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# 8.

4 Continuous Reactors

257
Run 3: 2021 steps in 0.15 seconds

120

-0.008 -0.007

100

1 I
80 I I - 60

1
1

## -0.006 -0.005 -0.004 O -0.003 -0.002 -0.001

-0

\ . ll '-, 1
1

. !i

40

\\

v i
120

1 i

20

|\ i I i '\ * \ \! \l ! \\ \
1

20

40

60

80

100

140

160

180

200

TIME Figure 3. Influence of initial KLa value from 100 to 160 h"^ on the S and O profiles.

8.4
8.4.1
System

Continuous Reactors

The steady state operation of a continuous fermentation having constant volume, constant flow rate and sterile feed is considered here (Fig. 1).

Biological Reaction Engineering, Second Edition. I. J. Dunn, E. Heinzle, J. Ingham, J. E. Pfenosil Copyright 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim ISBN: 3-527-30759-1

258

D,S

## Figure 1. Chemostat fermenter with model variables.

Model
The dynamic balance equations may be modified to apply only to the steady state by setting the time derivatives equal to zero. The corresponding equations are then: For biomass,
0 = - D X + rx

For substrate,
0 = D (S0 - S) + rs

Growth kinetics,
rx = ^ X

## Substituting into the biomass balance gives

\i = D where S is determined by the kinetics \i = f(S)

S =

X = Y(S 0 -S)

## 8.4 Continuous Reactors

259

The productivity of the reactor for biomass is X D. The above equations represent the steady state model for a chemostat with Monod kinetics. Using them it is possible to calculate the values of S and X, which result from a particular value of D, and to investigate the influence of the kinetic parameters.

Program
In Madonna programs, time can be used as a variable which will increase from the starting time. Here it is renamed D. Thus equations will be solved for increasing values of the dilution rate. Fortunately X and S can be explicitly solved for in this problem. If not, the ROOT FINDER facility of Madonna can be used. The program is found on the CD-ROM.

Nomenclature
The nomenclature is the same as the example CHEMO, Sec. 8.1.2.

Exercises

260

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Results
The steady state curves of X, S, and XD versus D are given Fig. 2. The results in Fig. 3 were obtained by varying K\$ in each run. An interesting effect can be observed on the position of the washout point.
Run 1:113 steps in 0 seconds

10 9876v5
f*r

-4
3.5

i
S" //
0
0.1 0.2 0.3 0.4

m ^r

*S**~\ ! I \ Y.-I 1i

-3
-2.5

~s;i
mm

\!

"

1 /{
0.5 0.6 0.7 0.8 0.9

2
1.5

43-

1
0.5

210-

0 1

## 8.4 Continuous Reactors

261
Run 5:113 steps in 0 seconds

8.4.2

## Continuous Culture with Inhibitory Substrate (CONINHIB)

System
Inhibitory substrates at high concentrations reduce the specific growth rate below that predicted by the Monod equation. The inhibition function may be expressed empirically as

where KI is the inhibition constant (kg/m3). If substrate concentrations are low, the term S2/Kj is lower in magnitude than KS and S, and the inhibition function reduces to the Monod equation. In batch cultures the term S2/Kj may be significant during the early stages of growth, even for higher values of K[. The inhibition function passes through a maximum at Smax = (Kg Ki)-5. A continuous inhibition culture will often lead

262

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

to two possible steady states, as defined by the steady state condition JLI = D and as in shown Fig. 1.

D=

## Figure 1. Possible steady states for a chemostat with inhibition kinetics.

One of these steady states (A) can be shown to be stable and the other (B) to be unstable. Thus, only state A and the washout state (S = SQ) are possible.

Model
A model of a chemostat with its variables is represented schematically in Fig. 2.

F,S 0

+-

F,S,X

263

## Cell material balance,

VdX jj- = ^ i V X - F X or,

## where D is the dilution rate = F/V. Substrate material balance,

VdS = F (S 0 -S) or, dS dT MX = D (S0 - S) - ^VX -~^f

## where Y is the yield factor.

Program
When the system equations are solved dynamically, one of two distinct steady state solutions are obtained, the stable condition A and the washout condition. The initial substrate and organism concentrations in the reactor will determine the result. This is best represented as a phase-plane plot X versus S. All results indicate washout of the culture when the initial cell concentration is too low; higher initial substrate concentrations increases the likelihood of washout.

Nomenclature Symbols
D KI KS Dilution rate Inhibition constant Saturation constant 1/h kg/m3 kg/m3

264
S

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Smax X Y

Substrate concentration Maximum in S for inhibition function Biomass concentration Yield coefficient Specific growth rate

Indices
0 I m

Exercises

## 8.4 Continuous Reactors

265

Results
Run 1:2000 steps in 0 seconds 1-1-2

10

15

20 TIME

25

30

35

40

## Run 10: 2000 steps in 0.0167 seconds

5 i

4.5 -

4
3.5

3
e/> 2.5

2
1.5

1
0.5

0
0.5 2.5

Figure 4. Phase-plane plot of X versus with varying ST from 0 to 5 kg/m3 using Batch Runs with overlay.

266

## Run 9: 2000 steps in 0 seconds

54.5-

X,

4 3.5-

^^^>

*" 3
</) 2 . 5 -

^ *t %%
\
"|

t**

* * C^1 v
"% % V

\ ^ x i ! l

2
1.5-

1
0.5-

/ /; ,--^, f/ // / ^' . *x
/
J

3:2 (0.5) 3:3 (0.5857) _ - - 3:4(0.6714) 3:5(0.7571) 3:6(0.8429) 3:7(0.9286) 3:8(1.014) 3:9(1.1)

j* S ^

f f / t / ,f !

rf'^T

"^ -s^^-

"^"^>*i'^' ** -^

## :' / / * / Jf*'^ * \ f 1 C & (

^^^*^^\$^S? ** ^^*^^ **

0.5

Figure 5. Phase plane plot of influence of the initial biomass Xi from 0.5 to 1.1 for Steady states upper left and lower right.
Run 20:2000 steps in 0.0167 seconds

= 0.0.

## Figure 6. Influence on the inhibition function made by varying KI between 1 and 3.

Reference
Edwards, V.H, Ko, R.C. and Balogh, S.A. (1972). Dynamics and Control of Continuous Microbial Propagators Subject to Substrate Inhibition Biotechnol. and Bioeng. 14, 939-974.

267

8.4.3
System

## Nitrification in Activated Sludge Process (ACTNITR)

Nitrification is the process of ammonia oxidation by specialized organisms, called nitrifiers. Their growth rate is much slower than that of the heterotrophic organisms which oxidize organic carbon, and they can be washed out of the reactors by the sludge wastage stream (Fs). In an activated sludge system (Fig. 1) when the organic load (F So/V) is high, then the high biomass growth rates require high waste rates. Nitrification will not be possible under these conditions because the concentration of nitrifiers (Ni) will become very low.

O,F O

2, F4

Reieto*

2, F3
Figure 1. Configuration and streams for the activated sludge system.

Model
The dynamic balance equations can be written for all components around the reactor and around the settler. The settler is simplified as a well-mixed system with the effluent streams reflecting the cell separation. Organic substrate balance for the reactor: = F 0 So + F 2 S 2 -

268

## Ammonia substrate balance in the reactor:

= FQ AQ + F2 A2 - FI A

R2Vt

## Reactor balance for the heterotrophic organisms:

pj7 = p2 O2 FI QI + RI Vj_

## Reactor balance for the nitrifying organisms:

l di

= F 2 N 2 - F i N i + R2Vi

## Organic substrate balance in the settler:

j^

V 2 dS 2

= F i S i - F3S2 - F4S2

3t

V 2 dA 2

= F A

I I -

- F4A2

= Fl l - F3 02

## Balance for nitrifying organisms in the settler:

V2 dN2 34 = F i N i - F 3 N 2
The equations for the flow rates are given below. Recycle flowrate:

F2 = F 0 R
where R is the recycle factor. Reactor outlet flow: Flow of settled sludge:

FI = F2 + F0 = F O R + FO

## 8.4 Continuous Reactors

269

where C is the concentration factor for the settler. Flow of exit substrate:

F4 = FI - F3>
Flow of exit sludge wastage:
F5 = F3 - ?2.

Note that C and R must be chosen so that F5 is positive. Monod-type equations are used for the growth rates of the two organisms.

l^2max
2 = ^Ni =

Program
The program is given on the CD-ROM.

Nomenclature Symbols
A C F Fo-5 KI K2 N O R Ammonia substrate concentration kg/m3 Concentrating factor for settler Flow rate m3/h Flow rates, referring to the figure m3/h Saturation constant of heterotrophs kg/m3 Saturation constant of nitrifying organisms kg/m3 Concentration of nitrifiers kg/m3 Concentration of heterotrophs kg/m3 Recycle factor -

270

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Rl
R2

V Y Hi

Growth rate of heterotrophics kg/m3h Growth rate of nitrifying organisms kg/m3h Organic substrate concentration kg/m3 Volumes m3 Yield coefficients kg/kg Specific growth rate of heterotrophs 1/h Specific growth rate of nitrifying organisms 1/h

Indices
Flow and concentration indices referring to Fig. 1 are as follows: 0 Refers to feed and initial values 1 Refers to reactor and organic oxidation 2 Refers to settler and ammonia oxidation 3 Refers to recycle 4 Refers to settler effluent 5 Refers to sludge wastage m Refers to maximum

Exercises

## 8.4 Continuous Reactors

271

Results
The results in Fig. 2 demonstrate the influence of flow rate on the effluent organics 82- The ammonia in the effluent A2 is seen, in Fig. 3, to respond similarly to FQ, but for a very high value of FQ = 1000 m3/h the nitrification ceases, and A2 becomes the same as the inlet value AQ. This corresponds to washout of the nitrifiers, which would be seen by plotting NI versus time.
Run 4: 405 steps in 0.0167 seconds
0.9 , 0.8 -I ,/"

M if
0.3- rr
02.

If J II / It

J II " 0-1 JM

## 82:1(20) 82:2(180) 82:3(340) 82:4(500)

" . 6

10

12

14

16

18

20

TIME Figure 2. Transient of S2 at various flow rates F0 (20 to 500m3/h, bottom to top).

## Run 4:405 steps in 0.0167 seconds

0.1 -I

0.090.08-

3 0.06 -|-_005 J* 1
0.040.03-

S.

## A2:1(20) A2:2(180) -_A2:3(340) I A2:4(5QO)

0.02J
0 2 4 6 8 10 TIME 12 14 16 18 20

Figure 3. Ammonia in the effluent (A2) at various flow rates F0 (5 to lOOOm^/h, bottom to top).

272

8.4.4 System

## Tubular Enzyme Reactor (ENZTUBE)

A tubular, packed-bed, immobilized-enzyme reactor is to be investigated by simulation. The flow is assumed to be ideal plug flow. The distribution of the enzyme is not uniform and varies linearly from the inlet to higher values at the outlet, as shown in Fig. 1.

## Enzyme concentration Enzyme distribution

Distance along reactor, Z Figure 1. Distribution of enzyme along the tubular reactor.

Model
The equations for steady state operation are given below. Substrate balance,
dS dZ
=

1 ~v

Kinetics,

The linear flow velocity is increased by the presence of the solid enzyme carrier particles according to

273

V7 L

F Ae
~

vm = KE

## and the linear distribution of enzyme distribution given by,

E = E0 + mZ

Program
The model is solved by renaming the independent variable, TIME, to be the reactor length coordinate Z. The program is given on the CD-ROM.

Nomenclature Symbols
A F K KM m r S vm vz Z e E Reactor tube cross section Flow rate Rate constant Michaelis-Menten constant Enzyme distribution constant Reaction rate Substrate concentration Maximum reaction velocity Linear flow velocity Reactor length Void volume fraction of packing Enzyme concentration m2 m3/h 1/h kg/m3 kg/m3 m kg/m3 h kg/m3 kg/m3h m/h m kg/m3

274

Indices
0 S

## Refers to inlet Refers to substrate

Exercises

Results
Flow rate is the primary operating variable, along with enzyme loading and inlet concentration. In Fig. 2 the influence of F is seen in the steady-state, axial, substrate profile.

## 8.4 Continuous Reactors

275
Run 6:1000 steps in 0.05 seconds

10

12

14

16

18

20

8.4.5
System

## Dual Substrate Limitation (DUAL)

In defined-nutrient growth media, one substrate can usually be made to be limiting by adjusting its concentration relative to those of the other medium components. In general, however, more than one substrate may limit the cell growth rate. In this case the yield coefficients for the various components, Yxsi> may vary depending upon the growth regime. This situation was discussed by Egli et al. (1989), who examined results at steady state with dual nutrient limitation. The present mathematical model simulates the transient behaviour of such a dual (Si -carbon, 82 -nitrogen) nutrient-limited system when carried out in a chemostat. The model assumes that the yield coefficients are each a function of the ratio 81/82, i.e. the ratio of the carbon-nitrogen substrate concentrations in the vessel. The original paper took the carbonnitrogen ratio in the feed stream as the controlling parameter. Here the concentrations in the reactor are assumed to be controlling.

Model
Assuming a perfectly mixed, constant volume continuous-flow stirred-tank reactor, the mass balance equations for the cells and for the two limiting substrates are as follows:

276

= D

(SlFeed - Si) -

D
where D = F/V.

## (o2pee(j o2) lv YQO J ]LlA

/O

O \

I ^__i^^_ I i i "V

## The specific growth rate is modelled as

Si V
S

The yield coefficients are assumed to vary with the carbon-nitrogen ratio in the reactor. Si RATIO = ^ The yield coefficients are varied according to RATIO using the following logic:
Y X Sl=Yi m i n where, _ Y2min Y 2max Bi = \r 1 - and Bo = 1v~,T"
Imax - 1mm

and and

if if

## RATIO < B i RATIO > B2

The boundaries of the three growth regimes in Fig. 1 are defined by the quantities BI and B2.

## 8.4 Continuous Reactors

277

C limitation

Double limitation

N limitation

10

XSi
r

0.8
XS1

B2

S2

Figure 1. Limitation regions for carbon and nitrogen showing influence on yield.

The yield coefficients for biomass on nitrogen and carbon take maximum or minimum values when only one substrate is limiting and vary linearly with opposing tendencies in the double-limitation region.

Program
Note that the programing of this example is rather more complicated than usual owing to the need to allow for the logical conditions of carbon limitation, nitrogen limitation or both substrates together causing limitation. A partial listing is seen below and the full program is on the CD-ROM.
(CALCULATION OF YIELD VALUES)

YXSl=if (RATIO < Bl) then YlMAX else ( if (RATIO > B2) then Y1MIN else (Y1MAX+(RATIO-B1)/(B2B1)*(Y1MIN-Y1MAX)) ) YXS2 = if (RATIO < Bl) then Y2MIN else ( if (RATIO > B2) then Y2MAX else (Y2MIN+(RATIO-B1)/(B2Bl)*(Y2MAX-Y2MIN)) )

278

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Nomenclature Symbols
Bi
B2 Cc Cn D F
Ratio of Y 2min /Yi max Ratio of Y 2max /Yi min Carbon source concentration Nitrogen source concentration Dilution rate Volumetric feed rate Affinity constant Reaction rates Carbon source concentration Nitrogen source concentration Biomass concentration Yield coefficient Specific growth rate

_
-

kg/m3 kg/m3

1/h
m3/h kg/m3 kg/m3 h kg/m3 kg/m3 kg/m3 kg/kg

KS
R

Si S2 X Y H

1/h

Indices
1
2

Exercises

## 8.4 Continuous Reactors

279

Results
The startup of a continuous culture is shown in Fig. 2. Note that the nitrogen level 82 in the reactor drops to a low level after 15 h and causes a change in the yield coefficients. The influence of dilution rate on the system was investigated by varying D from 0 to 1.5 as shown in Fig. 3.

3-c

X 1.5

280

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Reference
Egli, Th., Schmidt, Ch. R. (1989). "On Dual-Nutrient-Limited Growth of Microbes, with Special Reference to Carbon and Nitrogen Substrates", in Proceed. Microb. Phys. Working Party of Eur. Fed Biotech. Eds. Th. Egli, G. Hamer and M. Snozzi, Hartung-Goree, Konstanz, 45-53. This example was developed by S. Mason, ETH-Zurich.

8.4.6
System

## Dichloromethane in a Biofilm Fluidized Sand Bed (DCMDEG)

The process involves the removal of dichloromethane (DCM) from a gas stream and the subsequent degradation by microbial action. The reactor consists of biofilm sand bed column with circulation to an aeration tank, into which the substrate and oxygen enters in the gas phase, or the substrate can be fed in a liquid stream, as shown in Fig. 1. The column is approximated by a series of six stirred tanks. The reaction is treated with homogeneous, double saturation kinetics with dichloromethane (DCM) inhibition. The oxidation of one mole of DCM produces 2 moles of HC1, making a hydrogen ion balance for pH important. The yield with respect to oxygen is 4.3 mg DCM/mg 62. In practice, care must be taken to prevent stripping of DCM to the air stream.

## 8.4 Continuous Reactors

281

SR6>

SRin C jn , pH jn

## Figure 1. Schematic of fluidized bed column with external aeration vessel.

Model
The model does not include a gas balance on the aeration tank, since it is assumed that the gas phase dynamics are comparatively fast and hence an equilibrium with the inlet concentration of oxygen and DCM may be assumed. The biomass is assumed to grow slowly, and growth rates are therefore also not modelled. The model for pH changes does not include buffering effects. For the inlet section 1 at the bottom of the column the balances are as follows: O2 balance,

dCQ1 ^ dt

282

DCM balance,
dC

Srl _ dt

Srin~ C Srl

## H ion balance, dCHi

i -CHI

2r S i 84900

Here T is the residence time of the liquid in one section of the column. The constant 84,900 converts grams to moles and includes the stoichiometry. pHi = -0.434 log |Cm| Evaluation of rates for the inlet section 1:

maxCSrl

-01

KI )

## For the aeration tank the 62 and DCM balances are:

K L a 0 2(Co2eq-Coin)

-~ = (CSr6 - CSrin )

dC

at

R V

DCM (Cs2eq -

## Tr- (CSFO ~ CSrin

Program
The program constants describe DCM entering the reactor in the gas stream. The DCM concentration in the liquid feed is set to zero. The program is on the CD-ROM.

## 8.4 Continuous Reactors

283

Nomenclature
H+ ion concentration in section n kg mol/m3 Inlet dissolved oxygen concentration g/m3 Oxygen saturation constant g/m3 DCM saturation constant g/m3 DCM inlet concentration g/m3 DCM concentration in section n g/m3 Oxygen concentration in section n liquid g/m3 DCM concentration in feed g/m3 DCM gas concentration g/m3 Feed rate m3/h Inhibition constant g/m3 Transfer coefficients for DCM and 2 1/h Saturation constants g/m3 pH in n section n pH units Recirculation rate m3/h Oxygen uptake rate in section n g/m3 h Substrate uptake rate in section n g/m3 h Reactor volume m3 Volume of aeration tank m3 Maximum degradation rate g/m3 h Yield coefficient for DCM/oxygen Liquid residence time in one section h

oin

srin

CSFO

CSG F
KI KLa KS
pHn R

VR

VT
Vmax YSO

Exercises
1 1

284

## 8 Simulation Examples of Biological Reaction Processes Using Berkeley Madonna

Results
The concentrations in the stream leaving the top of the column (CSr6) during startup of the fluidized bed are shown in Fig. 2 for four values of F (0.5 to 10) The change of the pH for one flow rate (F = 0.5) is shown in Fig. 3.

## 8.4 Continuous Reactors

285
Run 4: 55 steps in 0.0167 seconds

0.05

0.1

0.15

0.2

0.25

0.3

0.35

0.4

0.45

0.5

TIME

Figure 2. Fluidized bed startup for four values of F (0.5 to 10, bottom to top).

3.5

3
2.5

2
1.5

## CSR6:1 ... PH6:1

1
0.5

0 0
0.05

0.1

0.15

0.2

0.25

0.3

0.35

0.4

0.45

0.5

TIME

Figure 3. Change of carbon substrate and pH in the top section 6 during startup.

Reference
D. Niemann Ph.D. Dissertation 10025, ETH, 1993.

286

8.4.7
System

## Two-Stage Chemostat with Additional Stream (TWOSTAGE)

Two chemostats are arranged in series (Fig. 1) with the intention that the first operates at a relatively high rate of cell growth, while the second operates at low growth rate, but high cell density, for secondary metabolite production. Additional substrate may be fed to the second stage.
, 810

X1.S!

Hi US

## Figure 1. Two-stage chemostat with two feed streams.

Model
The balance equations are written for each component in each reactor. Stage 1 with sterile feed,

= F[S O -S!] -

## 8.4 Continuous Reactors

287

Stage 2 with additional substrate feed and an input of cells and substrate from Stage 1,
V2

- [F + Fi]X2

KS + S2

v Y

## Productivity for biomass: First stage, Prodi = Both stages, Prod2 =

V,

Program
The program is on the CD-ROM.

Nomenclature Symbols
F

KS
Prod S V X Y

Volumetric feed rate Saturation constant Productivity for biomass Substrate concentration Reactor volume Biomass concentration Yield coefficient Specific growth rate

3

288

Indices
0 1 2 10 m

## Refers Refers Refers Refers Refers

to tank 1 inlet to tank 1 and inlet of tank 2 to tank 2 and outlet of system to separate feed for tank 2 to maximum

Exercises

Results
The results in Fig. 2 give biomass concentrations and productivities for both tanks during a startup with a constant feed stream to the first tank (F = 0.5). In Fig. 3 the influence on X2 of feed to the second tank Fl (0 to 1.0) with constant F is shown.

## 8.4 Continuous Reactors

289
Run 1: 805 steps in 0.0333 seconds
T5

35

40

Figure 2. Biomass (Xj X2) and productivities for both tanks (F = 0.5).

## Run 4: 805 steps in 0.0333 seconds

5
4.5

4
3.5

32.5
2
1.5

1
0.5

0 10 15 20 25 30 35 40

TIME

Figure 3. Influence on X2 of feed to the second tank (Ft = 0 to 1.0, curves right to left).