BACKGROUND Efficient production of (2R,3R)-2,3-butanediol (BDO) and acetoin using the GRAS strain Bacillus amyloliquefaciens from very high polarity (VHP) cane sugar and sugarcane molasses could lead to the development of a sustainable industrial process. RESULTS The effect of initial substrate concentration, nitrogen source, temperature, inoculum size and pH were evaluated in shake flask cultures. Batch bioreactor cultures showed that (2R,3R)-BDO production (up to 28 g L-1) with high yield (up to 0.43 g g(-1)) and productivity (up to 0.93 g L-1 h(-1)) was favored at low k(L)a values (17-49 h(-1)). Increasing k(L)a values (63-104 h(-1)) initiated acetoin production (up to 25.6 g L-1) with yields up to 0.4 g g(-1) and productivities up to 1.42 g L-1 h(-1). Fed-batch fermentation carried out at k(L)a value of 49 h(-1) with VHP cane sugar resulted in mixed production (127.3 g L-1) of (2R,3R)-BDO, (2R,3S)-BDO and acetoin at 122 h with high yield (0.50 g g(-1)) and productivity (1.04 g L-1 h(-1)). In a similar fed-batch fermentation with sugarcane molasses, mainly (2R,3R)-BDO production (48.7 g L-1 with purity of 87.8%) was achieved at 59 h with yield of 0.4 g g(-1) and productivity of 0.83 g L-1 h(-1). At 130 h, (2R,3R)-BDO was converted mainly into acetoin (55.4 g L-1). CONCLUSION This study demonstrates that k(L)a manipulation could be used in batch cultures to divert the bacterial metabolism towards either (2R,3R)-BDO or acetoin production. Fed-batch cultures with sugarcane molasses could lead to either (2R,3R)-BDO or acetoin production with negligible by-product formation. (c) 2019 Society of Chemical Industry