We have reviewed critical features of the key unit operations necessary to economically operate bioethanol production plants; however, these technologies cannot be used to plan or model actual production plants until the process is integrated at reasonable scale. This view is commonly overlooked by researchers and planners involved in the important details of unit operations. Pilot plant verifications of biomass to ethanol processes have "discovered" critical problems that result from fully integrated operation that were not realized at bench or simulated integrated operation, especially contamination of cellulase production and SSF unit operations (164, 73) and mass/02 transfer problems that result from lignocellulose utilization at large scale (73). Other unforeseeable scale-up issues may have equal impact on process economics; for example, the problems encountered in scaling pretreatment equipment are reflected (to some extent) only by experience in the pulp and paper industry.

For bioethanol production processes, in which yield is crucial to economic operation (78), all process-related diversions of fermentable sugars from ethanol are catastrophic. In the pretreatment section, for example, effects of wear and normal aging on equipment contacted by hot, acidic biomass are difficult to model Char buildup on piping and reactor surfaces may pose a threat to biomass yield from unpredicted adsorption or condensation reactions, as well as the better understood effects of heat transfer reduction.

Another aspect of fully integrated plant operation not always considered is the consequence of allowing sugars, especially mono — and disaccharides, extended residence times following initial prehydrolysis or enzyme saccharification (765). Reducing sugars are reactive under virtually all conditions encountered in a biomass conversion facility! Mildly acidic conditions and moderate temperatures (i. e., conditions consistent with those following dilute acid pretreatment and storage) permit reversion and transglycosylation of glucose to generate populations of all possible configurations of alpha — and beta-linked disaccharides (i. e., 1-1, 1-2, 1-3, 1-4, and 1-6) and even higher-order oligomers in yields that approach 10% (766). Many of these oligosaccharides are not fermentable, and the nonreducing sugars, such as 1-1 beta — and alpha-linked glucose (a — and p-trehalose), are quite stable even after dilution and neutralization. When enzymes are added to the process, as in SHF or hybrid SSF schemes, reversion and transglycosylation reactions produce all possible configurations of disaccharides consistent with the anomeric requirement of the enzyme; i. e., a-glucosidase produces only alpha products and P-glucanases produce only beta products, but many linkage combinations are possible. Clear precedent exists for efficient production (20% to 37% yield) of transglycosylation products from 10% solutions of glycosyl donor (cellobiose) using a Fusarium oxysporum P-glucosidase (767) and from 10% solutions of maltose using an A. niger a-glucosidase (168). Enzymatic transglycosylation is also possible at glycosyl donor concentrations as low at 2% (769). Because most amylases and cellulases have active sites that can tolerate some diversity in the glycosyl acceptor group, it may be possible to find disaccharides or higher forms of glucose-xylose, glucose-galactose, and even glucose-mannose in biomass hydrolysis process streams. Of further concern to bioethanol plant efficiency are the observations that transglycosylation reactions that involve glycosyl transferases have been reported that utilize non-carbohydrate species as glycosyl acceptors, including alcohols (methanol ethanol and propanol) (767) and lignin model compounds (veratryl and vanillyl alcohols) (170). An unfortunate conclusion easily drawn from the literature is that the precise nature (chemical composition) of reversion and transglycosylation products formed during proposed bioethanol process operations is not currently known, nor is the resultant impact of this loss on fermentable sugars taken into account in process models. One critical reason for this dilemma lies in the advanced level of analytical capability required to properly detect and identify these products in biomass processing streams. A related and contributing problem is the inability of most laboratories to adequately assess fermentation process mass balances; thus, failure to achieve theoretical yields is often attributed to microbial or enzymatic performance problems, not to the presence of nonfermentable and unassayable sugars.