Xylitol Production Research Articles

Construction and co-expression of plasmid encoding xylitol dehydrogenase and a cofactor regeneration enzyme for the production of xylitol from d-arabitol

The biotransformation of d-arabitol into xylitol was investigated with focus on the conversion of d-xylulose into xylitol. This critical conversion was accomplished using Escherichia coli to co-express a xylitol dehydrogenase gene from Gluconobacter oxydans and a cofactor regeneration enzyme gene which was a glucose dehydrogenase gene from Bacillus subtilis for system 1 and an alcohol dehydrogenase gene from G. oxydans for system 2. Both systems efficiently converted d-xylulose into xylitol without the addition of expensive NADH. Approximately 26.91g/L xylitol was obtained from around 30g/L d-xylulose within system 1 (E. coli Rosetta/Duet-xdh-gdh), with a 92% conversion yield, somewhat higher than that of system 2 (E. coli Rosetta/Duet-xdh-adh, 24.9g/L, 85.2%). The xylitol yields for both systems were more than 3-fold higher compared to that of the G. oxydans NH-10 cells (7.32g/L). The total turnover number (TTN), defined as the number of moles of xylitol formed per mole of NAD+, was 32,100 for system 1 and 17,600 for system 2. Compared with that of G. oxydans NH-10, the TTN increased by 21-fold for system 1 and 11-fold for system 2, hence, the co-expression systems greatly enhanced the NADH supply for the conversion, benefiting the practical synthesis of xylitol.

Enhanced Production of Bioxylitol from Corn Cobs by Candida magnoliae

An effective microbial process using Candida magnoliae was developed for xylitol production from xylose obtained from corn cob. The hydrolysate containing approximately 25 g-xylose/L and 3.0 g-gluc...

Production of xylitol and ethanol by Hansenula polymorpha from hydrolysates of sunflower stalks with phosphoric acid

This research analyses the effect exerted by the phosphoric acid concentration (CAo) in sunflower-stalk hydrolysis at 95°C, considering the results in relation to the yields in d-glucose and total reducing sugars (TRS). Besides to display the behavior of the yeast in generating bioalcohols from hydrolysates rich in nutrients such as phosphate, essential for the growth of yeasts in fermentation processes. The results show that the hydrolysis of the hemicellulose fraction was almost complete at CAo=2.67mol/L (25.3%), at which conversion reached 82.7%. The percentage of converted cellulose also increased with the acid concentration, reaching a value of 13.9% in the hydrolysis process of 2.67mol/L. The hydrolysis rate (rH) was calculated from the values of the concentration of total reducing sugars generated over the process. For a value of CAo=1.67mol/L, it was determined that (rH)max=3.5g TRS/(Lh) at the beginning of the hydrolysis process. After being detoxified, the hydrolysates were fermented with Hansenula polymorpha, ATCC 34438, registering better results in the production of ethanol that of xylitol. For an initial concentration of total reducing sugars of 13.3g/L, xylitol yield was 0.023g/g, and ethanol yield was 0.14g/g.

Evaluation of antioxidant activity of dilute acid hydrolysate of wheat straw during xylose production

Wheat straw, a lignocellulosic waste material, can be used as a raw material for the production of high-value products such as xylose for xylitol production or phenolic compounds that have antioxidant activity. There is a growing interest in the use of lignocellulosic wastes for conversion into various chemicals because of their low cost and the fact that they are renewable and abundant. The objective of this study was to determine the effects of H2SO4 concentration, temperature and reaction time on the production of sugars (xylose) and phenolic compounds. Response surface methodology (RSM) was used to optimize the hydrolysis process to obtain high xylose yield and phenolic compounds. The optimum reaction temperature, reaction time and acid concentration were 120°C, 45min and 4.7% respectively. Under these conditions, xylose and phenolic of the hydrolysate was found to be 0.16g/g-wheat straw and 0.014g-gallic acid/g-wheat straw, respectively.

Influence of the use of rice bran extract as a source of nutrients on xylitol production

Xylose-to-xylitol bioconversion using 2.5 or 10% (v/v) rice bran extract was performed to verify the influence of this source of nutrients on Candida guilliermondii metabolism. Semisynthetic medium (SM) and sugarcane bagasse hemicellulosic hydrolysate detoxified with ion-exchange resins (HIE) or with alteration in pH combined with adsorption onto activated charcoal (HAC) were fermented in 125 mL Erlenmeyer flasks at 30 ºC and 200 rpm for 72 hours. Activated charcoal supplemented with 2.5% (v/v) rice bran extract was fermented by C. guilliermondii in a MULTIGEN stirred tank reactor using pH 5.0 and 22.9/hour oxygen transfer volumetric coefficient. Higher values of xylitol productivity (0.70, 0.71, and 0.62 g.Lh-1) and xylose-to-xylitol conversion yield (0.71, 0.69, and 0.63 g.g-1) were obtained with 2.5% (v/v) rice bran in semisynthetic medium, ion-exchange resins, and activated charcoal, respectively. Moreover, during batch fermentation, the xylitol volumetric productivity and fermentation efficiency values obtained were 0.53 g.Lh-1 and 61.1%, respectively.

Exploiting the inter-strain divergence of Fusarium oxysporum for microbial bioprocessing of lignocellulose to bioethanol

Microbial bioprocessing of lignocellulose to bioethanol still poses challenges in terms of substrate catabolism. A targeted evolution-based study was undertaken to determine if inter-strain microbial variability could be exploited for bioprocessing of lignocellulose to bioethanol. The microorganism studied was Fusarium oxysporum because of its capacity to both saccharify and ferment lignocellulose. Strains of F. oxysporum were isolated and assessed for their genetic variability. Using optimised solid-state straw culture conditions, experiments were conducted that compared fungal strains in terms of their growth, enzyme activities (cellulases, xylanase and alcohol dehydrogenase) and yield of bioethanol and the undesirable by-products acetic acid and xylitol. Significant inter-strain divergence was recorded in regards to the capacity of studied F. oxysporum strains to produce alcohol from untreated straw. No correlation was observed between bioethanol synthesis and either the biomass production or microbial enzyme activity. A strong correlation was observed between both acetic acid and xylitol production and bioethanol yield. The level of diversity recorded in the alcohol production capacity among closely-related microorganism means that a targeted screening of populations of selected microbial species could greatly improve bioprocessing yields, in terms of providing both new host strains and candidate genes for the bioethanol industry.

Metabolic engineering of Saccharomyces cerevisiae for bioconversion of d-xylose to d-xylonate

An NAD+-dependent d-xylose dehydrogenase, XylB, from Caulobacter crescentus was expressed in Saccharomyces cerevisiae, resulting in production of 17±2g d-xylonate l−1 at 0.23gl−1h−1 from 23g d-xylose l−1 (with glucose and ethanol as co-substrates). d-Xylonate titre and production rate were increased and xylitol production decreased, compared to strains expressing genes encoding T. reesei or pig liver NADP+-dependent d-xylose dehydrogenases. d-Xylonate accumulated intracellularly to ∼70mgg−1; xylitol to ∼18mgg−1. The aldose reductase encoding gene GRE3 was deleted to reduce xylitol production. Cells expressing d-xylonolactone lactonase xylC from C. crescentus with xylB initially produced more extracellular d-xylonate than cells lacking xylC at both pH 5.5 and pH 3, and sustained higher production at pH 3. Cell vitality and viability decreased during d-xylonate production at pH 3.0. An industrial S. cerevisiae strain expressing xylB efficiently produced 43g d-xylonate l−1 from 49g d-xylose l−1.

Efficient utilization of hemicellulose hydrolysate for propionic acid production using Propionibacterium acidipropionici

Hemicellulose, which contains glucose, xylose, and arabinose as the 3 main sugars, is an important renewable source for biorefinery. In this study, propionic acid production from glucose, xylose, or arabinose using Propionibacterium acidipropionici ATCC 4875 was investigated. Using xylose, the predominant sugar in hemicellulose, a final propionic acid concentration of 53.2gl–1 was obtained via fed-batch fermentation. Using corncob molasses, a waste by-product from xylitol production as a representative of hemicellulose hydrolysate, the final concentration of propionic acid was 71.8gl–1, with a corresponding productivity of 0.28gl–1h–1. The present study suggests that hemicellulose hydrolysate is an excellent carbon source for efficient propionic acid production by this strain.

MIXTURE OF XYLOSE AND GLUCOSE AFFECTS XYLITOL PRODUCTION BY PICHIA GUILLIERMONDII: MODEL PREDICTION USING ARTIFICIAL NEURAL NETWORK

Production of several yeast products occur in presence of mixtures of monosaccharides. To study effect of xylose and glucose mixtures with system aeration and nitrogen source as the other two operative variables on xylitol production by Pichia guilliermondii, the present work was defined. Artificial Neural Network (ANN) strategy was used to athematically show interplay between these three controllable factors and the xylitol productivity response. In the first stage, model fitting was performed using Response Surface Methodology (RSM) and the appropriate fraction of this design then was applied for the ANN training step (Levenberg Marquardt ‘LM’ algorithm). The best ANN model configuration with the three test input variables composed of six neurons in the hidden layer and tangent sigmoid (TANSIG) and linear transfer function (PURELIN) were used as the activation functions for the data processing from inputs to the hidden layer and from the constructed neurons to the output nodes. The network performance was evaluated by Mean Squared Error (MSE) and the regression coefficient of determination (R2). These values respectively, for the RSM model fitting were 2.327× 10-4 and 0.9817, and for the ANN training data were 2.29 × 10-8 and 0.9999. While MSE and R2 values for the other two steps of ANN were 4.56 × 10-3 and 0.9741 (validating step) and1.52× 10-3 and 0.9325 (testing step), respectively. Positive synergism of ANN with RSM was confirmed.

Improved Ethanol and Reduced Xylitol Production from Glucose and Xylose Mixtures by the Mutant Strain of Candida shehatae ATCC 22984

A mutant Cs3512, which showed better fermentation of xylose and the mixtures of xylose and glucose, was obtained through mutation of Candida shehatae ATCC 22984 and screening with a medium containing antimycin A and TTC (2,3,5-triphenyltetrazolium chloride). Cs3512 produced 44.4g/l of ethanol from 121.3g/l of xylose, which was 13% higher than that by ATCC 22984. At the same time, xylitol production was reduced by 38% to 10.2g/l from 16.3g/l by ATCC 22984. Cs3512 also showed 8% increase in ethanol yield from 0.39 to 0.42g/g comparing to ATCC 22984 when fermenting the sugar mixture composed of 52.9g/l glucose and 21.2g/l xylose. When Cs3512 was used in the simultaneous saccharification and fermentation of lime pretreated rice straw via CaCCO (calcium capturing by carbonation) process, it produced ethanol at 77% of the theoretical yield. The results imply that Cs3512 is a potential non-recombinant yeast strain for ethanol production from lignocellulosic biomass.

Host cell and expression engineering for development of an E. coli ketoreductase catalyst: Enhancement of formate dehydrogenase activity for regeneration of NADH

BackgroundEnzymatic NADH or NADPH-dependent reduction is a widely applied approach for the synthesis of optically active organic compounds. The overall biocatalytic conversion usually involves in situ regeneration of the expensive NAD(P)H. Oxidation of formate to carbon dioxide, catalyzed by formate dehydrogenase (EC 1.2.1.2; FDH), presents an almost ideal process solution for coenzyme regeneration that has been well established for NADH. Because isolated FDH is relatively unstable under a range of process conditions, whole cells often constitute the preferred form of the biocatalyst, combining the advantage of enzyme protection in the cellular environment with ease of enzyme production. However, the most prominent FDH used in biotransformations, the enzyme from the yeast Candida boidinii, is usually expressed in limiting amounts of activity in the prime host for whole cell biocatalysis, Escherichia coli. We therefore performed expression engineering with the aim of enhancing FDH activity in an E. coli ketoreductase catalyst. The benefit resulting from improved NADH regeneration capacity is demonstrated in two transformations of technological relevance: xylose conversion into xylitol, and synthesis of (S)-1-(2-chlorophenyl)ethanol from o-chloroacetophenone.ResultsAs compared to individual expression of C. boidinii FDH in E. coli BL21 (DE3) that gave an intracellular enzyme activity of 400 units/gCDW, co-expression of the FDH with the ketoreductase (Candida tenuis xylose reductase; XR) resulted in a substantial decline in FDH activity. The remaining FDH activity of only 85 U/gCDW was strongly limiting the overall catalytic activity of the whole cell system. Combined effects from increase in FDH gene copy number, supply of rare tRNAs in a Rosetta strain of E. coli, dampened expression of the ketoreductase, and induction at low temperature (18°C) brought up the FDH activity threefold to a level of 250 U/gCDW while reducing the XR activity by just 19% (1140 U/gCDW). The E. coli whole-cell catalyst optimized for intracellular FDH activity showed improved performance in the synthesis of (S)-1-(2-chlorophenyl)ethanol, reflected in a substantial, up to 5-fold enhancement of productivity (0.37 g/gCDW) and yield (95% based on 100 mM ketone used) as compared to the reference catalyst. For xylitol production, the benefit of enhanced FDH expression was observed on productivity only after elimination of the mass transfer resistance caused by the cell membrane.ConclusionsExpression engineering of C. boidinii FDH is an important strategy to optimize E. coli whole-cell reductase catalysts that employ intracellular formate oxidation for regeneration of NADH. Increased FDH-activity was reflected by higher reduction yields of D-xylose and o-chloroacetophenone conversions provided that mass transfer limitations were overcome.

A Statistical Approach to Optimize Xylitol Production by <i>Debaryomyces nepalensis</i> NCYC 3413 <i>in Vitro</i>

Debaryomyces nepalensis NCYC 3413, halotolerant yeast isolated from rotten apple, was capable of utilizing components of hemicellulose hydrolysate such as glucose, galactose, mannose, xylose and arabinose. The organism utilizes xylose as a sole carbon source and produces xylitol. The Plackett-Burman design was applied to determine the specific medium components affecting xylitol production and found that xylose, K2HPO4, and ZnSO4 were critical in augmenting xylitol production. These significant parameters were further optimized using response surface methodology. The optimum concentrations of xylose, K2HPO4, and ZnSO4 were found to be 100 g/l, 10.6 g/l and 8.9 mg/l respectively. Under these optimal conditions the xylitol production increased from 27 g/l to 36 g/l with a yield of 0.44 g/g (57% increase in total yield). In addition, formation of the by product (glycerol) was decreased under optimal conditions.

Xylitol production from d-xylose and horticultural waste hemicellulosic hydrolysate by a new isolate of Candida athensensis SB18

This paper describes the production of xylitol from d-xylose and horticultural waste hemicellulosic hydrolysate by a new strain of Candida athensensis SB18. Strain SB18 completely consumed 250 and 300 g L −1 d-xylose and successful converted it to xylitol in the respective yield of 0.83 and 0.87 g g −1, resulting in 207.8 and 256.5 g L −1 of xylitol, respectively. The respective volumetric productivity were 1.15 and 0.97 g L −1 h −1. Approximately 100.1 g L −1 of xylitol was obtained from the bioconversion of detoxified horticultural waste hemicellulosic hydrolysate using strain SB18. The yield and productivity were 0.81 g g −1 xylose and 0.98 g L −1 h −1, respectively. Strain C. athensensis SB18 was able to completely utilize glucose, mannose, xylose and partially arabinose. This work demonstrates that stain C. athensensis SB18 is a promising strain for high-titer and high-yield xylitol production and it has great potential in bioconversion of hemicellulosic hydrolysate.

Acid hydrolysis of wheat straw: A kinetic study

Abstract Biotechnological xylitol production can be enhanced if the needed xylose solutions can be obtained from hydrolysis of low-cost lignocellulosic wastes. The hydrolysis of wheat straw to obtain xylose solutions has a double consequence, the elimination of a waste and the generation of a value-added product. The objective of this work was to study the xylose production from wheat straw by sulphuric acid hydrolysis at 130 °C. Several mass fraction of acid (1, 2, 3, 4 or 5%) were evaluated. Kinetic models were developed to explain the variation with time of xylose, glucose, arabinose, furfural, 5-(hydroxymethyl)-2-furaldehyde and acetic acid in the hydrolysates. Optimal conditions found were a H2SO4 mass fraction of 2% at 130 °C for 29 min, which yielded a solution with xylose, 18.9 kg m−3; glucose, 3.5 kg m−3; arabinose, 3.1 kg m−3; furfural, 0.6 kg m−3; 5-(hydroxymethyl)-2-furaldehyde, 0.3 kg m−3 and acetic acid, 2.3 kg m−3. In these conditions, 99% of the hemicelluloses and 11% of the glucan were hydrolysed.

Detoxification of Wheat Straw Formic Acid Hydrolysis and Xylitol Production

Xylitol can be obtained from hemicelullosic fraction of lignocellulosic materials containing D-xylose. Formic acid hydrolysis is widely used in lignocellulose pretreatment. However, formic acid hydrolysis wheat straw cannot be directly used as fermentation subsequently owing to various fermentation inhibitors, especially the formic acid in reaction system and released during pretreatment. This study describes main hydrolysis components and inhibitors prepared by hydrolysis of wheat straw with formic acid solution, and different detoxification methods ware used to romve the inhibitors. Study found treatment of wheat straw hydrolysate with overliming treatment and D311 ion-exchange resin had a good result on reduction formic acid and other main inhibitors, results showed which can eliminate 94% of residual formic acid and with only 15% reducion of xylose. To verify the effectiveness of different detoxification methods, hydrolysates of detoxification werr fermented by Candida.tropicalis AS2.1776, and the results also found that the hydrolysate treatmented with overliming treatment and D311 ion-exchange resin have the hightest xylitol yield.

Kinetic behavior of Candida tropicalis during xylitol production using semi-synthetic and hydrolysate based media

The effects of the xylose concentration and the air supply on the production of xylitol from D-xylose by the yeast Candida tropicalis (AY 2007) using semi-synthetic and hydrolysate based media were investigated in four fermentation batch runs. The xylitol formation was favored under oxygen-limited conditions (2nd experiment, air flow rate 0.3 vvm) with a maximum xylitol production (36.0gm/l) after 59h as compared with that obtained in the 1st experiment (30.99 g/L) after 66 h under relatively less oxygen limited condition (air flow rate 1.5 vvm). Increasing the initial xylose concentration from 45.5 g/L in the 2nd experiment to 96.9 g/L in the 3rdexperiment led to a great decrease in the xylitol accumulation rate (Qp) from 0.506 to 0.134 g/L, xylitol yield coefficient (Yp/s) from 0.704 to 0.180 g/g and a mild increase in the xylose consumption rate (Qs) from 0.70 to 0.75 g/L/h. The semi-synthetic medium (1st and 2nd experiments) had a maximum xylitol yield coefficient (Yp/s) of 0.701 and 0.704 g/g, respectively which was less than that obtained in the hydrolysate experiment (0.783 g/g). This is probably due to the low hydrolysate initial xylose content (29.8 g/L). Key words: Xylitol, Candida tropicalis, xylose, pH, aeration, kinetics.

Kinetic behavior of Candida tropicalis during xylitol production using semi-synthetic and hydrolysate based media

The effects of the xylose concentration and the air supply on the production of xylitol from D-xylose by the yeast Candida tropicalis (AY 2007) using semi-synthetic and hydrolysate based media were investigated in four fermentation batch runs. The xylitol formation was favored under oxygen-limited conditions (2nd experiment, air flow rate 0.3 vvm) with a maximum xylitol production (36.0gm/l) after 59h as compared with that obtained in the 1st experiment (30.99 g/L) after 66 h under relatively less oxygen limited condition (air flow rate 1.5 vvm). Increasing the initial xylose concentration from 45.5 g/L in the 2nd experiment to 96.9 g/L in the 3rdexperiment led to a great decrease in the xylitol accumulation rate (Qp) from 0.506 to 0.134 g/L, xylitol yield coefficient (Yp/s) from 0.704 to 0.180 g/g and a mild increase in the xylose consumption rate (Qs) from 0.70 to 0.75 g/L/h. The semi-synthetic medium (1st and 2nd experiments) had a maximum xylitol yield coefficient (Yp/s) of 0.701 and 0.704 g/g, respectively which was less than that obtained in the hydrolysate experiment (0.783 g/g). This is probably due to the low hydrolysate initial xylose content (29.8 g/L). Key words: Xylitol, Candida tropicalis, xylose, pH, aeration, kinetics.

Xylitol production by Candida tropicalis from corn cob hemicellulose hydrolysate in a two‐stage fed‐batch fermentation process

AbstractBACKGROUND: Xylitol, a sugar alcohol widely used in food and pharmaceutical industries, can be produced through biological reduction of xylose present in hemicellulose hydrolysates by Candida tropicalis. However, the aeration rate and by‐products originating from hemicellulose hydrolysis strongly inhibit the production of xylitol in a fermentation process. A two‐stage fed‐batch fermentation system was developed to reduce these inhibitory effects and to improve xylitol production from corn cob hemicellulose hydrolysates by C. tropicalis.RESULTS: Results of batch fermentations indicated that high xylitol production could be obtained from C. tropicalis at an initial xylose concentration of 80 g L−1 in corn cob hydrolysate medium at an aeration rate of 0.4 vvm at the micro‐aeration stage. In the two‐stage fed‐batch fermentation process, 96.5 g L−1 xylitol was obtained after 120 h, giving a yield of 0.83 g g−1 and a productivity of 1.01 g L−1 h−1, which were 12.16% and 65.57% higher than those in a batch fermentation.CONCLUSION: High xylitol production can be achieved in a two‐stage fed‐batch fermentation process, in which the negative effects of aeration rate and inhibitory compounds on xylitol formation can be considerably reduced. Copyright © 2011 Society of Chemical Industry

Xylitol production by Candida tropicalis under different statistically optimized growth conditions

Nutritional and environmental conditions of the xylose utilizing yeast Candida tropicalis were optimized on a shake-flask scale using a statistical factorial design to maximize the production of xylitol. Effects of the three growth medium components (rice bran, ammonium sulfate and xylose) on the xylitol production were studied, each at three different levels and the highest xylose to xylitol conversion ratio (57.2%) was achieved on using 20 gl-1 of xylose, 15 gl-1 of rice bran and 1 gl-1 of ammonium sulfate. By maintaining the pH of the growth medium at 5.5 using citrate buffer, the xylose to xylitol conversion ratio was greatly enhanced (93.75%) over the use of or unbuffered growth medium and phosphate buffered (88.3 and 83.8%).The least aeration rate was simulated by 40 ml of the growth medium at 250 ml-1. Erlenmeyer flasks result in maximum xylose to xylitol conversion ratio (98%), by using 25 gl-1 of xylose, while maximum xylitol production (36.25 gl-1) with a conversion ratio of 72.5% was achieved by using 50 gl-1 of xylose. Key words: xylitol, Candida tropicalis, xylose, yeast, pH, aeration.

Xylitol production by Candida tropicalis under different statistically optimized growth conditions

Nutritional and environmental conditions of the xylose utilizing yeast Candida tropicalis were optimized on a shake-flask scale using a statistical factorial design to maximize the production of xylitol. Effects of the three growth medium components (rice bran, ammonium sulfate and xylose) on the xylitol production were studied, each at three different levels and the highest xylose to xylitol conversion ratio (57.2%) was achieved on using 20 gl-1 of xylose, 15 gl-1 of rice bran and 1 gl-1 of ammonium sulfate. By maintaining the pH of the growth medium at 5.5 using citrate buffer, the xylose to xylitol conversion ratio was greatly enhanced (93.75%) over the use of or unbuffered growth medium and phosphate buffered (88.3 and 83.8%).The least aeration rate was simulated by 40 ml of the growth medium at 250 ml-1. Erlenmeyer flasks result in maximum xylose to xylitol conversion ratio (98%), by using 25 gl-1 of xylose, while maximum xylitol production (36.25 gl-1) with a conversion ratio of 72.5% was achieved by using 50 gl-1 of xylose. Key words: xylitol, Candida tropicalis, xylose, yeast, pH, aeration.