Although a number of high-value products have been produced using glycerol, production of ethanol from glycerol provides a unique opportunity for the biodiesel industry to generate another biofuel from biodiesel waste. E. aerogenes HU101 isolated from methanogenic sludge produced ethanol and H2 when glycerol was used as a substrate (Ito et al. 2005). Low amounts of lactate, acetate, formate, and 1,3-PDO were also detected as coproducts. A maximum ethanol yield of 0.85 mol/mol-glycerol and concentration of 90mM was reported following anaerobic fermentation of glycerol (Ito et al. 2005). Although promising, E. aero- genes needs yeast extract and tryptone for enhanced glycerol utilization. The highest yield and productivity were achieved in a packed-bed reactor that used porous carrier matrix as support material to immobilize microbial cells (Ito et al. 2005).

Escherichia coli has been reported to metabolize glycerol fermentatively under favorable culture conditions (Dharmadi et al. 2006). Cells growing fermentatively on glycerol exhibited exponential growth at a maximum specific growth rate of 0.040 per hour (Murarka et al. 2008). Cell growth was not affected despite blocking of several respiratory processes, demonstrating the fermentation of glycerol by E. coli.

Pathways responsible for fermentative metabolism of glycerol in E. coli have been revealed (Gonzalez et al. 2008). E. coli mutants with disrupted respiratory genes glpD and glpA have been shown to ferment glycerol, indicating the existence of alternative pathways for the metabolism of glycerol in the absence of electron acceptors such as oxygen and fumarate. E. coli encodes a type II glycerol dehydrogenase enzyme, GldA, which was earlier thought to be cryptic in nature without any significant physiological role in wild-type strains (Jin et al. 1983). Nonetheless, this enzyme has the potential to oxidize glycerol into dihydroxyacetone (DHA) that can further be converted to DHAP by DHA kinase. When gldA (encoding GldA) and dhaKLM (encoding DHA kinase) mutants were evaluated for their ability to ferment glycerol, they failed to do so. This observation underscores the active role of GldA and DHA kinase in the glycerol fermentation pathway. In addition to this oxidative pathway for metabo­lizing glycerol, a parallel reductive pathway acting as an electron sink was also discovered in the form of 1,2-PDO (Figure 6.6). Disruption of the 1,2-PDO synthesis pathway decreased cell growth on glycerol, while overexpression of this pathway led to cell growth without any supplementation of rich media (Gonzalez et al. 2008; Murarka et al. 2008).