Chemostat with Cell Recycle
- To keep the cell concentration higher than the normal
steady-state level in a chemostat. - To increase the cell and growth-associated product yield. - For low-product-value processes: e.g. waste treatment. fuel ethanol ,
X1, S
Chemostat with Cell Recycle
Cell mass balance (qp=0, kd 0, X0=0, Monod equation is applied):
where =net=g-kd
A chemostat can be operated at dilution rates higher than the specific growth rate when cell recycle is used.
Chemostat with Cell Recycle
When kd=0
Chemostat with Cell Recycle
Mass balance on growth-limiting substrate (qp=0, kd 0, X0=0, Monod equation is applied):
FS 0 FS V g X 1 1 YX / S
M
(1 ) FS V
dS dt
At steady state, dS/dt 0, D M X1 Y X / S (S0 S )
Since g [1 (1 C )]D, X1 Y X / S ( S0 S )
M
K s D(1 C ) m D(1 C )
1 C
, X1
YX / S [1 (1 C )]
[S0
K s D(1 C ) ] m D(1 C )
Chemostat with Cell Recycle
No recycle
m=1.00h-1, S0=2.0g/l, Ks=0.01 g/l, Yx/s=0.5 g/g, concentration factor C=2.0 and recycle ratio =0.5
Chemostat with Cell Recycle
Cell mass balance around the cell separator.
X1, S
(1 ) FX 1 FX 2 FCX 1 X 2
Example-Chemostat with Cell Recycle
Organisms are cultured in a chemostat with cell recycle. The system is operated under glucose limitation.
F 100 ml/h, V 1000ml, S 0 10 g glucose/L
M YX/S 0.5gdw cells/g substrate; m 0.2h 1 ,
Ks 1g/L, C 1.5, 0.7, X 0 0, K d 0
Determine specific growth rate net, S in the reactor effluent, cell concentration in the recycle stream (CX1) and in the concentrator effluent (X2)
Multi-stage Chemostat System
In some fermentations, the growth and productformation steps need to be separated.
e.g. secondary metabolite, culture of genetically engineered cells.
P10 Growth stage
P2 Product formation stage
At steady state, Vn, Xn,Sn,,Pn in the reactor of each stage dont change with time.
Genetically Engineered Cells (cont.)
Features of Genetically Engineered Cells:
have inserted recombinant DNA (plasmids) which allow for the production of a desired protein product. GE cells grow more slowly than original nonmodified strain (due to the extra metabolic burden of producing product). Genetic Instability causes the GE culture to (slowly) lose ability to produce product. The nonplasmid carrying cells or the cells with mutation in the plasmid (revertants) grow faster.
Genetically Engineered Cells (cont.):
In the first stage, only cell growth occurs and no inducer is added for product formation. The GE cells grow at the maximum rate and are not outcompeted in the first chemostat by revertant cells. When cell concentrations are high, an inducer is added in the latter (or last) chemostat to produce product at a very high rate.
Multi-stage Chemostat System
X0=0, Vi, i=1,2.n constant. Stage 1: cell growth condition, Kd=0, qp=0 dX Cell mass: FX 0 FX 1 1 X 1V1 V1 1 dt At steady state X0 1 D1 (1 ) X1 Limiting substrate: At steady state
FS 0 FS1 V1
1 X 1
Y XM/ S
V1
dS1 dt
S1 S 0
1 X 1
D1YXM/ S
1 , 2 ... and n are net specific growth rates in Stage 1, 2,..., and n, recspectively.
When Kd 0, they are equal to the respective specific gross growth rates in each stage.
Multi-stage Chemostat System
Stage 2 product formation conditions, Kd=0, F=0 dX FX 1 FX 2 2 X 2V2 V2 2 Cell mass: dt X1 At steady state 2 D2 (1 ) where D 2 F X2 V2 F
P10
P2
V2 are constant.
Multi-stage Chemostat System
Stage 2 product formation conditions, Kd=0, F=0 Limiting substrate: At steady state
FS1 FS 2 V2
2 X 2
Y
M X /S
V2
qp X 2 YP / S
V2
dS 2 dt
S 2 S1
2 X 2
D2YXM/ S
qp X 2 D2YP / S
Product: At steady state
FP FP2 V2 q p X 2 V2 1
dP2 dt
P2 P 1
qp X 2 D2
Multi-stage Chemostat System
Stage n product formation conditions, Kd=0, F=0
Similarly, equations could be obtained for nth stage. X n Dn (1 n 1 ) Xn
S n S n 1
n X n
DY
M n X /S
qp X n DnYP / S
Pn Pn 1
qp X n Dn
If n (e.g. Monod model) and qp are known functions, Xn , Pn, and Sn at nth stage could be determined by the above equations.
Fed-Batch
Nutrients are continuously or semi-continuously fed, while effluent is removed discontinuously.
- overcome substrate inhibition or catabolite repression by intermittent feeding of substrate by maintaining low substrate concentration. for production of secondary metabolites e.g. antibiotics, lactic acid, E. Coli making proteins from recombinant DNA technology.
Fed-Batch
Analysis of fed-batch with substrate continuously fed and no output: t=0, V0=0, F is constant.
dV F V Vo Ft dt At quasi steady state, S added=S consumed, X, S concentrations are constant.
Volume:
Cell mass balance:
since
FX O V net X
dV d ( XV ) dX V X , then dt dt dt
d ( XV ) dt
FX O V net X V
V net X X
1 dV F dV net D dt V dt V F F net
V
dX dV X dt dt
V0 Ft
Do 1 D0 t
Ks D
net D
m S
Ks S
if K d 0
Then S
m D
(Monod growth model applied)
Fed-Batch
where S0
assuming XXm
M
where Xt=Xt0 at t=0
Fed-Batch
(g) qp (i.e. g product/g cells-min) at Pt=Pt0, t=0
Fed-Batch
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Example: Fed- Batch
In a fed-batch culture operating with intermittent addition of glucose, the value of V is given at time t=2hr, when the system is at quasi-steady state.
F dV 200 ml/h, V 1000ml, S0 100 g glucose/L dt M YX/S 0.5gdw cells/g glucose; m 0.3h 1 ,
Ks 0.1g/L, X t 0 30 g cells
Determine V0 (initial volume of the culture). At t=2 hr, find S, X and Xt at quasi-steady state if qp=0.2 g product/g cells-h, P0=0.
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