Although isobutanol has been successfully produced from glucose-based media by various microbes, only a few studies have reported isobutanol production from lignocellulosic biomass. In this study, the feasibility of producing isobutanol from hydrothermally-pretreated empty fruit bunches (EFBs) was analyzed. Firstly, the Escherichia coli JK209 strain was constructed; subsequently, optimal fermentation conditions for isobutanol production using this strain were determined. Next, the raw EFBs were hydrothermally pretreated at 190 degrees C for 15 min, and the pretreated EFBs were hydrolyzed using 10 FPU of cellulase/g glucan. Finally, the resulting hydrolysate was fermented at 25 degrees C for 84 h, and isobutanol give rise to the yield of 0.26 g isobutanol/g glucose. These results imply that 80.1 g of isobutanol could be produced from 1 kg dry weight of EFBs. To our knowledge, this is the first feasibility study of isobutanol production from pretreated EFBs. (C) 2020 Elsevier Ltd. All rights reserved.