In order to obtain mutant strains showing higher solvent tolerance and butanol production than those of wild-type strains, the butanol-producing strain Clostridium beijerinckii L175 was subjected to mutagenesis using a combined method of low-energy ion beam implantation and N-methyl-N-nitro-N-nitrosoguanidine induction. With this effort, mutant strain MUT3 was isolated. When it was used for butanol fermentation in P2 medium, the production of butanol was 15.8 +/- 0.7 g/L 46% higher than the wild-type strain. Furthermore, after optimization of butanol production from cane molasses with MUT3, the maximum butanol production of 14.9 +/- 0.5 g/L were obtained in crew-capped bottles. When ABE production by MUT3 was carried out in a bioreactor, the production of butanol and total solvent were 15.1 +/- 0.8 g/L and 22.1 +/- 0.9 g/L, respectively. The remarkable butanol production and solvent tolerance of MUT3 make it promising for butanol production from cane molasses. (C) 2013 Elsevier Ltd. All rights reserved.