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# BioBio-Reactor Operation

Operating Modes
Batch
Fed-Batch
Continuous
Repeated Batch

CHNG 3804
Bio-Reactor Operation

Operating Conditions
Effects of Temperature
Feeding Regimes

mass in
through
system
boundaries

mass out
through
system
boundaries

mass
generated
within
system

mass
consumed
within
system

mass
accumulated
within
system

dM
= M i M o + RG Rc
dt

Batch Fermentation
Typical Model of a batch fermentation
Ignoring volume changes due to evaporation
S
dX
= X = max X
dt
km + S
dS
1 dX
=
mX
dt
Yxs dt
dS
1

X mX =
=
+m X
dt
Yxs
Yxs

## It may also be necessary to include equations for OUR,

product and by-product formation
If the cell death takes place in the reactor another term
need to be added to the first equation

Batch Fermentation
Oldest type of fermentation
fermentation
No broth withdrawal during
fermentation
Gas is vented
Commonly used for beer and wine
production as well as enzymatic
reactions

V
X
S
P

## Fed - Batch Fermentation

course of the fermentation
Typically the carbon source to
control the growth rate
Sometimes termed a semibatch fermentation
Need to include the effect of
substrate feed and change in
volume in the model equations

F
Xi
Si
Pi

V
X
S
P

## Application of Fed - Batch Fermentation

high growth rate.
This is critical in cultures where the oxygen
demand during fast growth is too high for mass
transfer capability of the reactor.
High substrate concentration may also be
inhibitory or switch on undesirable metabolic
pathways.
Fed-batch fermentation is used in production of
bakers yeast and penicillin production.

## Fed - Batch Fermentation

d (VX ) max S
=
X V
dt
km + S

d (VS )

= (
mX ) V + Fi Si
dt
Yxs
dV
= Fi i Evaporation
dt
density of the reactor contents, V is the liquid volume, Fi is the input mass flow rate.

Assuming no
evaporation
and the feed
and
fermenters
density are
equal

dV
= Fi
dt
d (VX )
dX
dV
=V
+X
dt
dt
dt
dX 1 d (VX )
dV
=
X
dt V
dt
dt
dX 1 max S
XV X Fi
=
dt V k m + S
dX
max S Fi
=

X
dt
km + S V

## Fed - Batch Fermentation

dV
= Fi
dt
d (VS )
dS
dV
=V
+S
dt
dt
dt
dS 1 d (VS )
dV
=
S
dt V
dt
dt
dS 1
=
dt V

Yxs

XV mX V + Fi Si S Fi

dS F

= (S i S )
+m X
dt V
Yxs

Dilution Rate
Dilution rate with the unit of T-1 D=F/V
In a fed-batch reactor V increases by time while F is
constant. Therefore D decreases as the reaction
proceeds.
dX
max S
=
D X
dt
km + S
dX
= ( D ) X
dt
dS
1 dX
= D (S i S )
+ mX
dt
Yxs dt
dS

= D (S i S )
+m X
dt
Yxs

dX
= ( D ) X = 0
dt
=D
From Monod model
s=

DK m

max D

dS

= D (S i S )
+m X =0
dt
Yxs

## As D is a function of time the integration of the above

equation is more complicated than equations for batch
fermentation.

Continuous Fermentation
Inflow of nutrients
Outflow of fermenter broth (and cells)
Industrial Fuel Alcohol fermentations
are commonly two or more continuous
fermenters in series
Generally regarded as most efficient
Not widely used

Continuous Fermentation

F
X
S
P

F
Xi
Si
Pi

Contamination problems
Regulation requirements for batch
traceability

V
X
S
P

Continuous Fermentation
Assuming Evaporation = 0 and Fi=Fo

dV
=0
dt
dX
max S F
=

X
dt
km + S V
dS F

= (S i S )
+m X
dt V
Yxs

d (VX ) max S
=
X V Fo X
dt
km + S

d (VS )
1 d (VX )
=
mX V + Fi Si Fo S
dt
Yxs dt
dV
= Fi Evaporatio n Fo
dt

## Continuous Operation of a Mixed reactor

Brewing, Production of bakers yeast and
waste water treatment, and enzyme
conversion
Assume the vessel is well mixed, the
product then has the same composition as
the reactor
Continuos reactor only used for enzyme
that are cheap, as the enzyme is removed
from the system.
For expensive enzymes immobilised form
is used in continuous reactors, then
enzyme retains in the reactor.

F
X
S
P

F
Xi
Si
Pi

V
X
S
P

Chemostat

Turbidostat

## CSTR containing microorganisms

Chemo Chemical, stat stationary or constant
The liquid volume is kept constant, by setting
inlet and outlet flow rates equal.
Can allow determination of many kinetic
parameters
Allows operator to control cell growth rate by
Maximum Productivity and Washout

## CSTR containing microorganisms

The liquid volume is kept constant, by setting
inlet and outlet flow rates equal.
The inlet flow rate is adjusted to keep the
biomass concentration constant.
corresponding to the set biomass concentration.
Turbidostats require more complex monitoring
and control system than chemostats and are not
used in large scale.

## Steady State Cell and Substrate Conc. as a

Function of Dilution rate in a Chemostat

## Continuous Operation of a Mixed reactor

X=

D(Si S )

Yxs

Dcrit =

+m

As D
D(Si S )
X=
D
+m
Yxs

max Si

K m + Si

QX = DX
Qx: is the volumetric productivity.
Volumetric rate of biomass production.

0.45

km

1.2

Yxs

0.5

0.005

Si

100

DK m
X = Si
Yxs
max D

D
D

## Steady State Cell and Substrate Conc. as a

Function of Dilution rate in a Chemostat

max

0.45

km

1.2

Yxs

0.5

0.005

Si

100

max

Washout region

0
max

S
X

0
0

## D Critical is the condition at which the steady

state biomass concentration just become zero

## To avoid wash out from the system the operating

dilution rate must always be less than D Critical

## Near washout the system is very sensitive to

small changes in D which cause relatively large
shifts in x and S

D critical

Qx = D ( S

KmD

max D

)Yxs

(kgm-3hr-1)

## Repeated Batch Fermentation

At the end of a batch fermentation a fraction of
the batch is left in the fermenter
New media is then added and a new batch is
commenced
Better conversion than batch fermentations
Used for

D optimum

## Waste water treatment (Sequencing Batch Reactor)

Beer making
Ethanol production

D optimum and critical are usually close. Therefore it is not practical to operate at D
opti.

## Evaluation of Kinetic and Yield

Parameters in Chemostat Culture
At steady state with sterile feed and negligible cell
death specific growth rate, , is equal to dilution rate,
D.
D=

Km
1
1
=
+
D max S max

max S
Km + S

## Chemostat operation is also convenient for

determining true yields and maintenance coefficient
for cell cultures.

1
1 m
Yxs

Yxs

Y is the observed biomass yield from substrate, Yxs is the true biomass
yield from substrate and m is the maintenance coefficient.

Example

## An entrobacter aerogenes auxotroph capable of overproducing threonine

has been isolated. The kinetic and yield parameters for this organism are
investigated using a 2 litre chemostat with 10 g/l glucose. Steady state cell
and substrate concentrations are measured as a function of flow rate.
Determine the maximum specific growth rate, the substrate constant Km, the
maintenance coefficient, and the true biomass yield for this culture.
Flow rate l/h

## Substrate Conc. g/l

1.0

3.15

0.010

1.4

3.22

0.038

1.6

3.27

0.071

1.7

3.26

0.066

1.8

3.21

0.095

1.9

3.1

0.477

Effects of Temperature
The growth rates of microorganisms vary with
temperature
There is typically a maximum, with a fairly sharp
decrease at higher temperatures
It may not always be best to run the fermentation
at this optimum value
Beer do not want excessive yeast growth
Maximum for product formation may be different to
cell growth
May want to reduce growth rate and hence oxygen
demand and heat duty

Temperature Profiles

Early
Growth
35-37oC

Late
Growth
32-33oC

Production
32oC

Time

Example: Beer

Temperature

Temperature

## Example: an IPTG induced Recombinant

E.coli producing a protein

Temperature Profiles

Early
Growth
18-20oC
Production
12-14oC

Time

## Substrate Concentrations and OURs

Substrate concentration effects growth
rate and hence oxygen uptake rate
High rates of inhibitory by-product formation
(small-medium scale)
High heat duties (large scale)

Feeding Regimes
Different methods for feeding substrates to
fermentation
DO-Stat
pH-Stat
Optimal Profiles

## Hence fed batch fermentations can be

used to avoid this problem

DODO-stat
Substrate feeding is linked to the dissolved
oxygen concentration.
If the dissolved oxygen concentration falls
below a threshold value then feeding is
stopped.
Feeding is not restarted until DO rises
above pre-specified value

Optimal Profiles
Mathematical models of fermentations can
be used to calculate Optimum feeding
profiles
In practice results have been mixed
largely due to plant model mismatches

## Need a model which is very accurate

across all of the likely range of operation
Or to include uncertainty in the
optimisation algorithm

pHpH-Stat
The exhaustion of readily available carbon
sources typically can lead to microorganisms
consuming proteins or amino acids for energy
This results in the production of ammonia
This results in a rise in the pH which can be
used to trigger substrate feeding
The consumption of carbon sources such as
glucose produces hydrogen ions which cause
the pH to drop

Summary
This lecture
Types of fermentations
Batch
Fed-Batch
Continuous
Repeated Batch

## Simple Model Equations

Reference: Bioprocess Engineering
Principle, P. Doran, Chapter 13.