Trichosporon Oleaginosus Research Articles

Lipid production from fed-batch fermentation of crude glycerol directed by the kinetic study of batch fermentations

Oleaginous yeast Trichosporon oleaginosus was cultivated in crude glycerol as substrate for lipid production in a 15L fermenter. The impact of weight carbon to nitrogen ratio (C/N) including 20, 30, 45 and 60 on lipid production from the strain was investigated in batch fermentations. The highest biomass and lipid concentration of 24.80g/L and 12.14g/L, respectively was observed during the fermentation at C/N ratio of 45. C/N ratio of 30 showed a comparable concentration of biomass (23.70g/L) and lipid (11.26g/L). Based on the kinetic study of the batch fermentations, fed-batch fermentation was designed and conducted. In the fed-batch fermentation, the maximum concentration of biomass and lipid was 43.82g/L and 21.87g/L, respectively at C/N ratio of 45.

Corn bran bioprocessing: Development of an integrated process for microbial lipids production

This study investigated the potential of corn bran as a feedstock for microbial lipid production using oleaginous yeast, Trichosporon oleaginosus ATCC20509. Different conditions (solid loading of biomass, acid loading, and pretreatment duration) were applied to optimize pretreatment processes using the Box-Behnken design. The highest sugar yield of 0.53g/g was obtained from corn bran hydrolysates at a pretreatment condition of 5% solid loading and 1% acid loading for 30min. Compared with synthetic media, up to 50% higher lipid accumulations in T. oleaginosus were achieved using corn bran hydrolysates during fermentation. Also, the direct effect of pretreatment condition on the lipid accumulation of T. oleaginosus was investigated using response surface methodology (RSM). Solid loading of biomass during the pretreatment process significantly affected the fermentation process for lipid accumulation of T. oleaginosus. The RSM model can provide useful information to design an integrated bioconversion platform.

Lipid Production for Biodiesel from Sludge and Crude Glycerol.

Oleaginous yeast Trichosporon oleaginosus was studied for lipid production using municipal sludge with or without fortification of crude glycerol in a 15-L fermenter. The maximum lipid content (concentration) was 32.0% w/w (9.35 g/L), 33.6% (10.13 g/L), 33.3% (9.13 g/L), and 33.1% (9.03 g/L) w/w with the addition of 25, 50, 100, and 150 g/L glycerol, respectively. Glycerol concentration had little effect on lipid accumulation. However, glycerol concentration substantially affected increase of biomass concentration and cell count. The suitable glycerol concentration was approximately 40 g/L for Trichosporon oleaginosus growing in sludge medium with initial suspended solids (SS) concentration 30 g/L. Addition of nitrogen to sludge-glycerol medium enhanced lipid and biomass concentration. The energy conversion efficiency was 1.78, 1.55, and 1.71 with no nitrogen added, with addition of 1 g/L urea, and 3.7 g/L peptone, respectively. The biodiesel production cost was estimated nearly 0.75 US$/L.

Characterization of lipid accumulation and lipidome analysis in the oleaginous yeasts Rhodosporidium azoricum and Trichosporon oleaginosus

The influence of cultural conditions on lipid production was investigated in two species, Trichosporon oleaginosus and Rhodosporidium azoricum. We showed that nitrogen limitation is not the main factor triggering the mechanism of lipid accumulation in T. oleaginosus. Moreover, a scarce availability of oxygen negatively affected lipid synthesis to a lesser extent in T. oleaginosus than in R. azoricum. This highlights how the importance of controlling fermentation parameters is strictly linked to the yeast species employed. We showed that these parameters affect the activity of important enzymes, influencing the metabolic fluxes into different pathways, in particular pentose phosphate pathway and cytoplasmic pyruvate bypass. Furthermore, T. oleaginosus exhibited wider substrate flexibility, faster growth and higher lipid accumulation in fed-batch cultivation. Microbial oils obtained from both yeasts proved a valuable feedstock, alternative to vegetable oils, for advanced diesel biofuel production.

The effects of TORC signal interference on lipogenesis in the oleaginous yeast Trichosporon oleaginosus

BackgroundOleaginous organisms are a promising, renewable source of single cell oil. Lipid accumulation is mainly induced by limitation of nutrients such as nitrogen, phosphorus or sulfur. The oleaginous yeast Trichosporon oleaginosus accumulates up to 70% w/w lipid under nitrogen stress, while cultivation in non-limiting media only yields 9% w/w lipid. Uncoupling growth from lipid accumulation is key for the industrial process applicability of oleaginous yeasts. This study evaluates the effects of rapamycin on TOR specific signaling pathways associated with lipogenesis in Trichosporon oleaginosus for the first time.ResultsSupplementation of rapamycin to nutrient rich cultivation medium led to an increase in lipid yield of up to 38% g/L. This effect plateaued at 40 μM rapamycin. Interestingly, the fatty acid spectrum resembled that observed with cultivation under nitrogen limitation. Significant changes in growth characteristics included a 19% increase in maximum cell density and a 12% higher maximum growth rate. T. oleaginosus only has one Tor gene much like the oleaginous yeast Rhodosporidium toruloides. Consequently, we analyzed the effect of rapamycin on T. oleaginosus specific TORC signaling using bioinformatic methodologies.ConclusionsWe confirm, that target of rapamycin complex 1 (TORC1) is involved in control of lipid production and cell proliferation in T. oleaginosus and present a homology based signaling network. Signaling of lipid induction by TORC1 and response to carbon depletion to this complex appear to be conserved, whereas response to nitrogen limitation and autophagy are not. This work serves as a basis for further investigation regarding the control and induction of lipid accumulation in oil yeasts.

Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions

Cocoa butter (CB) extracted from cocoa beans is the main raw material for chocolate production. However, growing chocolate demands and limited CB production has resulted in a shortage of CB supply. CB is mainly composed of three different kinds of triacylglycerols (TAGs), POP (C16:0–C18:1–C16:0), POS (C16:0–C18:1–C18:0), and SOS (C18:0–C18:1–C18:0). The storage lipids of yeasts, mainly TAGs, also contain relative high-level of C16 and C18 fatty acids and might be used as CB-like lipids (CBL). In this study, we cultivated six different yeasts, including one non-oleaginous yeast strain, Saccharomyces cerevisiae CEN.PK113-7D, and five oleaginous yeast strains, Trichosporon oleaginosus DSM11815, Rhodotorula graminis DSM 27356, Lipomyces starkeyi DSM 70296, Rhodosporidium toruloides DSM 70398, and Yarrowia lipolytica CBS 6124, in nitrogen-limited medium and compared their CBL production ability. Under the same growth conditions, we found that TAGs were the main lipids in all six yeasts and that T. oleaginosus can produce more TAGs than the other five yeasts. Less than 3% of the total TAGs were identified as potential SOS in the six yeasts. However, T. oleaginosus produced 27.8% potential POP and POS at levels of 378 mg TAGs/g dry cell weight, hinting that this yeast may have potential as a CBL production host after further metabolic engineering in future.

New kids on the block: emerging oleaginous yeast of biotechnological importance.

There is growing interest in using oleaginous yeast for the production of a variety of fatty acids and fatty acid-derived oleochemicals. This is motivated by natural propensity for high flux through lipid biosynthesis that has naturally evolved, making them a logical starting point for additional genetic engineering to improve titers and productivities. Much of the academic and industrial focus has centered on yeast that have significant genetic engineering tool capabilities, such as Yarrowia lipolytica, and those that have naturally high lipid accumulation, such as Rhodosporidium toruloides and Lipomyces starkeyi; however, there are oleaginous yeast with phenotypes better aligned with typically inhibitory process conditions, such as high salt concentrations and lignocellulosic derived inhibitors. This review addresses the foundational work in characterizing two emerging oleaginous yeast of interest: Debaryomyces hansenii and Trichosporon oleaginosus. We focus on the physiological and metabolic properties of these yeast that make each attractive for bioprocessing of lignocellulose to fuels and chemicals, discuss their respective genetic engineering tools and highlight the critical barriers facing the broader implementation of these oleaginous yeast.

Effect of carbon and nitrogen source concentrations on the growth and lipid accumulation of yeastTrichosporon oleaginosusin continuous and batch culture

BACKGROUND Biodiesel is an alternative to conventional fossil diesel fuel. Currently, it is mainly produced from vegetable oils and consequently it has an adverse impact on food price. Lipids from oleaginous microorganisms (e.g. microalgae, bacteria, fungi and yeasts) could be an alternative feedstock for biodiesel production and therefore the growth and lipid accumulation of Trichosporon oleaginosus was studied under different conditions. RESULTS The growth of T. oleaginosus on the xylose containing media enhanced the lipid accumulation and reduced cell growth, while on the glucose-containing media the opposite effect was observed. Trichosporon oleaginosus can accumulate above 50% of total lipids in dry cell mass during continuous and batch culture in the nitrogen-limited conditions. In the continuous culture relatively high productivity of total lipid accumulation (0.67 g dm−3 h−1) was observed on the glucose containing media. The type of substrate limitation has considerable impact on the fatty acid composition of yeast cell lipids. The predicted biodiesel properties (based on the fatty acids composition of lipids) were found to meet the CEN quality standards. CONCLUSION The results obtained show that T. oleaginosus has great potential for feasible biodiesel production, especially when sugars from industrial and agricultural waste are used as a substrate. © 2016 Society of Chemical Industry

Lipid production with Trichosporon oleaginosus in a membrane bioreactor using microalgae hydrolysate

Lipid production by Trichosporon oleaginosus was first studied in fed-batch operated stirred-tank bioreactors on a milliliter- and liter-scale making use of typical sugar monomers and a sugar mixture that may be derived from microalgae biomass hydrolysis after the extraction of lipids. 20.3gL−1 lipids (58% of dry cell mass) were produced with T. oleaginosus in a defined medium at nitrogen starvation in the fed-batch process with a typical microalgae derived carbohydrate mixture (60% glucose, 20% mannose, 20% galactose). Real microalgae hydrolysate resulted in superior growth of T. oleaginosus but no enhanced lipid formation was possible due to nitrogen and phosphorus excess in the hydrolysate. Phosphate precipitation and the application of a continuously operated membrane bioreactor with total cell retention due to the low sugar concentrations (∼40gL−1) in the microalgae hydrolysate resulted in the production of 30gL−1 lipids (53% of dry cell mass) with T. oleaginosus at high space-time-yields of 0.33g lipids L−1h−1. A high apparent lipid yield of 0.43g lipids g−1 sugars consumed (130% of the theoretical maximum) was achieved with the microalgae hydrolysate most likely due to the additional conversion of other carbon sources (e.g. uronic acids, peptides) in the hydrolysate.

Two consecutive step process for ethanol and microbial oil production from sweet sorghum juice

The juice extracted from sweet sorghum stalks has been previously explored to produce ethanol and also to grow oleaginous yeasts and algae with the objective of producing microbial oil. In this paper we propose a different process route in order to produce ethanol and microbial oil in two consecutive fermentation steps. Ethanol is produced first followed by the growth of oleaginous yeast employing the residual carbon and nitrogen left from the first step. Two yeasts were compared for ethanol production. Trichosporon oleaginosus was cultivated for lipid production. The yeast selection for the first step was the most important factor for achieving a high ethanol yield and the effect of inorganic nitrogen addition was not significant. The remaining sugars consisted of a mixture of sucrose and fructose and no residual glucose was detected in any of the runs. In the second step T. oleaginosus DSM 11815 grew in the pooled juices remaining from the first step for 168h and produced biomass with 28% lipid content. Glucose showed the highest uptake rate, sucrose was utilized until low glucose values prevailed, and fructose was slowly metabolized and a substantial amount remained. Although the two step process has flexibility in choosing the proper microorganism for each step, it is necessary to look for a rapid fructose uptake strain for both fermentations.

Genetic engineering and production of modified fatty acids by the non-conventional oleaginous yeast Trichosporon oleaginosus ATCC 20509

Establishing genetic engineering tools for sustainable production of tailor made fatty acids in the non-conventional, oleaginous yeastTrichosporon oleaginosus.

Ultrasonication aided biodiesel production from one-step and two-step transesterification of sludge derived lipid

Trichosporon oleaginosus was grown in wastewater sludge to produce lipid. The obtained biomass was employed to study ultrasonication effect on lipid extraction and in-situ transesterification. Besides the most utilized solvents (the mixture of chloroform and methanol, 1:1 v/v), other solvents including water, hexane, and methanol were also studied to explore their capacities in lipid extraction with ultrasonication. The maximum lipid recovery was 11.8, 35.3, 62.0, and 95.3% (w/w) with water, hexane, methanol, and the mixture of chloroform and methanol (1:1 v/v), respectively within 20 min. Chloroform and methanol gave the best performance. However, due to the adverse environmental concern related to the utilization of chloroform, in-situ transesterification (single step lipid extraction and transesterification to obtain biodiesel) was investigated.In case of in-situ transesterification, biodiesel yield of 95% (w/w) was obtained in 60 min with the assistance of ultrasonication; whereas, 24 h was required to achieve similar yield without ultrasonication.

Strategies of preserving lipids in microorganism after fermentation

Microbial oil is accumulated by microorganisms as stored energy during cultivation, and will be degraded to generate energy when they are not able to obtain external energy source. As the lipid is the desired production, the in situ degradation of oil by microorganisms after fermentation or during downstream processing has to be prevented. This study investigated the effect of pH, thermal, salinity, bead milling, ultrasonication, and microwave treatment on the viability of Trichosporon oleaginosus when it was used for lipid production. The cells in broth were completely inactivated with the treatment of pH 1 and 2 for 1h, temperature 80°C for 10min, ultrasonication for 8min, and microwave for 6min, respectively. It was observed that these treatments had no impact on final product (biodiesel) composition and were considered as safe and efficient methods to preserve lipid in cells.

Wastewater sludge as raw material for microbial oils production

Biodiesel is a promising alternative energy source as it is renewable, biodegradable, and environmental friendly. This study deals with oils production from wastewater sludge. Primary, secondary, and mixed municipal sludge and pulp and paper secondary sludge collected in Québec City, Canada has 5–11% oil content (based on weight) in initial sludge. The oil content is in the order of pulp and paper secondary sludge>municipal primary sludge>municipal mixed sludge>municipal secondary sludge. Different types of sludge were also used for lipid production by Pichia amethionina sp., Galactomyces sp., and Trichosporon oleaginosus. The results showed that maximum lipid content was obtained in municipal secondary sludge among all types. Further, the effect of initial sludge suspended solids concentration (10–30g/L) on lipid accumulation was investigated. The maximum lipid content of 30.17% and 32.42% w/w dry weight, was reached by P.amethionina sp. and Galactomyces sp., respectively, at 25g/L suspended solids concentration, and that of 37.69% w/w dry weight by T.oleaginosus at 30g/L suspended solids concentration all with municipal secondary sludge. The results showed that carbon to nitrogen ratio had more obvious effect on lower suspended solids concentration (10g/L) than on the higher one (30g/L) in lipid accumulation of the three strains.

Lipid production from Trichosporon oleaginosus cultivated with pre-treated secondary wastewater sludge

Wastewater sludge contains most of the nutrients required for growth of microorganism. In this study, thermal and chemical (acidic and alkaline)-thermal treatments were applied to different sludge suspended solids concentrations (10–30g/L) and the pre-treated sludge was used as raw material for lipid production by Trichosporon oleaginosus. The results showed that compared to 48h with thermal pre-treated sludge, only 42h was required in chemical–thermal pre-treated sludge to obtain maximum lipid content (around 39% w/w) by T. oleaginosus. The experiments using struvite formation to reduce nitrogen (ammonia nitrogen) amount and hence to increase C/N ratio showed that the formation was not an efficient way to enhance lipid accumulation as ammonia nitrogen contributes to a very small weight of total nitrogen. The biodiesel produced by pre-treated sludge in the study mainly contains C14, C16, and C18, which are also rich in currently commercial biodiesel produced from soybean seed, canola, and sunflower seed oils. It shows the new approach that wastewater sludge as a feedstock for the production of biodiesel.

Ultrasonication assisted lipid extraction from oleaginous microorganisms

Various solvents, including water, hexane, methanol, and chloroform/methanol (1:1 v/v), were tested to identify the efficiency of lipid extraction from Trichosporon oleaginosus and an oleaginous fungal strain SKF-5 under ultrasonication (520 kHz 40 W and 50 Hz 2800 W) and compared with the conventional chloroform methanol (2:1 v/v) extraction method. The highest lipid recovery 10.2% and 9.3% with water, 43.2% and 33.2% with hexane, 75.7% and 65.1% with methanol, 100% and 100% w/w biomass with chloroform/methanol were obtained from T. oleaginosus and SKF-5 strain, respectively, at ultrasonication frequency 50 Hz and power input 2800 W. Ultrasonication chloroform/methanol extraction recovered total lipid in a short time (15 min) and low temperature (25°C). Whereas the conventional chloroform methanol extraction to achieve total lipid recovery required 12h at 60°C. Ultrasonication chloroform/methanol extraction would be a promising method of lipid extraction from the microorganisms.

Characterization of oleaginous yeasts revealed two novel species: Trichosporon cacaoliposimilis sp. nov. and Trichosporon oleaginosus sp. nov.

Two new species in the anamorphic basidiomycetous genus Trichosporon (Tremellomycetes, Agaricomycotina) were uncovered in a DNA sequence-based molecular analysis of oleaginous yeasts maintained in the ATCC Mycology Collection. One yeast is named as Trichosporon cacaoliposimilis sp. nov. for its capability of synthesizing and accumulating a large amount of lipids having a composition equivalent to that of natural cacao butter. The type strain is ATCC 20505T, originally deposited as Trichosporon sp. The other can use food industry wastes and agricultural byproducts as the substrate for growth and accumulation of a high level of oil and accordingly is named Trichosporon oleaginosus sp. nov. The type strain is ATCC 20509T, previously identified as Cryptococcus curvatus. Molecular phylogenetic analyses indicate that T. cacaoliposimilis is a novel taxon in the Gracile clade of the genus, close to T. gracile and T. dulcitum, and that T. oleaginosus belongs to the Cutaneum clade, with T. jirovecii as the closest sister taxon. Other oleaginous yeasts were identified as new strains of known taxa, T. insectorum, Candida orthopsilosis and C. palmioleophila.