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Sebastián Sánchez - One of the best experts on this subject based on the ideXlab platform.
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acid and enzymatic fractionation of olive stones for ethanol production using Pachysolen tannophilus
Processes, 2020Co-Authors: Manuel Cuevas, Marwa Saleh, Juan Francisco Garciamartin, Sebastián SánchezAbstract:Olive stones are an abundant lignocellulose material in the countries of the Mediterranean basin that could be transformed to bioethanol by biochemical pathways. In this work, olive stones were subjected to fractionation by means of a high-temperature dilute-acid pretreatment followed by enzymatic hydrolysis of the pretreated solids. The hydrolysates obtained in these steps were separately subjected to fermentation with the yeast Pachysolen tannophilus ATCC 32691. Response surface methodology with two independent variables (temperature and reaction time) was applied for optimizing D-xylose production from the raw material by dilute acid pretreatment with 0.01 M sulfuric acid. The highest D-xylose yield in the liquid fraction was obtained in the pretreatment at 201 °C for 5.2 min. The inclusion of a detoxification step of the acid prehydrolysate, by vacuum distillation, allowed the fermentation of the sugars into ethanol and xylitol. The enzymatic hydrolysis of the pretreated solids was solely effective when using high enzyme loadings, thus leading to easily fermentable hydrolysates into ethanol. The mass macroscopic balances of the overall process illustrated that the amount of inoculum used in the fermentation of the acid prehydrolysates strongly affected the ethanol and xylitol yields.
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valorization of olive stones for xylitol and ethanol production from dilute acid pretreatment via enzymatic hydrolysis and fermentation by Pachysolen tannophilus
Biochemical Engineering Journal, 2014Co-Authors: Marwa Saleh, Manuel Cuevas, Juan F Garcia, Sebastián SánchezAbstract:Abstract Olive stones are an agro-industrial by-product abundant in the Mediterranean area that is regarded as a potential lignocellulosic feedstock for sugar production. Statistical modeling of dilute-sulphuric acid hydrolysis of olive stones has been performed using a response surface methodology, with treatment temperature and process time as factors, to optimize the hydrolysis conditions aiming to attain maximum d -xylose extraction from hemicelluloses. Thus, solid yield and composition of solid and liquid phases were assessed by empirical modeling. The highest yield of d -xylose was found at a temperature of 195 °C for 5 min. Under these conditions, 89.7% of the total d -xylose was recovered from raw material. The resulting solids from optimal conditions were assayed as substrate for enzymatic hydrolysis, while fermentability of hemicellulosic hydrolysates was tested using the d -xylose-fermenting yeast Pachysolen tannophilus. Both bioprocesses were considerably influenced by enzyme loading and inoculum size. In the enzymatic hydrolysis step, about 56% of cellulose was converted into d -glucose by using an enzyme/solid ratio of 40 FPU g−1, while in the fermentation carried out with a cell concentration of 2 g L−1 a yield of 0.44 g xylitol/g d -xylose and a global volumetric productivity of 0.11 g L−1 h−1 were achieved.
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fermentation of enzymatic hydrolysates from olive stones by Pachysolen tannophilus
Journal of Chemical Technology & Biotechnology, 2009Co-Authors: Manuel Cuevas, Sebastián Sánchez, Vicente Bravo, Nicolas Cruz, Juan Godoy F GarciaAbstract:BACKGROUND: Olive stones were pretreated with liquid hot water (LHW or autohydrolysis) at maximum temperatures between 175 and 225 °C (severity factors, logR0, between 2.73 and 4.39) to be subjected (both liquid and solid components) afterwards to enzymatic hydrolysis with cellulases from Trichoderma viride. Ethanol fermentation of hydrolysates was performed with the non-traditional yeast Pachysolen tannophilus ATCC 32691. RESULTS: After the enzymatic step, yields of hemicellulose solubilization reached 100%, while the cellulose was only partially hydrolysed (23%, logR0 = 4.39). The maximum yields in total reducing sugars and acetic acid, at the upper end of the severity range, was close to 0.25 and 0.04 g g−1 dry stone, respectively. During the fermentation stage, the increase in R0 reduced the maximum specific growth rate, biomass productivity, and overall biomass yield. The overall yields of ethanol and xylitol ranged, respectively, from 0.18 to 0.25 g g−1 and from 0.01 to 0.13 g g−1. CONCLUSIONS: The results demonstrate the possibility of producing ethanol from olive stones, making use of the cellulose and hemicellulose fraction of the waste. It was confirmed that the overall yield in xylitol strongly depended on severity factor, while the overall yield in ethanol remained practically constant for all the pretreatment conditions tested. Copyright © 2008 Society of Chemical Industry
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fermentation of acid hydrolysates from olive tree pruning debris by Pachysolen tannophilus
Bioprocess and Biosystems Engineering, 2008Co-Authors: Alberto J Moya, Vicente Bravo, Soledad Mateo, Sebastián SánchezAbstract:The influence of the type and concentration of acid in the hydrolysis process and its effect on the subsequent fermentation by Pachysolen tannophilus (ATCC 32691) to produce ethanol and xylitol was studied. The hydrolysis experiments were performed using hydrochloric, sulphuric and trifluoroacetic acids in concentrations ranging from 0.1 to 1.0 N, a temperature of 90 °C, and a time of 240 min. The fermentation experiments were conducted on a laboratory scale in a batch-culture reactor at pH 4.5 and 30 °C. The hydrolysis with the highest acid concentration produced the complete solubilization of hemicellulose to monosaccharides. The highest values for the specific rate of ethanol production were registered in cultures hydrolyzed with trifluoroacetic acid, and values were found to decrease as the acid concentration increased. The highest values of overall ethanol yields ( $$ Y^{{\text{G}}}_{{{\text{E/s}}}} $$ = 0.37 kg kg−1) were also found in the fermentation of the hydrolysates of trifluoroacetic acid.
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fermentation of olive tree pruning acid hydrolysates by Pachysolen tannophilus
Biochemical Engineering Journal, 2007Co-Authors: Inmaculada Romero, Sebastián Sánchez, Eulogio Castro, Manuel Moya, Encarnacion Ruiz, Vicente BravoAbstract:Abstract The influence of hydrolysis conditions on the fermentation of hydrolysates obtained from olive tree pruning (a renewable, low cost, largely available agricultural residue) is analyzed in this work. Hydrolysis was performed using sulfuric acid at atmospheric pressure (90 °C) in a concentration range of 0.5–4N for 240 min. The fermentation of hydrolysates was carried out by Pachysolen tannophilus in a discontinuous tank bioreactor at 30 °C and pH 3.5. The main fermentation parameters determined in this study include maximum specific growth rate, biomass productivity, specific substrate consumption rate, specific ethanol production rate, and ethanol and xylitol yields. The results show that ethanol yields are much higher than xylitol yields under all the conditions tested. The maximum ethanol yield (0.38 g/g) is reached with the hydrolysate obtained with 0.75N sulfuric acid. Under these conditions the conversion of the hemicellulose fraction is 92%.
Hung Lee - One of the best experts on this subject based on the ideXlab platform.
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transfer of plasmid into the pentose fermenting yeast Pachysolen tannophilus
Journal of Microbiological Methods, 2018Co-Authors: Guiying Mei, Paramjit K Bajwa, Mehdi Dashtban, Hung LeeAbstract:Abstract The pentose-fermenting yeast Pachysolen tannophilus can convert glucose and xylose in lignocellulosic hydrolysates to ethanol. However, it performs poorly in industrially relevant lignocellulosic hydrolysates containing mixed sugars and inhibitors. Efforts have been directed at improving the performance of this yeast to enable efficient lignocellulosic biomass conversion . While some successes have been reported using random mutagenesis and/or hybridization-based approaches, further genetic improvement of this yeast is hampered by the lack of efficient gene transfer methods as well as limited genetic information to guide further construction of robust strains of P. tannophilus. In this study, we aimed to address this short-coming by establishing the optimal conditions needed for efficient gene transfer into P. tannophilus. We ascertained that plasmids can be transferred into P. tannophilus through trans-kingdom conjugation or lithium acetate (LiAc) transformation. The efficiency of plasmid YEp13 (2-micron, LEU2) transferred into a P. tannophilus leucine auxotroph (Leu −) reached as high as 1.93 × 10−2 transconjugants per input recipient and 3.25 × 104 transformants per μg plasmid DNA through trans-kingdom conjugation and transformation, respectively. In trans-kingdom conjugation, the number of recipient P. tannophilus cells played an important role, while the ratio of donor (Escherichia coli) to recipient cells was less important. For efficient transformation in P. tannophilus , the use of PEG 3350 was essential, as no transformants were obtained in its absence. The transformation efficiency increased with the addition of single-stranded carrier DNA and incubation at 30 °C for >60 min. Plasmids with different replication origins or 2-micron plasmids with different CUG codon-optimized antibiotic resistance markers were unable to transform P. tannophilus under our experimental conditions. The results are of interest in the genetic manipulation and improvement of P. tannophilus.
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Determinants of tolerance to inhibitors in hardwood spent sulfite liquor in genome shuffled Pachysolen tannophilus strains
Antonie van Leeuwenhoek, 2015Co-Authors: Nicole K. Harner, Paramjit K Bajwa, Marc B. Habash, Jack T. Trevors, Glen D. Austin, Philip A. Formusa, Chi-kin Chan, Hung LeeAbstract:Genome shuffling was used to obtain Pachysolen tannophilus mutants with improved tolerance to inhibitors in hardwood spent sulfite liquor (HW SSL). Genome shuffled strains (GHW301, GHW302 and GHW303) grew at higher concentrations of HW SSL (80 % v/v) compared to the HW SSL UV mutant (70 % v/v) and the wild-type (WT) strain (50 % v/v). In defined media containing acetic acid (0.70–0.90 % w/v), GHW301, GHW302 and GHW303 exhibited a shorter lag compared to the acetic acid UV mutant, while the WT did not grow. Genome shuffled strains produced more ethanol than the WT at higher concentrations of HW SSL and an aspen hydrolysate. To identify the genetic basis of inhibitor tolerance, whole genome sequencing was carried out on GHW301, GHW302 and GHW303 and compared to the WT strain. Sixty single nucleotide variations were identified that were common to all three genome shuffled strains. Of these, 40 were in gene sequences and 20 were within 5 bp–1 kb either up or downstream of protein encoding genes. Based on the mutated gene products, mutations were grouped into functional categories and affected a variety of cellular functions, demonstrating the complexity of inhibitor tolerance in yeast. Sequence analysis of UV mutants (UAA302 and UHW303) from which GHW301, GHW302 and GHW303 were derived, confirmed the success of our cross-mating based genome shuffling strategy. Whole-genome sequencing analysis allowed identification of potential gene targets for tolerance to inhibitors in lignocellulosic hydrolysates.
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Mutants of the pentose-fermenting yeast Pachysolen tannophilus tolerant to hardwood spent sulfite liquor and acetic acid
Antonie van Leeuwenhoek, 2014Co-Authors: Nicole K. Harner, Paramjit K Bajwa, Marc B. Habash, Jack T. Trevors, Glen D. Austin, Hung LeeAbstract:A strain development program was initiated to improve the tolerance of the pentose-fermenting yeast Pachysolen tannophilus to inhibitors in lignocellulosic hydrolysates. Several rounds of UV mutagenesis followed by screening were used to select for mutants of P. tannophilus NRRL Y2460 with improved tolerance to hardwood spent sulfite liquor (HW SSL) and acetic acid in separate selection lines. The wild type (WT) strain grew in 50 % (v/v) HW SSL while third round HW SSL mutants (designated UHW301, UHW302 and UHW303) grew in 60 % (v/v) HW SSL, with two of these isolates (UHW302 and UHW303) being viable and growing, respectively, in 70 % (v/v) HW SSL. In defined liquid media containing acetic acid, the WT strain grew in 0.70 % (w/v) acetic acid, while third round acetic acid mutants (designated UAA301, UAA302 and UAA303) grew in 0.80 % (w/v) acetic acid, with one isolate (UAA302) growing in 0.90 % (w/v) acetic acid. Cross-tolerance of HW SSL-tolerant mutants to acetic acid and vice versa was observed with UHW303 able to grow in 0.90 % (w/v) acetic acid and UAA302 growing in 60 % (v/v) HW SSL. The UV-induced mutants retained the ability to ferment glucose and xylose to ethanol in defined media. These mutants of P. tannophilus are of considerable interest for bioconversion of the sugars in lignocellulosic hydrolysates to ethanol.
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Reversible inactivation of d-xylose utilization by d-glucose in the pentose-fermenting yeast Pachysolen tannophilus
FEMS Microbiology Letters, 1992Co-Authors: Hung LeeAbstract:A major problem in fermenting pentoses using lignocellulosic substrates is the presence of d-glucose which inhibits d-xylose utilization. We previously showed that d-glucose represses the induction of xylose reductase and xylitol dehydrogenase activities, thereby inhibiting d-xylose utilization in Pachysolen tannophilus. The question arose whether d-glucose can also inactivate d-xylose fermentation. P. tannophilus cells were grown on a defined d-xylose-containing liquid medium. At about 40 h, d-glucose was added to a final concentration of 3% (w/v). This led to a rapid cessation of d-xylose utilization, which resumed after 10–12 h before d-glucose was completely consumed. This suggests that d-glucose inactivated existing d-xylose catabolic enzymes and that inactivation was reversed at low d-glucose concentrations. This reversible inactivation was distinct from d-glucose repression. Addition of cycloheximide did not block the resumption of d-xylose consumption, suggesting that reactivation was independent of protein synthesis.
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plasma membrane mg 2 atpase of Pachysolen tannophilus characterization and role in alcohol tolerance
Applied and Environmental Microbiology, 1991Co-Authors: Maria F. S. Barbosa, Hung LeeAbstract:Following cell fractionation in sucrose density gradients, plasma membrane Mg(2+)-ATPase from Pachysolen tannophilus was studied. The ATPase displayed an apparent Km for ATP of 1.42 mM and was inhibited by high concentrations of Mg2+. The inhibitory effects of ethanol, 1-propanol, 1-butanol, and benzyl alcohol on Mg(2+)-ATPase were evaluated, and the concentration of each alcohol that inhibited ATPase activity by 50% (IC50) was determined. The IC50 decreased as the chain length of the alcohol increased. Moreover, the IC50 for ATPase activity was similar to the IC50 for growth rate, suggesting an association between impaired growth and ATPase inhibition. Almost complete inhibition of ATPase activity occurred at temperatures approaching 60 degrees C, and the optimal temperature was around 44 degrees C for ATPase from both control and ethanol-treated cells. Inclusion of 50 mM MgCl2 or CaCl2 in the medium did not rescue cells from the deleterious effects of ethanol.
Vicente Bravo - One of the best experts on this subject based on the ideXlab platform.
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fermentation of enzymatic hydrolysates from olive stones by Pachysolen tannophilus
Journal of Chemical Technology & Biotechnology, 2009Co-Authors: Manuel Cuevas, Sebastián Sánchez, Vicente Bravo, Nicolas Cruz, Juan Godoy F GarciaAbstract:BACKGROUND: Olive stones were pretreated with liquid hot water (LHW or autohydrolysis) at maximum temperatures between 175 and 225 °C (severity factors, logR0, between 2.73 and 4.39) to be subjected (both liquid and solid components) afterwards to enzymatic hydrolysis with cellulases from Trichoderma viride. Ethanol fermentation of hydrolysates was performed with the non-traditional yeast Pachysolen tannophilus ATCC 32691. RESULTS: After the enzymatic step, yields of hemicellulose solubilization reached 100%, while the cellulose was only partially hydrolysed (23%, logR0 = 4.39). The maximum yields in total reducing sugars and acetic acid, at the upper end of the severity range, was close to 0.25 and 0.04 g g−1 dry stone, respectively. During the fermentation stage, the increase in R0 reduced the maximum specific growth rate, biomass productivity, and overall biomass yield. The overall yields of ethanol and xylitol ranged, respectively, from 0.18 to 0.25 g g−1 and from 0.01 to 0.13 g g−1. CONCLUSIONS: The results demonstrate the possibility of producing ethanol from olive stones, making use of the cellulose and hemicellulose fraction of the waste. It was confirmed that the overall yield in xylitol strongly depended on severity factor, while the overall yield in ethanol remained practically constant for all the pretreatment conditions tested. Copyright © 2008 Society of Chemical Industry
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fermentation of acid hydrolysates from olive tree pruning debris by Pachysolen tannophilus
Bioprocess and Biosystems Engineering, 2008Co-Authors: Alberto J Moya, Vicente Bravo, Soledad Mateo, Sebastián SánchezAbstract:The influence of the type and concentration of acid in the hydrolysis process and its effect on the subsequent fermentation by Pachysolen tannophilus (ATCC 32691) to produce ethanol and xylitol was studied. The hydrolysis experiments were performed using hydrochloric, sulphuric and trifluoroacetic acids in concentrations ranging from 0.1 to 1.0 N, a temperature of 90 °C, and a time of 240 min. The fermentation experiments were conducted on a laboratory scale in a batch-culture reactor at pH 4.5 and 30 °C. The hydrolysis with the highest acid concentration produced the complete solubilization of hemicellulose to monosaccharides. The highest values for the specific rate of ethanol production were registered in cultures hydrolyzed with trifluoroacetic acid, and values were found to decrease as the acid concentration increased. The highest values of overall ethanol yields ( $$ Y^{{\text{G}}}_{{{\text{E/s}}}} $$ = 0.37 kg kg−1) were also found in the fermentation of the hydrolysates of trifluoroacetic acid.
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fermentation of olive tree pruning acid hydrolysates by Pachysolen tannophilus
Biochemical Engineering Journal, 2007Co-Authors: Inmaculada Romero, Sebastián Sánchez, Eulogio Castro, Manuel Moya, Encarnacion Ruiz, Vicente BravoAbstract:Abstract The influence of hydrolysis conditions on the fermentation of hydrolysates obtained from olive tree pruning (a renewable, low cost, largely available agricultural residue) is analyzed in this work. Hydrolysis was performed using sulfuric acid at atmospheric pressure (90 °C) in a concentration range of 0.5–4N for 240 min. The fermentation of hydrolysates was carried out by Pachysolen tannophilus in a discontinuous tank bioreactor at 30 °C and pH 3.5. The main fermentation parameters determined in this study include maximum specific growth rate, biomass productivity, specific substrate consumption rate, specific ethanol production rate, and ethanol and xylitol yields. The results show that ethanol yields are much higher than xylitol yields under all the conditions tested. The maximum ethanol yield (0.38 g/g) is reached with the hydrolysate obtained with 0.75N sulfuric acid. Under these conditions the conversion of the hemicellulose fraction is 92%.
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the fermentation of mixtures of d glucose and d xylose by candida shehatae pichia stipitis or Pachysolen tannophilus to produce ethanol
Journal of Chemical Technology & Biotechnology, 2002Co-Authors: Sebastián Sánchez, Vicente Bravo, Eulogio Castro, Alberto J Moya, F CamachoAbstract:The fermentation of mixtures of D-glucose and D-xylose by three non-traditional yeasts: Candida shehatae (ATCC 34887), Pachysolen tannophilus (ATCC 32691) and Pichia stipitis (ATCC 58376) have been studied to determine the optimal strain and initial culture conditions for the efficient production of ethanol. The comparison was made on the basis of maximum specific growth rate (µm), biomass productivity, the specific rates of total substrate consumption (qs) and ethanol production (qE) and the overall yields of ethanol and xylitol. All the experiments were performed in stirred-tank batch reactors at a temperature of 30 °C. The initial pH of the culture medium was 4.5. The highest values of µm (above 0.5 h−1) were obtained with P stipitis in cultures containing high concentrations of D-xylose. All three yeasts consumed the two monosaccharides in sequence, beginning with D-glucose. The values of qs diminished during the course of each experiment with all of the yeasts. The highest values of the specific rates of total substrate consumption and ethanol production were obtained with C shehatae (for t = 10 h, qs and qE were above 5 g g−1 h−1 and 2 g g−1 h−1, respectively), although the highest overall ethanol yields were fairly similar with all three yeasts, at around 0.4 g g−1. © 2002 Society of Chemical Industry
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comparative study of the fermentation of d glucose d xylose mixtures with Pachysolen tannophilus and candida shehatae
Bioprocess Engineering, 1999Co-Authors: Sebastián Sánchez, Vicente Bravo, Eulogio Castro, Alberto J Moya, F CamachoAbstract:We have performed a comparative analysis of the fermentation of the solutions of the mixtures of D-glucose and D-xylose with the yeasts Pachysolen tannophilus (ATCC 32691) and Candida shehatae (ATCC 34887), with the aim of producing bioethanol. All the experiments were performed in a batch bioreactor, with a constant aeration level, temperature of 30 °C, and a culture medium with an initial pH of 4.5. For both yeasts, the comparison was established on the basis of the following parameters: maximum specific growth rate, biomass productivity, specific rate of substrate consumption (qs) and of ethanol production (qE), and overall ethanol and xylitol yields. For the calculation of the specific rates of substrate consumption and ethanol production, differential and integral methods were applied to the kinetic data. From the experimental results, it is deduced that both Candida and Pachysolen sequentially consume the two substrates, first D-glucose and then D-xylose. In both yeasts, the specific substrate-consumption rate diminished over each culture. The values qs and qE proved higher in Candida, although the higher ethanol yield was of the same order for both yeasts, close to 0.4 kg kg−1.
C.a. García-burgos - One of the best experts on this subject based on the ideXlab platform.
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Immobilised derivatives of Williopsis californica, Williopsis saturnus, Pachysolen tannophilus: New biocatalysts useful in the stereoselective oxidation of 1-tetralol
Enzyme and Microbial Technology, 2006Co-Authors: J.d. Carballeira, C.a. García-burgos, M.a. Quezada, E. Álvarez, J.v. Sinisterra-gagoAbstract:Abstract Four new whole cell biocatalysts have been selected after a high throughput screening of microorganisms from public collections: Williopsis californica CBS 2158 , Williopsis saturnus NCYC 2313, Pachysolen tannophilus NCYC 1597 and Coniochaeta velutina CBS 981.68, in basis to the high yield and stereoselectivity in the oxidation of 1-tetralol. These microorganisms were immobilised in different supports by entrapment techniques, being especially remarkable the results obtained with different tailor made agar matrix from Hispanagar (Spain). The selection of the immobilization conditions for each strain was performed following a statistical design based in the response surface methodology. The immobilised biocatalysts – Williopsis californica and Williopsis saturnus – immobilised in tailor-made agar (2.5%) from Pterocladia and Gracilaria (Hispanagar S.A.) carry to 100% yield in the oxidation of cyclohexanol. The immobilised biocatalysts displayed a high S -stereoselectivity (ee > 98% and 50% yield) in the oxidation of ( R , S )-1-tetralol and a high yield in the oxidation of bulky substrates as 2-adamantanol (100%). The kinetic studies showed that the diffusion of reagents/products is the rate controlling step. The addition of glucose (0.5%) increases the oxidation yields.
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Williopsis californica, Williopsis saturnus, and Pachysolen tannophilus: novel microorganisms for stereoselective oxidation of secondary alcohols.
Biotechnology and bioengineering, 2004Co-Authors: J. D. Carballeira Rodríguez, C.a. García-burgos, M. A. Quezada Alvarez, E. Alvarez Ruiz, J. V. Sinisterra GagoAbstract:A screening of 416 microorganisms from different taxonomical groups (bacteria, actinomycetes, yeasts, and filamentous fungi) has been performed looking for active strains in the stereoselective oxidation of secondary alcohols. The working collection was composed of 71 bacterial strains, 45 actinomycetes, 59 yeasts, 60 basidiomycetes, 33 marine fungi, and 148 filamentous fungi. All microorganisms selected were mesophilic. Yeasts were the most active microbial group in the whole-cell-catalyzed oxidation. Williopsis californica, Williopsis saturnus, and Pachysolen tannophilus were the strains of greatest interest, both as growing cells and as resting cells. The oxidation of the alcohols takes place when cells are in the stationary growth phase (after 48 h of culture). These three strains are S-stereoselective for the oxidation of racemic secondary alkanols and show stereospecificity in the oxidation of menthol or neo-menthol, whereas iso-menthol is not oxidized. In the case of the 1-tetrahydronaphtol enantiomers, only the S-enantiomer is oxidized. The three strains were immobilized by entrapment using agarose and agar from algae of the Gracilaria genus. The agarose derivatives displayed significant improvement in the stereospecificity of the reactions.
J.v. Sinisterra-gago - One of the best experts on this subject based on the ideXlab platform.
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Immobilised derivatives of Williopsis californica, Williopsis saturnus, Pachysolen tannophilus: New biocatalysts useful in the stereoselective oxidation of 1-tetralol
Enzyme and Microbial Technology, 2006Co-Authors: J.d. Carballeira, C.a. García-burgos, M.a. Quezada, E. Álvarez, J.v. Sinisterra-gagoAbstract:Abstract Four new whole cell biocatalysts have been selected after a high throughput screening of microorganisms from public collections: Williopsis californica CBS 2158 , Williopsis saturnus NCYC 2313, Pachysolen tannophilus NCYC 1597 and Coniochaeta velutina CBS 981.68, in basis to the high yield and stereoselectivity in the oxidation of 1-tetralol. These microorganisms were immobilised in different supports by entrapment techniques, being especially remarkable the results obtained with different tailor made agar matrix from Hispanagar (Spain). The selection of the immobilization conditions for each strain was performed following a statistical design based in the response surface methodology. The immobilised biocatalysts – Williopsis californica and Williopsis saturnus – immobilised in tailor-made agar (2.5%) from Pterocladia and Gracilaria (Hispanagar S.A.) carry to 100% yield in the oxidation of cyclohexanol. The immobilised biocatalysts displayed a high S -stereoselectivity (ee > 98% and 50% yield) in the oxidation of ( R , S )-1-tetralol and a high yield in the oxidation of bulky substrates as 2-adamantanol (100%). The kinetic studies showed that the diffusion of reagents/products is the rate controlling step. The addition of glucose (0.5%) increases the oxidation yields.
