Chemical synthesis of 1,3-propanediol (1,3-PD) is environmentally unfriendly and hence its microbial production is preferred, especially for biomedical, cosmetic and textile applications. In this work, production of 1,3-PD by co-fermentation of glucose and glycerol by Lactobacillus reuteri was investigated under different cultivation conditions such as aeration, acetate concentration and different molar ratios of glucose/glycerol. The final concentration of 1,3-PD and yield attained under unaerated conditions was close to that obtained under anaerobic conditions. Addition of acetate in the initial medium at 5 g/l increased the productivity of 1,3-PD but above this concentration it was found to be inhibitory. Batch reactor experiments showed that the molar ratio of glucose and glycerol in the medium affected the fermentation pattern. The effect of molar ratios was further investigated in fed-batch fermentation and the optimum ratio was found to be 1.5. In repeated fed-batch fermentation with co-feeding of glucose and glycerol in the molar ratio of 1.5, 1,3-PD concentration reached up to 65.3 g/l, which is the highest 1,3-PD concentration reported so far for this strain. The yield (0.97 mol/mol) based on glycerol utilized also approached the theoretical value (1 mol/mol). (C) 2014, The Society for Biotechnology, Japan. All rights reserved.