The ability of Yarrowia lipolytica W29 immobilized by calcium alginate to degrade oil and chemical oxygen demand （COD） was examined. The degradation rules of oil and COD by immobilized cells with the cell density of...The ability of Yarrowia lipolytica W29 immobilized by calcium alginate to degrade oil and chemical oxygen demand （COD） was examined. The degradation rules of oil and COD by immobilized cells with the cell density of 6.65 × 10^6 CFU/mL degraded 2000 mg/L oil and 2000 mg/L COD within 50 h at 30℃ （pH 7.0, 150 r/min）, similarly to those of free cells, and the degradation efficiencies of oil and COD by immobilized cells were above 80%, respectively. The factors affecting oil and COD degradation by immobilized cells were investigated, the results showed that immobilized cells had high thermostability compared to that of free cells, and substrate concentration significantly affected degrading ability of immobilized cells. Storage stability and reusability tests revealed that the oil degradation ability of immobilized cells was stable after storing at 4~C for 30 d and reuse for 12 times, respectively, the COD degradation rate of immobilized cells was also maintained 82% at the sixth cycle. These results suggested that immobilized Y lipolytica might be applicable to a wastewater treatment system for the removal of oil and COD.展开更多
The production of extra cellular lipase in Solid State Fermentation (SSF) using Yarrowia lipolytica NCIM 3589 with Palm Kernal cake (Elaeis guineensis) has been studied. Different parameters such as incubation time, i...The production of extra cellular lipase in Solid State Fermentation (SSF) using Yarrowia lipolytica NCIM 3589 with Palm Kernal cake (Elaeis guineensis) has been studied. Different parameters such as incubation time, inoculum level, initial moisture content, carbon level and nitrogen level of the medium were optimized. Screening of various process variables has been accomplished with the help of Plackett-Burman design. The maximum lipase activity of 18.58 units per gram of dry fermented substrate (U/gds) was observed with the substrate of Palm Kernal cake in four days of fermentation.展开更多
The bioconversion of the hydrophobic and volatile limonene to perillic acid, a potential anticancer agent, by the yeast Yarrowia lipolytica was studied in two steps. Firstly, experimental design was used for process o...The bioconversion of the hydrophobic and volatile limonene to perillic acid, a potential anticancer agent, by the yeast Yarrowia lipolytica was studied in two steps. Firstly, experimental design was used for process optimization using high-purity limonene as substrate and secondly orange essential oil containing 89.1% limonene was used as substrate under the previously optimized conditions. Limonene concentration and pH were identified by fractional factorial design as significant factors and were optimized by central composite design. Under optimized process conditions (0.16% (v/v) limonene;pH 6.9), the 24 h biotransformation process resulted in the accumulation of 0.368 g·L-1 of perillic acid corresponding to a molar yield of 23.1%. A subsequent substrate addition under the same reaction conditions doubled perillic acid concentration to 0.793 g·L-1 and a molar yield of 24.2%. The use of orange essential oil under the optimized reaction conditions increased both perillic acid accumulation and yield to 0.872 g·L-1 and 29.7%, respectively. The robustness of Y. lipolytica allowed the efficient biotransformation of a crude by-product of the citrus industry into a valuable fine chemical.展开更多
3,4-Dihydroxy L-phenylalanine(L-DOPA)is considered a potent drug for the treatment of Parkinson disease.Physical and nutritional parameters where optimized by using Yarrowia lipolytica-NCIM 3450 to accomplished the hi...3,4-Dihydroxy L-phenylalanine(L-DOPA)is considered a potent drug for the treatment of Parkinson disease.Physical and nutritional parameters where optimized by using Yarrowia lipolytica-NCIM 3450 to accomplished the highest production of L-DOPA.Screenings of critical components were completed by using a Plackett–Burman design,while further optimization was carried out using the Box–Behnken design.The optimized factor levels predicted by the model were pH 6.1,1.659 g L^(-1)yeast extract,1.491 g L^(-1)L-tyrosine and 0.0290 g L^(-1)CuSO4.The predicted yield of L-DOPA with these levels was 1.319 g L^(-1),while actual yield obtained was 1.273 g L^(-1).The statistical analysis revealed that model is significant with F value 19.55 and R2 value 0.9514.This process resulted in a 3.594-fold increase in the yield of LDOPA.L-DOPA was confirmed by HPTLC and HPLC analysis.Thus,Yarrowia lipolytica-NCIM 3450 has potential to be a new source for the production of L-DOPA.展开更多
Growth rate of Yarrowia lipolytica NCIM 3589 was observed in a fermentation medium consisting of peptone, yeast extract, sodium chloride. Logistic equation was fitted to the growth data (time vs. biomass concentration...Growth rate of Yarrowia lipolytica NCIM 3589 was observed in a fermentation medium consisting of peptone, yeast extract, sodium chloride. Logistic equation was fitted to the growth data (time vs. biomass concentration) and compared with the prediction given by Artificial Neural Networks (ANN). ANN was found to be superior in describing growth characteristics. A single MATLAB programme is developed to fit the growth data by logistic equation and ANN.展开更多
Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica...Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica ATCC20688, which could produce a high yield of glutathione. First, the capability of glutathione production in the ATCC20688 strain was estimated. In comparison with other yeasts, the yield of this strain was higher than those in Pichia strains. Furthermore, this strain could produce glutathione by assimilating sodium oleate. We next performed mutation and gene cloning to improve the yield. After the yield of glutathione was improved in the isolated methylglyoxal-resistant mutant (MGR3), the glutathione synthetase gene was cloned into the MGR3 strain. By using this recombinant strain, we could reach the maximum yield and intracellular content of glutathione of 54 mg/L-medium and 30 mg/g-dry cell weight, respectively.展开更多
Transesterification is the most common production process for biodiesel. From this reaction, a glycerin phase is produced that is impure, thus lowering market value. However, because it is rich in carbon, it is an alt...Transesterification is the most common production process for biodiesel. From this reaction, a glycerin phase is produced that is impure, thus lowering market value. However, because it is rich in carbon, it is an alternative for generating bioproducts with a higher added value through bioconversion by microorganisms. The aim of this study was to screen parameters, such as pH （4, 5, 6, 7 and 8） and the initial glycerol concentration at 30 ± ℃ with agitation at 150 rpm for bioemulsifier and lipid synthesis in a submerged medium by Yarrowia lipolytica IMUFRJ 50678 from crude glycerin. The best conditions for bioemulsifier production were 30 ± ℃ at pH： 6 with 50 g/L of initial substrate, which produced 2.7 g/L of lipids, from which the optimum 300.5 mg/L of triglycerides was produced over 48 h of microorganism growth.展开更多
This work reports detergents production using biological surfactants, microbiologically synthesized, and compares its foaming power and emulsification capacity to those presented by a petroleum based surfactant. Both ...This work reports detergents production using biological surfactants, microbiologically synthesized, and compares its foaming power and emulsification capacity to those presented by a petroleum based surfactant. Both used microorganisms were capable to produce surfactants, been able to emulsify oil/water mixtures and cause decrease of surface tension of water. The biosurfactant produced from Yarrowia lipolytica has a critical micelle concentration lower than that obtained from Pseudomonas aeruginosa （10 and 30 mg·Lt, respectively）, but the later showed better results in foaming power and emulsification experiments, similar to the synthetic detergent.展开更多
Due to the scarcity of fossil fuels in the world, there is increasing interest in the commercial production of biodiesel, which leads to obtaining large amounts of glycerol as a byproduct. If not disposed of properly,...Due to the scarcity of fossil fuels in the world, there is increasing interest in the commercial production of biodiesel, which leads to obtaining large amounts of glycerol as a byproduct. If not disposed of properly, glycerol can generate environmental impact. One of the promises, the application of the crude glycerol is the production of citric acid by microbial fermentation. Citric acid is industrially produced by a submerged fermentation process with Aspergillus niger, using sucrose as carbon source, but due to increased demand for citric acid, alternative processes using renewable sources or waste materials as substrates and the cultivation of yeast strains are being studied. The aim of the study was to determine the best culture condition for maximum citric acid synthesis and lower isocitric acid production from crude glycerol through experimental design tool. For this purpose, the yeast strain Yarrowia lipolytica IMUFRJ-50682 was cultivated in nitrogen-limited glycerol-based media. Therefore, glycerol and yeast extract concentrations and agitation speed were evaluated as independent variables. With pure glycerol, the highest citric acid production achieved was 16.5 g/L with an isocitric acid production of 7.7% （in relation to citric acid）. With crude glycerol, citric acid production reduced to 6.7 g/L because of higher biomass yield. Therefore, an increase in the initial carbon to nitrogen molar ratio from 714 to 1,561 was necessary to increase citric acid production to 9.2 g/L, reducing isocitric acid production and to achieve a yield of 0.41 g of citric acid per glycerol consumed. In this condition, less nitrogen source was used, reducing production costs.展开更多
According literature, the induction of Yarrowia lipolytica alkaline protease promoter （PXPR2） is efficient in pH 〉 6.0 and with high peptone dose. To establish optimal pH and peptone concentration for induction of ...According literature, the induction of Yarrowia lipolytica alkaline protease promoter （PXPR2） is efficient in pH 〉 6.0 and with high peptone dose. To establish optimal pH and peptone concentration for induction of invertase gene （suc2 of Saccharomyces cerevisaie） under PXPR2 in new Y. lipolytica A-101 invertase positive （Suc＋） transformants their growth on Bioscreen C was analyzed. Minimal mineral medium with thiamine （MMT） and sucrose （1%）, adjusted to pH from 5.8 to 7.6 and supplemented by 0-0.1% of peptone was used. Biomass （OD）, maximal specific growth rate （μmax） and consumed sucrose were measured. Maximal yeasts growth, resulting from the optimal PXPR2 induction, was observed at pH 7.2 and with very low peptone doses （0.0025% and 0.01%）. For five clones （A-101 1356-5; A-101 B54-6; A-101 B57-4; A-101 A18 and W29 ura3-302） only 0.005% of peptone was needed. Amount of hydrolyzed sucrose varied from 24% to 83% and μmax from 0.06 to 0.28 hl. Suc^＋ clones differ in growth parameters, so the site of yeast cassette integration into genome influences expression level of suc2 under PXPR2. Designing large scale processes with Y. lipolytica Suc^＋ clones peptone concentration has to be 100 times smaller than recommended so far.展开更多
Longifolene(C15H24)is a tricyclic sesquiterpene widely utilized in the cosmetics and fragrances due to its versatile applications.Traditional extraction methods from plants suffer from low titer and lengthy production...Longifolene(C15H24)is a tricyclic sesquiterpene widely utilized in the cosmetics and fragrances due to its versatile applications.Traditional extraction methods from plants suffer from low titer and lengthy production cycles,while chemical synthesis is hampered by the compound’s complex structure,leading to high costs and insufficient market supply.This study aimed to develop a microbial cell factory for enhanced longifolene production.The strategy involved integrating longifolene synthase from Pinus sylvestris(PsTPS)into Yarrowia lipolytica and employing multiple metabolic engineering approaches.Initially,key genes in the mevalonate(MVA)pathway were overexpressed to enhance longifolene precursor availability for longifolene biosynthesis.Subsequently,protein engineering techniques were applied to optimize PsTPS(tPsTPS)for improved catalytic efficiency.Furthermore,co-expression of molecular chaperones was implemented to enhance the synthesis and secretion of PsTPS.The introduction of the isopentenol utilization pathway(IUP)further augmented the supply of C5 substrate.By optimizing the culture conditions,including a reduction in culture temperature,the efflux of longifolene was increased,and the dissolved oxygen levels were enhanced to promote the growth of the strain.These collective efforts resulted culminated in the engineered strain Z03 achieving a noteworthy production level of 34.67 mg/L of longifolene in shake flasks.This study not only demonstrates the feasibility of enhancing sesquiterpene production in Y.lipolytica but also highlights the potential of microbial platforms in meeting industrial demands for complex natural products.展开更多
基金supported by the National Key Technologies R & D Program of China (No. 2007BAC23B01)the Program of Department of Education, Jiangxi Province, China (No. 2007-41).
文摘The ability of Yarrowia lipolytica W29 immobilized by calcium alginate to degrade oil and chemical oxygen demand （COD） was examined. The degradation rules of oil and COD by immobilized cells with the cell density of 6.65 × 10^6 CFU/mL degraded 2000 mg/L oil and 2000 mg/L COD within 50 h at 30℃ （pH 7.0, 150 r/min）, similarly to those of free cells, and the degradation efficiencies of oil and COD by immobilized cells were above 80%, respectively. The factors affecting oil and COD degradation by immobilized cells were investigated, the results showed that immobilized cells had high thermostability compared to that of free cells, and substrate concentration significantly affected degrading ability of immobilized cells. Storage stability and reusability tests revealed that the oil degradation ability of immobilized cells was stable after storing at 4~C for 30 d and reuse for 12 times, respectively, the COD degradation rate of immobilized cells was also maintained 82% at the sixth cycle. These results suggested that immobilized Y lipolytica might be applicable to a wastewater treatment system for the removal of oil and COD.
文摘The production of extra cellular lipase in Solid State Fermentation (SSF) using Yarrowia lipolytica NCIM 3589 with Palm Kernal cake (Elaeis guineensis) has been studied. Different parameters such as incubation time, inoculum level, initial moisture content, carbon level and nitrogen level of the medium were optimized. Screening of various process variables has been accomplished with the help of Plackett-Burman design. The maximum lipase activity of 18.58 units per gram of dry fermented substrate (U/gds) was observed with the substrate of Palm Kernal cake in four days of fermentation.
文摘The bioconversion of the hydrophobic and volatile limonene to perillic acid, a potential anticancer agent, by the yeast Yarrowia lipolytica was studied in two steps. Firstly, experimental design was used for process optimization using high-purity limonene as substrate and secondly orange essential oil containing 89.1% limonene was used as substrate under the previously optimized conditions. Limonene concentration and pH were identified by fractional factorial design as significant factors and were optimized by central composite design. Under optimized process conditions (0.16% (v/v) limonene;pH 6.9), the 24 h biotransformation process resulted in the accumulation of 0.368 g·L-1 of perillic acid corresponding to a molar yield of 23.1%. A subsequent substrate addition under the same reaction conditions doubled perillic acid concentration to 0.793 g·L-1 and a molar yield of 24.2%. The use of orange essential oil under the optimized reaction conditions increased both perillic acid accumulation and yield to 0.872 g·L-1 and 29.7%, respectively. The robustness of Y. lipolytica allowed the efficient biotransformation of a crude by-product of the citrus industry into a valuable fine chemical.
文摘3,4-Dihydroxy L-phenylalanine(L-DOPA)is considered a potent drug for the treatment of Parkinson disease.Physical and nutritional parameters where optimized by using Yarrowia lipolytica-NCIM 3450 to accomplished the highest production of L-DOPA.Screenings of critical components were completed by using a Plackett–Burman design,while further optimization was carried out using the Box–Behnken design.The optimized factor levels predicted by the model were pH 6.1,1.659 g L^(-1)yeast extract,1.491 g L^(-1)L-tyrosine and 0.0290 g L^(-1)CuSO4.The predicted yield of L-DOPA with these levels was 1.319 g L^(-1),while actual yield obtained was 1.273 g L^(-1).The statistical analysis revealed that model is significant with F value 19.55 and R2 value 0.9514.This process resulted in a 3.594-fold increase in the yield of LDOPA.L-DOPA was confirmed by HPTLC and HPLC analysis.Thus,Yarrowia lipolytica-NCIM 3450 has potential to be a new source for the production of L-DOPA.
文摘Growth rate of Yarrowia lipolytica NCIM 3589 was observed in a fermentation medium consisting of peptone, yeast extract, sodium chloride. Logistic equation was fitted to the growth data (time vs. biomass concentration) and compared with the prediction given by Artificial Neural Networks (ANN). ANN was found to be superior in describing growth characteristics. A single MATLAB programme is developed to fit the growth data by logistic equation and ANN.
文摘Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica ATCC20688, which could produce a high yield of glutathione. First, the capability of glutathione production in the ATCC20688 strain was estimated. In comparison with other yeasts, the yield of this strain was higher than those in Pichia strains. Furthermore, this strain could produce glutathione by assimilating sodium oleate. We next performed mutation and gene cloning to improve the yield. After the yield of glutathione was improved in the isolated methylglyoxal-resistant mutant (MGR3), the glutathione synthetase gene was cloned into the MGR3 strain. By using this recombinant strain, we could reach the maximum yield and intracellular content of glutathione of 54 mg/L-medium and 30 mg/g-dry cell weight, respectively.
文摘Transesterification is the most common production process for biodiesel. From this reaction, a glycerin phase is produced that is impure, thus lowering market value. However, because it is rich in carbon, it is an alternative for generating bioproducts with a higher added value through bioconversion by microorganisms. The aim of this study was to screen parameters, such as pH （4, 5, 6, 7 and 8） and the initial glycerol concentration at 30 ± ℃ with agitation at 150 rpm for bioemulsifier and lipid synthesis in a submerged medium by Yarrowia lipolytica IMUFRJ 50678 from crude glycerin. The best conditions for bioemulsifier production were 30 ± ℃ at pH： 6 with 50 g/L of initial substrate, which produced 2.7 g/L of lipids, from which the optimum 300.5 mg/L of triglycerides was produced over 48 h of microorganism growth.
文摘This work reports detergents production using biological surfactants, microbiologically synthesized, and compares its foaming power and emulsification capacity to those presented by a petroleum based surfactant. Both used microorganisms were capable to produce surfactants, been able to emulsify oil/water mixtures and cause decrease of surface tension of water. The biosurfactant produced from Yarrowia lipolytica has a critical micelle concentration lower than that obtained from Pseudomonas aeruginosa （10 and 30 mg·Lt, respectively）, but the later showed better results in foaming power and emulsification experiments, similar to the synthetic detergent.
文摘Due to the scarcity of fossil fuels in the world, there is increasing interest in the commercial production of biodiesel, which leads to obtaining large amounts of glycerol as a byproduct. If not disposed of properly, glycerol can generate environmental impact. One of the promises, the application of the crude glycerol is the production of citric acid by microbial fermentation. Citric acid is industrially produced by a submerged fermentation process with Aspergillus niger, using sucrose as carbon source, but due to increased demand for citric acid, alternative processes using renewable sources or waste materials as substrates and the cultivation of yeast strains are being studied. The aim of the study was to determine the best culture condition for maximum citric acid synthesis and lower isocitric acid production from crude glycerol through experimental design tool. For this purpose, the yeast strain Yarrowia lipolytica IMUFRJ-50682 was cultivated in nitrogen-limited glycerol-based media. Therefore, glycerol and yeast extract concentrations and agitation speed were evaluated as independent variables. With pure glycerol, the highest citric acid production achieved was 16.5 g/L with an isocitric acid production of 7.7% （in relation to citric acid）. With crude glycerol, citric acid production reduced to 6.7 g/L because of higher biomass yield. Therefore, an increase in the initial carbon to nitrogen molar ratio from 714 to 1,561 was necessary to increase citric acid production to 9.2 g/L, reducing isocitric acid production and to achieve a yield of 0.41 g of citric acid per glycerol consumed. In this condition, less nitrogen source was used, reducing production costs.
文摘According literature, the induction of Yarrowia lipolytica alkaline protease promoter （PXPR2） is efficient in pH 〉 6.0 and with high peptone dose. To establish optimal pH and peptone concentration for induction of invertase gene （suc2 of Saccharomyces cerevisaie） under PXPR2 in new Y. lipolytica A-101 invertase positive （Suc＋） transformants their growth on Bioscreen C was analyzed. Minimal mineral medium with thiamine （MMT） and sucrose （1%）, adjusted to pH from 5.8 to 7.6 and supplemented by 0-0.1% of peptone was used. Biomass （OD）, maximal specific growth rate （μmax） and consumed sucrose were measured. Maximal yeasts growth, resulting from the optimal PXPR2 induction, was observed at pH 7.2 and with very low peptone doses （0.0025% and 0.01%）. For five clones （A-101 1356-5; A-101 B54-6; A-101 B57-4; A-101 A18 and W29 ura3-302） only 0.005% of peptone was needed. Amount of hydrolyzed sucrose varied from 24% to 83% and μmax from 0.06 to 0.28 hl. Suc^＋ clones differ in growth parameters, so the site of yeast cassette integration into genome influences expression level of suc2 under PXPR2. Designing large scale processes with Y. lipolytica Suc^＋ clones peptone concentration has to be 100 times smaller than recommended so far.
基金supported by the National Natural Science Foundation of China(No.42206137,32270118)the Science and Technology Planning Project of Guangzhou(2024A04J4129)the Natural Science Foundation of Guangdong Province(No.2019B1515120062).
文摘Longifolene(C15H24)is a tricyclic sesquiterpene widely utilized in the cosmetics and fragrances due to its versatile applications.Traditional extraction methods from plants suffer from low titer and lengthy production cycles,while chemical synthesis is hampered by the compound’s complex structure,leading to high costs and insufficient market supply.This study aimed to develop a microbial cell factory for enhanced longifolene production.The strategy involved integrating longifolene synthase from Pinus sylvestris(PsTPS)into Yarrowia lipolytica and employing multiple metabolic engineering approaches.Initially,key genes in the mevalonate(MVA)pathway were overexpressed to enhance longifolene precursor availability for longifolene biosynthesis.Subsequently,protein engineering techniques were applied to optimize PsTPS(tPsTPS)for improved catalytic efficiency.Furthermore,co-expression of molecular chaperones was implemented to enhance the synthesis and secretion of PsTPS.The introduction of the isopentenol utilization pathway(IUP)further augmented the supply of C5 substrate.By optimizing the culture conditions,including a reduction in culture temperature,the efflux of longifolene was increased,and the dissolved oxygen levels were enhanced to promote the growth of the strain.These collective efforts resulted culminated in the engineered strain Z03 achieving a noteworthy production level of 34.67 mg/L of longifolene in shake flasks.This study not only demonstrates the feasibility of enhancing sesquiterpene production in Y.lipolytica but also highlights the potential of microbial platforms in meeting industrial demands for complex natural products.
