AbstractThe rapidly expanding market for biodiesel is dramatically altering the cost and availability of glycerol, as typical biodiesel production processes generate about 10 wt % glycerol. Biodiesel producers have little financial incentive to purify the crude glycerol, and because it contains methanol, crude glycerol is considered a hazardous waste. Therefore, it is critical that new and innovative processes for managing and utilizing the glycerol co‐product be developed. In the longer term, as supplies continue to increase, glycerol will become a versatile building block chemical for the production of high value compounds within an integrated biorefinery.This research investigated the value‐added conversion of crude glycerol generated during biodiesel production using anaerobic fermentation. Pure cultures of Clostridium pasteurianum have been shown to produce significant amounts of butanol and 1,3‐propanediol when utilizing purified glycerol as the sole substrate. Butanol is of particular interest as a renewable biofuel, as it has a higher heating value, higher octane number, lower vapor pressure, and higher miscibility than ethanol, making butanol preferable to ethanol for blending with petroleum fuels.Preliminary experiments compared growth and product formation of C. pasteurianum (ATCC®6013™) utilizing both purified and biodiesel‐derived crude glycerol as the sole carbon source in batch culture. Cultures utilizing crude glycerol demonstrated significant growth and production of butanol, 1,3‐PDO, and ethanol at concentrations up to 25 g/L crude glycerol. This work is significant in that it is the first report of butanol production from crude glycerol using this organism. The maximum yield of butanol produced from pure glycerol was 0.36 g/g, while the maximum butanol yield produced from crude glycerol was 0.30 g/g. These yields are substantially higher than the 0.15–0.20 g/g butanol yield typically achieved during the fermentation of glucose using C. acetobutylicum. These results indicate that biodiesel‐derived crude glycerol is a promising, low‐cost, renewable feedstock for butanol production. © 2009 American Institute of Chemical Engineers Environ Prog, 2009