium toruloides

Mass–energy balance of lipid fermentation by oleaginous yeast Rhodosporidium toruloides Y4 with glucose as sole carbon source was investigated. The elemental composition of the freeze-dried cell samples obtained at different time points was determined using a Vario EL3 analyzer to estimate the biomass energetic yield (η). Then this work established the biomass energetic yield (η) to sets of biochemical variables and a new equation was developed to determine η without elemental composition of biomass. Biomass energetic yield estimated by the new equation was highly in accordance with that based on elemental composition. Bomb-calorimetric measurements were shown to be a direct method of quantifying the energy content of oleaginous yeasts. Combustion heat (Q c) of biomass determined experimentally was in consistent with those calculated according to its elementary contents. The relationship between lipid content and Q c of the cells was simulated and a new practical equation was developed based on lipid content (Y L/S) to evaluate Q c of biomass. Biomass energetic yield of R. toruloides Y4 could reach higher than 0.8. Combustion heat of biomass obtained after 116 h was 33 (kJ g−1) that was about 73% of the combustion heat of diesel. The results revealed that R. toruloides Y4 was an efficient “energy-converter” in lipid production with glucose as the substrate. The results also implied the approaches to estimating η of fermentation, and Q c of single oil cells developed in this study should be valuable to address the overall techno-economical analysis of bio-energy production.

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.

Lipid production by Rhodosporidium toruloides Y4 using different substrate feeding strategies

Microbial lipid is a potential alternative feedstock for the biodiesel industry. New culture strategies remain to be developed to improve the economics of microbial lipid technology. This work describes lipid production by the oleaginous yeast Rhodosporidium toruloides Y4 using a 15-l bioreactor with different substrate feeding strategies. Among these strategies, the intermittent feeding mode gave a lipid productivity of 0.36 g l(-1) h(-1), whereas the constant glucose concentration II (CC-II) mode gave the highest lipid productivity of 0.57 g l(-1) h(-1). The repeated fed-batch mode according to the CC-II mode was performed with a duration time of 358 h, and the overall lipid productivity was 0.55 g l(-1) h(-1). Our results suggested that substrate feeding modes had a great impact on lipid productivity and that the repeated fed-batch process was the most appealing method by which to enhance microbial lipid production.

Phosphate-limitation mediated lipid production by Rhodosporidium toruloides

Nitrogen-limited conditions have been routinely prepared for efficient lipid production by oleaginous microorganisms. However, it is difficult to attain high cellular lipid contents with natural nitrogen-rich substrates. In the present study, we showed that lipid accumulation by Rhodosporidium toruloides Y4 was directly linked to the carbon to phosphorus (C/P) molar ratios of the culture media. Moreover, such lipid accumulation phenomena were effective regardless of the presence of high amounts of nitrogen sources. Thus, cellular lipid content and lipid yield were 62.2% and 0.205g/g glucose, respectively, using a medium with a carbon to nitrogen (C/N) molar ratio of 6.1 and a C/P molar ratio of 9552. This work suggested that phosphorus limitation can be equally effective and efficient to mediate lipid accumulation, which in turn, provides opportunities to produce microbial lipid more economically using natural or waste materials with high nitrogen content.

Lipid production from Jerusalem artichoke by Rhodosporidium toruloides Y4

Jerusalem artichoke (JA) is a perennial herbaceous plant widely available as non-grain raw material. Microbial lipid has been suggested as a potential feedstock for large scale biodiesel production. This paper describes lipid production using JA tuber processed by oleaginous yeast Rhodosporidium toruloides Y4. Batch and fed-batch modes were tested with feeding of concentrated JA extracts or JA hydrolysates. Cultivation of R. toruloides Y4 with JA extracts gave a moderate cellular lipid content of 40% (w/w), whereas lipid titer and cellular lipid content reached 39.6 g l(-1) and 56.5% (w/w), respectively, when JA hydrolysates were fed. Our results suggested that JA tubers may be further explored as raw material for large scale microbial lipid production.

Effects of biomass hydrolysis by-products on oleaginous yeast Rhodosporidium toruloides

Lignocellulosic biomass hydrolysis inevitably coproduces byproducts that may have various affects on downstream biotransformation. It is imperative to document the inhibitor tolerance ability of microbial strain in order to utilize biomass hydrolysate more effectively. To achieve better lipid production by Rhodosporidium toruloides Y4, we performed fermentation experiments in the presence of some representative inhibitors. We found that acetate, 5-hydroxymethylfurfural and syringaldehyde had slightly inhibitory effects; p-hydroxybenzaldehyde and vanillin were toxic at a concentration over 10 mM; and furfural and its derivatives furfuryl alcohol and furoic acid inhibited cell growth by 45% at around 1 mM. We further demonstrated that inhibition is generally additive, although strong synergistic inhibitions were also observed. Finally, lipid production afforded good results in the presence of six inhibitors at their respective concentrations usually found in biomass hydrolysates. Fatty acid compositional profile of lipid samples indicated that those inhibitors had little effects on lipid biosynthesis. Our work will be useful for optimization of biomass hydrolysis processes and lipid production using lignocellulosic materials.

High-density cultivation of oleaginous yeast Rhodosporidium toruloides Y4 in fed-batch culture

Microbial lipid production by the oleaginous yeast Rhorosporidium toruloides Y4 was studied using glucose as carbon source, in order to realize high-density cell culture. Batch cultures demonstrated that there was little inhibitory effect with a substrate concentration of up to 150 g l −1. Flask fed-batch cultures were run for 25 days and reached a dry biomass and cellular lipid content of 151.5 g l −1 and 48.0% (w/w), respectively. Using pilot-scale fed-batch cultures in a 15-l stirred-tank fermenter for 134 h resulted in dry biomass, lipid content and lipid productivity of 106.5 g l −1, 67.5% (w/w) and 0.54 g l −1 h −1, respectively. The fed-batch culture model used here featured initial nutrient-rich media and a pure carbon source with discontinuous feeding. Gas chromatography analysis revealed that lipids from R. toruloides Y4 contained mainly long-chain fatty acids with 16 and 18 carbon atoms. The four major constituent fatty acids were oleic acid, palmitic acid, stearic acid and linoleic acid. A slight increase in stearic acid production was found during the culture process. Based on these compositional data, microbial lipids from R. toruloides Y4 are a potential alternative oil resource for biodiesel production.

Optimization of Culture Conditions for Lipid Production by Rhodosporidium toruloides

The effects of culture conditions on the lipid production by Rhodosporidium toruloides Y4 were investigated using uniform design principles and single-factor experiments. The optimal medium was obtained as follows: 70 g/L glucose, 0.1 g/L (NH 4) 2SO 4, 0.75 g/L yeast powder, 1.5 g/L MgSO 4·7H 2O, 0.4 g/L KH 2PO 4, sterilized at 121 °C for 15 min, and then supplemented with 1.91×10 −6 mmol/L ZnSO 4, 1.50 mmol/L CaCl 2, 1.22×10 −4 mmol/L MnCl 2, and 1.00×10 −4 mmol/L CuSO 4. The optimal fermentation conditions were also developed as: 50 mL of medium (pH 6.0) in a 250-mL Erlenmeyer flask with 10% inoculum (28-h-old) under orbital shaking at 200 r/min for 120 h at 30 °C. Under the optimized conditions, yeast accumulated lipids up to 76% on a cellular biomass basis with biomass yield of 18.2 g/L.