Scale-Up Methodologies For Yeast Fermentation Processes: Escherichia Coli and

JOURNAL OF BIOSCIENCE AND BIOENGINEERING
Vol. 97, No. 6, 347–364. 2004
Scale-Up Methodologies for Escherichia coli and

Yeast Fermentation Processes
BETH HELENE JUNKER1
Merck Research Laboratories, Bldg. 810-127, PO Box 2000, Rahway, NJ 07065, USA 1
Received 22 August 2003/Accepted 15 March 2004
Scale-up techniques from the literature have been compiled and reviewed for applicability to
Escherichia coli and yeast processes. The consistency of design and operating parameters for
the pilot scale vessels in an existing fermentation pilot plant, ranging in nominal volume from
100 l to 19,000 l, was established and compared favorably with approaches found in the
literature. Differ- ences were noted as a function of parameters such as fermentor scale, vessel
geometry, agitator type/size and ungassed/gassed power input. Further analysis was conducted
using actual fermentation data for historical and recent development processes collected over
a 10-year-period, focus- sing on operating conditions at peak culture oxygen uptake rates.
Scale-up estimates were performed based on geometric similarity, agitator tip speed, gassed
power per unit volume and mixing time. Generally, scale-up calculations from the 280 l
scale were most similar to the parameters of installed equipment. Scale-up from the 30 l
laboratory scale typically underpredicted parameters with scale-up from the 280 l scale being
most appropriate. The 19,000 l fermentor installation was notably different in geometric
similarity from the 280 l–1900 l scales since its design was meant to accommodate a wide
range of operating volumes. Analysis of historical and recent processing performance was
conducted for single cell bacterial or yeast fermentations which challenged peak operating
conditions of the fermentors. Identification of key issues associated with scale-up for these
specific pilot plant vessels was believed to be critical to efficient process development,
clinical material production, and expected process transfer to a manufacturing facility.
[Key words: scale-up, Escherichia coli, yeast, pilot plant, fermentor]
General concerns for recombinant DNA scale-up have well. Additionally, the role of scale-down studies can be
been addressed by Van Brunt (1). Although the three main significant in identifying and evaluating problems at the
scales for bioprocess development are laboratory, pilot plant large scale (8–11).
and production (2), Votruba and Sobotka have added the Biological factors affected by scale include the number
shake-flask scale to this list (3). The scale-up ratio is typi- of generations associated with the inoculum development
cally about 1 : 10 for biotechnological processes up to and production phases, mutation probability, contamination
100,000 l (3), but lower ratios of about 1 : 5 often have vul- nerability, pellet formation and selection pressure (8,
been used for increased comfort levels (i.e., decreased risk 12). Chemical factors affected by scale include (i) pH
of unexpected performance on scale-up). Production scale control agents (i.e., type and concentration of acid and/or
for many recombinant DNA products is likely to be about base), medium quality (i.e., purity of components) and
10,000 l which is more traditionally pilot scale for other water quality (8); (ii) carbohydrate (e.g., oil), nitrogen (e.g.,
types of products (4). The exact methodology used for ammonia), phosphorus and product concentrations (6); (iii)
scale- up is largely dependent on process conditions and redox potential and foam formation due to surface tension
whether preliminary data exist to show that the procedure changes (3). Physical factors affected by scale include tank
chosen is applicable (5). configuration, aeration, agitation, back-pressure (and
Some authors maintain that the total environment (Young hydrostatic pressure), medium sterilization, temperature
[6] coined this term to encompass all the chemical and control/heat transfer and removal, and mixing (3, 8, 12).
physical variables relating to the fermentor broth) of the There is a comprehensive paper outlining general ap-
cell needs to be considered (6). A complete catalog of proaches to scale-up (13) and it includes comments from
factors affected by scale is detailed extensively by Reisman several other authors on the implications of general trends
(7) with key items being emphasized by other authors (1– in scale-up. Unless specifically maintained constant with
6) as scale-up to the larger fermentor, dissolved carbon dioxide
(dCO2) is higher in the larger vessel than the smaller vessel
e-mail: [email protected] due to the added head pressure, shear force variation is
phone: +1-732-594-7010 fax: +1-732-594-7698
347
348 JUNKER J. BIOSCI. BIOENG.,
higher, and bulk mixing is less efficient due to longer circu- gen transfer. For the scale-up of toyocamycin production by
lation times and larger stagnant regions (8). In addition, for a shear-sensitive mutant of Streptomyces
the larger vessel, the liquid height to tank diameter ratio can chrestomyceticus, neither the constant Pg/VL method nor
be greater, gas moves upwards with more limited backmix-
ing, vertical dissolved oxygen (DO) gradients exist if bulk the constant OTR method could be used, and thus scale-up
mixing is slower than mass transfer rates, and radial DO was done at the lowest possible tip speed for the
gradients emerge since the shear rate declines rapidly with geometrically similar larger vessel (20). Mixing and
distance from the impeller (14). Other trends in scale-up in- circulation times were found to be more important than
clude decreased heat transfer surface to volume ratio, de- using constant oxygen uptake rate (OUR) for the scale-up
creased quality of mixing, generally higher superficial air of a 2,3-butanediol fermentation by Enterobacter
velocity, and lower average shear force although peak shear aerogenes under microaerobic conditions in which
forces are higher (14, 15). Furthermore, if cheaper medium homogeneity was important (21). Although, in general, the
components have been selected with variable composition scale-up based on dCO2 would be even more feasi- ble as
for different bulk lots, a previously unnoticed auxotrophic more reliable dCO2 sensors become available (16), the
growth pattern may appear (15), but this occurrence is mini- specific carbon dioxide evolution rate was currently still felt
mized by the use of defined medium. to be useful for scale-up of secondary metabolite cultures
There are several published descriptions of scale-up stud- (22).
ies and a few interesting examples are noteworthy. Oxygen The purpose of this paper is to briefly describe and evalu-
transfer often can be most important upon scale-up due to ate various scale-up methods and approaches which can be
its low solubility in medium (16). Key to scale-up using applied to example E. coli and yeast processes. The first
constant oxygen transfer rate (OTR) is the ability to mea- step involved cataloging characteristic measurements for
sure or estimate the volumetric mass transfer coefficient, existing laboratory (30 l scale) and pilot plant (100 l–
KLa, and the gassed power per unit liquid volume, Pg/VL. 19,000 l scale) fermentors (Tables 1 and 2). Next, relevant
Published correlations can generate significant errors as in parameters were calculated and compared as a function of
the example of KLa factors for a commercial-scale scale based on geometry, power input per unit volume, gas
penicillin fermentation (17). The scale-up of bialaphos flow rate and mixing time (Tables 3–5). Various scale-up
production by Streptomyces hygroscopicus based on KLa scenar- ios were explored based on maintaining geometric
and Pg/VL was not successful due to the large DO similarity (Table 6), constant impeller speed (Table 7), and
constant power input per unit liquid volume (Tables 8 and
concentration gradients in the fermentor (18). In this 9). A comparison was also completed based on medium
example, the culture was not sensitive to impeller tip speed heat stress during sterilization (Table 10). A listing of
changes upon scale-up. Scale- up was successful when the historical and recent achievable processing conditions was
target DO in the laboratory fermentor was used to control developed (Tables 11 and 12), and the relationship of OUR
the large scale fermentor DO using a probe located at the to KLa and Pg/VL to KLa was quantified for various
bottom of the large scale fermentor. Similarly, scale-up
deteriorated production syndrome (SUDS) was observed processes (Table 13). For consistency, all volumes quoted in
during the scale-up of milbemycin production by S. this text are nominal vessel volumes (rather than total or
hygroscopicus when agitation rate was used to control DO working volumes) unless otherwise stated. Units were
as was done at the small scale (19). SUDS syndrome selected to minimize rounding errors where possible.
included culture morphological changes which affected
packed cell volume and viscosity, carbon source uptake I. SCALE-UP METHODS AND APPROACHES
changes, and decreased productivity. An alternative DO
control strategy using aeration rate, back-pressure, tem- Several scale-up methods are described below, and se-
perature, in addition to agitation rate, was developed to lected scale-up approaches were evaluated using two scales
maintain laboratory yields upon scale-up. as a basis: the 280 l pilot scale which is a common first step
pilot plant fermentor scale and the 30 l laboratory scale
Additional examples focus on parameters other than oxy- which is common to use for pre-pilot plant (laboratory)
stud-
TABLE 1. Characteristics of laboratory and pilot scale vessels

Vessel Vessel
Scale Working Installed Max. Vessel total height width, Max.
(nominal volume) tank volume, motor power, agitator speed, tangent/tangent, (with top and OD, air flow rate,
VL (l) Po (hp) Nmax (rev/min) HTT (m) bottom dish), DT (m) Qmax (l/min)
HT (m)
30 l 20 1 875 0.66 0.742 0.31 30
100 l 75 3 400 0.785 0.96 0.41 (ID) 120
280 l 180 7.5 460 1.12 1.34 0.56 300
800 l 600 7.5 330 1.63 2.08 0.81 600
1000 l 750 10 300 1.52 1.88 0.86 (ID) 1200
1200 l 900 15 282 1.83 2.185 0.92 1200
1900 l 1500 15 230 2.13 2.57 1.07 1500
19000 l 15000 60 155 5.87 6.58 1.98 15000
The 280 l scale fermentor was originally designed with a 3 hp motor which was enlarged to 7.5 hp for replacement convenience. OD, Outer di-
ameter; ID, inner diameter.
TABLE 2. Characteristics of laboratory and pilot-plant scale impellers
Scale Impeller type Impeller diameter, Impeller tip speed, Number of
(nominal volume) (upper/lower) DI (m) nNmaxDI (m/s) impellers, NI
30 l Rushton 0.102 4.7 4
100 l Hydrofoil 0.184 3.8 2
A315
280 l Rushton 0.205 4.9 2
Hydrofoil 0.28 6.7 3
Maxflo T
800 l Rushton 0.305 5.4 2
Maxflo T
A315
Smith Top, 0.343 Top, 6.1 2
CD-6 Btm, 0.356 Btm, 6.3
Hydrofoil/Smith Top, 0.406 Top, 7.2 2
Maxflo T/CD-6 Btm, 0.394 Btm, 7.0
HE-3/Smith Top, 0.533 Top, 9.4 2
HE-3/CD-6 Btm, 0.394 Btm, 7.0
1000 l Rushton 0.305 4.8 2
A315
1200 l Hydrofoil 0.46 6.8 2
A315
1900 l Rushton 0.42 5.1 3
Maxflo T
19000 l Rushton 0.685 5.6 4
Maxflo T
Hydrofoil Top/Mid, 0.94 7.6 3
A315 (Btm, 8.4) (Btm, 8.4)
Impeller data for top/bottom impellers. Units of length selected to minimize rounding errors. Top, Top impeller; Mid, middle impeller; Btm, bot-
tom impeller.
TABLE 3. Geometric comparisons for laboratory and pilot scale fermentors

Scale H /D D /(V )1/3 at V (l), Re× 106 VL/VT
(nominal volume) TT DI/DT T L L (water)
(HTT/DT) geometric similarity
30 l 2.4 0.33 (R) 1.14 at 20 0.48 (R) 0.57
(2.1)
100 l 2.4 0.45 (A) 0.97 at 75 0.71 (A) 0.75
(1.9)
280 l 2.4 0.36 (R) 0.99 at 180 1.0 (R) 0.60
(2.0) 0.5 (M) 1.9 (M)
800 l 2.6 0.38 (R) 0.96 at 600 1.6 (R) 0.704
(2.0) 0.5 (M) 2.9 (M)
0.5 (A) 2.9 (A)
1000 l 2.1 0.35 (R) 0.95 at 750 1.5 (R) 0.75
(1.7) 0.43 (A) 2.2 (A)
1200 l 2.4 0.5 (A) 0.95 at 900 3.1 (A) 0.75
(2.0)
1900 l 2.4 0.39 (R) 0.935 at 1500 2.1 (R) 0.79
(2.0) 0.5 (M) 0.96 at 1400 3.4 (M)
19000 l 3.3 0.346 (R) 0.80 at 15000 3.8 (R) 0.79
(2.9) 0.462 (M) 0.95 at 9000 6.8 (M)
0.47 (A) 7.2 (A)
(Btm, 0.56) (Btm, 8.8)
VT is total tank capacity not nominal volume. R, Rushton; M, Maxflo T; A, A315; Btm, bottom impeller.
ies. Details of some of these vessels and their impellers have

been discussed elsewhere (23–25). Figure 1 shows a typical Geometric similarity of reactor geometry [DT/VT)1/3 or
(DT/VL)1/3] Geometric similarity is expressed as
follows:
fermentor diagram with key locations identified. D /D = (V /V )1/3 (1)
T2 T1 T2 T1
where DTi is the tank diameter and VTi is the total tank vol- Fermentors with a standard geometry are beneficial since
ume of vessel, i. Alternatively, the liquid volume, VLi, several published scale-up correlations assume geometry to
might be used. It assumes reasonably constant impeller be constant (8). However, lab fermentors may have substan-
geometry (i.e., impeller diameter, DI, and number of tially lower HT/DT ratios of 1:1 compared to 3:1 for pilot
impellers, NI) as specified below. Based on target scale fermentors, so laboratory scale data may be deceiving
working/total volumes obtained from geometric similarity (8). Larger pilot scale and production fermentors can be de-
(Table 6), the desired working volume in the fermentor signed for variable working volumes to accommodate the
may be altered during experi- mentation. larger range of batch sizes expected in a multi-use facility.
Experiments to obtain data for several empirical correla- Table 6 shows the scale-up parameters of expected total
tions utilized geometrically similar vessels. Although geo- and working volumes calculated based on geometrical simi-
metric similarity is a pre-requisite for applying established larity. For the 280 l scale-up basis, the calculated numbers
scale-up relationships, it is rarely achievable in practice (26, differed dramatically from installed equipment for the 30 l
27). Subsequently, a range of acceptance of equivalency and 19,000 l scales, but were reasonably close (within
was proposed (26) to include the following: (i) DI/DT = 2– 17%) between the 100 l and 1900 l scales. Although the
0.3– 0.45, where DI/DT is the ratio of the impeller to tank 19,000 l expected values were lower by 30–47%, the
diameters; (ii) HL/DT is less than or equal to 2 (no range 30 l value was higher by 46–55%. For the 30 l scale-up
was given), where HL is the height of the liquid in the basis, all the calculated numbers were substantially lower
vessel; and than the installed values by 19–45% between the 100
(iii) either two or three impellers. As shown in Table 3, l and 1900 l scales and 52–65% for the 19,000 l scale.
DI/DT values ranged from 0.33–0.5 for all vessels in this fa- Reasons for these differences were evident by comparing
cility. Specifically, the range was 0.33–0.39 for vessels values of the geometric similarity parameter, (DT/VL)1/3,
with Rushton radial flow impellers (Lightnin, Avon, NY, listed in Table 3. Val- ues for the 100 l to 1900 l scale
USA), 0.462–0.5 for vessels with Maxflo T axial flow ranged from 0.935–0.99 (average of 0.958 ± 0.019);
impellers (Chemineer, Dayton, OH, USA), and 0.43–0.56 values for the 30 l and 19,000 l
for vessels with A315 axial flow impellers (Lightnin). In
the case of the vessels studied, HT/DT ratios ranged from
2.1–3.3 where HT is the total tank height. The range of the
tangent-to-tangent tank height to tank diameter ratio,
HTT/DT (which was felt to be more useful than HL/DT to
compare since fermentor working volumes may be
variable), was 1.7–2.9, with the value of 2.9 for the 19,000
l fermentor being well above the remainder which ranged
from 1.7–2.1. All vessels had two or three impellers except
for two of the three 19,000 l vessels (those with Rushton
and Maxflo T impellers) which had four impellers.
FIG. 2. Decline of installed, ungassed and gassed power draw per

unit working volume as a function of scale for fermentors under study
at operating conditions listed in Table 1 and for impeller geometries
listed in Table 2. For all scales, 1.7< HTT/DT <2.9 and 1.0<Qmax/VL
FIG. 1. Schematic of fermentor showing nomenclature. <1.67.
(a) For Rushton (R) geometry, 0.33 < DI/DT< 0.39, 4.7 < ITS < 5.6.
(b) For Maxflo T (M) hydrofoil geometry, 0.462< DI/DT< 0.5,
6.4<ITS< 7.4. For A315 (A) axial flow geometry, 0.43 < DI/DT< 0.56,
5.8 < ITS < 7.6.
TABLE 4. Mixing time, power input and gas flow comparisons for laboratory and pilot scale fermentors
Mixing time, Po/V L Pg/VL at Qmax Q /V Superficial Gas flow
Scale T from Design and Nmax max L gas velocity,
(nominal (observed (vvm at number,
volume)
mix
Eq. 10h Po/VL in water) (observed working Vs =
in water) 4Qmax/(nD 2T), NA =
(s) (hp/1000 (hp/1000 l at V min–1) Qmax/NmaxDI 3
l) (hp/1000 l at lpm) L
(cm/s)
rpm)
30 l 3.4 50.0 NI NI 1.5 0.66 0.032 (R)
100 l 13.4 40.0 NI NI 1.6 1.5 0.050 (A)
280 l 20.1 41.7 23.9 at 450 (R) 18.8 at 300 (R) 1.67 2.0 0.078 (R)
18.9 at 450 (M) 16.7 at 300 (M) 0.030 (M)
800 l 29.2 12.5 15.0 at 335 10 at 600 (R) 1.0 2.0 0.063 (R)
12.5 at 300 (R)
16.3 at 335* 8.8 at 600 (M) 0.027 (M)
12.1 at 300 (M)
11.2 at 335 7.8 at 600 (A) 0.027 (A)
8.7 at 300 (A)
1000 l 30.9 13.3 7.7 at 300 (R) 4.3 at 1200 (R) 1.6 3.4 0.140 (R)
5.1 at 300 (A) 4.9 at 600 0.079 (A)
3.5 at 1200 (A)
4.6 at 600
1200 l 32.3 16.7 10.1 at 282 (A) 6.8 at 1200 (A) 1.33 3.0 0.044 (A)
7.1 at 600
1900 l 36.2 10.0 13.9 at 225 (R) 6.8 at 1500 (R) 1.0 2.8 0.088 (R)
10.0 at 225 (M) 8.0 at 1500 (M) 0.043 (M)
19000 l 53.7 4.0 4.4 at 142 (R) 2.0 at 15000 (R) 1.0 8.1 0.300 (R)
4.4 at 150 (M) 4.1 at 15000 (M) 0.130 (M)
4.4 at 150 (A) 3.2 at 15000 (A) 0.120 (A)
Power measurements performed at 25°C in water and at ambient pressure. Unaerated conditions not maintainable at top speed for certain fer-
mentors marked with an asterisk due to drive overheating. No power measurement devices installed at 30 l or 100 l scales. R, Rushton; M, Maxflo
T; A, A315. NI, Power measuring device not installed.
TABLE 5. Mass transfer comparisons for laboratory and pilot scale fermentors
Scale Measured (Po /VL)a (V s)b (Pg /VL)a (V s)b
Vs at Qmax Qmax/VL
(nominal –1 Pg /P
o at using Eq. 9a and 9b using Eq. 9a and 9b
(cm/s) (vvm, min ) Qmax and Nmax and design Po and measured Pg
volume/designation)
30 l 0.66 1.5 NA 31.1 NI
100 l 1.5 1.6 NA 15.5 NI
280 l 2.0 1.67 0.79 (R) 19.4 11.3 (R)
0.88 (M) 10.5 (M)
800 l 2.0 1.0 0.67 (R) 8.6 7.4 (R)
0.54 (M) 6.8 (M)
0.70 (A) 6.3 (A)
1000 l 3.4 1.6 0.55 (R) 12.8 6.0 (R)
0.69 (A) 5.3 (A)
1200 l 3.0 1.33 0.67 (A) 13.8 7.5 (A)
1900 l 2.8 1.0 0.49 (R) 9.3 7.2 (R)
0.80 (M) 8.0 (M)
19000 l 8.1 1.0 0.45 (R) 10.3 6.5 (M)
The 1000 l scale fermentor was typically run at 600 lpm rather than at 1200 0.93
lpm maximum
(M) air flow rate which was used to obtain
10.4calculated
(M) and
measured values. Equation 9a used for 30 l scale and Eq. 9b used for 100 l–19,000
0.74 l scales. Power measurements performed at 25°C
(A) 8.8 in
(A)water and
at ambient pressure. No power measurement devices installed at 30 l or 100 l scales. R, Rushton; M, Maxflo T; A, A315. NI, Power measuring
device not installed. NA, Calculation not applicable.
scales were notably different (greater than 4.5–9.5 SD from peller speed and impeller diameter for vessel i (8). Table 2
the mean). Operation of the 19,000 l scale fermentor at a shows that typical installed impeller tip speeds ranged from
lower working volume brings its geometrical similarity 3.8 to 7.6 m/s with no clear trend observed with scale, but
closer to the average value of the smaller vessels. about a 25–37% increase with the hydrofoil (A315
Constant impeller tip speed (ITS) or shear ITS is and Maxflo T) impellers than the disk turbine (Rushton)
expressed as impellers for the same fermentor scale primarily due to
their
N2/N1 = = T1/VT2)1/3 larger diameters.
(2) Tip speed is used as a rule for scale-up when there is not a
DT1/DT2 (V
which assumes that ITS= nNi DIi where Ni and DIi are the good understanding of the relationship between shear and
im-
morphology for mycelial cultures. A rough rule of thumb is late the values in Table 7, this explains why the 30 l and
that cell damage can occur at tip speeds above 3.2 m/s, but 19,000 l values were substantially different than installed
the exact value is influenced by many factors such as broth
rheology (28). Calculated tip speeds usually are greater than values and why the values for the 30 l scale-up basis were
3 m/s for production fermentors (29) and ranged from 5–7 not in agreement as well.
m/s from a survey of industrial fermentors (Einsele, A., Constant ungassed or (more often) gassed power input
Abstr. 5th Intern. Ferment. Symp., p. 69, 1976). A constant per liquid volume (Po/VL or Pg/VL) Scale-up designs
tip velocity in the range of 5–7 m/s was found to be used may also include the ungassed power input, Po, which is ex-
for scale-up for antibiotic fermentations (Einsele, Abstr. 5th pressed as
Intern. Ferment. Symp., 1976). Although useful in branched Po = NP N 3DI5q (3)
yeast, filamentous bacterial and fungal fermentations for
estimating the potential for hyphae breakage and thus alter- where NP, the power number, is a proportionality factor
ation of broth morphology, tip speed is less useful for single based on the impeller design (among other factors) and q is
cell bacterial or yeast fermentations. the broth density (8) which is typically considered to be
If scale-up is conducted using constant tip speed (with slightly greater than water. The power number generally re-
geometric similarity), then often the value of Pg/VL is low- mains constant with scale-up if the same impeller type is
ered which can adversely affect aeration efficiency. It is used.
possible to recover from this deficiency using more impel- Constant ungassed power, (Po/VL)i, for vessel, i, is ex-
pressed as
lers in the larger vessel and in this manner it may be pos- (P /V ) /(P /V ) = (V /V )c or P /V = f V c (4)
sible to maintain both tip speed and Pg/VL constant (30). Tip o L2 o L1 L1 L2 o L 1 L
speed influences impeller shear which is proportional to2 where c was found to be –0.37 in practice based on a
product of impeller tip speed and impeller diameter, NDI , survey of industrial plants of various scales and processes
for turbulent flow conditions (29). (Einsele, Abstr. 5th Intern. Ferment. Symp., 1976). A
As shown in Table 7, for the 280 l scale-up basis, there similar relationship can be applied for gassed3 power
was reasonable agreement (within 2–24%) between ex- inputs. General values of Po/VL are 1–3 kW/m (1.3–4
pected and actual total and working volumes for the 280 hp/1000 l) for vessels up to 300 m3 (300,000 l) working
l– 1900 l scales, but expected values were substantially volume (Einsele, Abstr. 5th Intern. Ferment. Symp., 1976),
lower (58–69%) at the 19,000 l scale and grossly and a rule of thumb is that Po/VL is 2–4 kW/m3 (2.7–5.4
higher (97– 355%) for the 30 l and 100 l scales. For the 30 hp/1000 l) at the production scale with no volume range
l scale-up basis, all estimated values were substantially given by Kossen and Oosterhuis (29).
lower (40– 85%) than installed values, except for the 100 l If scale-up is conducted using constant Po/VL with geo-
vessel which was substantially higher (131–175%) due to
its higher peak agitation rate. Since geometric similarity metric similarity, then circulation time, mixing time and im-
was used to calcu-
TABLE 7. Scale-up based on constant impeller speed
TABLE 6. Scale-up based on geometric similarity a. Base case 280 l (pilot scale)
a. Base case 280 l (pilot scale) Scale Total Total Working Working
Scale Total Total Working Working (nominal volume, volume, volume, volume,
(nominal volume, volume, volume, volume, volume) actual, expected, actual, expected,
volume) actual, expected, actual, expected, VT (l) VT (l) VL (l) VL (l)
VT (l) VT (l) VL (l) VL (l) 30 l 35 69 20 42
30 l 35 51 20 31 100 l 100 455 75 274
100 l 100 117 75 71 280 l 299 Basis 180 Basis
280 l 299 Basis 180 Basis 800 l 852 754 600 454
800 l 852 905 600 545 1000 l 1000 1078 750 649
1000 l 1000 1100 750 661 1200 l 1271 1298 900 781
1200 l l
19000 1271
18887 1304
13216 900
15000 785
7956 1900 l 2044 2392 1500 1440
1900 l 2044 2086 1500 1256 Base lcase 30 l (laboratory
b.19000 18887 7815
scale) 15000 4705
b. Base case 30 l (laboratory scale) Scale Total Total Working Working
Scale Total Total Working Working (nominal volume, volume, volume, volume,
(nominal volume, volume, volume, volume, volume) actual, expected, actual, expected,
volume)
30 l actual,
35 expected,
Basis actual,
20 expected,
Basis 100 l VT100
(l) VT231
(l) VL (l)
75 VL132
(l)
VT100
(l) VT 81
(l) VL (l) VL 46
(l) 30 l 35 Basis 20 Basis
100 l 75 280 l 299 152 180 87
280 l 299 206 180 118 800 l 852 382 600 218
800 l 852 624 600 357 1000 l 1000 547 750 312
1000 l 1000 758 750 433 1200 l 1271 658 900 376
1200 l 1271 900 900 514 1900 l 2044 1214 1500 693
1900 l 2044 1439 1500 822 19000 l 18887 3965 15000 2266
19000 l 18887 9120 15000 5211 Relationship: N /N2 1= D /D = (V /V )1/ 3
= (V /V )1/3 (assumes geo-
Relationship: D /D metric similarity). T1 T2 T1 T2 L2 L1
=(V /V )1/3=(V /V )1/3.

T2 T1 T2 T1 L2 L1
peller tip speed increase (30), but the size of eddies does not TABLE 8. Scale-up based on constant power per unit volume for
change (29). The Po/VL required to provide a certain OTR Rushton radial flow impellers
generally decreases with scale (2). Scale-up on the basis of a. Base case 280 l (pilot scale)
constant Po/VL can result in a larger than necessary motor
size. In addition, it is difficult to have high power per unit Scale Po/VL Po/VL Pg/VL Pg/VL
volume inputs at the large scale (27) due to practical limita- (nominal measured expected measured at expected at
tions in motor size. volume) at Nmax at Nmax Nmax and Qmax Nmax and Qmax
Assuming geometrically similar vessels in which D /D (hp/1000 l) (hp/1000 l) (hp/1000 l) (hp/1000 l)
=(V T1 T2 100 l NI 33.0 NI 26
/V )1/3 30 l NI 53.9 NI 42.4
T1 T2
(5), then scale-up based on constant ungassed 280 l 23.9 Basis 18.8 Basis
or gassed power per unit volume (Po/VL or Pg/VL, respec- 800 l 15.0 15.3 10.0 12.0
tively) simplifies to 1000 l 7.7 14.1 4.3 11.1
N /N = (V /V )2/9 (5) 1200 l NA 13.2 NA 10.4
2 1 T1 T2 1900 l 13.9 10.9 6.8 8.6
This expression is very similar to N2/N1 =(VT1/VT2)1/3 used in 19000 l 4.4 4.65 2.0 3.7
Table 7 which was derived for scale-up on the basis of con- b. Base case 30 l installed hp (laboratory scale)
stant tip speed and geometric similarity (5). Although the Scale Po/VL Po/VL
gassing rate influence is incorporated into the value of Pg (nominal measured expected
based on measurements or expected decreases in power loss volume) at Nmax at Nmax
upon gassing, it can be challenging to accurately account (hp/1000 (hp/1000 l)
for l)
the gassing (aeration) efficiency. In addition, this relation- 30 l NI Basis
ship is most effective for turbulent flows observed during 100 l NI 30.7
single cell E. coli and yeast cultures and less effective for 280 l 23.9 22.2
high viscosity mycelial cultures (5). 800 l 15.0 14.2
As seen in Table 4, measured values of Pg/VL at maxi- 1000 l 7.7 13.1
mum aeration (Qmax) and agitation (Nmax) rates for stated 1200 l NA 12.2
working volumes ranged from nearly 19 hp/1000 l down to 1900 l 13.9 10.1
19000 l 4.4 4.3
2 hp/1000 l as the scale increased from 280 l to 19,000 l. Relationship: (V /V )0.37=(P /V ) /(P /V ) or (P /V ) /(P /V ) . Mea-
L1 L2 oL2 o L 1 gL2 gL1
Measured values of Po/VL were higher than those for Pg/VL sured Po and Pg values used in calculations except for 30 l base case
due to power decrease upon gassing, and the ratio of Pg/Po where installed value for Po of 1 hp used. NI, Power measuring device
not installed. NA, Calculation not applicable.
varied according to impeller type, fermentor scale and
sparger/impeller geometry (Table 5; 24). Tables 8a (for
Rushton radial flow impellers) and 9 (for Maxflo T and TABLE 9. Scale-up based on constant power per unit volume
A315 hydrofoil axial flow impellers) show that scale-up for hydrofoil axial flow impellers
based on measured Po/VL and measured Pg/VL values agreed
reasonably well (within 26%) with expected values for the Base case 280 l (pilot scale)
800 l, 1900 l and 19,000 l scales. The notable exception Scale Po/VL Po/VL Pg/VL Pg/VL
was the 19,000 l scale with Rushton impellers for which the (nominal measured expected measured at expected at
expected value was higher than the measured value by volume) at Nmax at Nmax Nmax and Qmax Nmax and Qmax
85%. Expected values for the 1000 l and 1200 l scale (hp/1000 l) (hp/1000 l) (hp/1000 l) (hp/1000 l)
fermentors were notably higher by about 83–180% due 30 l NI 42.6 NI 37.7
to the lower than typical measured Pg/VL for this scale 100 l NI 26.1 NI 23.1
vessel (versus installed motor power). Measured values for 280 l 18.9 (M) Basis 16.7 (M) Basis
the 30 l and 100 l scales were not able to be compared since 800 l 16.3 (M) 12.1 8.8 (M) 10.7
no power measurement device was installed. 11.2 (A) 7.8 (A)
Table 8b shows the use of installed values for Po/VL for 1000 l 5.1 (A) 11.2 3.5 (A) 9.8
the 30 l fermentor as a basis for scale-up. Agreement of the 1200 l 10.1 (A) 10.4 6.8 (A) 9.2
expected and actual values to within about 30% was evident 1900 l 10.0 (M) 8.6 8.0 (M) 7.6
for all scales except the 1000 l scale which was higher by 19000 l
Relationship: (V 4.4 (M)=(P /Vo L) 2/(P
0.37
/VL1 )L2 3.7/Vo ) or (P /V g) L/(P
L 1 4.1 (M) 2
/V )g L. 1Mea-
3.3
70%. Using the 1000 l scale as a basis (data not shown), the o g
4.4 (A) 3.2 (A)
sured P and P values used in calculations. M, Maxflo T; A, A315.
expected values were lower for all scales by at least 40%, a
trend which is also due to the lower than typical measured For the maximum operating conditions of agitation (Table6
Pg/VL. 1), impeller Reynold’s numbers ranged from 0.48–8.8 × 10
Similar impeller Reynold’s number (NRe) Scale-up for water (or for single cell E. coli and yeast broths with
can be accomplished based on a constant impeller Reynold’s viscosities similar to water), suggesting that for filamentous
number, NRe: broths with higher viscosities flow may switch from turbu-
NRe = qNDI2/4 (6) lent to laminar. The use of constant Reynold’s number gen-
erally has not worked well for fermentation scale-up since
where 4 is the broth viscosity (8, 31). For the same broth, the effect of aeration on the process was not incorporated
this expression simplifies to (2), and the impeller Reynolds number generally increases
N2/N1 = (DI1/DI2)2 (7) for successful scale-up designs (26). Other dimensionless
TABLE 10. Comparison of typical heat up, hold time and cool down times as well as Fo and Ro values
for medium sterilizations as a function of scale
Heat up Hold Hold Cool down Fo Ro
Scale (nominal volume/jacket type) time Heat up time time time Cool down total total
(area%) (area%)
(min) (min) (area%) (min) (min) (min)
100 l (dimple, jacket loop) 29 24.3 40 65.5 18.6 10.2 75.0 61.6
280 l (straight) 15.5–16.2 15–16 40 74.9 26–29.4 9.6 74.9 57.0
800 l (dimple) 20–21 13–15.6 45 74.3–78.6 17.7–18.6 8.2–10.0 73.7 63.6
1000 l (dimple, jacket loop) 34–38 19–20 45 70–71 27–27.5 9–10 108.8 74.1
1200 l (half pipe coil) 24–30 19–22 45 65–66 33–35 12–14.5 85.4 66.2
1900 l (half pipe coil) 35–42 24–27 45 61.8–63.3 30–32 11–13 86.7 72.8
19000 l (half pipe coil) 83–88 37 45 46.4 95–110 16.4 91.2 86.6
Medium hold temperature of 123.5°C for batch sterilization. Hold temperatures of 124°C for 1000 l and 123°C for 100 l scale. Fo and Ro values
calculated according to Junker et al. (38). Only temperatures above 60°C were included in summation. Heat up and cool down times calculated
from 40°C. A 19,000 l scale vessel used cooling tower water for cool down and not chilled water. Area% is the area under the temperature versus
time curve for the stage divided by the total area. Heat up and cool down area percentages calculated for temperatures above 60°C.
groups also have been examined for scale-up with limited scale-up, but mainly between the pilot scale and production
success, often resulting in technically unrealistic equipment stages. Most commonly when the power is changed only by
and operating parameters. As it is difficult to maintain all increasing agitation speed, then Eq. 9c simplifies to
dimensionless parameters constant upon scale-up, those
most important to the process must be identified accurately. KLa = f2(Pg/VL)0.50 for production vessels (33) (9d)
Constant oxygen uptake rate (OUR), mass transfer
coefficient (KLa) or dissolved oxygen (DO) Scale-up In laboratory scale fermentors where measurements of
based on constant OUR assumes that the OUR is equal to Pg/VL are difficult to accurately obtain, then the key
the OTR: components of the power number (N, DI) may be used:
KLa = f2 (N 3DI 2)0.42 for laboratory vessels (34) (9e)
OTR = KLa(Csat – CL) = OUR = µX/YX/O2 (8) where, although the exponent of 0.42 was established for a
where Csat is the broth DO concentration at saturation, CL is 0.6 l working volume, the literature range is 0.16–0.68.
the measured broth DO concentration, µ is the specific Since they are generally most applicable at or near the con-
growth rate, X is the measured cell density and YX/O2 is the ditions used to determine the exponents, these correlations
calculated cell yield per amount of oxygen consumed (32). might best be used for only qualitative guidance in scale-up
Changes in back-pressure and hydrostatic pressure with calculations (28).
scale-up influence the values of Csat (and subsequently CL). As shown in Table 5, calculated values of (Pg/VL)aVsb
It is also possible to use the log mean of the DO concentra- using measured values of Pg/VL and Eq. 9a and 9b are
tion difference. Since for most aerobic fermentations the reasonably similar (average [ave]: 7.8 ± 1.9; relative stan-
critical DO concentration which adversely affects growth dard deviation [rsd]: 24%) from the 280 l to 19,000 l (180 l–
rate is very low, CL is assumed to be zero. 15,000 l working volume) scales. When designa values of
Several correlations of the general form P /V are used, much higher values of (P /V ) V b are ob-
o L g L s
K a = f (P /V )aV b (9) tained as expected, with the values being reasonably similar
L 2 g L s
(ave: 11.0 ± 2.2; rsd: 20%) from the 800 l to 19,000 l scales
exist to estimate KLa using gassed power per liquid volume, and much higher below the 800 l scale (600 l working vol-
Pg/VL, and gas superficial velocity, Vs, for fermentations ume). Only a fraction of these design values actually can be
(similar to the Van Riet correlation for mass transfer): delivered to the fermentation broth with the amount depen-
dent on vessel/agitator geometry, rheology, agitator speed,
KLa = f2(Pg/VL)0.95Vs 0.67 and superficial velocity (28).
for laboratory scale (8 l) vessels (5) (9a) From a survey of industrial fermentors up to 100,000 l in
volume, KLa values ranged from 400–800 h–1 (Einsele,
KLa = f2(Pg/VL) Vs 0.67 0.67 Abstr. 5th Intern. Ferment. Symp., 1976). The KLa values
for pilot scale (400 l) vessels (5) (9b) obtained in Tables 11 and 12 are generally within this range
assuming a Henry’s law constant of 1 mmol/l-atm. The
KLa = f2(Pg/VL) Vs 0.50 0.50 KLa varies roughly with the inverse of the apparent broth
viscosity (15). Surface aeration can be a substantial
for production (23,000 l–46,000 l) vessels (5) (9c) contributor for smaller-scale fermentors, specifically about
33% for 5 l vessels and about 10% for 50 l vessels (15). Pilot
where f2 is a proportionality constant in all equations whose scale fermentors below 250 l in volume usually have
value varies depending upon the specific process and the significant surface aeration compared to larger scale vessels, so
units of KLa, Pg/VL and Vs used in the calculations. All vol- it is generally best to scale-up from larger vessels of
umes cited in Eq. 9a–c are operating volumes. For about 500 l–1000 l (8). Surface aeration decreases markedly
these correlations, the dependence of KLa on Pg/VL is lower with scale (27); thus for larger vessels, KLa values can be
as the vessel scale increases (i.e., the exponent decreases). more easily inter- preted. Scale-up based on KLa is
Simi- larly, the dependence of KLa on Vs also decreases complicated by the fact
upon
TABLE 11a. Representative historical (1992 through mid-2001) comparison of achievable processing conditions
as a function of scale for E. coli cultivations
Peak oxygen
Fermentation Air flow Impeller Gassed Calculated Value of f2
Cultivation/Scale uptake rate, Pressure speed, power, Pg/VL KLa at using Eq. 9b
(nominal volume) volume (l) rate Q (kg/cm2) N P (hp/1000 ) peak OUR K a/[(P /V )0.67
g l L gL
(impeller type) (l/min) (rpm) (hp) (mmol/l-h-atm) ×(V0.67
)
(mmol/l-h) ]s
E. coli DH5/280 l 180 (R) 260 300 1.8 400 NI NI 685 NA
E. coli DH5/280 l 150 (M) 172 190 1.25 400 NI NI 624 NA
E. coli PF436/280 l 175 (M) 65 80 0.6 383 NI NI 346 NA
E. coli RR1/280 l 160 (M) 77 80 0.3 409 NI NI 472 NA
E. coli OP50/280 l 175 (R) 39 100 0.5 213 NI NI 186 NA
E coli K12/800 l 605 (M) 94 400 0.7 287 6.3 10.4 434 76
E. coli DH5/1000 l 600 (M) 141 600 1.0 300 4.1 6.8 738 143
E. coli PF436/1900 l 800 (M) 38 400 0.7 178 5.6 7.0 167 55
E. coli RR1/1900 l 760 (M) 95 450 0.3 229 11.2 14.7 596 111
E. coli OP50/1900 l 810 (R) 49 500 0.7 125 2.7 3.3 303 143
E. coli Polym./1900 l 1080 (M) 101 600 1.2 225 9.0 8.3 486 110
P. aeruginosa/1900 l 1200 (M) 47 510 0.3 132 7.4 6.2 186 57
Values bolded are at/near (within 20%) of maximum conditions for that particular scale. R, Rushton; M, Maxflo T; A, A315. NI, Power measure-
ment device was not installed. NA, Calculation not applicable.
TABLE 11b. Representative historical (1992 through mid-2001) comparison of achievable processing conditions
as a function of scale for yeast cultivations
Fermentation Peak Air flow Impeller Gassed Calculated Value of f2

Cultivation/Scale oxygen rate, Pressure speed, power, Pg/VL K a at using Eq. 9b
(nominal volume) volume (l) Q (kg/cm2) N P (hp/1000 l) peakL OUR K a/[(P /V )0.67
uptake
rate ×
g L g L
(impeller type) (mmol/l-h) (l/min) (rpm) (mmol/l-h-atm) (Vs ]
0.67
(hp)
)
C. sorbophila/280 l 150 (R) 54 100 0.7 223 NI NI 226 NA
C. chilensis/280 l 180 (R) 141 180 1.5 400 NI NI 297 NA
S. cerevisiae QC2B/280 l 180 (R) 74 80 0.7 354 NI NI 350 NA
S. cerevisiae 1375/280 l 180 (R) 64 90 0.5 315 NI NI 324 NA
C. sorbophila/800 l 475 (R) 105 505 0.7 320 >7.5* >15.8 465 NA
475 (M) 90 475 0.7 286 >7.5* >15.8 386 NA
C. chilensis/800 l 600 (R) 106 335 1.2 245 4.2 7.7 325 77
500 (M) 117 490 1.7 220 3.2 7.0 318 62
S. cerevisiae QC2B/800 l 570 (M) 93 220 0.7 265 2.4 4.2 463 218
S. cerevisiae 1375/800 l 530 (R) 63 250 0.5 266 5.2 9.8 344 84
S. cerevisiae 1375/1000 l 600 (M) 60 400 0.34 308 4.2 7.0 844 211
C. chilensis/1900 l 1500 (M) 49 530 0.83 175 3.1 2.1 252 155
S. cerevisiae 1375/1900 l 1400 (M) 53 700 0.5 173 5.5 3.9 264 89
Values bolded are at/near (within 20%) of maximum conditions. Values marked with an asterisk denote saturated power measurement readings.
R, Rushton; M, Maxflo T; A, A315. NI, Power measurement device was not installed. NA, Calculation not applicable.
that it is process-specific and that it changes over the course

of the fermentation (35), making it difficult to reliably Constant gas flow number (NA), volumetric gas flow
quan- tify. rate per unit volume of liquid (vvm or Q/VL), or superfi-
Alternatively, an optimal value for CL may be determined cial velocity (Vs) Scale-up also can be performed based
from laboratory studies and used for scale-up (2). Equation on the flow (aeration) number, NA= Q/NDI3, where Q is the
8 then can be rearranged to calculate KLa values during the volumetric flow rate. A typical range for the flow number
fermentation at various ages. Scale-up based on constant for production fermentors is 0.1–0.15 (29) which compares
DO can be an attractive method. Although KLa and OUR favorably with the values of 0.12–0.13 for the 19,000 l fer-
can change dramatically over the course of the fermenta- mentors with hydrofoil impellers and is much lower than
tion, the minimum acceptable value of CL is usually known the value of 0.3 obtained for the 19,000 l fermentor with a
from laboratory studies. A CL above 30–70% saturation Rushton impeller (Table 4). Aeration numbers for other
pilot scale vessels (100 l–1900 l) ranged from 0.027–
usually assures adequate DO in less mixed regions of a vis- 0.14 with an aeration number of 0.032 obtained for the 30 l
cous mycelial fermentation; for a less viscous E. coli or laboratory vessel (Table 4).
yeast fermentation, a CL above a lower limit of 10–30% sat- Scale-up based on the volumetric gas flow rate per unit
uration can be adequate. Scale-up based on constant DO volume of liquid (Q/VL or vvm) also should be examined to
must consider the scale-up of methods to control DO such ensure that resulting values of superficial velocity are rea-
as agitation, pressure and air flow rate (19). Cascade control sonable. For a specific process, a balance must be achieved.
of DO by agitation, pressure and air flow rate can be effec- If the volumetric gas flow rate per unit volume of liquid,
tive in maintaining DO above critical values (35). Q/VL, remains constant upon scale-up, then Vs may increase
to the point of flooding in production-scale tanks ( 2 ) .
Simi-
TABLE 12a. Comparison of recent (mid-2001 to 2002) typical achievable processing conditions as a function of scale for E. coli cultivations
Air flow Impeller Gassed Calculated Value of f2
Scale Fermentation Peak rate,
(nominal volume (l) OUR Pressure speed, power Pg/VL KLa at using Eq. 9b
Q (kg/cm )2
N Pg (hp/1000 l) peak OUR K a/[(P /V )0.67
volume) (impeller type) (mmol/l-h) (l/min) (rpm) (hp) (mmol/l-h-atm) L g L
×(V s)0.67]
280 l 180 (R) 155 300 1.2 460 3.25 18.1 572 51
180 (M) 158 300 1.2 460 3.4 18.9 475 41
800 l 600 (R) 142 500 1.4 330 9.2 15.3 391 46
600 (M) 115 500 1.5 330 9.2 15.3 354 42
600 (A) 126 470 1.5 330 6.2 10.3 330 40
600 (HE) 127 500 1.2 338 8.6 14.3 393 48
600 (MC) 124 500 1.2 338 9.0 15.0 364 40
600 (CC) 136 500 1.25 330 7.2 12.0 527 70
1200 l 900 (A) 116 1200 1.3 280 5.5 6.1 296 42
1900 l 1500 (R) 96 1500 1.0 230 11.4 7.6 354 46
1500 (A) 116 1500 0.9 230 13.2 8.8 465 55
Values bolded are at/near (within 20%) of maximum conditions for that scale. R, Rushton; M, Maxflo T; A, A315; HE, HE-3/Maxflo T; MC,
Maxflo T/CD-6; CC, CD-6/CD-6.
TABLE 12b. Comparison of recent (mid-2001 to 2002) typical achievable processing conditions as a function of scale for yeast cultivations
Peak oxygen Air flow Impeller Gassed Calculated Value of f2
Scale Fermentation uptake
(nominal volume (l) rate, Pressure speed, power, Pg/VL KLa at using Eq. 9b
g
rate Q (kg/cm2) N P (hp/1000 l) peak OUR KLa/[(Pg /V
L
)0.67
volume) (Impeller type) (l/min) (rpm) (hp) (mmol/l-h-atm) ×(Vs)0.67]
(mmol/l-h)
280 l 180 (M) 98 235 1.0 338 1.7 9.6 430 69
800 l 600 (R) 93 500 0.7 325 7.7 12.8 453 60
500 (M) 97 268 1.1 276 5.0 9.9 490 114
600 (A) 82 500 0.7 327 5.0 8.3 577 101
600 (MC) 92 500 0.7 271 4.6 7.7 461 85
1200 l 900 (A) 94 1000 1.1 226 2.8 3.1 390 99
1900 l 1500 (R) 79 925 0.97 172 5.0 3.3 385 120
1500 (M) 88 980 0.97 170 6.0 4.0 450 119
19000 l 14600 (M) 78 7800 0.89 96 16.2 1.1 380 136
14600 (A) 82 9300 0.95 106 16.2 1.1 343 109
Values bolded are at/near (within 20%) of maximum conditions for that scale. R, Rushton; M, Maxflo T; A, A315; HE, HE-3/Maxflo T; MC,
14600 (R)
Maxflo T/CD-6; CC, CD-6/CD-6. 74 8400 0.90 115 29.5 2.0 343 78
larly, if it is desired to maintain Vs constant upon scale-up, accomplished by maintaining estimated mixing times con-
then larger values of Q/VL must be implemented at the stant. Several relationships have been derived or empirically
smaller scale (36). Typical values of Q/VL range from 1.67 developed
down to 1.0, with the values generally decreasing at the
larger scale (Table 4). Although the volumetric air flow rate Tmix = f3VL0.3 (10a)
has relatively little effect on the KLa value, higher air flow
rates can cause dramatic foaming (27) and higher gas holdup from a survey of industrial fermentors from 50 l to 100,000
in the fermentor. If the superficial velocity, Vs, and Pg/VL are l in volume (26; Einsele, Abstr. 5th Intern. Ferment. Symp.,
constant, the gas holdup does not change with scale-up (29). 1976) and
Flooding happens when the superficial air velocity, Vs, Tmix = f3(Pg/VL)–0.37 (10b)
approaches 25–50% of the bubble rise velocity since
the fermentor gas holdup volume only can be about 20% or for a tetracycline process up to 52,000 l in volume (Einsele,
less ( 1 5 ) . In the case of water with an average bubble Abstr. 5th Intern. Ferment. Symp., 1976), where k is a pro-
rise velocity of 22 cm/s, flooding occurs at 5–10 cm/s portionality constant in both equations. For geometrically
(15). When similar vessels in the region of turbulent flow:
flooding occurs, the impeller cannot disperse all the sup-
plied gas, the gas rises as big bubbles to the liquid surface Tmix N 2/3DI–1/6 = constant (5) (10c)
and the impeller pumping action diminishes (2). As shown Thus, solving Eq. 10c assuming equal mixing times gives
in Table 5, Vs ranges from 0.66–3.4 cm/s for vessels Eq. 10d (5):
between 30 l and 1900 l in total volume, jumping to 8.1
cm/s for the 19,000 l scale. Based on these guidelines, N2/N1 = (DI2/DI1)1/4 (10d)
possibly the 19,000 l scale Qmax conditions may be prone to and the corresponding power inputs are given by Eq. 10e
flooding. Note that flooding also may occur at low (5):
agitation speeds
with too high of an air flow rate for all scales. (P /V ) /(P /V ) =
3
(N 2
)/(N 3D 2) (10e)
D
oL2 oL1 2 I2 1 I1
Constant mixing time Alternatively, scale-up can
be
For geometrically similar vessels in the region of turbulent
flow where both Po/VL and mixing time are constant, Eq.
10e becomes (12)
N1/N2 = (DI2/DI1)2/3 (10f)
The volumetric power input required to maintain equal mix-
ing time increases as VL2/3. Thus, using the relationship of
Eq. 10f, Pg/VL can become prohibitively high upon scale-up
and is usually overestimated. This technique generally has
not worked well for fermentations (2).
A mixedness index, Im, also has been proposed (7) which
does not assume geometric similarity:
Im = f3(ITS)(DT/HT)(DT)–2/3 (10g)
This expression may be particularly useful for scale-up for
vessels already constructed in which geometric similarity
may not have been maintained. Mixing times need to be
evaluated relative to nutrient mass transfer rates for fed- FIG. 3. Linear regression of literature mixing times (Eq. 10h) re-
batch cultures in the presence of cellular nutrient uptake. ported for laboratory, pilot scale and production processes (4, 14, 20).
For a culture (not specified but likely to be a yeast) with a
cell density of 20 g/l growing at a rate of 0.2 h–1 (doubling
time of 3.5 h), if the DO (CL) is at 20% saturation, oxygen ported (14). Finally, mixing times of 15 s for 120 l, 40 s for
1200 l and 60 s for 12,000 l vessels have been reported
in the broth would be depleted in 2.5 s (32). In the case (20). These mixing times were reported for both Newtonian
where glucose was the limiting nutrient, glucose could be and non-Newtonian fluids/broths.
depleted as fast as 4.5 s (32). Longer mixing times can These mixing time estimates may be graphed (Fig. 3).
cause locally Linear regression of these available data points from the lit-
erature yields the equation
high glucose rates which in turn can cause locally low DO
levels owing to higher cell metabolism. As a consequence, Tmi = 17.5 (VL) – 19.4
x log10
local areas of mixed acid E. coli fermentations can result, (10 l to 60,000 l, r2 = 0.876) (10h)
producing overall lower biomass yields upon scale-up (9).
Thus, scale-up based on mixing time can be relevant for Tmix = 223.5 log10(VL) – 1004.6
fed-batch E. coli and yeast cultures, especially at high cell (60,000 l to 120,000 l, r2 = 0.902) (10i)
densities.
The characteristic time analysis above can be extended where Tmix is the mixing time in seconds and VL is the liquid
further and divided into characteristic times for transport working volume in liters. Clearly the mixing time increase
phenomena (transfer or supply) and characteristic times for with volume is steeper once the working volume is above
conversion (consumption or reaction) rates (11). Transport 60,000 l. Equation 10 h was used to estimate mixing times
phenomena include oxygen transfer (including oxygen for each existing fermentor scale (Table 4). The relative
transfer from the gas bubble to the liquid), liquid magnitudes of these times can be compared with those ob-
circulation, gas residence, and heat transfer. Conversion tained from the times estimated using Eq. 10a and 10b as-
rates include the zero and first order rates for oxygen and suming the proportionality constant, f3, remains constant
substrate consumption, cell growth and heat production. with scale.
It can be important to compare nutrient consumption and Other influences After the initial strategies outlined
transfer rates with each other as well as with circulation above are implemented, the resulting vessel design needs to
times to establish whether gradients are likely to exist (11). consider other influences to assure optimal performance.
Specifically, for gluconic acid fermentation in a stirred tank Heat transfer rates Heat evolution is the combination
by a fungal culture, oxygen limitation can occur if con- of mechanical heat from the agitator and metabolic heat
sumption and transfer rates are similar in magnitude. If liq- from the culture. The ratio of metabolic heat evolution to
uid circulation times also are this same order of magnitude, oxygen consumption is approximately 115 kcal heat
then oxygen gradients are favored. In contrast, temperature evolved per mole of oxygen consumed for selected E. coli,
gradients are less likely to occur since, although heat pro- yeast and fungal cultures (32, 37). The total amount of
duction and transfer rates are similar, they often are oxygen consumed by the culture can be estimated by
substantially greater than liquid circulation times. This integration of the OUR value versus time curve then
analysis is applicable to high cell density fed-batch E. coli multiplication by the tank working volume. The desired
and yeast cultures. heat transfer coefficient and jacket area then can be
Observed mixing times of several seconds exist for labo- evaluated using expected cooling fluid flow rates and
ratory fermentors, 20–30 s for 1000–2000 l pilot scale fer- inlet/outlet temperatures. Require- ments for the jacket type
mentors and 70–140 s for 60,000 l–120,000 l production (e.g., dimple, half-coil) and jacket area then are established.
fermentors (4). Specific mixing times for these scales of 5 For large fermentors over 5000 l, it can be difficult to
s for 10 l, 20 s for 1000 l, 29 s for 1800 l, 67 s for 60,000 l, sustain OURs above 300 mmol/l-h since heat transfer
100 s for 100,000 l and 140 s for 120,000 l vessels have problems arise.
been re- Medium sterilization effects As the fermentor scale in-
creases, the heat up and cool down times become a larger
proportion of the overall sterilization time cycle as shown
in Table 10. The heat up time portion increases from about from Pg/VL and KLa correlations.
15% to about 37%, and the cool down portion increases An application of this method is the use of KLa for scal-
from 10% to 16% as the scale rises from 280 l to 19,000 l. ing-up a Bacillus thuringiensis fermentation. This scale-up
The 100 l and 1000 l scales are exceptions to this trend as was accomplished using the equation, KLa = constant N aV b
they required slightly longer than the expected ranges be- (obtained by combining Eq. 9, the expected ratio of Pg/VLs ,
cause they are equipped with a recirculating jacket loop and Eq. 3) with scale-up on the basis of KLaPT where PT is
with indirect heating and cooling via heat exchangers. The the total pressure in the vessel including head pressure (40).
values of Fo increased from about 75 to about 91 min and Another application is to use an iterative strategy based on
the values of Ro increased from about 57 to about 87 min incremental air flow rates to calculate resulting scale-up pa-
over this same range of scales (Table 10; 38). These in- rameters and then to examine each set of parameters for a
creases in Fo and Ro directly translate to a rise in the level of particular flow rate to further optimize compressor and agi-
sterilization overkill and medium heat stress respectively as tator power requirements (36).
scale increases. The adverse impact of medium heat stress Method of Ju and Chase/Diaz and Acedevo This
is process dependent and can be minimized with the use of method (2, 41) focuses on scaling-up based on equivalent
OTR not KLa. First, the scale-up volume is selected, then
continuous (high temperature, short time) media steriliza-
tion. geometric similarity is used to obtain DT. Constant impeller
Biological factors Biological factors, such as the num- tip speed isc assumed to obtain N. Using the relationship,
ber of generations through which the organism will Pg/VL= f1VL (Eq. 4), Pg/VL then is estimated and, using em-
progress over the course of the seed and production pirical relationships between Pg/VL and KLa (Eq. 9a–c),
fermentations, need to be considered in the selection of the the KLa values for the scale-up fermentor are determined.
final production scale. Assuming exponential growth, the Final- ly, the OTR rate for the large fermentor is matched
number of generations, Ng, can be expressed as (12) with that of the small scale fermentor and the back-pressure
for the large fermentor is calculated assuming CL is 0
Ng = 1.44 (lnVL + ln XN – ln XNo) (11) mmol/l.
where XN is the final cell number (or mass) and XNo is the The key element of this method is that there are three de-
initial inoculum cell number (or mass). The stability of the grees of freedom for scale-up. These are commonly N, Q
recombinant DNA insert and the requirement for an agent and DI/DT (i.e., reactor geometry). Thus, only three items
to exert selective pressure need to be evaluated for the may be specified to be constant for scale-up. If another de-
number of expected generations in the process plus a gree of freedom is added such as the gas phase partial pres-
comfortable safety margin. sure, then this value also can be specified to maintain the
Generalized approaches to scale-up Several authors have maximum OTR constant. To determine the effect on the
recommended approaches or combinations of approaches cultivation, it is generally useful to run small scale
to scale-up based on the equations and principals described fermentations using gas blending to vary this inlet oxygen
above. No single method appears to be suitable for all partial pressure.
fermentations with a high probability of success (5), and the Method of Wang et al. In this method (42), two key
applicability of one method over another is process- values are maintained constant on scale-up: KLa (calculated
specific. Success often depends on the reliability of empiri- by Eq. 9a–c) and ITS. The ratio of DI/DT then is adjusted to
cal scale-up correlations and the integrity of small-scale within reasonable limits to complete the design. Although
data. A few examples are highlighted below. geometric similarity is not maintained, the resulting varia-
Method of Hubbard This method (39) was developed tion of the DI/DT ratio with scale often can still provide ade-
as a combination of approaches from other authors. During quate gas dispersion. This technique adds flexibility for in-
the laboratory scale fermentation, air flow rate, impeller stallations of existing fermentors.
speed, yield, and fermentor geometric dimensions are mea- Proposals for improved scale-up calculations This
sured. Basic broth properties such as density, viscosity, sur- simplified method (27) evaluates key quantities of scale-up
face tension, and oxygen diffusion coefficient are deter- interest based on two variables: N and DI. The impeller
mined. (Of these broth properties, it is probably most im- Reynold’s number is represented by NDI2, the Froude num-
portant to understand the broth rheological behavior for ber by N 2DI, the Weber number by N 2DI3, the gassed
mycelial cultures.) Several quantities, such as Q/VL, NRe, power per unit liquid volume by N 3DI2, the impeller tip
ITS, and NA, are calculated from the laboratory scale fer- speed by NDI, and Q/HT by DI/N (where HT is specifically
mentor conditions based on measured quantities. the height of the fermentor and not the height of the liquid
For the plant scale fermentor, a volume is selected based volume). It combines relevant dimensionless group analysis
on product yield and plant capacity. Geometric similarity with other empirically applicable parameters.
then is used to calculate vessel dimensions. Scale-up is Scale-up analysis using one or more of these above meth-
based on matching KLa values and then proceeding in either ods has suggested methods for operating laboratory fermen-
one of two ways. The first way is to calculate the flow rate, tors for the best scale-down performance. For example,
Q, by either holding Q/VL or superficial velocity, Vs, con- nitrogen-diluted air is recommended to be used at the 10 l
stant, then to calculate the impeller speed, N, based on scale to appropriately mimic scale-up to the 10,000 l scale
Pg/VL versus KLa correlations (Eq. 9a–c). The second way is (2). Scale-down studies designed to simulate large-scale
to set N using constant impeller tip speed, then to operating conditions have been highly successful in some
calculate Q cases in predicting behavior and identifying causes for sub-
optimal large-scale performance (43–46).
Current scale-up practices for E. coli and yeast cultures
are largely based on maintaining equivalent CL, KLa or III. SCALE-UP RESULTS FOR
OTR performance as a first pass. In these fermentations, PRIOR FERMENTATIONS
oxygen supply is limiting upon scale-up due to high cellular (1992 THROUGH MID-2001)
oxygen demand and not due to high broth viscosity as in A survey of processing was conducted in this fermenta-
the case of fungal cultures. Subsequently an evaluation of tion pilot plant from 1992 through mid-2001 to obtain in-
the impact of mixing time as it affects DO and nutrient formation about achievable process conditions for E. coli
distribution can be conducted, which is especially (Table 11a) and yeast (Table 11b) fermentations. Bolded
recommended for high cell density fed-batch cultures. values represent operation within 20% of maximum condi-
tions for agitation and aeration for the pilot plant fermentors
II. FERMENTATION PROCESS CONDITIONS shown in Table 1. In the case of fermentor back-pressure,
the desired maximum value was fixed at 1.5 kg/cm2,
To evaluate the operating performance of existing pilot although during this period of operation (early 1990s) some
scale vessels, fermentor conditions (working volume, air fermentations were conducted at pressures as high as 1.8
flow rate, pressure, impeller speed, gassed power draw, kg/cm2, a back-pressure subsequently found to adversely
mass transfer coefficient, OUR) were compiled based on affect the agitator double mechanical seal longevity. Also
historical records for several E. coli (Table 11a) and yeast during this period, power draw readings were not available
cultures (Table 11b). OUR was calculated based on mea- at 280 l scale and the watt transducers installed at 800 l
surements of off-gas composition by a mass spectrometer scale, which were not corrected for gearbox losses (24),
(47). Data was taken at the time of peak OUR which did became saturated at higher power draws.
not necessarily correspond to the time of peak mass transfer Table 11a illustrates that historical E. coli cultivations
coefficient. Cultivations were conducted at the 280 l to predominantly challenged the maximum impeller speed pri-
1900 l scales over the past decade. In addition, specific marily at the 280 l and 1000 l scales. Only a few 280 l cul-
recent studies were performed at the various scales (280 tivations simultaneously approached peak conditions of air
l–19,000 l) for one example E. coli (E. coli DH5, Table flow rate, agitation and pressure. Peak OURs ranged from
12a) and one example yeast process (Candida chilensis, 38 to 260 mmol/l-h with OURs above 90 mmol/l-h reached
Table 12b) using fermentation media designed to promote for each of the four scales tested. Pg/VL and KLa values cal-
high-peak OTR. Tabulated data are representative of typical culated at peak OUR ranged from 3.3–14.7 hp/1000 l
fermentations of that specific process at that selected scale. and 167–738 mmol/l-h-atm, respectively. Although there
Control parameters for each fermentation varied accord- was extensive experience at the 280 l and 1000 l scales
ing to development objectives. After its initial decline from with high OUR fermentations, the degree of challenge to
pre-inoculation levels, DO was generally controlled be- the operating capacity of the vessels at the other scales was
tween 30% and 80% of saturation, calibrated to air at am- limited. KLa values achieved at the smaller 280 l scale were
bient pressure, using various strategies. In some cases, the
fermentation was initiated with the agitation, back-pressure generally achievable upon scale-up to the 1000 l or 1900 l
and/or air flow rates in cascade control with DO. In other scales for those processes for which this was a development
cases, when the need to control DO at its set point resulted goal (E. coli DH5, E. coli RR1). Values of the
in the agitation and air flow rates reaching maximum set- proportionality constant, f2, from Eq. 9b ranged from 55 to
tings, these two parameters were placed in automatic mode 143, indicating an expected variability in performance of
at their maximum set points and then back-pressure was any empirical scale- up correlation due to the specific
placed in cascade control. Other strategies were utilized de- process, scale-up and operating parameters.
pending on the specific process requirements. For example, Table 11b summarizes comparable historical data for yeast
if foaming occurred in the fermentation or a higher dCO 2 cultivations. As noted in the case of the E. coli processing,
concentration was desired, the back-pressure was typically few challenges to peak operating conditions were made ex-
raised to 1–1.5 kg/cm2 earlier in the process. cept at the 280 l and 1000 l scales. Peak OURs were
Specific process descriptions have been published for notably lower than in the case of the E. coli fermentations,
some of the processes described (E. coli RR1 [48, 49], E. ranging from 49 to 141 mmol/l-h with OURs above 50
coli OP-50 [50], E. coli PF436 [51], Saccharomyces cere- mmol/l-h reached for each of the four scales tested. Pg/VL
visiae QC2B [52], S. cerevisiae 1375 [53], Pseudomonas and KLa values calculated at peak OUR ranged from 2.1
aeruginosa MB5001 [54], and Candida sorbophila [55– hp/1000 l to over 15 hp/1000 l and 226–844 mmol/l-h-atm,
57]). Other process descriptions are not published (E. coli respectively. These ranges were similar to those noted for
DNA polymerase, E. coli K12, E. coli DH5, C. chilensis), E. coli processes. Again the extensive experience at the
but are consistent with current state-of-the-art procedures. 280 l and 1000 l scales was not matched at the other scales.
Exact processes run were based on these descriptions, but In the case of these yeast processes (C. sorbophila, C.
they de- viated significantly in many cases to achieve chilensis, and S. cerevisiae 1375), the KLa values achieved
process development goals. at the 280 l scale were generally able to be obtained at the
larger 800 l, 1000 l and 1900 l scales. Values of the
proportionality constant, f2, from Eq. 9b generally ranged
from 62 to 218, again indicating expected variability in
performance due to the process, scale-up and operating
parameters. For a specific process at a specific scale, the
range was substantially smaller.
IV. SCALE-UP RESULTS FOR CURRENT STUDIES conditions from typical process development data sets.
Based on this historical analysis of process performance, Often the range of process conditions that is needed
recent studies (mid-2001 to present) were conducted to im- conflicts with how to optimally execute the processes,
prove the challenge to fermentor peak operating conditions making data difficult to obtain and back-calculate. It can
for E. coli and yeast fermentations. In the case of E. coli be difficult to vary conditions within a single cultivation as
fermentations, initial carbon and nitrogen concentrations was done previously (58) without adversely affecting
were raised to increase the peak OUR beyond that which subsequent data. Conse- quently, not all processes/scales
might normally be observed with this process. It was de- were able to be analyzed if the operating conditions at peak
sired to conduct specific studies at the 800 l, 1200 l, 1900 l OUR values for individual cultivations did not vary
and 19,000 l scales to obtain more complete information sufficiently.
Table 13a summarizes the relationship between peak OUR
and KLa:
about the capabilities of the installed pilot scale equipment. OUR = A K a + (12a)
B
Upgraded variable frequency drives (VFDs) were installed 1L 1
on all, but the 19,000 l scale vessels (which retained their for selected historical and current processes at various
watt transducers) in which the power draw was monitored scales for which the broth DO levels were similar. The
directly from the VFD electrical current (Altivair 66; slope, A1, varies from 0.075 to 0.46, indicating a large
Schneider Electric, Palatine, IL, USA) This resulted in the dependence on process and operating conditions on mass
new capability to measure power draw at the 280 l scale transfer. For a specific process, however, the values when
and an expanded measurement range for the 800 l scale. attainable were reasonably constant upon scale-up.
Table 12a summarizes the achievable processing condi- Prior scale-up studies in pilot scale vessels have been
tions for the E. coli DH5 process for a process scaled-up based on secondary metabolite fermentations in which gen-
based on maintaining similar minimum DO levels. For eral agreement was found with established KLa and Pg/VL
these aerobic processes, DO was required to be above its trends (58). Since the more recent focus of pilot plant stud-
critical value to maximize growth and productivity. Peak ies has been on scale-up of E. coli and yeast cultures, peak
OURs decreased slightly from about 155–158 mmol/l-h to data from recent scale-up studies have been analyzed and
about 96– 116 mmol/l-h upon scale-up from the 280 l to results presented in Table 13b. The exponent of Pg/VL in Eq.
1900 l scales. Pg/VL decreased with scale as expected 9b, A2, was calculated for selected historical and current
ranging from about 18–19 hp/1000 l at the 280 l scale to processes at various scales according to
6.1–8.8 hp/1000 l at the
1200 l and 1900 l scales. Calculated KLa values declined log K a = A log(P /V ) + B (12b)
only slightly with scale, decreasing from 475–572 mmol/l- L 2 g L 2
h-atm at the 280 l scale to 354–465 mmol/l-h-atm at The values obtained appear to be consistent for both E. coli
the 1900 l scale. Values of f2 in Eq. 9b were substantially and yeast processes and are about 0.96–0.98, compared
more uniform than found with the historical data, ranging with 0.58–0.8 found previously for a filamentous bacterial
from 40 to 70, matching the lower range found with the and a fungal cultivation (23, 58). The higher values reflect
historical the greater impact of a KLa change with OUR for these E.
E. coli process data. Values were relatively consistent with coli and yeast cultures, most likely due to their lower broth
scale and operating conditions, suggesting a reliable empiri- vis- cosity and largely single cell morphology. Interestingly,
cal relationship may be obtainable. the higher value of the exponent appears to be more
Achievable processing conditions for the yeast C. consistent with the laboratory scale correlation (Eq. 9a) than
chilensis process are summarized in Table 12b, again the pilot scale correlation (Eq. 9b).
based on scaling-up using similar minimum DO levels. For mycelial broths, rheological characteristics influence
Upon scale- up from the 280 l to 19,000 l scale, peak OURs scale-up performance. Heat and oxygen transfer rates are 5–
decreased slightly from about 98 mmol/l-h to about 74– 50% of bacterial fermentations and bulk mixing is poorer
82 mmol/l-h. As expected, Pg/VL decreased with scale leading to lower broth homogeneity (28). As a consequence,
ranging from about ITS often has been a useful rule of thumb for scale-up (27).
9.6 hp/1000 l at the 280 l scale to 1.1–2.0 hp/1000 l at Rheology is less important for single cell bacterial and yeast
the fermentations compared with the relative magnitudes of nu-
19,000 l scales. Both peak OUR and Pg/VL values were no- trient uptake and supply rates. Thus, scale-up based on DO,
tably lower for the yeast process than for the E. coli OUR or KLa often has been more useful for these cultiva-
process. Calculated KLa values declined only slightly with tions. For facilities with existing installed equipment, relia-
scale, decreasing from about 430 mmol/l-h-atm at the 280 ble scale-up is most readily achieved based on selecting op-
l scale to 343–380 mmol/l-h-atm at the 19,000 l scale. erating conditions such that the minimum DO remains simi-
Values of f2 in Eq. 9b ranged from 60 to 136 and were less lar between the two scales. Implications for other key pa-
uniform with scale and operating conditions than found rameters such as OUR and KLa then can be easily evaluated.
with current E. coli process data. This difference may have Owing to the availability of recombinant E. coli and yeast
been due to the tendency of this culture to clump and even cultures with high specific productivities, typical production
branch during growth. The range of f2 values also was lower volumes for recombinant proteins often do not range above
than that found with the yeast historical data. 5000 l and/or may not require high cell density processes.
As demand for capacity increases, additional efforts may be
V. TRENDS AND IMPLICATIONS FOR SCALE-UP made to improve volumetric productivity and thus push the
Established relationships for scale-up can be difficult to
use due to lack of sufficient data over a range of processing
TABLE 13a. Comparison of relationship between OUR and KLa for selected historical and current cultivations
Scale
Cultivation OUR range KLa range
(nominal Regression used for used for Number
A1 coefficient, of
volume) regression regression
(atm) r2 (mmol/l-h) cultivations
(mmol/l-h-atm)
Historical (1992 to mid-2001)
C. sorbophila 280 l 0.22 0.97 30–53 120–230 11
800 l 0.22 0.94 50–105 230–470 11
C. chilensis 280 l 0.46 0.99 75–140 160–300 4
800 l 0.32 0.99 65–118 155–320 3
S. cerevisiae QC2B 800 l 0.19 0.98 40–92 180–460 22
S. cerevisiae 1375 280 l 0.19 0.91 25–66 140–340 33
800 l 0.17 0.84 53–67 285–370 11
1900 l 0.17 0.99 51–60 258–308 4
E. coli RR1 280 l 0.16 0.90 31–86 190–500 20
1900 l 0.18 0.88 25–97 151–596 5
E. coli DH5 1000 l 0.17 0.99 80–150 412–780 16
E. coli OP50-1 1900 l 0.075 0.95 39–49 177–303 3
Recent (mid-2001 to 2002)
C. chilensis 280 l 0.31 0.97 71–158 230–535 7
1200 l 0.18 0.92 73–95 296–445 5
E. coli DH5 280 l 0.39 0.96 50–98 250–373 4
Equation 12a: OUR = A1KLa + B1. Based on peak OUR measured during fermentation (not necessarily peak OUR capacity of equipment).
TABLE 13b. Relationship between KLa (mmol/l-h-atm) and Pg/VL (hp/1000 l) for historical and current results
Scale Superficial Regression Pg/VL range KLa range Number

Cultivation (nominal volume) velocity, used for used for of
B2 coefficient, regression regression
(impeller type) (hp/1000 l) (mmol/l-h-atm) cultivations
r2
Single scale results A V 2
s
(cm/s)
S. avermitilis a 800 l (R) 0.667 0.58 1.8 NA 0.67–8.0 35–90 1
(converted to hp/1000 l 800 l (M) 0.667 0.59 2.2 NA 0.33–2.0 50–180 1
from kw/1000 l)
A. terreus b 800 l (R) 0.5–1.0 0.6 1.9 NA 0.38–5.6 30–200 1
(converted to hp/1000 l 0.8 1.2 NA 0.31–4.4 4–45 1
from kw/1000 l)
S. cerevisiae 1375 1000 l (R) 1.13–1.7 0.96 0.97 0.92 3.7–7 485–897 6
E. coli RR1 1900 l (M) 0.75–0.84 0.96 1.6 0.81 4.1–14.7 150–600 5
E. coli DH5 280 l (M) 2.0 0.98 1.5 0.89 8.3–18.9 230–535 7
Multi-scale equation
Equation 9b as applied to 100 l–19000 l 1.5–8.1 0.67 NA NA 1–20 NA NA
facility vessels
Equation 12b: logKLa = A2log(Pg/VL)+ B2. Range of superficial velocities assumed constant for calculated values. Common (base 10) logarithm
utilized. R, Rushton; M, Maxflo T; A, A315. NA, Calculation not applicable.
a
See Ref. 58.
b
See Ref. 23.
limits of existing and future fermentor design capacity.

When more scale-up experience is gained over a broad c : exponent (Eq. 4)
range of volumes, the utility of various scale-up CC : Chemineer CD-6 top/CD-6 bottom impeller con-
methodologies can better be evaluated. Throughout these figuration
evaluations, an effective scale-down model is key to CL : measured concentration of dissolved oxygen in
effective trouble shooting at the large scale, and its broth, mmol/l or %saturation
development should be a priority. Csat : concentration of dissolved oxygen in broth at sat-
uration, mmol/l or % saturation
dCO2 : dissolved carbon dioxide
NOMENCLATURE DI : diameter of impeller, m
a : exponent (Eq. 9) DO : dissolved oxygen
b : exponent (Eq. 9) DT : diameter of vessel, OD, m
DTi : diameter of vessel, i, OD, m
A1, B1: slope and intercept constants (Eq. 12a)
f1 : proportionality constant (Eq. 4)
A2, B2: slope and intercept constants (Eq. 12b)
A : Lightnin A315 impeller (pumping downwards) f2 : proportionality constant (Eqs. 9, 9a–e)
configuration f3 : proportionality constant (Eqs. 10a–g)
Fo : equivalent kill for observed sterilization tempera-
ture relative to that at 121°C, min µ : growth rate, h–1
gc : gravitational acceleration conversion factor, 1 kg q : density, g/cm3
m/Ns2 4 : viscosity, g/cm-s (poise)
HE : Chemineer HE-3 top/CD-6 bottom impeller con-
figuration
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JOURNAL OF BIOSCIENCE AND BIOENGINEERING

Vol. 97, No. 6, 347–364. 2004

Scale-Up Methodologies for Escherichia coli and

Received 22 August 2003/Accepted 15 March 2004

TABLE 1. Characteristics of laboratory and pilot scale vessels

TABLE 3. Geometric comparisons for laboratory and pilot scale fermentors

ies. Details of some of these vessels and their impellers have

FIG. 2. Decline of installed, ungassed and gassed power draw per

=(V /V )1/3=(V /V )1/3.

Fermentation Peak Air flow Impeller Gassed Calculated Value of f2

that it is process-specific and that it changes over the course

Scale Superficial Regression Pg/VL range KLa range Number

limits of existing and future fermentor design capacity.

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