Some bacteria have the capacity to produce ethanol in significant amounts mak­ing them potential microorganisms for industrial purposes (see Table 6.2). Among bacteria, the most promising microorganism is Zymomonas mobilis. This facul­tative anaerobe presents higher ethanol yields than yeasts, which is related to the metabolic pathways involved. Z. mobilis makes use of the Entner-Doudoroff pathway converting 1 mol of hexose into 2 mol of ethanol, but releasing only 1 mol of ATP (Jeffries, 2005; Figure 6.3), unlike S. cerevisiae that uses the Embden — Meyerhoff-Parnas pathway (i. e., the glucolytic way) but forms two molecules of ATP (see Figure 6.1). This fact implies a lower cell yield due to the lower energy yield of this bacterium, increasing the amount of ethanol that can be obtained from the same amount of substrate compared to yeasts. There have been reported ethanol yields of 97% of the theoretical yield from glucose. Furthermore, this bacterium has a more rapid fermentation due to its higher ethanol production rate (three to five times higher than in S. cerevisiae) and substrate consumption

rate. Additionally, this bacterium has a high ethanol tolerance (100 g/L) and a higher optimal production temperature. Z. mobilis requires no controlled addition of oxygen and is much more susceptible to genetic manipulations to enhance its yield or transfer new traits than is the case of yeasts.

Among the drawbacks of Z. mobilis, the too narrow range of fermentable sub­strates (glucose, fructose, and sucrose) should be noted (Claassen et al., 1999; Hawgood et al., 1985). Other disadvantage of the use of this bacterium for fermen­tation of sugarcane syrup and other sucrose-based media is the formation of the polysaccharide levan (made up of fructose units), which increases the viscosity of fermentation broth, and of sorbitol, a product of fructose reduction that reduces the efficiency of the conversion of sucrose into ethanol (Doelle and Doelle, 1989; Grote and Rogers, 1985; Lee and Huang, 2000). In addition, preculture conditions have a significant influence on bacterium performance, especially on sucrose hydrolysis rate. Hence, the addition of invertase to the culture medium has been proposed (Doelle and Doelle, 1989). Lee and Huang (2000) studied batch ethano — lic fermentation using Z. mobilis through nonstructured models based on meta­bolic analysis. These models allowed the use of ethanol and sorbitol formation during cultivation on a medium containing glucose and fructose or on a sucrose medium supplemented with immobilized invertase.

Other bacteria that have been investigated in order to implement processes of direct conversion of lignocellulosic biomass into ethanol are thermophilic and saccharolytic clostridia. Clostridium thermohydrosulfuricum, C. thermosaccha- rolyticum, and C. thermocellum can synthesize up to 2 mol EtOH/mol hexose. Likewise, these bacteria may transform pentoses and amino acids into ethanol. Having saccharolytic properties, these microorganisms have the ability to grow on a wide variety of nontreated wastes. C. thermocellum can even directly con­vert lignocellulosic materials into ethanol (McMillan, 1997). In this way, ethanol can be directly obtained from pretreated lignocellulosic biomass without the need of adding costly cellulases. Moreover, the cultivation of these microorganisms at high temperatures offers the possibility of an easier ethanol removal by distilla­tion or pervaporation and reduced cooling expenditures (Claassen et al., 1999). Clostridium thermocellum has been the most studied thermophilic clostridium because it has the capacity to produce cellulases, hydrolyzing the cellulose and fermenting the glucose forming ethyl alcohol. On the other hand, the possibility of employing C. thermosaccharolyticum to produce ethanol from pentoses result­ing from hemicellulose degradation during the pretreatment of biomass has been shown (Wyman, 1994).

The main drawback of these bacteria consists in their very low ethanol toler­ance compared to yeasts. Consequently the maximum reached ethanol concentra­tions are lower than 30 g/L. In addition, they exhibit a reduced ethanol yield due to the formation of fermentation by-products like acetic acid and lactate that make the final ethanol concentrations very low and cultivation times prolonged (3 to 12 days) (Baskaran et al., 1995; McMillan, 1997; Szczodrak and Fiedurek, 1996; Wyman, 1994). Process integration can play a crucial role for evaluating the most optimal configurations in order to implement them at an industrial level.