Optimization of Biotransformation of l-Tyrosine to l-DOPA by Yarrowia lipolytica-NCIM 3472 Using Response Surface Methodology.
Ontology highlight
ABSTRACT:
SUBMITTER: Gurme ST
PROVIDER: S-EPMC3626946 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Abstract3,4-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 R(2) value 0.9514. This process resulted in a 3.594-fold increase in the yield of l-DOPA. 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.
| S-EPMC4050310 | biostudies-literature
Project description:L-DOPA (3,4-dihydroxyphenyl-L-alanine) is an extensively used drug for the treatment of Parkinson's disease. In the present study, optimization of nutritional parameters influencing L-DOPA production was attempted using the response surface methodology (RSM) from Brevundimonas sp. SGJ. A Plackett-Burman design was used for screening of critical components, while further optimization was carried out using the Box-Behnken design. The optimized levels of factors predicted by the model were pH 5.02, 1.549 g l(-1) tryptone, 4.207 g l(-1) L-tyrosine and 0.0369 g l(-1) CuSO(4) , which resulted in highest L-DOPA yield of 3.359 g l(-1). The optimization of medium using RSM resulted in a 8.355-fold increase in the yield of L-DOPA. The anova showed a significant R(2) value (0.9667), model F-value (29.068) and probability (0.001), with insignificant lack of fit. The highest tyrosinase activity observed was 2471 U mg(-1) at the 18th hour of the incubation period with dry cell weight of 0.711 g l(-1). L-DOPA production was confirmed by HPTLC, HPLC and GC-MS analysis. Thus, Brevundimonas sp. SGJ has the potential to be a new source for the production of L-DOPA.
| S-EPMC3815894 | biostudies-literature
Project description:Yarrowia clade contains yeast species morphologically, ecologically, physiologically and genetically diverse in nature. Yarrowia lipolytica NCIM 3590 (NCIM 3590), a biotechnologically important strain, isolated from Scottish sea waters was reinvestigated for its phenotypic, biochemical, molecular and genomic properties as it exhibited characteristics unlike Y. lipolytica, namely, absence of extracellular lipolytic activity, growth at lower temperatures (less than 20 °C) and in high salt concentrations (10% NaCl). Molecular identification using ITS and D1/D2 sequences suggested NCIM 3590 to be 100% identical with reference strain Yarrowia bubula CBS 12934 rather than Y. lipolytica CBS 6124 (87% identity) while phylogenetic analysis revealed that it clustered with Y. bubula under a separate clade. Further, whole genome sequencing of NCIM 3590 was performed using Illumina NextSeq technology and the draft reported here. The overall genome relatedness values obtained by dDDH (94.1%), ANIb/ANIm (99.41/99.42%) and OrthoANI (99.47%) indicated proximity between NCIM 3590 and CBS 12934 as compared to the reference strain Y. lipolytica. No extracellular lipase activity could be detected in NCIM 3590 while LIP2 gene TBLASTN analysis suggests a low 42% identity with e value 2 e?77 and 62% coverage. Hence molecular, phylogenetic, genomics, biochemical and microbial analyses suggests it belongs to Yarrowia bubula.
| S-EPMC7943819 | biostudies-literature
Project description:Response surface methodology was used to enhance the production of ?-galactosidase from Fusarium moniliforme NCIM 1099 in solid-state fermentation. Plackett-Burman design was employed for selection of critical media constituents which were optimized by central composite rotatable design. Wheat bran, peptone and FeSO4·7H2O were identified as significant medium components using PB design. Further CCRD optimized medium components as wheat bran; 4.62 ?g, peptone; 315.42 ?g, FeSO4·7H2O; 9.04 ?g. RSM methodological optimization increased the enzyme production from 13.17 to 207.33 U/g showing 15.74-fold enhancement. The ?-galactosidase was purified by 70% fractionation followed by DEAE anion exchange column chromatography which yields 23.33% with 28.68-fold purification. The molecular weight of ?-galactosidase was 57 kDa which was determined by SDS-PAGE analysis. Purified enzyme has optimum pH of 4.0 and was found to be stable in wide pH range of 3.0-9.0. Its optimum temperature was 50 °C, whereas its activity remains above 50% up to 2 h at 75 °C. Hg2+ was found to be a potent inhibitor and Mg2+ acted as an activator of enzyme. No significant change was observed in enzyme activity for galactose concentration, ranging from 1 to 100 mM. The K m values of enzyme for substrates p-nitrophenyl-?-D-galactopyranoside, melibiose and raffinose were 0.20, 1.36, and 3.66 mM, respectively. Low K m and stability to various physiological conditions of enzyme represents its potential which can be exploited in various industrial applications.
| S-EPMC5148754 | biostudies-literature
Project description:Oleaginous yeasts are fast emerging as a possible feedstock for biodiesel production. Yarrowia lipolytica, a model oleaginous yeast is known to utilize a variety of hydrophobic substrates for lipid accumulation including waste cooking oil (WCO). Approaches to increase lipid content in this yeast include metabolic engineering which requires manipulation of multiple genes in the lipid biosynthesis pathway. A classical and cost-effective approach, namely, random chemical mutagenesis on the yeast can lead to increased production of biodiesel as is explored here.In this study, chemical mutagenesis using the alkylating agent, N- methyl-N'-nitro-N-nitrosoguanidine (MNNG) as well as an additional treatment with cerulenin, a fatty acid synthase inhibitor generated 800 mutants of Y. lipolytica NCIM 3589 (761 MNNG treated and 39 MNNG + cerulenin treated). A three-stage screening using Sudan Black B plate technique, Nile red fluorimetry and total lipid extraction using solvent was performed, which enabled selection of ten high lipid yielding mutants. Time course studies of all the ten mutants were further undertaken in terms of biomass, lipid yield and lipid content to select three stable mutants (YlB6, YlC7 and YlE1) capable of growing and accumulating lipid on WCO, with lipid contents of 55, 60 and 67% as compared to 45% for the wild type. The mutants demonstrated increased volumetric lipid productivities (0.062, 0.044 and 0.041 g L-1 h-1) as compared to the wild type (0.033 g L-1 h-1). The fatty acid profile of the three mutants consisted of a high content of C16 and C18 saturated and monounsaturated fatty acids and was found to be suitable for biodiesel production. The fuel properties, namely, density, kinematic viscosity, total acid number, iodine value of the three mutants were evaluated and found to lie within the limits specified by internationally accepted standards. Additionally, it was noted that the mutants demonstrated better cetane numbers and higher heating values than the wild type strain.The chemical mutagenesis strategy adopted in this study resulted in the successful isolation of three stable high SCO yielding mutants. The mutants, namely, YlB6, YlC7 and YlE1 exhibited a 1.22, 1.33 and 1.49-fold increase in lipid contents when grown on 100 g L-1 waste cooking oil than the parental yeast strain. The fatty acid methyl ester (FAME) profiles of all the three mutants was determined to be suitable for biodiesel suggesting their potential applicability while simultaneously addressing the management of waste cooking oil.
| S-EPMC5655982 | biostudies-literature
Project description:BACKGROUND:Yarrowia lipolytica is a non-conventional yeast that is extensively investigated for its ability to excrete citrate or to accumulate large amounts of storage lipids, which is of great significance for single cell oil production. Both traits are thus of interest for basic research as well as for biotechnological applications but they typically occur simultaneously thus lowering the respective yields. Therefore, engineering of strains with high lipid content relies on novel concepts such as computational simulation to better understand the two competing processes and to eliminate citrate excretion. RESULTS:Using a genome-scale model (GSM) of baker's yeast as a scaffold, we reconstructed the metabolic network of Y. lipolytica and optimized it for use in flux balance analysis (FBA), with the aim to simulate growth and lipid production phases of this yeast. We validated our model and found the predictions of the growth behavior of Y. lipolytica in excellent agreement with experimental data. Based on these data, we successfully designed a fed-batch strategy to avoid citrate excretion during the lipid production phase. Further analysis of the network suggested that the oxygen demand of Y. lipolytica is reduced upon induction of lipid synthesis. According to this finding we hypothesized that a reduced aeration rate might induce lipid accumulation. This prediction was indeed confirmed experimentally. In a fermentation combining these two strategies lipid content of the biomass was increased by 80%, and lipid yield was improved more than four-fold, compared to standard conditions. CONCLUSIONS:Genome scale network reconstructions provide a powerful tool to predict the effects of genetic modifications and the metabolic response to environmental conditions. The high accuracy and the predictive value of a newly reconstructed GSM of Y. lipolytica to optimize growth conditions for lipid accumulation are demonstrated. Based on these findings, further strategies for engineering Y. lipolytica towards higher efficiency in single cell oil production are discussed.
| S-EPMC4623914 | biostudies-literature
Project description:BackgroundOdd chain fatty acids (odd FAs) have a wide range of applications in therapeutic and nutritional industries, as well as in chemical industries including biofuel. Yarrowia lipolytica is an oleaginous yeast considered a preferred microorganism for the production of lipid-derived biofuels and chemicals. However, it naturally produces negligible amounts of odd chain fatty acids.ResultsThe possibility of producing odd FAs using Y. lipolytica was investigated. Y. lipolytica wild-type strain was shown able to grow on weak acids; acetate, lactate, and propionate. Maximal growth rate on propionate reached 0.24?±?0.01 h-1 at 2 g/L, and growth inhibition occurred at concentration above 10 g/L. Wild-type strain accumulated lipids ranging from 7.39 to 8.14% (w/w DCW) depending on the carbon source composition, and odd FAs represented only 0.01-0.12 g/L. We here proved that the deletion of the PHD1 gene improved odd FAs production, which reached a ratio of 46.82% to total lipids. When this modification was transferred to an obese strain, engineered for improving lipid accumulation, further increase odd FAs production reaching a total of 0.57 g/L was shown. Finally, a fed-batch co-feeding strategy was optimized for further increase odd FAs production, which generated 0.75 g/L, the best production described so far in Y. lipolytica.ConclusionsA Y. lipolytica strain able to accumulate high level of odd chain fatty acids, mainly heptadecenoic acid, has been successfully developed. In addition, a fed-batch co-feeding strategy was optimized to further improve lipid accumulation and odd chain fatty acid content. These lipids enriched in odd chain fatty acid can (1) improve the properties of the biodiesel generated from Y. lipolytica lipids and (2) be used as renewable source of odd chain fatty acid for industrial applications. This work paves the way for further improvements in odd chain fatty acids and fatty acid-derived compound production.
| S-EPMC5991449 | biostudies-literature
Project description:Oyster (Crassostrea talienwhanensis) protein was hydrolyzed by trypsin to produce peptides with different response values, and response surface methodology (RSM) was applied to optimize the hydrolysis conditions. The highest degree of hydrolysis (DH) of the oyster peptide (OP) was obtained at an enzyme concentration of 1593.2 U/g, a pH of 8.2, a hydrolysis temperature of 40.1 °C, a hydrolysis time of 6.0 h, and a water/material ratio of 8.2. The greatest hydroxyl-radical-scavenging activity of OP was obtained at an enzyme concentration of 1546.3 U/g, a pH of 9.0, a hydrolysis temperature of 50.2 °C, a hydrolysis time of 5.1 h, and a water/material ratio of 5.6. The largest branched-chain amino acid (BCAA) content of OP was obtained at an enzyme concentration of 1323.8 U/g, a pH of 8.3, a hydrolysis temperature of 41.7 °C, a hydrolysis time of 6.7 h, and a water/material ratio of 4.8. The three experimental values were significantly in agreement with the predicted values within the 95% confidence interval. Furthermore, ultrafiltration and chromatographic methods were used to purify the OP, and 13 peptides that were rich in Lys, Arg, His, and Thr were identified by LC-MS/MS. The results of this study offer different optimum hydrolysis conditions to produce target peptides from oyster protein by using RSM, and this study provide a theoretical basis for the high-value utilization of oyster protein.
| S-EPMC7357005 | biostudies-literature
Project description:BACKGROUND:Insect wax is a famous biological resource for the role in economic production in China. Insect wax is a good source of policosanol, which may is a candidate supplement in foodstuff and pharmaceuticals that has important physiological activities. Therefore, this work aims to investigate a high-yield and rapid method for policosanol fabrication from insect wax. RESULTS:The conditions for policosanol fabrication were optimized as follows: an oil bath temperature of 112.7°C and reductant dosage of 0.97 g (used for the reduction of 10.00 g of insect wax). The yield of policosanol reached 83.20%, which was 4 times greater than that of existing methods, such as saponification. The total content of policosanol obtained under the optimal conditions reached 87%. In other words, a high yield of policosanol was obtained from insect wax (723.84 mg/g), that was 55 times higher than that generated from beeswax-brown via saponification. The concentrations of metal residues in policosanol were within the limits of the European Union regulations and EFSA stipulation. The LD50 values for oral doses of insect wax and policosanol were both > 5 g/kg. CONCLUSION:Policosanol was fabricated via solvent-free reduction from insect wax using LiAlH4 at a high yield. The fabrication conditions were optimized. Policosanol and insect wax showed high security, which made them potential candidates as supplements in foods, pharmaceuticals and cosmetics. The rapid and high-yield method has great potential for commercial manufacturing of policosanol.
| S-EPMC5953464 | biostudies-literature
Project description:The aim of this work was to demonstrate an optimization methodology to reliably obtain stable macrodispersions (i.e., for ≥24 h) of carbon nanotubes in water using sonication. Response surface methodology (RSM) was utilized to assess and optimize the sonication parameters for the process. The studied input parameters were (i) sonication time (duration), (ii) amplitude (of vibration), and (iii) pulse-on/off (duration) of the sonicator. The analyzed responses were mean diameter and size distribution of multiwalled carbon nanotube (MWNT) aggregates in water, which were measured by the dynamic light scattering technique. A semiempirical model was developed and statistically tested to estimate the magnitude of sonicator parameters required to obtain specified MWNT macrodispersions (i.e., aggregates' mean diameter and distribution) in water. The results showed that MWNT aggregates of 2 ± 0.5 μm can be obtained by optimizing sonicator parameters to a sonication time of 89 s, amplitude of 144 μm, and pulse-on/off cycle of 44/30 s. These process settings for 100 mg/L MWNTs in a 30 mL aliquot of deionized water would consume 863 J/mL of sonication energy. Contrary to the popular belief, "sonication time" and/or "sonication energy input" were not found to be proportional to the degree of dispersion of MWNTs in water. This might be the reason for the frequent disparity and nonreproducibility of sonication results reported in scientific literature, especially for dispersing nanomaterials in a number of different systems. The amplitude of vibration was noted to be the most sensitive parameter affecting MWNT aggregates' diameter and distribution in water. The characterization of MWNTs was performed using electron microscopy, surface area analyzer, thermogravimetric analyzer, and zeta potential analyzer. This study can be helpful in evaluating sonication dispersion of particulate matter in other incompressible fluids such as graphene dispersion in organic solvents.
| S-EPMC6648579 | biostudies-literature