Figure 1. Reactor configuration for the starch liquefaction process. (Termamyl is a bacterial thermophilic α-amylase) (Olsen, 1995).

Figure 2. Flow scheme of glucose isomerization process with parallel fixed bed reactors, charcoal absorber, ion exchange column, and evaporator(with kind permission of Liese et al. 2000/2006, 2006)

Figure 3. Whole dry grind process for bioethanol manufacture (DDGS: Dried Distillers Grains with Solubles) (Kunz, 2007).

Figure 4. Production facility for bioethanol manufacture. The figure shows ethanol tanks (left), milling, saccharification (behind), fermentation station (background, left), and grain silos (background, middle).with kind permission of Suedzucker AG, Germany.

Figure 5. Five-step process for the conversion of biomass to ethanol(Merino and Cherry, 2007, reproduced with permission from Springer Science + Business Media).

Figure 6. Positioning of the cellulose fibrils in wood (left) and cotton fibers (right) (from outer to inner position). Wood fibers: (M) middle lamella (lignin and hemicelluloses); (P) primary wall (fibril position unarranged); (S ₁) secondary wall I (two or more fibrillar layers crossing one another and positioned spirally along the fiber axis); (S ₂) secondary wall II (fibrils wound spirally around the fiber axis); (S ₃) secondary wall III (fibrils tightly interlaced). Cotton fibers: (P) primary wall (interlaced fibrils); (S) secondary wall (fibrils wound spirally around the fiber axis; at distinct distances along the fiber axis the spiral reverses direction) (Krässig et al., 2004).

Figure 7. Schematic view of the primary T. reesei enzymes involved in hydrolysis of cellulose. Cellulose is represented as stacked chains of black circles with reducing (R) and nonreducing (NR) ends indicated. There are two major cellobiohydrolases that attack the cellulose chain ends processively from the reducing (CBH I) and nonreducing (CBH II) ends of the chain, releasing the glucose disaccharide cellobiose. In addition, there are three major endoglucanases depicted (EGI, II, and III) that attack the cellulose chain randomly, and two ß-glucosidases (BG) that hydrolyze cellobiose released by the CBHs to glucose. Triangles represent cellulose binding motifs, and the arrow represents an additional hypothetical protein component that may assist in cellulase action by disrupting the cellulose crystal structure(Merino and Cherry, 2007, reproduced with permission from Springer Science + Business Media).

Figure 8. Conversion-time behavior of nonpretreated Avicel at optimal ratio of activities of β-glucosidase/cellulase 1 : 20(Reproduced with permission of Elsevier ©2008, from Bommarius et al., 2008); T = 50 °C, pH 5.0; V = 8 mL. (Filled circle) 1.5 U cellulase; (filled square) 15 U cellulase; (filled triangle) 30 U cellulase. I, II, and III denote the three kinetic phases identified.

Figure 9. Schematic flowsheet for the production of ethanol using acid-catalyzed steam pretreatment followed by enzymatic hydrolysis/SSF of cellulose-containing materials(Galbe and Zacchi, 2007, reproduced with permission from Springer Science + Business Media).

Figure 10. Biosynthesis of penicillins and cephalosporins used for the production of semisynthetic β-lactam antibiotics. Note the change in isomer structure of the amino acids given in brackets (Ingolia and Queener, 1989; Crawford et al., 1995; Weil et al., 1995).

Figure 11. Enzymatic and chemical production of semisynthetic penicillins and cephalosporins from 6-APA, 7-ACA, and 7-ADCA, by hydrolysis of the fermentation products given in Figure 10. The by-products phenylacetate and adipate can be recycled in the fermentation. The amounts produced are estimated from literature data (Bruggink, 2001; Elander, 2003).

Figure 12. Alternative processes for the production of (R)-7-ADCA from isopenicillin N in metabolically engineered cells. (a) In cells where the oxidation of (R)-aminoadipyl-7-ADCA has been blocked, only the part of the ring that is expanded is shown (Adrio et al., 2002). (b) In cells with expandase but without the racemase that catalyzes isomerization of the aminoadipyl-group in isopenicillin N (Crawford et al., 1995; Bruggink, 2001).

Figure 13. Flowsheet for the production of 7-ACA, with concentrations of the precursors after each processing step. The increase in concentration after the chromatographic step is due to the use of displacement chromatography or nanofiltration. The recycled and nonrecycled waste is shown; d-CephC is deacetoxy-cephalosporin C and (R)-AAA is (R)-amino-adipic acid.

Figure 14. Chemical (left) and enzyme technological new one-step (center) and older (right) two-step processes to produce 7-aminocephalosporanic acid (7-ACA) from cephalosporin C. (Note that the charges in the acidic and basic functional groups of the compounds at the process pH are not given.) The one-step enzyme process has now replaced the two-step enzyme process.

Figure 15. Temperature and pH dependence of the apparent equilibrium constant (K _app = [glutarate][7-ACA]/[Glu-7-ACA]) for the hydrolysis of 100 mM Glu-7-ACA. (a) pH dependence at 30 °C with phosphate buffers with NaCl (I = 1.5 M); (b): temperature dependence at pH 8.0 in phosphate buffer (I = 0.2 M) (Spieß et al., 1999). The estimated K _app for the hydrolysis of CephC at pH 8.5 and 25 °C is given by X, and its pH dependence by the dotted line (Shin et al., 2005).

Figure 16. pH and temperature dependence of the apparent first-order rate constant (k ₁ + k ₂[7-ACA]) with [7-ACA]) = 100 mM for the reactions (1)–(3) that reduce the yield of 7-ACA.

Figure 17. The half-life of wild-type glutaryl amidase (blank circle) and penicillin amidase (filled circle) produced in E. coli as function of pH (25 °C, buffer with I = 0.2 M).

Figure 18. The pH-T process window for the hydrolysis of 100 mM CephC in 1 h is the overlapping part of the optimal biocatalyst and reaction windows (³ 95% hydrolysis). The upper temperature is limited by the inactivation of 7-ACA and not by the biocatalyst properties. The process temperature can be increased by reducing the hydrolysis time, to keep the 7-ACA loss constant.

Figure 19. A differential reactor with a high content of immobilized enzyme and minimal abrasion. The system consists of a mixing vessel for pH control and one or more parallel fixed bed reactors through which the substrate solution is pumped until the end point of the reaction is reached. The temperature in both the mixing vessel and the packed beds must be controlled. With this reactor system, short hydrolysis times, to reduce the loss of 7-ACA, can be kept within a desired time interval that cannot be achieved with a single reactor. The productivity (kg product/used kg immobilized biocatalyst) is also increased. This illustrated for 20 reactors each filled with (V _max,0/10), where V _max,0 is the biocatalyst amount required to hydrolyze a given amount of CephC solution in a given time. At first only 10 reactors are used. When they have lost 10% of their activity, the 11th reactor is added for the hydrolysis, when these 11 reactors have lost 10% of their activity, the 12th reactor is added, and so on. When after the addition of the 20th reactor, the reactors have lost 10% of their activity, the first of the first 10 reactors used is replaced by a reactor with unused biocatalyst. When this reactor system has lost 10% of its activity, the second of the first 10 reactors is replaced, and so on. With this reactor system, the hydrolysis times for each batch differ maximally by 10%. When the first used 10 reactors have been replaced by reactors with unused biocatalyst, the immobilized biocatalyst is used for more than two half-lifetimes. The interval within which the hydrolysis time varies is increased and the productivity reduced when the number of reactors is decreased.

Figure 20. Structure of atorvastatin calcium salt.

Figure 21. Route to optically pure (R)-4-cyano-3-hydroxy-butyric acid starting from readily available epichlorohydrin using a nitrilase.

Figure 22. An aldolase (DERA) catalyzes the direct formation of two stereocenters from acetaldehyde and chloroacetaldehyde with subsequent formation of a hemiacetal.

Figure 23. Route to optically pure (R)-4-cyano-3-hydroxy-butyric acid starting from a prochiral ketone using a combination of a ketoreductase (KRED), a glucose dehydrogenase (GDH), and a halohydrin dehalogenase (HHDH). The reduction was performed with 240 g ketone at pH 7 in a buffer (570 ml) containing 100 mM triethanolamine and butyl acetate (370 ml) for 8 h at 25 °C. Both enzymes (854 mg KRED and 578 mg GDH) were added as lyophilized powders. After addition of Celite and filtration, the product was extracted with butyl acetate yielding 96% alcohol. The subsequent cyanation step was performed in an airtight reactor (due to the presence of gaseous HCN) at pH 7.3 and 40 °C with 1.03 g HHDH (supplied as aqueous solution) with 70 g alcohol in a ~400 ml volume. After complete conversion and removal of residual HCN, the mixture was treated with Celite and bleach followed by filtration and product extraction with ethyl acetate yielding 93% final product (Ma et al., 2010).