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Abstract: A two-stage pretreatment method was proposed and tested for deconstruction of recalcitrant Miscanthus. During a 1st pretreatment at 50°C, 1.0–4.0% alkaline peroxide solutions were used to partially remove hemicellulose and lignin. The remaining solids were subjected to a 2nd pretreatment at 121°C with electrolyzed water. Using 15 filter paper units of cellulase per gram cellulose, a digestibility of 95% was achieved by the two-stage method, which was higher than 81% obtained from a 1% H2SO4 pretreatment (200°C, 8min). A mass balance for the two-stage process showed that 63% hemicellulose and 64% lignin were removed from the Miscanthus samples after the 1st pretreatment. The Scanning Electron Microscopy images and Fourier Transform Infrared Spectroscopy spectra revealed that after the 1st pretreatment, crystalline cellulose fibers were partially exposed and the linkages between residual lignin and hemicellulose disappeared. After the 2nd pretreatment, cellulose fibers were completely exposed even with cracks developed. [Copyright &y& Elsevier]

[Display omitted] • A ball-mill assisted alkaline peroxide pretreatment process was developed. • 70% of XOS alone with high yield of fermentable sugars was achieved. • The pretreatment process was environmental-friendly. • The mechanism of enhanced enzymatic digestibility by BAP pretreatment was revealed. The aim of this work was to develop a feasible ball mill-assisted alkaline peroxide pretreatment followed by stepwise hydrolysis to improve the yield of xylo -oligosaccharides (XOS) and fermentable sugars. The hydrogen peroxide charge, ball-milling time, and solid-to-liquid ratio affected the compositions, particle sizes, morphology, and crystallinity of the corn stover, directly improving the following hydrolytic efficiency. The optimal pretreatment was with 0.45 g/g (H 2 O 2 : substrate) and 1:3 solid-to-liquid ratio (w/v) for 1.0 h ball-milling, resulting in 84.29% delignification. Physicochemical properties of the pretreated samples were characterized and their correlations to the enzymatic hydrolysis were revealed. Compared with one-step cellulase hydrolysis, the two-step xylanase-cellulase hydrolysis of the pretreated corn stover showed significant advance in preparing XOS, producing 69.65% (on the base of xylan content in pretreated sample) of XOS, along with 20.55% xylose, 68.94% glucose, and 21.15% gluco -oligosaccharides. The yield of XOS was 2–7 times higher than those in previous studies. [ABSTRACT FROM AUTHOR]