Governments want to replace fossil fuels with biofuels like ethanol and biodiesel due to concerns about pollution and price instability. However, the biodiesel industry must find a market for glycerol to be profitable. The large quantities and reduced state of its carbon have made glycerol an attractive choice for ethanol production. While Escherichia coli can produce ethanol anaerobically, cell growth requires expensive complex nutrients. To bypass this issue, we used microaerobic conditions and maintained 91% recovery of carbon into products. We identified the pathways involved in the microaerobic metabolism of glycerol using genetic and biochemical tools and further engineered the optimized anaerobic E. coli strains for microaerobic conditions. Greater cell growth and product synthesis were achieved by overexpressing glycerol dehydrogenase and dihydroxyacetone kinase. The final strain DeltafrdADeltaptaDeltaldhA (EH05) pZSKLMg1dA was tested at high glycerol levels where it produced almost exclusively ethanol in minimum media at superior volumetric rates.