Xylose isomerization

Measurement of Xylose Isomerization Kinetics and Equilibrium All experiments were carried out at 34 °C in a volume of 25 ml in 50-ml shake flasks agitated at 130 rpm in an incubated shaker. Each experiment was conducted in duplicate. All experiments used 60 g/ 1 xylose, and unless otherwise noted, 5.2 g/1 of enzyme pellets (0.13 g) was used for each experiment. Buffered solutions used in making the isomerization media were 0.01 M Tris buffer (pHed to 7.5 using 0.01 M NaOH) and 0.05 M sodium citrate buffer (pHed to 4.5 using citric acid). The pH was measured at the beginning of the experiments but was not monitored throughout. We have observed a small drift in pH of less than 1 unit over the course of 48 hrs. Even with this drift, the pH of the bulk solution stayed within the range suitable for fermentation and well-below the pH optimum of XI (pH 7.5). In experiments with co-inunobilized pellets, urea concentration was 0,0.01, or 0.1 M. 

The interior pellet pH is a function of the urease loading and the urea concentration profile in the pellet. The for urease hydrolysis of urea is 2.9 mM, [29] so with 0.01 M (10 mM) urea, we are initially consuming urea at approximately 78% of Fmax at the surface of the pellet. Increasing the bulk concentration of urea will result in increased ammonia production and an increase in the interior pellet pH. Depending upon whether the interior pH is above or below the pH for optimum XI activity, an increase in interior pH will decrease or increase the rate of xylose isomerization. To achieve optimal isomerization in the co-immobilized pellet system, the urea concentration in the bulk solution can be optimized for a specific urease loading and should be maintained at a constant concentration throughout the isomerization to allow maximal, constant XI activity. 

Effect of Sodium Tetraborate Addition on Xylose Isomerization... 

As seen in these the two experiments, the rate of xylose isomerization is very similar for the first 4 h. For both cases, the concentration of urea is significantly higher than the Xm for urease so the internal pH profiles within the pellet are likely to be similar. As the pellets also have the same XI loading, xylulose production is equivalent in both. However, by 8 h, urea consumption in A results in a decrease in reaction velocity for urea hydrolysis. With reduced ammonia production, the internal volume of the pellet with active XI decreases, and a drop in xylulose production relative to B is observed. Based on the results shown for A, urea hydrolysis is no longer effective at maintaining the two pH microenvironments by 24 h. 

For B, with a much higher initial urea concentration, the active zone for xylose isomerization is maintained for a much longer period of time (>48 h). The final xylulose concentration measured at 48 h was 52 g/1, corresponding to a xylose/xylulose ratio of 1 6.5. 

Simultaneous isomerization and fermentation (SIF) is expected to drive the isomerization forward and lead to higher xylose utilization. Unfortunately, the pH optima for the isomerization ( 7.5) and the fermentation ( 4.5) steps are vastly different. The approach we have taken is to develop a novel technique for SIF that is capable of sustaining two different pH-microenvironments in a single vessel—one optimal for xylose isomerization and the other optimal for fermentation of xylulose. The technique involves co-immobilization of urease with xylose isomerase. We have shown that it is possible to sustain a significant pH gradient between the bulk liquid and the core region of the pellet by adding urea to the fermentation broth. 

The first step of xylose catabolism is its conversion to xylulose. In bacteria, it takes place by the direct isomerization catalysed by xylose isomerase. In PeniciUium chrysogenum, a sequence of enzymes in the initial steps of pentose metabolism was observed that differs from xylose isomerization in bacteria [91, 92]. These enzymes were common in yeast and filamentous fungi. In this oxido-reductive pathway, xylose is first reduced to the xyhtol in the presence of NAD(P)-linked xylose reductase, which is then reoxidized by NAD(P)-hnked dehydrogenase to give xylulose (Fig. 1). It has been assumed that this oxido-reductive pathway is common among fungi [93]. Both the enzymes involved, xylose reductase and xylitol dehydrogenase, were found to be inducible and relatively specific for the D-xylose and xyhtol in F. oxysporum, whereas D-xylose isomerase was not detected. 

Of this group of enzymes, glucose isomerse, which is used in the production of starch syrup with a high content of fructose (cf. 19.1.4.3.5), is very important. The enzyme used industrially is of microbial origin. Since its activity for xylose isomerization is higher than for glucose, the enzyme is classified under the name xylose isomerase (cf. Table 2.4). 

Choudhary V, Pinar AB, Sandler SI, Vlachos DG, Lobo RE. Xylose isomerization to xylulose and its dehydration to furfural in aqueous media. ACS Catal 2011 1 1724—8. 

Figure 8.9 External mass transfer resistance—xylose hydrogenation and isomerization to xylitol and by-products on sponge Ni (based on the results of Mikkola et al. [22]).

The enzymic synthesis of D-plant polysaccharides. Xylose isomerase has been found in Lactobacillus pento-... 

D-[l- C]Xylose was subjected to boiling in 4 M aqueous sodium hydroxide. The resulting mixture contained 2,4-dihydroxybutanoic acid, lactic acid, and D-a,j8-xylometasaccharinic acid. The almost uniform distribution of the C label among the carbon atoms of 2,4-dihydroxybutanoic acid indicated that this acid is probably formed by the recombination of completely isomerized, two-carbon fragments. Fragmentation of D-xylose occurred mainly at one of the central bonds, C-2-C-3 or C-3-C-4. 

Interest in the bacterial ens me xylose/glucose isomerase has been driven by its use in the isomerization of ucose to fructose to produce high>fructose corn syrups, and in the isomerization of xylose to xylulose for the conversion of the more fermentable xylulose to ethanol In this work, a brief historical perspective is presented, followed by a summary of the current understanding of the enzyme s major features. Also, a useful compilation of available xylose isomerase DNA sequences is presented with annotation of some of the major areas identified as being of functional significance. The extent of homology between the xylose isomerases is discussed with reference to differences in their function. 

A true xylose isomerase, that did not require arsenate for its activity, was found in strains of Lactobacillus, especially L. brevis (25). In this study, it was found that the activity of isomerization of glucose and xylose were essentialfy equal, and that ribose was isomerized to ribulose at a reduced rate 24). The xylose isomerases from this strain, like those from other species, requires divalent cations for activity. In this case, Mn and Co were found to be required for activity. In other studies, Co has been found to increase xylose isomerase stability 25,26,21. ... 

Industrially relevant consecutive-competitive reaction schemes on metal catalysts were considered hydrogenation of citral, xylose and lactose. The first case study is relevant for perfumery industry, while the latter ones are used for the production of sweeteners. The catalysts deactivate during the process. The yields of the desired products are steered by mass transfer conditions and the concentration fronts move inside the particles due to catalyst deactivation. The reaction-deactivation-diffusion model was solved and the model was used to predict the behaviours of semi-batch reactors. Depending on the hydrogen concentration level on the catalyst surface, the product distribution can be steered towards isomerization or hydrogenation products. The tool developed in this work can be used for simulation and optimization of stirred tanks in laboratory and industrial scale. 

DL-Dihydrostreptose and its ribo isomer were similarly obtained. Birch reduction of 2-methyl-3-furoic acid, followed by addition of methanol, bromination, and dehydrobromination, gave 402 as a mixture of the isomers. Hydroxylation of 402 with osmium tetraoxide-so-dium chlorate, and subsequent treatment with acetone-sulfuric acid afforded three isomeric acetals (403-405). The structures of these compounds were assigned on the basis of their H-n.m.r. spectra. In addition, the relationship between 403 and 404 was established by hydrolysis and reglycosidation. The methyl esters 403-405 were quantitatively reduced to the corresponding alcohols. The mixture of alcohols obtained from 403 and 404 was converted into crystalline 5-deoxy-3-C-(hydroxymethyl)-l,2-0-isopropylidene-a-DL-ribofuran-ose (406), which was compared directly with a sample prepared from D-xylose. Methyl 5-deox y-3-C-(hydroxy methyl)-2,3-O-isopropy lidene-/3-DL-lyxofuranoside (407), obtained by reduction of 405 with lithium aluminum hydride, was hydrolyzed with dilute hydrochloric acid, to give a,/3-DL-dihydrostreptose.2,ifi... 

These enzymes vary widely in secondary and tertiary structure.1273 Mannose-6-phosphate isomerase is a 45 kDa Zn2+-containing monomer. The larger 65 kDa L-fucose isomerase, which also acts on D-arabinose, is a hexameric Mn2+-dependent enzyme.1273 L-Arabinose isomerase of E. coli, which interconverts arabinose and L-ribulose, is a hexamer of 60-kDa subunits128 while the D-xylose isomerase of Streptomyces is a tetramer of 43-kDa subunits.129 The nonenzymatic counterpart of the isomerization catalyzed by the enzyme is the base-catalyzed Lobry deBruyn-Alberda van Ekenstein transformation (Eq. 13-25).130... 

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