Fed-batch fermentation is a technique that is applied to processes in which a high substrate concentration is toxic to the culture. In such a case, the reactor is initiated in a batch mode with a low substrate concentration (usually 60-100 gL-1) and low fermentation medium volume, usually less than half the volume of the fermentor. The reactor is inoculated with the culture and the fermentation pro­ceeds. As the substrate is utilized by the culture, it is replaced by adding a concentrated substrate solution at a slow rate, thereby keeping the substrate concentration in the fermentor below the toxic level to the culture (Ezeji et al., 2004). When using this approach, the culture volume increases over time unless culture fluid is removed. The culture is harvested when the liquid volume is approximately 75% of the volume of reactor. Since butanol is toxic to C. aceto- butylicum and C. beijerinckii cells, the fed-batch fermentation technique cannot be applied in this case unless one of the novel simultaneous fermentation and product recovery techniques is applied. In a number of studies (Ezeji et al., 2004; Qureshi et al., 1992), this technique has been applied successfully to the ABE fermentation. In fed-batch fermentation, Ezeji et al. (2004) were able to utilize 500 g glucose in 1 L culture volume (500 gL-1) as compared to 60 gL-1 in a control batch process.

Qureshi et al. (1992) used a concentrated substrate of whey permeate (350 gL-1) to produce butanol in a fed-batch reactor of C. acetobutylicum. In this process, three different techniques of butanol separation were compared including perstraction, gas stripping, and pervaporation. In the three processes, 57.8, 69.1 and 42.0 gL-1 ABE were produced, respectively. ABE yield of 0.37, 0.38, and 0.34 were obtained, respectively. Overall, application of these three techniques suggested that fed-batch fermentation technique can be applied to the ABE fermentation provided ABE is removed from the culture broth simultaneously. In another study, Qureshi et al. (2001) produced ABE in a fed-batch fermentation of C. acetobutylicum and removed these solvents using a silicone-silicalite syn­thesized pervaporation membrane. The reactor was fed with 700 gL-1 glucose solution. In this system, 155 gL-1 ABE was produced with an average yield of 0.31-0.35 and productivities ranging from 0.13 to 0.26 gL-1h-1. Using the fed — batch technique, fermentation of more sugars (2-3 times) was possible when novel product removal techniques were applied to the process.