AbstractBamboo is a potential lignocellulosic biomass for the production of bioethanol because of its high cellulose and hemicellulose contents. An acid-free ethanosolv process was proposed to overcome the problems caused by the acid catalysts commonly used in organosolv processes. In this research, ethanosolv pretreatment catalyzed by NaOH was used to enhance the enzymatic saccharification of moso bamboo. The addition of 10% (w/w on bamboo) NaOH in 75% (v/v) ethanol was demonstrated to be effective in the pretreatment and fractionation of bamboo. The pretreatment yielded a solid fraction with 60.1% cellulose. The cellulose-to-glucose conversion yield was 28.9% to 45.1%, depending on pretreatment conditions, after enzymatic hydrolysis of the solid fraction at 50 °C for 48 h using enzyme loading (15 filter paper units of cellulase/g cellulose and 30 IU b-glucosidase/g cellulose). The concentrations of fermentation inhibitors such as 5-hydroxy-2-methyl furfural (HMF) and furfural were negligible in the spent liquor after the ethanosolv pretreatment and were much lower than those in the spent liquor from H2SO4-water or ethanosolv only treatment.