Cell Kinetics

Cell Kinetics and
Fermenter Design
Dr. Sivashankar Raja
1
Cell Kinetics and Fermenter
Design:Introduction
Understanding the growth kinetics of microbial,
animal, or plant cells is important for the design and
operation of fermentation systems employing them.
Cell kinetics deals with the rate of cell growth and how
it is affected by various chemical and physical
conditions.
Cell kinetics is the result of numerous complicated

networks of biochemical and chemical reactions and
transport phenomena, which involves multiple phases
and multicomponent systems.
2
Design:Introduction
During the course of growth, the
heterogeneous mixture of
young and old cells is
continuously changing and
adapting itself in the media
environment which is also
continuously changing in
physical and chemical
conditions.
3
Design:Introduction
4
Design:Introduction
Design:Introduction
6
Design:Introduction
7
Design:Definitions
A few definitions:
The growth rate definitions:
8
Design:Definitions
Growth rates definitions:
9
Design:Definitions
Growth Rates:
10
Design:Definitions
Division Rates:
11
Design:Definitions
12
Growth Cycle for Batch Cultivation
Six phases of growth and death
of unicellular microorganisms:
13
Six phases of growth and death
of unicellular microorganisms:
14
The Lag Phase:
• The lag phase occurs because the cells must

adjust to the new medium.
• When the transfer is from low to high nutrient
medium, the lag period is longer. This is because
the cell must produce necessary enzymes for
metabolism.
• Another important factor affecting the length of
the lag phase is the size of the inoculum. If a small
amount of cells are inoculated into a large volume,
they will have a long lag phase.
15
The Lag Phase:
For large-scale operation of the cell culture, it is

our objective to make this lag phase as short as
possible. Therefore, to inoculate a large fermenter,
we need to have a series of progressively larger
seed tanks to minimize the effect of the lag phase.
16
Exponential Growth Phase
In unicellular organisms, the progressive

doubling of cell number results in a
continually increasing rate of growth in the
population.
The rate of cell population increase at any

particular time is proportional to the number
density (CN) of bacteria present at time.
17
18
19
20
Factors Affecting the Specific Growth Rate
1. Substrate concentration
2. Product concentration
3. Others
(pH of the medium, Temperature and
Oxygen supply
21
Factors Affecting the Specific Growth Rate
Limiting Substrate Concentration:
Monod Equation:
22
Limiting Substrate Concentration:
23
24
STATIONARY AND DEATH PHASE
25
Stirred-tank Fermenter
26
For a large-scale operation, the stirred-tank

fermenter (STF) is the most widely used design in
industrial fermentation. It can be employed for both
aerobic or anaerobic fermentation of a wide range
of cells including microbial, animal, and plant cells.
Figure 6.3 shows a diagram of a fermenter used for

penicillin production (Aiba et al., 1973).
27
• The mixing intensity can be varied widely by choosing a

suitable impeller type and by varying agitating speeds.
The mechanical agitation and aeration are effective for
the suspension of cells, oxygenation, mixing of the
medium, and heat transfer. The STF can be also used for
high viscosity media.
• The agitator is effective in mixing the fermenter content,

but it consumes a large amount of power and can
damage a shear-sensitive cell system such as
mammalian or plant cell culture.
28
29
BATCH OR PLUG FLOW FERMENTER
• In a tubular-flow fermenter, nutrients and
microorganisms enter one end of a cylindrical
tube and the cells grow while they pass through.
• Since the long tube and lack of stirring device

prevents complete mixing of the fluid, the
properties of the flowing stream will vary in both
longitudinal and radial direction. However, the
variation in the radial direction is small
compared to that in the longitudinal direction.
• The ideal tubular-flow fermenter without radial

variations is called a plug-flow fermenter (PFF).
30
31
32
33
34
The Monod kinetic parameters 𝝁max, KS can’t be estimated as we did in
case of Michaelis-Menten kinetics. The initial reaction rate is always zero
due to lag phase, during which Monod kinetics doesn’t apply. It should
be noted that even though the monod equation has the same form as the
Michaelis-Menten equation, the rate equation is different. In the
Michaelis –Menten equation,
35
Ideal Continuous Stirred-tank Fermenter
36
Continuous culture systems can

be operated as chemostat or as
turbidostat. In a chemostat the
flow rate is set at a particular value
and the rate of growth of the
culture adjusts to this flow rate.
37
In a turbidostat the turbidity is set at a

constant level by adjusting the flow
rate.
It is easier to operate chemostat than
turbidostat, because the former can be
done by setting the pump at a constant
flow rate, whereas the later requires an
optical sensing device and a controller.
38
However, the turbidostat is recommended

when continuous fermentation needs to
be carried out at high dilution rates
near the washout point, since it can
prevent washout by regulating the flow
rate in case the cell loss through the
output stream exceeds the cell growth in
the fermenter.
39
40
41
The above expression is valid only when 𝛕m 𝛍max >1.

If 𝛕m 𝛍max <1, all the cells will wash out and eq. 6.31
not valid.
42
43
Evaluation of Monod Kinetic Parameters
By measuring
44
Evaluation of Monod Kinetic Parameters
45
Multiple Fermenters Connected in Series
46
47
48
CSTF AND PFF IN SERIES
49
For CSTF we can calculate:
50
For PFF we can calculate:
51
the growth kinetics in a PFF can

be significantly different from that in a CSTF.
52
MULTIPLE CSTFs IN SERIES
53
54
55
56
57
CELL RECYCLING
58
PFF WITH CELL RECYCLING
A PFF requires the initial presence

of microorganisms in the inlet stream as a
batch fermenter requires initial inoculum.
The most economical way to provide cells
in the inlet stream is to recycle the part of the
outlet stream back to the inlet with or
without a cell separation device.
59
Figure 6.17 shows the schematic diagram

of a PFF with cell recycling. Unlike the CSTF,
the PFF does not require the cell separator
in order to recycle, though its presence
increases the productivity of the fermenter
slightly.
60
61
62
63
64
CSTF WITH CELL RECYCLING
65
The cellular productivity in a CSTF

increases with an increase in the
dilution rate and reaches a maximum
value. If the dilution rate is increased
beyond the maximum point,
66
the productivity will be decreased

abruptly and the cells will start to
be washed out because the rate of
cell generation is less than that of
cell loss from the outlet stream.
67
One way to improve the reactor
productivity is to recycle the cell
by separating the cells from the
product stream using a cross-flow
filter unit (Figure 6.19).
The high cell concentration maintained

using cell recycling will increase
the cellular productivity since the
growth rate is proportional to the cell
concentration.
68
However, there must be a limit in the increase

of the cellular productivity with increased cell
concentration because in a high cell
concentration environment, the nutrient-
transfer rate will be decreased due to over
crowding and aggregation of cells.
The maintenance of the extremely high cell

concentration is also not practical because
the filter unit will fail more frequently at the
higher cell concentrations.
69
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73
ALTERNATIVE FERMENTERS
74
75
76
ALTERNATIVE FERMENTERS:
COLUMN FERMENTERS
77
ALTERNATIVE FERMENTERS:
LOOP FERMENTERS
78
STRUCTURED KINETIC MODEL
TO REPRESENT CELL KINETICS
79
Structured models
• A much more general approach with much greater a priori

predictive power is a model capturing the important kinetic
interactions among cellular subcomponents. Initially,
chemically structured models were based on two
components, but at least three components appear
necessary to give good results.
• More sophisticated models with 20 to 40 components are

being used in many laboratories. A schematic of one such
model is given in Fig. 6.14.
An idealized sketch of the model E. coli B/rA growing in a glucose–ammonium salts medium with
glucose or ammonia as the limiting nutrient. At the time shown the cell has just completed a round
of DNA replication and initiated cross-wall formation and a new round of DNA replication. Solid
lines indicate the flow of material, while dashed lines indicate flow of information. The symbols
are: A1, ammonium ion; A2, glucose (and associated compounds in the cell); W, waste products
(CO2, H2O, and acetate) formed from energy metabolism during aerobic growth; P1, amino acids;
P2, ribonucleotides; P3, deoxyribonucleotides; P4, cell envelope precursors; M1, protein (both
cytoplasmic and envelope); M2RTI, immature ―stable‖ RNA; M2RTM, mature ―stable‖ RNA (r-RNA
and t-RNA—assume 85% r-RNA throughout); M2M, messenger RNA; M3, DNA; M4, nonprotein
part of cell envelope (assume 16.7% peptidoglycan, 47.6% lipid, and 35.7% polysaccharide); M5,
glycogen; PG, ppGpp; E1, enzymes in the conversion of P2 to P3; E2, E3, molecules involved in
directing cross-wall formation and cell envelope synthesis; GLN, glutamine; E4, glutamine
synthetase; * indicates that the material is present in the external environment.
In eq. 6.63, the rfi term must be in terms of intrinsic concentrations and the term µnetCi
/X represents dilution by growth.
The model indicated in Fig. 6.14 is that for a single cell. The single-cell model
response can be directly related to culture response if all cells are assumed to behave
identically. In this case each cell has the same division cycle. A population will have,
at steady state, twice as many cells at ―birth‖ as at division. The average
concentrations in the culture will be at the geometric mean (a time equal to multiplied
by the division time) for each cell component.

Cell Kinetics

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Cell Kinetics

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Cell Kinetics and

Dr. Sivashankar Raja

Cell kinetics is the result of numerous complicated

The growth rate definitions:

• The lag phase occurs because the cells must

For large-scale operation of the cell culture, it is

In unicellular organisms, the progressive

The rate of cell population increase at any

For a large-scale operation, the stirred-tank

Figure 6.3 shows a diagram of a fermenter used for

• The mixing intensity can be varied widely by choosing a

• The agitator is effective in mixing the fermenter content,

• Since the long tube and lack of stirring device

• The ideal tubular-flow fermenter without radial

Continuous culture systems can

In a turbidostat the turbidity is set at a

However, the turbidostat is recommended

The above expression is valid only when 𝛕m 𝛍max >1.

the growth kinetics in a PFF can

A PFF requires the initial presence

Figure 6.17 shows the schematic diagram

The cellular productivity in a CSTF

the productivity will be decreased

The high cell concentration maintained

However, there must be a limit in the increase

The maintenance of the extremely high cell

• A much more general approach with much greater a priori

• More sophisticated models with 20 to 40 components are

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