Biosynthetic strategies to produce xylitol: an economical venture.

Xylitol is a natural five-carbon sugar alcohol with potential for use in food and pharmaceutical industries owing to its insulin-independent metabolic regulation, tooth rehardening, anti-carcinogenic, and anti-inflammatory, as well as osteoporosis and ear infections preventing activities. Chemical and biosynthetic routes using D-xylose, glucose, or biomass hydrolysate as raw materials can produce xylitol. Among these methods, microbial production of xylitol has received significant attention due to its wide substrate availability, easy to operate, and eco-friendly nature, in contrast with high-energy consuming and environmental-polluting chemical method. Though great advances have been made in recent years for the biosynthesis of xylitol from xylose, glucose, and biomass hydrolysate, and the yield and productivity of xylitol are substantially improved by metabolic engineering and optimizing key metabolic pathway parameters, it is still far away from industrial-scale biosynthesis of xylitol. In contrary, the chemical synthesis of xylitol from xylose remains the dominant route. Economic and highly efficient xylitol biosynthetic strategies from an abundantly available raw material (i.e., glucose) by engineered microorganisms are on the hard way to forwarding. However, synthetic biology appears as a novel and promising approach to develop a super yeast strain for industrial production of xylitol from glucose. After a brief overview of chemical-based xylitol production, we critically analyzed and comprehensively summarized the major metabolic strategies used for the enhanced biosynthesis of xylitol in this review. Towards the end, the study is wrapped up with current challenges, concluding remarks, and future prospects for designing an industrial yeast strain for xylitol biosynthesis from glucose.

Xylitol production and furfural consumption by a wild type Geotrichum sp

Background: Xylitol is a five carbons polyol with promising medical applications. It can be obtained from chemical D-xylose reduction or by microbial fermentation of Sugarcane Bagasse Hemicellulosic Hydrolysate. For this last process, some microbial inhibitors, as furfural, constitute severe bottleneck. In this case, the use of strains able to produce xylitol simultaneously to furfural neutralization is an interesting alternative. A wild-type strain of Geotrichum sp. was detected with this ability, and its performance in xylitol production and furfural consumption was evaluated. Furthermore, were analyzed its degradation products. Results: Geotrichum sp. produced xylitol from D-xylose fermentation with a yield of 0.44 g-g-1. Furfural was fully consumed in fermentation assay and when provided in the medium until concentration of 6 g-L-1. The furfural degradation product is not an identified molecule, presenting a molecular weight of 161 g-mol-1, an uncommon feature for the microbial metabolism of this product. Conclusion: This strain presents most remarkable potential in performing furfural consumption simultaneous to xylitol production. Subsequent efforts must be employed to establish bioprocess to simultaneous detoxification and xylitol production by Geotrichum sp.

Sugarcane straw as a feedstock for xylitol production by Candida guilliermondii FTI 20037

Abstract Sugarcane straw has become an available lignocellulosic biomass since the progressive introduction of the non-burning harvest in Brazil. Besides keeping this biomass in the field, it can be used as a feedstock in thermochemical or biochemical conversion processes. This makes feasible its incorporation in a biorefinery, whose economic profitability could be supported by integrated production of low-value biofuels and high-value chemicals, e.g., xylitol, which has important industrial and clinical applications. Herein, biotechnological production of xylitol is presented as a possible route for the valorization of sugarcane straw and its incorporation in a biorefinery. Nutritional supplementation of the sugarcane straw hemicellulosic hydrolyzate as a function of initial oxygen availability was studied in batch fermentation of Candida guilliermondii FTI 20037. The nutritional supplementation conditions evaluated were: no supplementation; supplementation with (NH4)2SO4, and full supplementation with (NH4)2SO4, rice bran extract and CaCl2·2H2O. Experiments were performed at pH 5.5, 30 °C, 200 rpm, for 48 h in 125 mL Erlenmeyer flasks containing either 25 or 50 mL of medium in order to vary initial oxygen availability. Without supplementation, complete consumption of glucose and partial consumption of xylose were observed. In this condition the maximum xylitol yield (0.67 g g-1) was obtained under reduced initial oxygen availability. Nutritional supplementation increased xylose consumption and xylitol production by up to 200% and 240%, respectively. The maximum xylitol volumetric productivity (0.34 g L-1 h-1) was reached at full supplementation and increased initial oxygen availability. The results demonstrated a combined effect of nutritional supplementation and initial oxygen availability on xylitol production from sugarcane straw hemicellulosic hydrolyzate.

Enhanced Xylitol Production by Mutant Kluyveromyces marxianus 36907-FMEL1 Due to Improved Xylose Reductase Activity.

A directed evolution and random mutagenesis were carried out with thermotolerant yeast Kluyveromyces marxianus ATCC 36907 for efficient xylitol production. The final selected strain, K. marxianus 36907-FMEL1, exhibited 120 and 39 % improvements of xylitol concentration and xylitol yield, respectively, as compared to the parental strain, K. marxianus ATCC 36907. According to enzymatic assays for xylose reductase (XR) activities, XR activity from K. marxianus 36907-FMEL1 was around twofold higher than that from the parental strain. Interestingly, the ratios of NADH-linked and NADPH-linked XR activities were highly changed from 1.92 to 1.30 when K. marxianus ATCC 36907 and K. marxianus 36907-FMEL1 were compared. As results of KmXYL1 genes sequencing, it was found that cysteine was substituted to tyrosine at position 36 after strain development which might cause enhanced XR activity from K. marxianus 36907-FMEL1.

Bioprocessing of bagasse hydrolysate for ethanol and xylitol production using thermotolerant yeast.

Fermentation of xylose-rich and glucose-rich bagasse hydrolysates, obtained from the two-stage acid hydrolysis was studied using the thermotolerant yeast Kluyveromyces sp. IIPE453. The yeast could grow on xylose-rich hydrolysate at 50 °C with the dry cell weight, cell mass yield and maximum specific growth rate of 5.35 g l(-1), 0.58 g g(-1) and 0.13 h(-1), respectively. The yeast was found to be very promising for ethanol as well as xylitol production from the sugars obtained from the lignocellulosic biomass. Batch fermentations of xylose-rich and glucose-rich hydrolysates yielded 0.61 g g(-1) xylitol and 0.43 g g(-1) ethanol in the broth, respectively based on the sugars present in the hydrolysate. Overall ethanol yield of 165 g (210 ml) and 183 g xylitol per kg of bagasse was obtained, when bagasse hydrolysate was used as a substrate. Utilization of both the glucose and xylose sugars makes the process most economical by producing both ethanol and xylitol based on biorefinery concept. On validating the experimental data of ethanol fermentation, the modified Luong kinetic model for product inhibition as well as inhibition due to inhibitory compounds present in hydrolysate, the model was found to be the best fit for ethanol formation from bagasse hydrolysate using Kluyveromyces sp. IIPE453.

Coupled production of single cell oil as biodiesel feedstock, xylitol and xylanase from sugarcane bagasse in a biorefinery concept using fungi from the tropical mangrove wetlands.

This work evaluates sugarcane bagasse (SCB) conversion, in a biorefinery approach, to coproduce biodiesel and high value products using two novel mangrove fungi. On acid pre-treatment, sugarcane bagasse hydrolysate (SCBH) resulted in a xylitol yield of 0.51 g/g xylose consumed in 72 h by Williopsis saturnus. After SCB pretreatment, sugarcane bagasse residue (SCBR) was utilized using Aspergillus terreus for production of xylanase (12.74 U/ml) and cell biomass (9.8 g/L) which was extracted for single cell oil (SCO; 0.19 g/g) and transesterified to biodiesel. The FAME profile exhibited long chain SFAs and PUFAs with predicted biodiesel properties lying within the range specified by international standards. This biorefining approach of SCB utilization for co-production of xylitol, xylanase and SCO gains importance in terms of sustainability and eco-friendliness.

A rare sugar xylitol. Part II: biotechnological production and future applications of xylitol.

Xylitol is the first rare sugar that has global markets. It has beneficial health properties and represents an alternative to current conventional sweeteners. Industrially, xylitol is produced by chemical hydrogenation of D-xylose into xylitol. The biotechnological method of producing xylitol by metabolically engineered yeasts, Saccharomyces cerevisiae or Candida, has been studied as an alternative to the chemical method. Due to the industrial scale of production, xylitol serves as an inexpensive starting material for the production of other rare sugars. The second part of this mini-review on xylitol will look more closely at the biotechnological production and future applications of the rare sugar, xylitol.

Fermentation performance of engineered and evolved xylose-fermenting Saccharomyces cerevisiae strains.

Lignocellulose hydrolysate is an abundant substrate for bioethanol production. The ideal microorganism for such a fermentation process should combine rapid and efficient conversion of the available carbon sources to ethanol with high tolerance to ethanol and to inhibitory components in the hydrolysate. A particular biological problem are the pentoses, which are not naturally metabolized by the main industrial ethanol producer Saccharomyces cerevisiae. Several recombinant, mutated, and evolved xylose fermenting S. cerevisiae strains have been developed recently. We compare here the fermentation performance and robustness of eight recombinant strains and two evolved populations on glucose/xylose mixtures in defined and lignocellulose hydrolysate-containing medium. Generally, the polyploid industrial strains depleted xylose faster and were more resistant to the hydrolysate than the laboratory strains. The industrial strains accumulated, however, up to 30% more xylitol and therefore produced less ethanol than the haploid strains. The three most attractive strains were the mutated and selected, extremely rapid xylose consumer TMB3400, the evolved C5 strain with the highest achieved ethanol titer, and the engineered industrial F12 strain with by far the highest robustness to the lignocellulosic hydrolysate.

Carbon material and bioenergetic balances of xylitol production from corncobs by Debaryomyces hansenii.

The effect of oxygenation on xylitol production by the yeast Debaryomyces hansenii has been investigated in this work using the liquors from corncob hydrolysis as the fermentation medium. The concentrations of consumed substrates (glucose, xylose, arabinose, acetate and oxygen) and formed products (xylitol, arabitol, ethanol, biomass and carbon dioxide) have been used, together with those previously obtained varying the hydrolysis technique, the level of adaptation of the microorganism, the sterilization procedure and the initial substrate and biomass concentrations, in carbon material balances to evaluate the percentages of xylose consumed by the yeast for the reduction to xylitol, alcohol fermentation, respiration and cell growth. The highest xylitol concentration (71 g/L) and volumetric productivity (1.5 g/L.h) were obtained semiaerobically using detoxified hydrolyzate produced by autohydrolysis-posthydrolysis, at starting levels of xylose (S(0)) and biomass (X(0)) of about 100 g/L and 12 g(DM)/L, respectively. No less than 80% xylose was addressed to xylitol production under these conditions. The experimental data collected in this work at variable oxygen levels allowed estimating a P/O ratio of 1.16 mol(ATP)/mol(O). The overall ATP requirements for biomass production and maintenance demonstrated to remarkably increase with X(0) and for S(0) >or= 130 g/L and to reach minimum values (1.9-2.1 mol(ATP)/C-mol(DM)) just under semiaerobic conditions favoring xylitol accumulation.

Cloning of the xylitol dehydrogenase gene from Gluconobacter oxydans and improved production of xylitol from D-arabitol.

Xylitol dehydrogenase (XDH) was purified from the cytoplasmic fraction of Gluconobacter oxydans ATCC 621. The purified enzyme reduced D-xylulose to xylitol in the presence of NADH with an optimum pH of around 5.0. Based on the determined NH2-terminal amino acid sequence, the gene encoding xdh was cloned, and its identity was confirmed by expression in Escherichia coli. The xdh gene encodes a polypeptide composed of 262 amino acid residues, with an estimated molecular mass of 27.8 kDa. The deduced amino acid sequence suggested that the enzyme belongs to the short-chain dehydrogenase/reductase family. Expression plasmids for the xdh gene were constructed and used to produce recombinant strains of G. oxydans that had up to 11-fold greater XDH activity than the wild-type strain. When used in the production of xylitol from D-arabitol under controlled aeration and pH conditions, the strain harboring the xdh expression plasmids produced 57 g/l xylitol from 225 g/l D-arabitol, whereas the control strain produced 27 g/l xylitol. These results demonstrated that increasing XDH activity in G. oxydans improved xylitol productivity.

Obtenção e cinética da xilose redutase e xilitol desidrogenase de Candida guilliermondii cultivada em hidrolisado hemicelulósico de bagaço de cana-de-açúcar / Obtainment and kinetics of xylose reductase and xylitol dehydrogenase of Candida guillirmondii cultivated in sugarcane bagasse hemicellulosic hydrolysate

O metabolismo da D-xilose em leveduras inicia-se com a redução da D-xilose a xilitol pela xilose redutase (XR), a qual requer como cofator o NADPH e/ou NADH. O xilitol é oxidado a xilulose pela xilitol desidrogenase (XD), a qual emprega NA`D IND. +ï ou NAD`P IND. +ï como cofator. No presente trabalho, a bioconversão de D-xilose em xilitol por Candida guilliermondii FTI 20037 foi avaliada sob os aspectos enzimático e fermentativo, durante o cultivo em hidrolisado hemicelulósico de bagaço de cana-de-açúcar sob variadas condições de pH inicial, temperatura e aeração. Frente aos resultados obtidos nos cultivos em frascos agitados, encontrou-se a máxima atividade da XR (874,2 U.`mg IND. prot POT. -1ï) em pH inicial 6,0 e temperatura de 35ºC, condição esta na qual foram também verificados os máximos parâmetros fermentativos da produção de xilitol...