This study aims to efficiently use hemicellulose-based biomass for ABE (acetone-butanol-ethanol) production by co-fermentation with starch-based biomass. Two processes were investigated: (I) co-fermentation of sugars derived from hemicellulose and starch in a mixture of barley straw and grain that was pretreated with dilute acid; (II) co-fermentation of straw hemicellulosic hydrolysate and gelatinized grain slurry in which the straw was pretreated with dilute acid. The two processes produced 11.3 and 13.5 g/L ABE that contains 7.4 and 7.8 g/L butanol, respectively. In process I, pretreatment with 1.0% H2SO4 resulted in better ABE fermentability than with 1.5% H2SO4, but only 19% of pentoses were consumed. In process II, 95% of pentoses were utilized even in the hemicellulosic hydrolysate pretreated with more severe condition (1.5% H2SO4). The results suggest that process II is more favorable for hemicellulosic biomass utilization, and it is also attractive for sustainable biofuel production due to great biomass availability. (C) 2014 Elsevier Ltd. All rights reserved.