Increasing concern on fossil fuels depletion and global environmental deterioration has recently renewed interest in butanol production from lignocellulose by acetone-butanol-ethanol (ABE) fermentation. Unfortunately, there was barely microorganism possessing the compound ability of lignocellulose decomposition and solvents production natively. Comparing with complicated genetic and metabolic engineering strategy to engineer native cellulolytic or solventogenic bacteria, mixed culture is a convenient and feasible approach for ABE fermentation by consolidated bioprocessing (CBP) from lignocellulose. But few communities of microorganisms succeeded in producing butanol of high titer or productivity without adding butyrate. Here a novel strategy for sequential co-culture of Clostridium thermocellum ATCC 27405 and Clostridium beijerinckii NCIMB 8052 was proposed to produce solvents efficiently in one pot reaction with alkali extracted corn cobs (AECC), a low-cost renewable feedstock, as the sole carbon source. In this strategy, soluble sugars accumulation by C. thermocellum hydrolyzing AECC was considered to be paramount for the CBP and was promoted considerably by contrast with previous co-culture studies. Under the combinatorial optimal culture parameters for sugars accumulation and ABE production, the CBP decomposed 88.9 g L-1 of AECC and manufactured ABE 19.9 g L-1 (acetone 3.96, butanol 10.9 and ethanol 5.04 g L-1) in 200 h without feeding butyrate. (C) 2014 Elsevier Ltd. All rights reserved.