To date, although the high-carbohydrate (HC) feed has been extensively adopted in the aquaculture industry, its effects on the intestinal function and development of aquatic animals still remain unclear. In addition, the corresponding nutritional intervention is still barely reported. This study aimed to evaluate the influence of xylooligosaccharides (XOS) on the intestinal health of Megalobrama amblycephala subjected to a HC feeding. Fish (average weight: 44.55 ± 0.15 g) were randomly offered 3 diets, including a control one (29 % carbohydrate), a HC one (41 % carbohydrate), and a XOS supplemented one (HC + 1.0 % XOS, HCX) respectively for 12 weeks. The HC feeding caused morphological abnormalities of intestine, an increased intestinal permeability, and the intestinal immunosuppression, all of which were markedly reversed by XOS administration. In addition, compared with the HC group, HCX feeding remarkably promoted the intestinal activities of digestive and brush border enzymes, and the expressions of cell proliferation-related proteins (Wnt10b and Cyclin D1). The 16s rDNA sequencing also revealed that XOS administration increased the abundance of beneficial bacteria, and decreased that of pathogenic ones. In conclusion, dietary supplementation of XOS improved the intestinal histomorphology, barrier function, cell proliferation and bacterial communities of carbohydrate-overloaded fish Megalobrama amblycephala.

We propose a closed-loop pretreatment process, wherein volatiles produced during steam explosion pretreatment were recovered and reintroduced as acid catalysts into the pretreatment system. The volatiles were separated through a drastic decompression process followed by a steam explosion process and recovered as a liquified catalyst (LFC) through a heat exchanger. The LFC effectively served as an acid catalyst for hemicellulose hydrolysis, significantly decreasing residence time from 90 min to 30 min to achieve 80 % conversion yield at 170 °C. Hydrolysates with high content of lower molecular weight oligomeric sugars were obtained using LFC, and were considered advantageous for application as prebiotics. These results are attributed to the complementary features of acetic acid and furfural contained within the LFC. Computational simulation using Aspen Plus was used to investigate the effects of recycling on LFC, and it demonstrated the feasibility of the catalyst-recirculating system. A validation study was conducted based on simulation results to predict the actual performance of the proposed pretreatment system. Based on these results, the recirculating system was predicted to improve the conversion yield and low-molecular weight oligomers yield by 1.5-fold and 1.6-fold, respectively.

Enhancing root development is pivotal for boosting crop yield and augmenting stress resilience. In this study, we explored the regulatory effects of xylooligosaccharides (XOSs) on lettuce root growth, comparing their impact with that of indole-3-butyric acid potassium salt (IBAP). Treatment with XOS led to a substantial increase in root dry weight (30.77%), total root length (29.40%), volume (21.58%), and surface area (25.44%) compared to the water-treated control. These enhancements were on par with those induced by IBAP. Comprehensive phytohormone profiling disclosed marked increases in indole-3-acetic acid (IAA), zeatin riboside (ZR), methyl jasmonate (JA-ME), and brassinosteroids (BRs) following XOS application. Through RNA sequencing, we identified 3807 differentially expressed genes (DEGs) in the roots of XOS-treated plants, which were significantly enriched in pathways associated with manganese ion homeostasis, microtubule motor activity, and carbohydrate metabolism. Intriguingly, approximately 62.7% of the DEGs responsive to XOS also responded to IBAP, underscoring common regulatory mechanisms. However, XOS uniquely influenced genes related to cutin, suberine, and wax biosynthesis, as well as plant hormone signal transduction, hinting at novel mechanisms of stress tolerance. Prominent up-regulation of genes encoding beta-glucosidase and beta-fructofuranosidase highlights enhanced carbohydrate metabolism as a key driver of XOS-induced root enhancement. Collectively, these results position XOS as a promising, sustainable option for agricultural biostimulation.

OBJECTIVE: New characterized carbohydrate-active enzymes are needed for use as tools to discriminate complex carbohydrate structural features. Fungal glycoside hydrolase family 3 (GH3) β-xylosidases have been shown to be useful for the structural elucidation of glucuronic acid (GlcA) and arabinofuranose (Araf) substituted oligoxylosides. A homolog of these GH3 fungal enzymes from the bacterium Segatella baroniae (basonym Prevotella bryantii), Xyl3C, has been previously characterized, but those studies did not address important functional specificity features. In an interest to utilize this enzyme for laboratory methods intended to discriminate the structure of the non-reducing terminus of substituted xylooligosaccharides, we have further characterized this GH3 xylosidase.
RESULTS: In addition to verification of basic functional characteristics of this xylosidase we have determined its mode of action as it relates to non-reducing end xylose release from GlcA and Araf substituted oligoxylosides. Xyl3C cleaves xylose from the non-reducing terminus of β-1,4-xylan until occurrence of a penultimate substituted xylose. If this substitution is O2 linked, then Xyl3C removes the non-reducing xylose to leave the substituted xylose as the new non-reducing terminus. However, if the substitution is O3 linked, Xyl3C does not hydrolyze, thus leaving the substitution one-xylose (penultimate) from the non-reducing terminus. Hence, Xyl3C enables discrimination between O2 and O3 linked substitutions on the xylose penultimate to the non-reducing end. These findings are contrasted using a homologous enzyme also from S. baroniae, Xyl3B, which is found to yield a penultimate substituted nonreducing terminus regardless of which GlcA or Araf substitution exists.

Xylooligosaccharides (XOs) have shown high potential as prebiotics with nutritional and health benefits. In this work, XOs were obtained from highly purified, carboxy-reduced glucuronoarabinoxylans by treatment with Driselase®. The mixtures were fractionated, and the structures were elucidated by methylation analysis and NMR spectroscopy. Antioxidant activity was determined by the methods of DPPH and β-carotene/linoleic acid. It was found that the most active oligosaccharides (P3 and G3) comprised 4 or 5 xylose units, plus two arabinoses and one 4-O-methylglucose as side chains, their sequence of units was determined. The optimal concentration for their use as antioxidants was 2 mg/mL. The synthetic antioxidant butylated hydroxytoluene (BHT, 0.2 mg/mL) showed a percentage of inhibition 15% higher than P3. Although its concentration was ∼10 times higher, P3 is non-toxic, and could have great advantages as food additive. These results show that pure XOs exert significant antioxidant activity, only due to their carbohydrate nature.

Arabinoxylan is a major hemicellulose in the sugarcane plant cell wall with arabinose decorations that impose steric restrictions on the activity of xylanases against this substrate. Enzymatic removal of the decorations by arabinofuranosidases can allow a more efficient arabinoxylan degradation by xylanases. Here we produced and characterized a recombinant Bifidobacterium longum arabinofuranosidase from glycoside hydrolase family 43 (BlAbf43) and applied it, together with GH10 and GH11 xylanases, to produce xylooligosaccharides (XOS) from wheat arabinoxylan and alkali pretreated sugarcane bagasse. The enzyme synergistically enhanced XOS production by GH10 and GH11 xylanases, being particularly efficient in combination with the latter family of enzymes, with a degree of synergism of 1.7. We also demonstrated that the enzyme is capable of not only removing arabinose decorations from the arabinoxylan and from the non-reducing end of the oligomeric substrates, but also hydrolyzing the xylan backbone yielding mostly xylobiose and xylose in particular cases. Structural studies of BlAbf43 shed light on the molecular basis of the substrate recognition and allowed hypothesizing on the structural reasons of its multifunctionality.

Rice husks are rich in xylan, which can be hydrolyzed by xylanase to form xylooligosaccharides (XOS). XOS are a functional oligosaccharide such as improving gut microbiota and antioxidant properties. In this study, the structure and functional characteristics of XOS were studied. The optimal xylanase hydrolysis conditions through response surface methodology (RSM) were: xylanase dosage of 3000 U/g, hydrolysis time of 3 h, hydrolysis temperature of 50 °C. Under this condition, the yield of XOS was 150.9 mg/g. The TG-DTG curve showed that XOS began to decompose at around 200 °C. When the concentration of XOS reached 1.0 g/L, the clearance rate of DPPH reached 65.76 %, and the scavenging rate of OH reached 62.10 %, while the clearance rate of ABTS free radicals reached 97.70 %, which was equivalent to the clearance rate of VC. XOS had a proliferative effect on four probiotics: Lactobacillus plantarum, Lactobacillus brucelli, Lactobacillus acidophilus, and Lactobacillus rhamnosus. However, the further experiments are needed to explore the improvement effect of XOS on human gut microbiota, laying a foundation for the effective utilization of XOS. XOS have a wide range of sources, low price, and broad development prospects. The reasonable utilization of XOS can bring greater economic benefits.

Production of value-added compounds and sustainable materials from agro-industrial residues is essential for better waste management and building of circular economy. This includes valorization of hemicellulosic fraction of plant biomass, the second most abundant biopolymer from plant cell walls, aiming to produce prebiotic oligosaccharides, widely explored in food and feed industries. In this work, we conducted biochemical and biophysical characterization of a prokaryotic two-domain R. champanellensis xylanase from glycoside hydrolase (GH) family 30 (RcXyn30A), and evaluated its applicability for XOS production from glucuronoxylan in combination with two endo-xylanases from GH10 and GH11 families and a GH11 xylobiohydrolase. RcXyn30A liberates mainly long monoglucuronylated xylooligosaccharides and is inefficient in cleaving unbranched oligosaccharides. Crystallographic structure of RcXyn30A catalytic domain was solved and refined to 1.37 Å resolution. Structural analysis of the catalytic domain releveled that its high affinity for glucuronic acid substituted xylan is due to the coordination of the substrate decoration by several hydrogen bonds and ionic interactions in the subsite -2. Furthermore, the protein has a larger β5-α5 loop as compared to other GH30 xylanases, which might be crucial for creating an additional aglycone subsite (+3) of the catalytic site. Finally, RcXyn30A activity is synergic to that of GH11 xylobiohydrolase.

BACKGROUND: Dietary supplementation of xylooligosaccharides (XOS) has been found to influence gut health by manipulating cecal microbiota and producing microbe-origin metabolites. But no study investigated and compared the effect of in ovo feeding of xylobiose (XOS2) and xylotriose (XOS3) in chickens. This study investigated the effect of in ovo feeding of these XOS compounds on post-hatch gut health parameters in chickens. A total of 144 fertilized chicken eggs were divided into three groups: a) non-injected control (CON), b) XOS2, and c) XOS3. On the 17th embryonic day, the eggs of the XOS2 and XOS3 groups were injected with 3 mg of XOS2 and XOS3 diluted in 0.5 mL of 0.85% normal saline through the amniotic sac. After hatching, the chicks were raised for 21 d. Blood was collected on d 14 to measure plasma immunoglobulin. Cecal digesta were collected for measuring short-chain fatty acids (SCFA) on d 14 and 21, and for microbial ecology and microbial metabolic pathway analyses on d 7 and 21.
RESULTS: The results were considered significantly different at P < 0.05. ELISA quantified plasma IgA and IgG on d 14 chickens, revealing no differences among the treatments. Gas chromatography results showed no significant differences in the concentrations of cecal SCFAs on d 14 but significant differences on d 21. However, the SCFA concentrations were lower in the XOS3 than in the CON group on d 21. The cecal metagenomics data showed that the abundance of the family Clostridiaceae significantly decreased on d 7, and the abundance of the family Oscillospiraceae increased on d 21 in the XOS2 compared to the CON. There was a reduction in the relative abundance of genus Clostridium sensu stricto 1 in the XOS2 compared to the CON on d 7 and the genus Ruminococcus torques in both XOS2 and XOS3 groups compared to the CON on d 21. The XOS2 and XOS3 groups reduced the genes for chondroitin sulfate degradation I and L-histidine degradation I pathways, which contribute to improved gut health, respectively, in the microbiome on d 7. In contrast, on d 21, the XOS2 and XOS3 groups enriched the thiamin salvage II, L-isoleucine biosynthesis IV, and O-antigen building blocks biosynthesis (E. coli) pathways, which are indicative of improved gut health. Unlike the XOS3 and CON, the microbiome enriched the pathways associated with energy enhancement, including flavin biosynthesis I, sucrose degradation III, and Calvin-Benson-Bassham cycle pathways, in the XOS2 group on d 21.
CONCLUSIONS: In ovo XOS2 and XOS3 feeding promoted beneficial bacterial growth and reduced harmful bacteria at the family and genus levels. The metagenomic-based microbial metabolic pathway profiling predicted a favorable change in the availability of cecal metabolites in the XOS2 and XOS3 groups. The modulation of microbiota and metabolic pathways suggests that in ovo XOS2 and XOS3 feeding improved gut health during the post-hatch period of broilers.

The capacity of different Bacillus species to produce large amounts of extracellular enzymes and ability to ferment various substrates at a wide range of pH and temperature has placed them among the most promising hosts for the industrial production of many improved and novel products. The global interest in prebiotics, for example, xylooligosaccharides (XOs) is ever increasing, rousing the quest for various forms with expanded productivity. This article provides an overview of xylanase producing bacilli, with more emphasis on their capacity to be used in the production of the XOs, followed by the purification strategies, characteristics and application of XOs from bacilli. The large-scale production of XOs is carried out from a number of xylan-rich lignocellulosic materials by chemical or enzymatic hydrolysis followed by purification through chromatography, vacuum evaporation, solvent extraction or membrane separation methods. Utilization of XOs in the production of functional products as food ingredients brings well-being to individuals by improving defense system and eliminating pathogens. In addition to the effects related to health, a variety of other biological impacts have also been discussed.