32 α l arabinofuranosyl xylobiose

The oligosaccharide profile after enzymatic hydrolysis of different substrates was investigated applying HPAEC–PAD with Dionex ICS-5000 system (Thermo Fisher Scientific) using a Dionex CarboPac PA200 (250 × 3 mm, 5.5 μm) analytical and a guard (50 × 3 mm) columns (Thermo Fisher Scientific). The separation was performed using a constant mobile phase composition of 100 mM NaOH at 0.5 mL/min, and a linear gradient from 0 to 30 min of 0–120 mM sodium acetate and thereafter a constant concentration of 160 mM sodium acetate until 40 min. AXOS standards xylobiose (X²), xylotriose (X³), xylotetraose (X⁴), xylopentaose (X⁵), xylohexaose (X⁶), 3²-α-L-arabinofuranosylxylobiose (A³X), 2³-α-L-arabinofuranosyl-xylotriose (A²XX), 3³-α-L-arabinofuranosyl-xylotetraose (XA³XX), 2³-α-L-arabinofuranosyl-xylotetraose (XA²XX) and 2³,3³-di-α-L-arabinofuranosyl-xylotriose (A²⁺³XX) were purchased from Megazyme.

All buffering chemicals and substrates including glucose, xylose, mannose, galactose, cellobiose, lactose, mannobiose, β-(1 → 4)-_D-galactobiose, isomaltose and maltose were purchased from Sigma-Aldrich (Germany). The other substrates including cellotriose, cellotetraose, xylobiose, xylotriose, xylotetraose, 3²-α-l-arabinofuranosyl-xylobiose (A³X), 2³-α-l-arabinofuranosyl-xylotriose (A²XX), 2²-(4-O-methyl-α-d-glucuronyl)-xylobiose (U^4m2X), 2³-(4-O-methyl-α-d-glucuronyl)-xylotriose (U^4m2XX), chitosanbiose, 3²-β-d-glucosyl-cellobiose, β-(1 → 4)-_D-glucosyl-_D-mannose, gentiobiose, kojibiose, sophorose, laminaribiose, and nigerose were purchased from Megazyme (Ireland). The laccase from T. versicolor (38429, Sigma-Aldrich, Germany) was used in oxidation reactions to recycle the electron acceptor, 1,4-benzoquinone (BQ, PHR1028, Sigma-Aldrich, Germany). Methylated cellobiose and lactose was produced in house (details in Additional File 2).

Xylobiose (XOS2) was purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan), and xylotriose (XOS3) was purchased from FUJIFILM Wako Pure Chemical Corporation (Osaka, Japan). The other oligosaccharides used in this study were purchased from Megazyme (Wicklow, Ireland): xylotetraose (XOS4), xylopentaose (XOS5), xylohexaose (XOS6), 3²-α-l-arabinofuranosyl-xylobiose (AXOS1), 2³,3³-di-α-l-arabinofuranosyl-xylotriose (AXOS2), 2³-α-l-arabinofuranosyl-xylotriose (AXOS3), and 3³-α-l-arabinofuranosyl-xylotetraose (AXOS4). A schematic of these structures is represented in Fig. S2 in the supplemental material. AXH were prepared as follows: 50 ml of 2% (wt/vol) AX (Megazyme) in 100 mM phosphate buffer (pH 6.5) was supplemented with 75 U of endo-xylanase derived from Cellvibrio mixtus (Megazyme); this solution was incubated at 40°C for 16 h and boiled for 10 min to inactivate the enzyme. The boiled solution was added to 200 ml of ethanol, which was then evaporated to obtain the AXH. XOS-95P, which included XOS2 (33.2%), XOS3 (13.78%), and XOS with a degree of polymerization of ≧4 (46.29%) (31 ), was obtained from B Food Science Co., Ltd. (Aichi, Japan).