Produksi dan Pemurnian Xilooligosakarida dari Xilan Tongkol Jagung menggunakan Xilanase Streptomyces P26B4 dan Khamir IP4

Xylooligosaccharides (XOS) are sugar oligomers from xylan that can be used as prebiotics to improve digestive tract health. Xylan can be extracted from corncobs which are a by-product of agriculture. The purpose of this study was to produce XOS through hydrolysis of corn cobs xylan using Streptomyces P26B4 xylanase. The products of hydrolysis also consisted of monomer xylose; for that xilooligosaccharides were purified using yeast IP4. The xylan hydrolysis products are quantitatively analyzed based on the value of reducing sugars and degree of polymerization (DP), strengthened qualitatively with TLC. Sugar component was analyzed after the addition of yeast by HPLC. P26B4 xylanase isolates had the highest activity on the 7th day incubation, pH 5,5 buffer citrate and temperature of 50°C. The lowest DP value of xylan hydrolysis was 2.49 at a concentration of 6%, and the 24th hour of incubation. TLC chromatograms showed that xylose and XOS products were produced. Purification of XOS at 6%, showed a decreasing in the area of xylose before and after receiving yeast respectively 1.87% and 1.41%.Therefore, yeast IP4 has the potential to consume xylose amnd purify the XOS. Keywords: corncobs xylan, IP4 yeast, purification, Streptomyces P26B4 xylanase, xylooligosaccharides

Optimization of β-1,4-Endoxylanase Production by an Aspergillus niger Strain Growing on Wheat Straw and Application in Xylooligosaccharides Production

Plant biomass constitutes the main source of renewable carbon on the planet. Its valorization has traditionally been focused on the use of cellulose, although hemicellulose is the second most abundant group of polysaccharides on Earth. The main enzymes involved in plant biomass degradation are glycosyl hydrolases, and filamentous fungi are good producers of these enzymes. In this study, a new strain of Aspergillus niger was used for hemicellulase production under solid-state fermentation using wheat straw as single-carbon source. Physicochemical parameters for the production of an endoxylanase were optimized by using a One-Factor-at-a-Time (OFAT) approach and response surface methodology (RSM). Maximum xylanase yield after RSM optimization was increased 3-fold, and 1.41- fold purification was achieved after ultrafiltration and ion-exchange chromatography, with about 6.2% yield. The highest activity of the purified xylanase was observed at 50 °C and pH 6. The enzyme displayed high thermal and pH stability, with more than 90% residual activity between pH 3.0–9.0 and between 30–40 °C, after 24 h of incubation, with half-lives of 30 min at 50 and 60 °C. The enzyme was mostly active against wheat arabinoxylan, and its kinetic parameters were analyzed (Km = 26.06 mg·mL−1 and Vmax = 5.647 U·mg−1). Wheat straw xylan hydrolysis with the purified β-1,4 endoxylanase showed that it was able to release xylooligosaccharides, making it suitable for different applications in food technology.

Feruloyl esterase (FAE-1) sourced from a termite hindgut and GH10 xylanases synergy improves degradation of arabinoxylan

Cereal feedstocks have high arabinoxylan content as their main hemicellulose, which is linked to lignin by hydroxycinnamic acids such as ferulic acid. The ferulic acid is linked to arabinoxylan by ester bonds, and generally, the high substitution of ferulic acid leads to a loss of activity of xylanases targeting the arabinoxylan. In the current study, a feruloyl esterase (FAE-1) from a termite hindgut bacteria was functionally characterised and used in synergy with xylanases during xylan hydrolysis. The FAE-1 displayed temperature and pH optima of 60 ℃ and 7.0, respectively. FAE-1 did not release reducing sugars from beechwood xylan (BWX), wheat arabinoxylan (WAX) and oat spelt xylan (OX), however, displayed high activity of 164.74 U/mg protein on p-nitrophenyl-acetate (pNPA). In contrast, the GH10 xylanases; Xyn10 and XT6, and a GH11 xylanase, Xyn2A, showed more than two-fold increased activity on xylan substrates with low sidechain substitutions; BWX and OX, compared to the highly branched substrate, WAX. Interestingly, the FAE-1 and GH10 xylanases (Xyn10D and XT6) displayed a degree of synergy (DS) that was higher than 1 in all enzyme loading combinations during WAX hydrolysis. The 75%XT6:25%FAE-1 synergistic enzyme combination increased the release of reducing sugars by 1.34-fold from WAX compared to the control, while 25%Xyn10D:75%FAE-1 synergistic combination released about 2.1-fold of reducing sugars from WAX compared to controls. These findings suggest that FAE-1 can be used in concert with xylanases, particularly those from GH10, to efficiently degrade arabinoxylans contained in cereal feedstocks for various industrial settings such as in animal feeds and baking.

Optimization of β-1,4-endoxylanase production by a new Aspergillus niger strain growing on wheat straw and application in xylooligosaccharides production

Plant biomass constitutes the main resource of renewable carbon in the planet and its valorization has traditionally been focused on the use of cellulose, although hemicellulose is the second most abundant group of polysaccharides on earth. Enzymes involved in its degradation are usually glycosyl hydrolases and filamentous fungi are good producers of these enzymes. In this study, a new strain of Aspergillus niger was utilized for hemicellulase production under solid state fermentation using wheat straw as a single carbon source. Physicochemical parameters for production of an endoxylanase were optimized by using one factor at a time approach and response surface methodology (RSM). Maximum xylanase yield after RSM optimization was increased 3-fold. The enzyme was purified by ultrafiltration and ion-exchange chromatography1.41-fold, with 6.2 % yield. Highest xylanase activity was observed at 50 °C and pH 6. A high pH and thermal stability were found, greater than 90% residual activity between pH 3.0-9.0 and between 30-40°C, after 24 h of incubation, presenting half-lives of 30 min at 50 and 60°C. Enzyme was mostly active for wheat arabinoxylan, and displayed the following kinetic parameters Km of 26.06 mg•ml-1 and Vmax of 5,647 U•mg-1min-1.Wheat straw xylan hydrolysis with the purified β-1,4 endoxylanase showed that it was able to release xylooligosaccharides, making it suitable for different applications in food technology.

Autohydrolysis pretreatment of castor plant pruning residues to enhance enzymatic digestibility and bioethanol production

Castor plant is used commonly for oil extraction and biodiesel synthesis. However, the residues during pruning are not being used effectively. These residues have the potential to be used as feedstock to produce bioethanol and other by-products. The present work assessed the eco-friendly autohydrolysis pretreatment of castor plant pruning residues at different severity factors (R0), applying a range of temperatures from 100 °C to 200 °C. The hydrolysis of pretreated solids was carried out using a commercial cellulases complex at different solid and enzyme loadings. The enzymatic hydrolysate with a higher glucose concentration was further subjected to fermentation using Saccharomyces cerevisiae ATCC 4126. The results showed an efficient xylan hydrolysis (77.5%) and a preservation of glucan up to 83% in the solids pretreated at an R0 of 5.78. The enzymatic hydrolysis of the pretreated solids at an R0 of 5.78 showed a glucose release of 2.9-fold higher than non-pretreated material. In the hydrolysate fermentation, a maximum ethanol production of 50.5 g/L was achieved (equivalent to 6.4% v/v), corresponding to a conversion efficiency of 98% and a biomass-to-ethanol conversion yield of 93.0 g of ethanol per kilogram of feedstock.

The GH10 and GH48 dual-functional catalytic domains from a multimodular glycoside hydrolase synergize in hydrolyzing both cellulose and xylan

Background Regarding plant cell wall polysaccharides degradation, multimodular glycoside hydrolases (GHs) with two catalytic domains separated by one or multiple carbohydrate-binding domains are rare in nature. This special mode of domain organization endows the Caldicellulosiruptor bescii CelA (GH9-CBM3c-CBM3b-CBM3b-GH48) remarkably high efficiency in hydrolyzing cellulose. CbXyn10C/Cel48B from the same bacterium is also such an enzyme which has, however, evolved to target both xylan and cellulose. Intriguingly, the GH10 endoxylanase and GH48 cellobiohydrolase domains are both dual functional, raising the question if they can act synergistically in hydrolyzing cellulose and xylan, the two major components of plant cell wall. Results In this study, we discovered that CbXyn10C and CbCel48B, which stood for the N- and C-terminal catalytic domains, respectively, cooperatively released much more cellobiose and cellotriose from cellulose. In addition, they displayed intramolecular synergy but only at the early stage of xylan hydrolysis by generating higher amounts of xylooligosaccharides including xylotriose, xylotetraose, and xylobiose. When complex lignocellulose corn straw was used as the substrate, the synergy was found only for cellulose but not xylan hydrolysis. Conclusion This is the first report to reveal the synergy between a GH10 and a GH48 domain. The synergy discovered in this study is helpful for understanding how C. bescii captures energy from these recalcitrant plant cell wall polysaccharides. The insight also sheds light on designing robust and multi-functional enzymes for plant cell wall polysaccharides degradation.