Standard sugars

The samples were ground and extracted with 5 mL 80% (v/v) ethanol for 30 min at 80°C, and the extraction was repeated 3 times. The extracts were pooled and dried at 40°C in an oven, and then decolorized with active carbon. The residues were re-dissolved in 1-mL distilled water and were filtered through the 0.45-μm filter. A high-performance liquid chromatography (HPLC) (Agilent 2100 system; Palo Alto, CA, USA) was used to analyze the sugar content. The retention times and peak heights of standard sugars (Sigma) were used to identify and quantify the eluted sugars. The precipitate was used to extract the starch. The residue was incubated with 10 mL of 30% (v/v) perchloric acid for 15 min and centrifuged at 3,500 g for 10 min. After 3 times centrifugation, the combined supernatants were analyzed with anthrone-H 2 SO 4 reagent, and starch content was calculated by the produced glucose content.

Compound 1 (0.3 mg) was heated in 1 M HCl (0.1 mL) at 90°C for 3 hours. After neutralization with NH 4 OH followed by extraction with CHCl 3 , the aqueous layer was evaporated in vacuo to give a crude sugar residue. The resulting residue was analysed by TLC using EtOAc-MeOH-H 2 O-AcOH (70-20-10-0.5) in comparison with standard sugars (Sigma, Germany). The spots of the product on TLC were identical to those of D-glucose (Rf 0.36), D-xylose (Rf 0.62) and L-arabinose (Rf 0.46)

Ion Chromatography was performed using a Dionex 5000+ fitted with a 4 x 250mm analytical CarboPac PA1 column. Flow rate was set to 1.0ml / min running 0-15min: 25mM NaOH, 15-20min: linear gradient of 25-75mM NaOH, 20-30min: 75mM NaOH with linear gradient of 0-260mM NaOAc, 32-34min: 75mM NaOH with 260mM NaOAc, 34-42min: 200mM NaOH, 42-52min: 25mM NaOH (Adapted from Kuhnel, 2012) .
Calibration was performed with standard sugars obtained from Sigma and made up to the desired concentration in ultrapure H2O. Monomeric sugar concentrations were calculated directly from the soluble fraction. Oligomeric sugars were first hydrolysed into their constituent monomers by the addition of 106µL of 72% H2SO4 to 3ml of each soluble fraction (in triplicate) and autoclaving for 1hr at 121°C. Samples were neutralised with solid CaCO3 and filtered before analysis. To calculate the carbohydrate composition of the insoluble residue from each pre-treatment; 100-300mg of the washed and dried material was acid hydrolysed by the addition of 3ml 72% H2SO4 for 1hr at 30°C, 150rpm. After 1hr, 84ml of ultrapure H2O was added and the sample autoclaved for 1hr at 121°C. Samples were neutralised with solid CaCO3 and filtered before analysis. Hydrolysis and analysis of all samples was repeated in triplicate.