Arabinoxylan is a significant component of cereal hemicelluloses. It has become a functional food ingredient of interest with beneficial health benefits linked to its prebiotic effects. Understanding the enzymatic mechanisms for the complete degradation of this polysaccharide is also essential for the biofuels industry by allowing the efficient conversion and utilisation of plant biomass. An article recently published by McClearyet al. discusses the action of the various enzymes involved in the hydrolysis of wheat arabinoxylan.1
Structurally, arabinoxylan consists of a β-1,4 linked xylan backbone with the xylose monomers bearing a mixture of either 3- monosubstituted or 2,3-disubstituted α-L-arabinose. Some arabinofuranosyle residues are further substituted with ferulic acid, which under certain circumstances can crosslink to other ferulic acid residues, forming di-ferulate bridges.
Figure: Arabinoxylan contains D-xylosyl monomeric units linked β-1,4 and extensively modified with α-L-arabinofuranosyl residues on positions 2 and 3.
Assay Kits
Megazyme currently provides kits for the assay of xylanase activity and is developing an enzymatic assay kit for the determination of wheat flour arabinoxylan. This will be the first commercially available kit of its kind.
| Product Code | Product Name |
| K-XYLOSE | Xylose (can be used for the measurement Wheat Flour Arabinoxylan) |
| K-XYLS | Xylanase (Xylazyme AX) |
| K-AZOWAX |
Xylanase (Azo-Wax) |
Carbohydrates
Oligosaccharides
Specific xylo-oligosaccharides (XOS) and arabino-xylooligosaccharides (AXOS) have been prepared with high purity for use, as chromatographic standards, and in enzyme characterisation and binding studies.
| Product Code | Product Name | Chemical Structure |
| 1,4-β-D-Xylo-oligosaccharides | ||
| O-XBI | Xylobiose |  |
| O-XTR | Xylotriose | |
| O-XTE | Xylotetraose | |
| O-XPE | Xylopentaose | |
| O-XHE | Xylohexaose | |
| Arabinoxylo-oligosaccharides (AXOS) | ||
| O-A3X | 32-α-L-Arabinofuranosyl -xylobiose (A3X) |
 |
| O-A2XX | 23-α-L-Arabinofuranosyl -xylotriose (A2XX) |
 |
| O-XA3XX | 33-α-L-Arabinofuranosyl -xylotetraose (XA3XX ) |
 |
| O-XAXXMIX | 33-α-L- plus 23-α-L- Arabinofuranosyl- xylotetraose (XA3XX/XA2XX) mixture |
 |
Polysaccharides
Arabinoxylan polysaccharides obtained from rye and wheat are available from Megazyme. High, medium and low viscosity grades have been prepared with these viscosities indicating the molecular weight range of each product. Acid and enzyme debranched arabinoxylans have also been produced providing alternative substrates with varying degrees of arabinose substitution. Xylan sourced from beechwood is available as a substrate for reducing sugar assays of endo-1,4-β-xylanase. Beechwood xylan contains approximately 10% 4-O-methylglucuronate substitution, which is exactly the same as that in birchwood xylan.
| Product Code | Product Name |
| P-RAXY | Arabinoxylan (rye flout; high viscosity) |
| P-WAXYL | Arabinoxylan (wheat flour; low viscosity) |
| P-WAXYM | Arabinoxylan (wheat flour; medium viscosity) |
| P-WAXYH | Arabinoxylan (wheat flour; high viscosity) |
| P-WAXYI | Arabinoxylan (wheat flour; insoluble) |
| P-ADWAX22 | Arabinoxylan (acid debranched; 22% ara)NEW |
| P-ADWAX26 | Arabinoxylan (acid debranched; 26% ara)NEW |
| P-EDWAX30 |
Arabinoxylan (enzymically debranched using an α-L-arabinofuranosidase that removes 1,3-linked α-L-arabinofuranose from doubly substituted D-xylose residues, 30% ara)NEW |
| P-XYLNBE | Highly purified xylan from beechwood. Suitable as a replacement for birchwood xylan as a substrate for β-xylanase in DNSA reducing sugar assaysNEW |
| P-WAXYNS | Arabinoxylan (wheat flour; low viscosity; specifically for use in Nelson-Somogyi reducing sugar assays for β-xylanase)COMING SOON |
Enzymes
endo-1,4-β-Xylanases (EC 3.2.1.8) have the ability to hydrolyse the interlinking β-1,4 glycosidic bonds present in the xylan backbone to varying degrees based on 1) the identity of the endo-xylanase in question and 2) the neighbouring arabinose substitution present.
exo-1,4-β-Xylosidases (EC 3.2.1.37) sequentially hydrolyse xylose residues, one unit at a time, from the non-reducing end of xylo-oligosaccharides.
α-L-Arabinofuranosidases (EC 3.2.1.55) can cleave the arabinofuranose residues from the xylan backbone with strict specificity in some cases for 3-mono-substituted or 2,3-di-substituted arabinofuranose bearing xylose residues.
Feruloyl esterases (EC 3.1.1.73) cleave the ferrulate crosslinks between arabinoxylan and other polymeric structures within hemicellulose.
Figure: Schematic representation of the hydrolytic action of arabinoxylan degrading enzymes.
| Product Code | Product Name |
| endo-1,4-β-Xylanases | |
| E-XYTR1 | endo-1,4-β-Xylanase M1 (Trichoderma viride) |
| E-XYTR3 |
endo-1,4-β-Xylanase M3 (Trichoderma longibrachiatum) |
| E-XYAN4 | endo-1,4-β-Xylanase M4 (Aspergillus niger) |
| E-XYRU6 | endo-1,4-β-Xylanase M6 (rumen microorganism) |
| E-XYNAP | Recendo-1,4-β-Xylanase (Aeromonas punctata) |
| E-XYNBS | Recendo-1,4-β-Xylanase (Bacillus stearothermophilus T6) |
| E-XYNACJ | Recendo-1,4-β-Xylanase (Cellvibrio japonicus) |
| E-XYNBCM |
Recendo-1,4-β-Xylanase (Cellvibrio mixtus) |
| E-XYLNP | Recendo-1,4-β-Xylanase (Neocallimastix patriciarum) |
| E-XYLATM |
Recendo-1,4-β-Xylanase (Thermotoga maritima) |
| exo-1,4-β-Xylosidases |
|
| E-BXSEBP | Recβ-D-Xylosidase (Bacillus pumilus) |
| E-BXSR |
Recβ-D-Xylosidase (Selenomonas ruminantium) |
| α-L-Arabinofuranosidases |
|
| E-AFASE | α-L-Arabinofuranosidase (Aspergillus niger) |
| E-ABFAN | α-L-Arabinofuranosidase (Aspergillus nidulans) |
| E-AFAM2 | Recα-L-Arabinofuranosidase (Bifidobacterium adolescentis) |
| E-ABFCJ |
Recα-L-Arabinofuranosidase (Cellvibrio japonicus) |
| E-ABFCT | Recα-L-Arabinofuranosidase (Cellvibrio thermocellum) |
| E-ABFUM |
Recα-L-Arabinofuranosidase (Ustilago maydis)NEW |
| E-ABFBO17 | Recα-L-Arabinofuranosidase (BACOVA 03417) (Bacteroides ovatus)NEW |
| E-ABFBO21 | Recα-L-Arabinofuranosidase (BACOVA 03421) (Bacteroides ovatus)NEW |
| E-ABFBO25 |
Recα-L-Arabinofuranosidase (BACOVA 03425) (Bacteroides ovatus)NEW |
| Feruloyl Esterases | |
| E-FAEZCT | RecFeruloyl Esterase (Clostridium thermocellum) |
| E-FAERU | RecFeruloyl Esterase (rumen microorganism) |
| Rec (recombinant enzyme) | |
Enzyme Substrates
Colourimetric oligosaccharides with a chemically-defined structure are available for the measurement of the activity of endo-xylanases, exo-xylosidases and α-L-arabinofuranosidases. These are suitable for use in both manual and high throughput automated analysis applications. The activity of the enzyme assayed can be determined directly by measuring the absorption increase at 400 nm which corresponds to the release of the nitrophenolate ion in solution.
| Product Code | Product Name | Chemical Structure | ||
| endo-1,4-β-Xylanase substrates | ||||
| O-PNPX2 | 4-Nitrophenyl-β- xylobioside |
 |
||
| O-PNPX3 | 4-Nitrophenyl-β- xylotriosideNEW |
|||
| exo-1,4-β-Xylanase substrates | ||||
| O-4MUX2 | 4-Methylumbelliferyl-β- xylobioside |
 |
||
| O-4MUX3 |
4-Methylumbelliferyl-β- xylobiosideNEW |
|||
| O-PNPX | 4-Nitrophenyl-β-D- xylopyranosideNEW |
 |
||
| α-L-Arabinofuranosidase substrates | ||||
| O-PNPAF | 4-Nitrophenyl-α- L-arabinofuranosideNEW |
 |
||
Insoluble and soluble chromogenic enzyme substrates derived from arabinoxylan and xylan (birchwood) have been prepared for the detection and specific measurement of endo-xylanase activity. These substrates produce a colourimetric response upon incubation with a suitable xylanase that can be related to enzyme activity through the standard curve provided (soluble substrates only) or used for screening on agar plates. The insoluble substrate is also available in tablet form to increase accuracy and convenience for the end-user for quantitative assays.
Figure: Schematic representation of the use of dyed polysaccharides (soluble) for the assay of endo-hydrolase activity.
| Product Code | Product Name |
| Tablets | |
| T-XAX | Xylazyme AX (very versatile, highly sensitive and widely used substrates for the assay of endo-xylanase) |
| T-XYZ | Xylazyme (very versatile, highly sensitive and widely used substrates for the assay of endo-xylanase) |
| Soluble Substrates | |
| S-AWAXL | Azo-Wheat Arabinoxylan (liquid) |
| S-AWAXP | Azo-Wheat Arabinoxylan (powder) |
| S-AXBL | Azo-Xylan (birchwood) (liquid) |
| S-AXBP | Azo-Xylan (birchwood) (powder) |
| Insoluble Substrates | |
| I-AZWAX | AZCL-Arabinoxylan (wheat) (useful for screening of endo-xylanase in gels and on agar plates) |
| I-AZXBW | AZCL-Xylan (birchwood) (useful for screening of endo-xylanase in gels and on agar plates) |
1 McCleary, B. V., McKie, V. A., Draga, A., Rooney, E., Mangan, D. & Larkin, J. (2015). Hydrolysis of wheat flour arabinoxylan, acid-debranched wheat flour arabinoxylan and arabino-xylo-oligosaccharides by b-xylanase, α-L-arabinofuranosidase and b-xylosidase. Carbohydrate Research, 407, 75-96.
