Xylazyme AX Tablets

endo-1,4-β-Xylanase (EC 3.2.1.8) is employed across a broad range of industries including animal feed, brewing, baking, biofuels, detergents and pulp (paper). Despite its importance, a rapid, reliable, reproducible, automatable assay for this enzyme that is based on the use of a chemically defined substrate has not been described to date. Reported herein is a new enzyme coupled assay procedure, termed the XylX6 assay, that employs a novel substrate, namely 4,6-O-(3-ketobutylidene)-4-nitrophenyl-β-4⁵-O-glucosyl-xylopentaoside. The development of the substrate and associated assay is discussed here and the relationship between the activity values obtained with the XylX6 assay versus traditional reducing sugar assays and its specificity and reproducibility were thoroughly investigated.

Arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis releases only C3-linked arabinose residues from double-substituted xylose residues. A genomic library of B. adolescentis DSM20083 was screened for the presence of the axhD3 gene. Two plasmids were identified containing part of the axhD3 gene. The nucleotide sequences were combined and three open reading frames (ORFs) were found. The first ORF showed high homology with xylanases belonging to family 8 of the glycoside hydrolases and this gene was designated xylA. The second ORF was the axhD3 gene belonging to glycoside hydrolase family 43. The third (partial) ORF coded for a putative carboxylesterase. The axhD3 gene was cloned and expressed in Escherichia coli. Several substrates were employed in the biochemical characterization of recombinant AXHd3. The enzyme showed the highest activity toward wheat arabinoxylan oligosaccharides. In addition, β-xylanase from Trichoderma sp. was able to degrade soluble wheat arabinoxylan polymer to a higher extent, after pretreatment with recombinant AXHd3. Arabinoxylan oligosaccharides incubated with a combination of recombinant AXHd3 and an α-L-arabinofuranosidase from Aspergillus niger did not result in a higher maximal release of arabinose than incubation with these enzymes separately.

A fine balance exists between enzyme activity and the adverse effects associated with feed processing. Accurate estimation of enzyme activity in the feed is a pre-requisite to optimising the response.

Various procedures for the measurement of xylanase in fermentation broths, commercial enzyme mixtures, bread improver mixtures and feed samples are described. Problems associated with the routine use of reducing-sugar based methods axe highlighted and the advantages and limitations of viscometric and dye-labelled substrate procedures for measurement of trace levels of activity in feed samples are discussed.

Hydrolysis of mannan-type polysaccharides by β-mannanase is dependent on substitution on and within the main-chain as well as the source of the β-mannanase employed. Characterisation of reaction products can be used to define the sub-site binding requirements of the enzymes as well as the fine-structures of the polysaccharides. Action of endo-arabinanase and endo-galactanase on arabinans and arabinogalactans is described. Specific assays for endo-arabinanase and arabinan (in fruit-juice concentrates) are reported.

Enzymic degradation of carbohydrates is of major significance in the industrial processing of cereals and fruits. In the production of beer, barley is germinated under well defined conditions (malting) to induce maximum enzyme synthesis with minimum respiration of reserve carbohydrates. The grains are dried and then extracted with water under controlled conditions. The amylolytic enzymes synthesized during malting, as well as those present in the original barley, convert the starch reserves to fermentable sugars. Other enzymes act on the cell wall polysaccharides, mixed-linkage β-glucan and arabinoxylan, reducing the viscosity and thus aiding filtration, and reducing the possibility of subsequent precipitation of polymeric material. In baking, β-amylase and α-amylase give controlled degradation of starch to fermentable sugars so as to sustain yeast growth and gas production. Excess quantities of α-amylase in the flour result in excessive degradation of starch during baking which in turn gives a sticky crumb texture and subsequent problems with bread slicing. Juice yield from fruit pulp is significantly improved if cell-wall degrading enzymes are used to destroy the three-dimensional structure and water binding capacity of the pectic polysaccharide components of the cell walls. Problems of routine and reliable assay of carbohydrate degrading enzymes in the presence of high levels of sugar compounds are experienced with such industrial process.

This study was designed to test graded supplementation of a thermostable xylanase in pelleted, wheat-based diets fed to broiler chickens over a 28-d period. A total of 600 Ross 708 male broilers were allotted to 1 of 5 dietary treatments: positive control (PC), negative control (NC; 125 kcal of AME/kg diet reduction relative to PC), and NC supplemented with 10, 15, or 30 g/ton of xylanase. Wheat-soybean meal-based diets were pelleted and fed in 2 feeding phases (14-d each). Study outcomes included growth performance, AME, and ileal digesta viscosity with 20 battery cages of 6 birds per treatment. Data were analyzed by 1-way ANOVA along with estimation of Pearson correlation coefficients. Whereas no difference between NC and PC was observed for BW gain, NC birds exhibited increased (P < 0.05) feed intake during each feeding phase and overall, which caused improvements (P < 0.05) in feed conversion ratio (FCR) for PC vs. NC birds. The analyzed AME of PC birds was 112 kcal/kg of diet greater (P < 0.05) than for NC birds, though no differences in digesta viscosity were observed. Xylanase supplementation of the NC diet at 15 or 30 g/ton elicited overall improvements (P < 0.05) in BW gain beyond the PC, while the 30 g/ton level equalized feed intake with the PC. Regardless of level, xylanase supplementation improved (P < 0.05) the FCR relative to the NC, thereby equalizing the response with the PC. Similarly, supplementation with any xylanase level increased (P < 0.05) AME over the NC, making all treatments synonymous with the PC. Digesta viscosity of all xylanase-supplemented treatments was decreased relative to both the NC and PC treatments. Overall, this study provided clear evidence that addition of a thermostable xylanase to pelleted wheat-based diets elicited improvements in growth performance of broilers concomitant with a reduction in digesta viscosity and elevation of analyzed dietary AME content.

An experiment was conducted to test the hypothesis that an enzyme premix containing xylanase and glucanase improves the apparent total tract digestibility (ATTD) of energy and total dietary fiber (TDF) and the concentration of digestible energy (DE) and metabolizable energy (ME) in diets fed to growing pigs. A corn-soybean meal diet and a sorghum-soybean meal diet were formulated, and 4 additional diets were formulated by adding 400 g/kg distillers dried grains with solubles (DDGS) or 400 g/kg wheat middlings to the corn-based diet and the sorghum-based diet. Six additional diets were prepared by adding an enzyme premix including xylanase and β-glucanase to each of these diets. One hundred and forty-four growing pigs (61.7 ± 5.3 kg) were allotted to a randomized complete block design with 12 diets and 12 replicate pigs per diet. Pigs were adapted to the diets for 12 days before being moved to metabolism crates. Individual pig weights and feed consumption were recorded, and average daily feed intake (ADFI), average daily gain (ADG), and gain to feed ratio (G:F) were calculated for each dietary treatment. After 4 days of adaptation to the metabolism crates, urine and fecal materials were collected during the following 4 days according to the marker to marker approach. Results indicated that the ATTD of GE, and DE and ME increased (P < 0.01) if enzymes were added to the diets regardless of grain source or co-product inclusion, but no effect of enzymes on ATTD of TDF was observed. However, ATTD of TDF was greater in the corn based diet containing DDGS compared with the diet containing wheat middlings, but in the sorghum-based diet, no difference in ATTD of TDF was observed (grain source × co-product interaction, P < 0.05). However, growth performance were not affected by inclusion of enzymes, but inclusion of co-products to the diets decreased (P < 0.05) the ADG and G:F in pigs. In conclusion, mixture of enzyme including xylanase and β-glucanase used in this experiment has the potential to increase the ATTD of GE, and DE and ME, in both corn-based and sorghum-based diets without or with fiber containing co-products.

Treating ruminant feeds with exogenous fibrolytic enzymes may potentially increase forage cell wall degradability and thus feed efficiency. In nature, fungi biosynthesize lignocellulolytic enzymes that can break down lignocellulosic material into its sugar components, thereby providing ready fermentable substrates. This work showed the in vitro fibrolytic activity of three Trichoderma strains (T. atroviride strain P1, T. afroharzianum strain T22, T. reesei strain T67). Total protein concentration and enzymatic (e.g., glucanase, cellulase, and xylanase) activities were determined in fungal culture filtrates after 7 and 14 days of growth on different fiber-based media. The enzymatic mixtures produced by Trichoderma spp. showed the highest concentration of fibrolytic enzymes and were added to industrial feed to test their ability to hydrolyze insoluble fibers. The supplementation of industrial feeds containing medium-fiber or low-fiber concentrates with T22 enzymes produced in the presence of lyophilized mushrooms and durum wheat fiber reduced hemicellulose concentration up to 33% and 24%, respectively. These results may offer novel opportunities to develop livestock feeds with improved fiber digestibility.

Exogenous carbohydrases are commonly added to monogastric animal feed to degrade non-starch polysaccharides (NSP) to improve zootechnical performance and nutrient digestion. In the current study, the effects of dietary supplementation of a new, intrinsically thermostable, monocomponent xylanase on intestinal parameters, zootechnical performance and carcass traits of broilers were evaluated. A total of 720 1-day-old broilers were randomly allotted to four wheat-based diets with different levels of xylanase (T1: control; T2: 30,000 U/g; T3: 45,000 U/g; T4: 90,000 U/g) and fed from 1 to 35 days of age. The data showed that xylanase supplementation reduced the intestinal viscosity significantly, especially in the ileum. Duodenum and cecum pH was not influenced by xylanase supplementation, while T3 treatment showed the lowest pH value in jejunum and ileum. Xylanase supplementation reduced significantly feed conversion ratio (FCR) without affecting feed intake (FI). T2 treatment exhibited a higher body weight gain (BWG) compared the other treatments. Live weight, carcass weight, legs, and liver weight were significantly higher in T3 and T4 treatments compared to T1. No significant differences on foot pad lesions were observed among the four treatments. In conclusion, dietary supplementation of this new xylanase significantly improves intestinal tract viscosity and affects beneficially broilers’ performance and carcass traits.

Anti-nutritional compounds such as non-starch polysaccharides (NSP) are present in viscous cereals used in feed for poultry. Therefore, exogenous carbohydrases are commonly added to monogastric feed to degrade these NSP. Our hypothesis is that xylanase not only improves laying hen performance and digestibility, but also induces a significant shift in microbial composition within the intestinal tract and thereby might exert a prebiotic effect. In this context, a better understanding on whether and how the chicken gut microbial population can be modulated by xylanase is required. To do so, the effects of dietary supplementation of xylanase on performance, apparent total tract digestibility (ATTD) and cecal microbiome in laying hens were evaluated in the present study. A total of 96 HiSex laying hens were used in this experiment (3 diets and 16 replicates of 2 hens). Xylanase was added to the diets at concentrations of 0, 45,000 (15 g/t XygestTM HT) and 90,000 U/kg (30 g/t Xygest HT). The diets were based on wheat (~55%), soybean and sunflower meal. The lowest dosage, 45,000 U/kg, significantly increased average egg weight and improved feed efficiency compared to the control treatment (P<0.05). Egg quality parameters were significantly improved in the experiment in response to the xylanase addition. For example, during the last 28 days of the trial, birds receiving the 45,000 U/kg and the 90,000 U/kg treatments exhibited an increase in Haugh units and albumin heights (P<0.05). Compared with the control, the ATTD of organic matter and crude protein were drastically improved in the 45,000 U/kg treatment group (P<0.05). Furthermore, gross energy and the ATTD of crude fat were improved significantly for birds fed 90,000 U/kg group compared to the control. Importantly, 16S rRNA gene analysis revealed that xylanase at 45,000 U/kg dosage can exert a change in the cecal microbiome. A significant increase in beneficial bacteria (Bacilli class; Enterococcaceae and Lactobacillales orders; Merdibacter, Enterococcus and Nocardiopsis genera; Enterococcus casseliflavus species) was documented when adding 45,000 U/kg xylanase to the diet of laying hens. In conclusion, dietary supplementation of xylanase 45,000 U/kg significantly improved laying hen performance and digestibility. Furthermore, microbiome data suggest that xylanase modulates the laying hen bacterial population beneficially, thus potentially exerting a prebiotic effect.

The human gut can be viewed as a flow-through system with a short residence time, a high turnover rate and a spatial gradient of physiological conditions. As a consequence, the gut microbiota is exposed to highly fluctuating environmental determinants presented by the host and diet. Here, we assessed the fermentation and colonisation of insoluble wheat bran by faecal microbiota of three individuals at an unprecedented sampling intensity. Time-resolved 16S rRNA gene amplicon sequencing, revealed a dynamic microbial community, characterised by abrupt shifts in composition, delimiting states with a more constant community, giving rise to a succession of bacterial taxa alternately dominating the community over a 72 h timespan. Early stages were dominated by Enterobacteriaceae and Fusobacterium species, growing on the carbohydrate-low, protein rich medium to which wheat bran was supplemented. The onset of wheat bran fermentation, marked by a spike in short chain fatty acid production with an increasing butyrate proportion and an increased endo-1,4-β-xylanase activity, corresponded to donor-dependent proportional increases of Bacteroides ovatus/stercoris, Prevotella copri and Firmicutes species, which were strongly enriched in the bran-attached community. Literature and database searches provided novel insights into the metabolic and growth characteristics underlying the observed succession and colonisation, illustrating the potency of a time-resolved analysis to increase our understanding of gut microbiota dynamics upon dietary modulations.

Objective: Our objectives were to evaluate milk production and constituent responses to changes in the diet for pens of cows over time and whether differences in response were attributable to fibrolytic enzymes and dairy. Materials and Methods: A multiherd trial used 7,507 cows in 8 control and enzyme-treated (750 mL/t of DM feed) replicates (16 pens) on 3 dairies. Feed composition and milk production and constituents by pen (n = 12) were analyzed weekly. Time-series cross-correlation estimates by pen of feed component intakes (kg/d) and milk responses were pooled to produce effect size (ES) estimates. Results and Discussion: We observed differences between treatment and control pens for soluble protein (ES = 0.249) in the same week, acid detergent–insoluble CP (ES = 0.293) and lignin (ES = 0.237) 1 wk before with milk protein percentage, and acid detergent–insoluble CP (ES = 0.276) and lignin (ES = 0.246) 1 wk before with milk protein yield. These differences are consistent with enzymes improving feed digestibility, particularly for protein and fiber fractions. Differences in production responses to intake of feed components among dairies were observed. More significant and larger differences occurred among dairies than for treatments. The dairy that increased milk production most with treatment had an estimated MP excess from the diet, whereas the least responsive had an estimated MP-deficit diet and was the highest producing. Implications and Applications: We provide evidence for variability in enzyme response and that changes in dietary feed components influence production outcomes immediately and up to 3-wk later.

Recent outbreaks traced to contaminated flour have created a need in the milling industry for a process that reduces pathogens in wheat while maintaining its functional properties. Vacuum steam treatment is a promising technology for treatment of low-moisture foods. Traditional thermal treatment methods can compromise wheat functionality due to high temperatures; thus, maintaining the functional quality of the wheat protein was critical for this research. The objective of this study was to evaluate the effect of vacuum steam treatment of hard red spring (HRS) wheat kernels on final flour quality and the overall efficacy of vacuum stream treatment for reducing pathogens on HRS wheat kernels. HRS wheat samples were treated with steam under vacuum at 65, 70, 75, and 85°C for 4 and 8 min. Significant changes in dough and baked product functionality were observed for treatments at ≥ 70°C. Treatment time had no significant effect on the qualities evaluated. After determining that vacuum steam treatment at 65°C best preserved product quality, HRS wheat was inoculated with Escherichia coli O121 and Salmonella Enteritidis PT 30 and processed at 65°C for 0, 2, 4, 6, or 8 min. The treatments achieved a maximum average reduction of 3.57 ± 0.33 log CFU/g for E. coli O121 and 3.21 ± 0.27 log CFU/g for Salmonella. Vacuum steam treatment could be an effective pathogen inactivation method for the flour milling industry.

Our objectives were to characterize responses in the field to a mix of fibrolytic enzymes using large commercial dairy herds and sufficient study power to evaluate milk production and reproductive responses to an enzyme treatment started during the precalving period. We hypothesized that the use of the enzyme treatment would increase milk production when provided to dairy cows precalving and for approximately 200 d of lactation. The study was conducted on 7,507 cows, in 8 replicates and 16 pens, at 3 dairies in the United States. Eight pens were randomly allocated as control pens and received no enzyme, and another 8 pens received enzyme treatment at a dose of 750 mL/t of dry matter feed. Milk production and energy-corrected milk yield were increased with the enzyme treatment by 0.70 and 0.80 kg/d, respectively, across a 5-month period. Milk fat percentage was not significantly increased by enzyme treatment, but milk fat yield was significantly increased by 0.040 kg/d, compared with controls. Milk protein yield increased 0.010 kg/d with enzyme treatment despite a small reduction of 0.020 percentage units in milk protein percentage. We found no evidence of an increase in the ln somatic cell count for the enzyme-treated cows. Body weight overall was not increased for enzyme-treated cows, but we did observe a numerical increase in dry matter intake (0.20 kg/head per day) for enzyme-treated cows. Most production responses to the enzyme treatment were influenced by dairy. Compared with controls, milk yield in enzyme-treated cows was significantly higher by 3.6 kg/d in dairy 2 and numerically higher by 0.60 and 0.20 kg/d in dairies 1 and 3, respectively. Reproduction, health, and risk of removal or death were not significantly influenced by treatment, apart from a reduced time to first breeding. Production responses to the enzyme treatment varied by dairy from substantial to minor increases, but variation among dairies was not evident in differences in dry matter intake or in partitioning of body weight among enzyme-treated and control pens and cows. It appears likely that the increase in production reflected increased digestibility of feed; however, further work is needed to identify factors influencing the variation in production responses to enzymes.

We investigated the effects of adding up to 11% rice bran (RB) in corn-soybean meal diets fed to broiler chickens without or with a multi-enzyme supplement (MES). The MES supplied xylanase, β-glucanase, invertase, protease, cellulase, α-amylase and mannanase with targeted activity of 2,500, 300, 700, 10,000, 1,200, 24,000, and 20 U/kg of feed, respectively. The study used a two-phase feeding program (starter, d 0 to 24; finisher, d 25 to 35) with RB added at 5% and 11%, respectively creating 4 diets in each phase. Diets were iso-caloric and iso-nitrogenous and contained phytase (500 FTU/kg) and TiO₂ as a digestibility marker. Three hundred and sixty d-old male Ross 708 broiler chicks were placed in cages based on BW (15 birds/cage) and allocated to 4 diets (n = 6). Birds had free access to feed and water. Body weight and feed intake were recorded. Excreta samples were collected 3 d prior to the end of each phase for apparent retention (AR) of components. Samples of birds were sacrificed on d 24 and 35 for gut weight and ceca digesta for organic acid content. There was no interaction (P > 0.10) between RB and MES on BWG and FCR in starter or finisher phase. In finisher phase, birds fed MES had better BWG (961 versus 858 g) and FCR (1.69 versus 1.86) than birds fed non-MES diets (P < 0.01). Feeding RB reduced (P = 0.02) BWG in finisher phase resulting in lower d 35 BW. Birds fed RB had higher (P ≤ 0.01) gizzard weight on d 24 and 35 than non-RB birds. An interaction (P ≤ 0.01) between RB and MES on concentrations of propionic and iso-butyric acids in ceca digesta showed that MES reduced these acids in non-RB diet. The AR of gross energy was higher (P < 0.02) for MES versus non-MES birds in starter and finisher phases. In conclusion, independently, RB increased gizzard weight and reduced final BW whereas MES improved growth and energy utilization.

Unlike wheat bread, the dough of which has a visco-elastic network and high gas-holding capacity, oat bread generally has a low volume and a dense structure. We showed earlier that including rye water-extractable components in an oat bread batter recipe increases loaf volume by ca. 30% (Pauly and Delcour, submitted as back-to-back publication). We here report on efforts to identify the active factor(s). Anion exchange chromatography allowed enriching the active factor(s). This and the fact that only a limited volume increase was observed when oat batter was supplemented with boiled rye extract indicate that proteins are likely the most important components responsible for the volume increase. While the most active factor(s) had a pI below 4.5, components with pI values between 4.5 and 8.5 also contributed to oat loaf volume. Alkaline rye components (pI > 8.5) or rye arabinoxylan had no impact. Rye water-extractable components smaller than 6–8 kDa also had a positive impact on loaf volume.

Loaf volume and crumb structure of oat bread are not comparable to those of bread from wheat flour. Hydrocolloids, surfactants and/or enzymes are often included in oat batter recipes for quality enhancement reasons. In this study, we examined the impact of water-extractable components from barley, oat, rye and wheat flour on oat bread quality. We speculated that such water extracts contain components which also would enhance the quality of oat bread. As expected, extract protein, non-starch polysaccharide, lipid and enzyme levels varied widely amongst the different cereal flours used. The extracts also varied in foaming properties and extract viscosities. Rye flour contained the highest level of water-extractable components. Inclusion of rye aqueous extract resulted in the largest loaf volume increase and in softer crumb than noted for control oat bread. Rheofermentometer analyses showed that the moment of gas cell opening was delayed when rye extract was added, indicating improved batter gas cell stabilization, while collapse during baking was not affected. The oat bread improving effect of the rye extract is likely due to a combination of the impact of different of its constituents such as enzymes and surface active components.

The effect of a combination of carbohydrase and phytase enzymes on growth performance, insulin-like growth factor 1 gene expression, insulin status, and insulin receptor sensitivity in broiler chickens fed wheat-soybean meal diets containing 6% (starter) and 12% (grower-finisher) of full-fat rapeseed (canola type; low glucosinolate, low erucic acid) from 1 to 42 d of age was studied. A total of 510 one-day-old male broiler chickens were randomly assigned to 3 dietary treatments, with 17 pens per treatment and 10 birds per pen. The dietary treatments consisted of a control diet and P- and Ca-deficient diets supplemented with either phytase (500 U/kg) or a combination of phytase and a multi-carbohydrase enzyme (Superzyme OM). The diets were pelleted at 78°C and were fed ad libitum throughout the starter (9 d), grower (18 d), and finisher (15 d) phases of the experiment. Over the entire trial, growth performance of birds fed the phytase-supplemented diet did not differ from birds fed the control diet. The use of phytase in combination with a multicarbohydrase enzyme improved (P = 0.007) the feed conversion ratio from 1.90 to 1.84. Insulin liver receptor sensitivity increased by 9.3 and 12.3% (P = 0.004) for the phytase- and the carbohydrase-phytase-supplemented diets, respectively. There was no effect of phytase alone or carbohydrase and phytase supplementation on total plasma cholesterol, high-density lipoprotein cholesterol, and blood glucose levels. However, low-density lipoprotein cholesterol decreased (P = 0.007) for the phytase-carbohydrase treatment. Gene expression of insulin-like growth factor 1 tended to decrease by 32% (P = 0.083) after phytase-carbohydrase supplementation. The combination of carbohydrase and phytase enzymes may serve as an attractive means of facilitating nutrient availability for digestion and thus enhance the feeding value of wheat-soybean meal-based diets containing full-fat rapeseed. However, the extent to which the effects of enzyme addition on insulin receptors are associated with growth performance of broiler chicken requires further research.

Hypocrea jecorina is an important, filamentous fungus due to its effective production of hydrolytic enzymes. Gene expression studies provide deeper insight into environment sensing and cellular response mechanisms. Reverse transcription-quantitative PCR is a gene-specific and powerful tool to measure even minor changes in mRNA composition. An accurate normalization strategy is absolutely necessary for appropriate interpretation of reverse transcription-quantitative PCR results. One frequently applied strategy is the usage of a reference gene. Adequate reference genes for Hypocrea have not been published so far. By using the NormFinder and geNorm softwares, we evaluated the most stable genes amongst six potential reference genes in 34 samples from diverse cultivation conditions. Under those experimental conditions, sar1 encoding for a small GTPase was found to be the most stable gene, whereas act encoding for actin was not amongst the best validated ones. The influence of the reference system on the expression data is demonstrated by analysis of two target genes, encoding for the Xylanase regulator 1 and for Xylanase II. We further validated obtained xylanase 2 transcription rates with the corresponding enzyme activity.