127 Example 6.2 A chemostat study was performed with yeast. The medium flow rate was varied and the steady-state concentration of cells and glucose in the fermenter were measured and recorded. The inlet concentration of glucose was set at 100 g/é. The volume of the fermenter contents was 500 mé. The inlet stream was sterile. Flow rate Cell Concentration Substrate Concentration F, mé/br Cx, g/t Cs, g/t 31 5.97 0.5 50 5.94 1.0 71 5.88 2.0 91 5.76 4.0 200 0 100 a. Find the rate equation for cell growth. b. What should be the range of the flow rate to prevent washout of the cells? Solution: a, Let’s assume that the growth rate can be expressed by Monod kinetics. If this assumption is reasonable, the plot of 1/y versus 1/Cs will result in a straight line according to Eq. (6.35). The dilution rate for the chemostat is F v The plot of 1/D versus 1/Cs is shown in Figure 6.9 which shows a straight line with intercept D= = 3.8 maz and slope As 55 maz Therefore, mer = 0.26hr-!, and As = 1.37g/é. The rate equation of cell growth is _ 0.26 Cs Cx ™ = T3T+Cs b. To prevent washout of the cells, the cell concentration should be main- tained so that it will be greater than zero. Therefore, from Eq. (6.33) Cx = Yxys (cs - att) >0 maz — 1142 alr (a) (b) Figure 6.20 Loop fermenters: (a) air-lift, (b) ICI pressure cycle, (c) stirred loop, and (4) jet loop. can maintain a high air-flow rate per unit area at the lower section of the fermenter where the cell concentration is high. Several sieve plates can be in- stalled in the column [Figure 6.19(c)] for the effective gas-liquid contact and the breakup of the coalesced bubbles. The cylindrical column can be divided into multiple stages which are equipped with stirrers [Figure 6.19(d)]. This configuration will be analogous to the stirred-tank fermenter connected in se- ties as explained in a-earlier section. To enhance the mixing without internal moving parts, the fermentation broth can be pumped out and recirculated by using an external liquid pump [Figure 6.19(e) and (£)]. 6.8.2 Loop Fermenter A loop fermenter is a tank or column fermenter with a liquid circulation loop, which can be a central draft tube or external loop. Depending on how the liquid circulation is induced, it can be classified into three different types: air-lift, stirred loop, and jet loop (Figure 6.20). The liquid circulation of the air-lift fermenter is induced by sparged air which creates a density difference between the bubble-rich part of the liquid in the riser and the denser bubble-depleted part of the liquid in the downcomer143 as shown in Figure 6.20(a). The ICI pressure cycle fermenter (Imperial Chemical Industries Ltd. England) is an air-lift fermenter with an outer loop. which was developed for the aerobic fermentation requiring heat removal such as the single-cell protein production from methanol. Medium and air are introduced into the upper and lower parts of the loop as shown in Figure 6.20(b). The air serves two purposes: It provides the oxygen needed for the growth of the microorganisms and the rising air creates natural circulation of the liquid in the fermenter through the loop. A heat exchanger to cool the liquid medium is installed in the loop. It was claimed that the fermenter gives a high rate of oxygen absorption per unit of volume, that it uses a high proportion of oxygen in the air passed through the fermenter, and that the high circulation of the fermentation liquor provides good mixing (Technical Brochure, ICI Ltd.). The liquid circulation and mixing can be enhanced by installing a pro- peller or by circulating liquid externally using a pump as shown in Fig- ure 6.20(c) and (d). However, adding the propeller or pump diminishes the real advantages of an air-lift fermenter for being simple and energy efficient.Nomenclature Q2aQb Ci, Cr Fs baATOO 4 & ca ARE EKO SS flow rate of bleeding stream, m*/s concentration, mass per unit volume of culture, kg/m* intracellular concentration, mass per unit volume of biotic phase, kg/m* extracellular concentration, mass per unit volume of abiotic phase, kg/m? constants cell number density, number of cells/m* dilution rate, s~! flow rate, m°/s system coefficient for the Monod kinetics, kg/m* average mass of cell in a system, kg flow rate of filtrate stream, m3/s mass of biotic phase, kg number of cells rate of growth per unit volume, kg/m’s rate of jth component formed from ith reaction per unit volume of a system, intracellular property, kg/m*s rate of jth component formed from ith reaction per unit volume of a system, extracellular property, kg/m's time, doubling time, s specific volume of a system containing only biotic phase, m?/kg working volume of fermenter, m* yield constant bleeding ratio, defined as B/F average cell division rate, s~? specific growth rate, s? or kg/m?s density, kg/m* residence time, s SUBSCRIPT SuM sae batch fermenter input stream mixed fermenter cell in number basis cell in dry weight basis product plug-flow fermenter145 s substrate Problems 6.1 Derive the relationship giving the change with respect to time of the cell concentration in a batch fermenter, Eq. (6.24). 6.2 Aiba et al. (1968) reported the results of a chemostat study on the growth of a specific strain of baker’s yeast as shown in the following table. The inlet stream of the chemostat did not contain any cells or products. Dilution Inlet Steady-state Steady-state Steady-state Rate | Glucose Cone. Glucose Conc. Ethanol Conc. Cell Conc. D, bro? Cs, g/t Cs, g/t Co, g/é Cx, g/é 0.084 215 0.054 7.97 2.00 0.100 10.9 0.079 4.70 1.20 0.160 21.2 0.138 8.57 2.40 0.198 20.7 0.186 8.44 2.33 0.242 10.8 0.226 4.51 1.25 a. Find the rate equation for cell growth b. Find the rate equation for product (ethanol) formation. 6.3 Andrews (1968) proposed the following model for the growth of microor- ganisms utilizing inhibitory substrates. Assume that a chemostat study was performed with a microorganism. The volume of the fermenter content was 1. The inlet stream was sterile. The flow rate and inlet substrate concentration were varied and the steady-state concentration of glucose in the fermenter was measured146 and recorded as follows (the data are arbitrary) Flow Inlet Rate Glucose Concentration Glucose Concentration F.t/hr 0.20 30 0.5 0.25, 30 0.7 0.35 30 1 0.50 30 16 0.70 30 3.3 0.80 30 10 0.50 60 30 0.60 60 22 0.70 60 15 a. Determine the kinetic parameters (fmez. Ks, and Ky) of this mi- croorganism. b. If the cell yield, ys, is 0.46 g/g, what is the steady-state cell con- centration when the flow rate is 0.20 ¢/h? c. Andrews concluded in his paper that the primary result of sub- strate inhibition in a continuous culture may be process instability. Explain what might happen if you suddenly increase the substrate concentration from 30 to 60.¢/é and why. 6.4 Rate equations for the cells (yeast), substrate (glucose), and product in the ethanol fermentation process are given as follows: 1 &e\" (6s (0-) ra) 7 = Os 1 ax S “dt ~~ Yxj5 dt = ie = 2 tx pe “dt Yujp dt _ where Ks =1.6g/0, ptmaz = 0.24bt™, Yuyp = 0.16, Yxys = 0.08, Chg = 100 g/2, Ca, =0, Cx, = 0.1/2, andn=2 a. Calculate the change of Cx, Cp, and Cs as a function of time when Cs = 100 ¢/8. b. Show the effect of the initial substrate concentration on the Cx versus ¢ curve.147 c. Show the effect of the maximum growth rate (jimez) on the Cx versus t curve (Cs, = 100 g/2) 6.5 Derive Eqs. (6.39), (6.41), and (6.42) 6.6 The growth rate of E. coli in synthetic medium can be expressed by 6.7 6.8 Monod kinetics as nw 0:985CsCx * 0.714 Cs where Cs is the concentration of a limiting substrate, glucose. You are going to cultivate E. coli in a steady-state CSTF (working volume: 10 ¢) with a flow rate of 7é/hr. The initial substrate concentration is 10 g/é and the cell yield constant (Yx,s) is 0.6. The feed stream is sterile. (g/¢hr] a. What will be the doubling time and the division rate of the cells in the CSTF? b. What will be the cell and substrate concentrations of the outlet stream? c. If you connect one more 10-€ CSTF to the first one, what will be the cell and substrate concentrations in the second fermenter? d. If you increase the flow rate from 7 to 10/hr for these two fer- menters connected in series, what will happen and why? Make a recommendation to avoid the problem if there is any. Suppose that the growth rate of a microorganism can be expressed as the following equation: Ty = Mmar(1 ~ e795/Ks) Cy where fmaz = 0.365 hr! and K's = 6.8¢/é. The cell yield Yx/s is found to be 0.45 a. If you cultivate this microorganism in a 10 CSTR with the flow rate of 2.8£/hr, what will be the steady-state cell concentration of the outlet stream? The substrate concentration of the inlet stream is 13g/€. The inlet stream is sterile. b. Explain the difference between this model and Monod model by using ji versus Cs graph. Herbert et al. (1956) reported that the growth kinetics of Aerobacter cloacae in a chemically defined medium (glycerol as a limiting substrate) could be expressed by Monod kinetics as follows: — dx _ HmazCsCx at Ks + Cs148 where plmar = 0.85hr-! and Ks = 1.23 x 107 g/f. The yield was found to be 0.53g dry weight of organism/g glycerol used You are a biochemical engineer who has been assigned the task of de- signing the most effective continuous fermentation system to grow the microorganism (Aerobacter cloacae) with glycerol as its limiting sub- strate. For the following three questions, the concentration of glycerol in the feed stream and that of glycerol in the outlet stream should be 3g/é and 0.1 g/2, respectively. a. Since you have learned that the 1/r, versus Cs curve for Monod kinetics has 2 U shape, you have recommended that the most effec- tive system would be the combination of a continuous stirred-tank fermenter (CSTF) and a plug-flow fermenter (PFF). You were quite sure of this because the substrate concentration in the outlet stream has to be so low. However, your boss is insisting that the use of second PFF in addition to the first CSTF will not improve the pro- ductivity very much. Who is right? Prove whether you are right or wrong by drawing the 1/ry versus Cs curve for this microorganism. Does it have a U shape? Discuss why you are right or wrong. (If you are right, think about how you can nicely correct you boss's. wrong idea. If you are wrong, it will teach you that you have to be careful not to make a quick conclusion without adequate analysis.) b. Recommend the best fermenter system (fermenter type and vol- ume) which can handle 100 ¢/hr of feed stream. The best fermenter system is defined as that which can produce the maximum amount of cells per unit time and volume c. If Ks = 1.23g/E instead of 1.23 x 10-?g/¢, what is the best fer- menter system (fermenter type, volume) which can handle'100 £/hr in the feed stream. Draw the block diagram of the fermenter system with the concentrations of the substrate and the cells in the inlet and outlet streams of each fermenter. How is this case different from the case of part (a) and why? 6.9 Suppose you have an organism that obeys the Monod equation: dCx _ UmezCsCx dt Ks+Cs where [maz = 0.5 hr-} and Ks = 2g/¢ The organism is being cultivated in a steady-state CSTF, where F = 100 £/hr, Cs, = 50 g/2, and Yxjs = 0.5. a. What size vessel will give the maximum total rate of cell produc- tion?6.10 6.12 149 b. What are the substrate and cell concentrations of the optimum fermenter in part (a)? c. If the exiting flow from the fermenter in part (a) is fed to a second fermenter (CSTF), what should be the size of the second fermenter to reduce the substrate concentration to 1 g/é? d. If the exiting flow from the first fermenter in part (a) is fed to a second fermenter whose size is the same as the first, what will be the cell and substrate concentrations leaving the second fermenter? | You are going to cultivate yeast, Saccharomyces cerevisiae, by using a 10 m3-fermenter your company already owns. You’want to find out the amount of ethanol the fermenter can produce. Therefore, a chemostat study was carried out and the Monod kinetic parameters for the mi- croorganism grown in the glucose medium at 30°C, pH 4.8, were found to be: Ks = 0.025 ¢/2 and pimar = 0.25h-). The ethanol yield (Yp)s) is 0.44 (g/g) and cell yield (Yx,s) is 0.019 (g/g). The inlet substrate concentration is 50 g/ a. What flow rate will give the maximum total ethanol production in the continuous fermenter and what is the maximum ethanol pro- duction rate? b. If you want to convert 95 percent of the incoming substrate, what must the ethanol production rate be for the continuous fermenter? c. If you have two 5m?-fermenters instead of one 10 m*-fermenter, what is your recommendation for the use of these fermenters to con- vert 95 percent of the incoming substrate? Would you recommend connecting two fermenters in series to improve the productivity? Why or why not? You are a biochemical engineer in a pharmaceutical company. Your com- pany is a major producer of penicillin. Currently, what kind of fermenter is your company using for penicillin production? Why? Your boss asked you to study the possibility of using an air-lift fermenter as a replacement since it has many advantages. What is your recommendation? Consider an organism with the following data for a 1-¢ chemostat, using150 an inlet substrate concentration Cs, of 30 g/£: Concentration Flowrate Substrate Cell Product F(mé/hr) Cs (g/£) Cx (g/2) Cp (¢/8) 27.5 10.0 12.0 1.04 24.2 5.56 14.7 1.27 22.1 3.70 15.8 1.37 18.8 2.32 16.7 1.44 a. In a continuous, perfectly mixed vessel at steady state with no cell death, if inlet substrate concentration Cs, now equals 25.0 g/¢ and cell concentration Cx, is 0.0 g/£, what dilution rate D will give the maximum total rate of cell production? What are the outlet cell and substrate concentrations at this dilution rate? b. Using the preceding organism, your boss would like you to design a continuous reactor system with an inlet flow and substrate concen- tration of 250 é/hr and 25 g/£, respectively, which will produce an overall yield of product of 2100 kg/yr, given an operating time of 300 day/yr at 24 hr/day. Assume no cells or product in the system inlet. Would you recommend a single fermenter of two fermenters in series? Design a system which will meet production constraints while minimizing total fermenter volume. Report reactor volumes and effluent cell, substrate, and product concentrations for the pro- posed fermenter(s). [Contributed by Brian S. Hooker, TriState Uni- versity.] 6.13 A strain of yeast is being cultivated in a 30-@ CSTF with a cell recycling system (cell settler) as shown in the following figure. The cell settler was designed so that the cell concentration of its outlet stream is 30 percent of that of its inlet stream, whereas the substrate concentrations of the two streams are the same. The growth rate of the cells can be represented by the Monod kinetics with the parameters: Ks = 0.05¢/£, Umar = 0.3h-?, and Yx/s = 0.025. Calculate the steady-state substrate and cell concentrations in the fermenter. The inlet substrate concentration is 100 g/é and the flow rate is 20 £/hr. The feed stream is sterile.151 hy 0 € far, 00g/e one 6.14 A plug-flow fermenter is to be used to cultivate microbial cells. It has been determined that the fermenter efficiency can be improved by recy- cling a portion of the product stream so that it returns to the entrance for an additional pass through the fermenter. The recycle rate (R) is defined as olume of fluid returned to entrance vi R= volume leaving the system a. Show that an optimal recycle rate must satisfy nLtRU = X54) _ R+i RQ —Xs;) RIL + RIL — Xsy)] where Xs; is the fraction of a limiting substrate S that is converted to cell mass. The optimal recycle rate corresponds to the minimum- sized reactor needed to attain a desired level of conversion. b. Determine the recycle ratio needed to minimize reactor size for frac- tional conversion of Xs = 0.995. 6.15 By using the structured model proposed by Ramkrishna et al. (1967), show the change of the concentrations [in g dry weight/é] of G-mass, H-mass, and inhibitor, and the fraction of G-mass with time during a batch cultivation of a microorganism which has the following parameters152 and initial conditions: 0.5 hr" a's = 2 2.5hr7? ar =0 Ks =0.2¢/€ an = 0.2 x 10-4 Ks = 0.1g/e ar, = 0.0267 K = 150 ¢/ghr K' = 70 /ghe Ko =3.0 x 10 g/t g/é Kg =0.5% 10% g/€ Oxy = 8.0 X 10-8 g/l as =8 Cxgq = 1.0 x 10° g/t Compare your simulation result with Figure 14 of the paper by Ram- krishna et al. (1967). If you showed the change of the cell concentrations in g dry weight/liter, the shape of the curves are quite different from those in the paper. What are the differences? Explain. Do the param- eter values predict realistic growth curves? What new features can this model predict which the Monod model cannot?Chapter 7 Genetic Engineering The central tool for the new biotechnology is the recombinant DNA tech- nique.! It allows direct manipulation of genetic material of individual cells. By inserting foreign genetic information into fast-growing microorganisms, we can produce foreign gene products (proteins) with higher rates and yields that have not been possible with any other cellular systems. This technology is also known as genetic engineering because it involves the manipulation of genetic materials.? In this chapter, basic principles involved in recombi- nant DNA technology and problems involved in cultivating the genetically engineered cells are briefly described. 7.1 DNA and RNA Deoxyribonucleic acid (DNA) is the most important molecule in living cells and contains all of the information that specifies the cell. DNA and ribonu- cleic acid (RNA) are macromolecules that are linear polymers built up from simple subunits, nucleotides.? The monomeric unit, nucleotide, has the fol- lowing three components (Figure 7.1): 1. A cyclic five-carbon (pentose) sugar: deoxyribose for DNA, and ribose for RNA. 2. A nitrogenous base of either purine or pyrimidine derivation, covalently attached to the 1'-carbon atom of the sugar by an N-glycosylic bond as shown in Figure 7.1. a. The purines: adenine (A) and guanine (G). b. The pyrimidines: cytosine (C), thymine (T) for DNA only, and uracil (U) for RNA only. 3. ‘A phosphate attached to the 5' carbon of the sugar by phosphoester linkage. 1 Read Chapter 1 for a general introduction to the new biotechnology. 2 ‘The name genetic engineering should not mislead the readers that it is a field of engineering; it is a field of biological science. 3 Macromolecules: a polymer, especially one composed of more than 100 repeated monomers.Figure 6.9 The plot of 1/D versus 1/Cs for Example 6.2 Solving the preceding equation for Tm yields V _ Kst+Cs, m= >> > FO Cs;tmaz Therefore, VC5;pmaz _ 0.5(100)(0.26) F =e Ox, Figure 6.15 Schematic diagram of the multiple CSTFs connected in series. is also better than one CSTF. Figure 6.15 shows the schematic diagram of the multiple CSTFs connected in series. For the nth steady-state CSTF, the material balance for the microorganisms can be written as F(Cx,_1 — Cxn) + Vatx, = 0 (6.49) where HmazCSnCXn eee 6.50) Tm = Ks + Os, (6.50) Growth yield can be expressed as Cx, — Cx, Yus = Ge Cs ao By solving Eqs. (6.49), (6.50), and (6.51) simultaneously, we can calculate either dilution rate with the known cell concentration, or vice versa. The estimation of the cell or substrate concentration with the known dilution rate can be done easily by using graphical technique as shown in Figure 6.16 (Luedeking, 1967). From Eq, (6.49), the dilution rate of the first reactor when the inlet stream is sterile is F_ it, Dasa 6.52 eae (6.52 which can be represented by the slope of the straight line connecting the origin and (Cx, ,ry,) in Figure 6.16. Similarly, for the second fermenter Ee ey ~ Cx, — Cx, Cx, which is the slope of the line connecting (Cx,,0) and (Cx,,ry,). Therefore, by knowing the dilution rate of each fermenter, you can estimate the cell concentration of each fermenter, or vice versa. (6.51) Dr= | (6.53) SSF135 2 4Cx, Oxr6 Cx Figure 6.16 Graphical solution for a two-stage continuous fermentation. The line represents the Monod model with maz = 0.935, Ks = 0.71 8/8, ¥xys = 0.6, Cs, = 10¢/2, Cx, = 0. Example 6.3 Suppose you have a microorganism that obeys the Monod equation: dCx _ UmazCsCx dt Ks+Cs where mez = 0.7hr-! and Ks = 5g/£. The cell yield (Yx;s) is 0.65. You want to cultivate this microorganism in either one fermenter or two in series. The flow rate and the substrate concentration of the inlet stream should be 500 é/hr and 85 g/é, respectively. The substrate concentration of the outlet stream must be 5/8. a. If you use one CSTF, what should be the size of the fermenter? What is the cell concentration of the outlet stream? b. If you use two CSTFs in series, what sizes of the two fermenters will be most productive? What are the concentration of cells and substrate in the outlet stream of the first fermenter? c. What is the best combination of fermenter types and volumes if you use two fermenters in series? Solution: a. For a single steady-state CSTF with a sterile feed, the dilution rate is equal to specific growth rate: F _ pmezCs _ 0.7(5) P= T= Kas 545The cell concentration of the outlet stream is Cx = Yx/s(Cs, — Cs) = 0.65(85 — 5) = 52g/é b. For two CSTFs in series, the first fermenter must be operated at Cx,op and Csopt. Cs, B cx eb Cs, v Lj ly Cs, Therefore, from Eq. (6.39) through Ex, (6.42), a= [Ks+Cs, _ 5+85 _ 49 Ks 5 Cx, = Cope = YaysCs.2 = 0.65(85) oS z= ‘elt Os, = Csint = wy - re = 16e/e Ti = Tmopt = maleny = Wey = 1.9hr V; = tm = 1.9(500) = 950£ For the second fermenter, from Eq. (6.49), F(Cx, - Cx) + a =0 By rearranging the preceding equation for V2, F(Cx,—Cx,) ____ 500(52 — 45) Va = 192£ mazOs,Cx,](Ks + Cs) 0.7(5)(52)/(5 +5) The total volume of the two CSTFs is V=VY+V, = 950 +192 = 1,1422137 which is 20 percent smaller than the volume required when we use a single CSTF. c. The best combination is a CSTF operated at the maximum rate followed by a PFF. The volume of the first fermenter is 950 ¢ as calculated in part (b). For the second PFF, from Eq. (6.48) 1 [( KsYxis +1) ln Cx KsYxis r | Ty maz (\Cx, + Cs, Yx/s Cx, * Oy + Os Yas Cs, _ 1 ff 5(0.65) 52, 5(0.65) | 16] _ 07 (= + 16(0.65) * 1) nig + wae 5 | ~o Therefore, Ve = traF = 0.32(500) = 1606 The total volume of the CSTF and PFF is V=VY+ = 950+ 160 =1,1108 which is 22 percent smaller than the volume required when we use a sin- gle CSTF. The additional saving by employing the second PFF instead of a second CSTF is not significant in this case. 6.7 CSTF with Cell Recycling The cellular productivity in a CSTF increases with an increase in the dilution rate and reaches a maximum value. If the dilution rate is increased beyond the maximum point, the productivity will be decreased abruptly and the cells will start to be washed out because the rate of cell generation is less than that of cell loss from the outlet stream. Therefore, the productivity of the fermenter is limited due to the loss of cells with the outlet stream. One way to improve the reactor productivity is to recycle the cell by separating the cells from the product stream using a cross-flow filter unit (Figure 6.17). ‘The high cell concentration maintained using cell recycling will increase the cellular productivity since the growth rate is proportional to the cell concentration. However, there must be a limit in the increase of the cel- lular productivity with increased cell concentration because in a high cell concentration environment, the nutrient-transfer rate will be decreased due to overcrowding and aggregation of cells. The maintenance of the extremely high cell concentration is also -not practical because the filter unit will fail more frequently at the higher cell concentrations. If all cells are recycled back into the fermenter, the cell concentration will increase continuously with time and a steady state will never be reached138 Figure 6.17 Schematic diagram of CSTF with cell recycling. Therefore, to operate a CSTF with recycling in a steady-state mode, we need to have a bleeding stream, as shown in Figure 6.17. The material balance for cells in the fermenter with a cell recycling unit is aCx dt It should be noted that actual flow rates of the streams going in and out of the filter unit do not matter as far as overall material balance is concerned. For a steady-state CSTF with cell recycling and a sterile feed, FCy, -— BCx +V Cx =V (6.54) pp=fa=p (6.55) Tm where 8, the bleeding ratio, is defined as pad (6.58) Now, BD instead of D is equal to the specific growth rate. When # = 1, cells are not recycled, therefore, D=p. If the growth rate can be expressed by Monod kinetics, substitution of Eq, (6.11) into Eq, (6.55) and rearrangement for Cs yields BKs Cs = ————_, 6.57 5 Finbmas — B (6.57) which is valid when Tmimaz > 9. The cell concentration in the fermenter can be calculated from the value of Cs as Cx = HE(Ca,— Cs) (6.58) Figure 6.18 shows the effect of bleeding ratio on the cell concentration and productivity. As @ is reduced from 1 to 0.5, the cell concentration and productivity is doubled.139 Cell Cone., ¢/€ hr Cellular Prod., g/€ Figure 6.18 The effect of bleeding ratio on the cellular productivity. 6.8 Alternative Fermenters Many alternative fermenters have been proposed and tested. These fer- menters were designed to improve either the disadvantages of the stirred tank fermenter — high power consumption and shear damage, or to meet a specific requirement of a certain fermentation process, such as better aera- tion, effective heat removal, cell separation or retention, immobilization of cells, the reduction of equipment and operating costs for inexpensive bulk products, and unusually large designs. Fermenters are usually classified based on their vessel type such as tank, column, or loop fermenters. The tank and column fermenters are both con- structed as cylindrical vessels. They can be distinguished based on their height-to-diameter ratio (H/D) as (Schiigerl, 1982): H/D <3 for the tank and H/D>8 for the column fermenter A loop fermenter is a tank or column fermenter with a liquid circulation loop, which can be a central draft tube or an external loop. Another way to classify fermenters is based on how the fermenter con- tents are mixed: by compressed air, by a mechanical internal moving part, or by external liquid pumping. Representative fermenters in each category are listed in Table 6.3 and the advantages and disadvantages of three basic fermenter types are listed in Table 6.4. .140 Table 6.3 Classifications of Fermenters Primary Source of Mixing Vessel Compressed Internal External Type Air | Mowng Parts Pumping Tank - | stirred-tank - Column | bubble column | multistage sieve tray tapered column | (or cascade) | packed-bed Loop | air-lift | propeller loop | jet loop pressure cycle | ' Table 6.4 Advantages and Disadvantages of Three Basic Fermenter Configurations r T Type Advantages | Disadvantages 1. Flexible and adaptable Stirred- 2. Wide range of mixing | tank intensity cells 3. Ability to handle high 3. High equipment costs viscosity media High power consumption we Damage shear sensitive 1. No moving parts | 1. Poor mixing Bubble 2. Simple | 2. Limited to low Column | 3. Low equipment costs | viscosity system 4. High cell concentration | 3. Excessive foaming. 1. No moving parts | 1. Poor mixing 2. Simple 2. Limited to low efficiency 3. Excessive foaming 4. Good heat transfer | Atr-lift 3. High gas absorption | viscosity system | 6.8.1 Column Fermenter The most simple fermenter is the bubble column fermenter (or tower fer- menter), which is usually composed of a long cylindrical vessel with a sparg-141 / | | 1 7 alr \ (a) (o) Figure 6.19 Column fermenters: (a) bubble column, (b) tapered bubble col- ‘umn, (c) sieve-tray bubble column, (d) multistage stirred column, (e) sieve- tray column with external pumping, and (f) packed-bed column with ex- ternal pumping. {c) {d) (e) (f ing device at the bottom [Figure 6.19(a) ~ (c)]. The fermenter contents are mixed by the rising bubbles which also provide the oxygen needs of the cells. Since it does not have any moving parts, it is energy efficient with respect to the amount of oxygen transfer per unit energy input. As the cells settle, high cell concentrations can be maintained in the lower portion of the column without any separation device. However, the bubble column fermenter is usually limited to aerobic fer- mentations and the rising bubbles may not provide adequate mixing for opti- mal growth. Only the lower part of the column can be maintained with high cell concentrations, which leads to the rapid initial fermentation followed by a slower one involving less desirable substrates. As the cell concentration increases in a fermenter, high air-flow rates are required to maintain the cell suspension and mixing. However, the increased air-flow rate can cause exces- sive foaming and high retention of air bubbles in the column which decreases the ptoductivity of the fermenter. As bubbles rise in the column, they can coalesce rapidly leading to a decrease in the oxygen-transfer rate. Therefore, column fermenters are inflexible and limited to a relatively narrow range of operating conditions. To overcome the weaknesses of the column fermenter, several alternative designs have been proposed. A tapered column fermenter [Figure 6.19(b)}