The objective of this work was the convertion of glycerol to dihydroxyacetone (DHA) by a fed-batch process at different glycerol concentrations, 50 and 100 g/l, using Gluconobacter oxydans.The fermentation was conducted during 72 h, at a pH 5.3 and 30°C under a flow of 1.66 vvm aeration.The amounts of produced DHA and consumed glycerol, were determined using gas chromatography and UV-VIS spectrophotometry.Product-substrate yields were obtained to 91% and 79% for concentrations of glycerol 50 g/l and 100 g/l, respectively, which showed the inhibition phenomenon to high substrate concentrations and reduced biomass growth.This research shows that the production of dihydroxyacetone from glycerol is feasible and inexpensive Therefore, it is possible to solve, future problems about pollution and accumulation of glycerol.