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[Display omitted] • Glycerol organosolv pretreatment could efficiently deconstruct corn stover. • The yield of glucose/xylose reached ∼80 % with glycerol pretreatment by cellulase. • l -cysteine-assisted glycerol pretreatment was developed for enhanced enzymolysis. • Fermentable sugars production reached 26.08 g/100 g by the proposed pretreatment. • Effects of raw materials prices on economic feasibility of processes were studied. Deconstruction of lignocellulose via efficient pretreatment is crucial for producing fermentable sugars. In this study, effects of glycerol organosolv pretreatment (GOP) on main chemical composition of corn stover were investigated. Results indicate that the residual corn stover after 80 wt% glycerol pretreatment (at 220 °C for 0.5 h) yielded 75.97 % glucose and 78.21 % xylose after enzymatic hydrolysis, which were enhanced by 3.39- and 6.08-fold compared to the untreated corn stover. Subsequently, an l -cysteine-assisted GOP was proposed with higher yields of glucose (86.20 %) and xylose (91.13 %). When pretreating corn stover with 80 wt% glycerol containing 0.07 wt% l -cysteine at 220 °C for 0.5 h, higher fermentable sugars of 26.08 g were produced from 100 g feedstock after enzymolysis. Intrinsic mechanisms of the proposed pretreatment for enhancing enzymatic digestibility were elucidated by physiochemical characterization technologies and techno-economic analysis was also studied. This study provides guidance for fermentable sugars production from renewable lignocellulose. [ABSTRACT FROM AUTHOR]