Cell Growth Models

Lecture 6
Fall 2007
Quantifying Growth Kinetics
• Structured vs Unstructured -
– S - model divides the cell mass into components
– U - assumes fixed cell composition - exponential
growth phase of batch or in continuous
• Segregated vs Nonsegregated
– S - assumes different types of cells exist
– N - all cells are the same type

Unstructured, Nonsegregated Models
(Monod)

• Assumptions
– One limiting substrate
– Semi empirical relationship
– Single enzyme system with
Michaelis-Menten kinetics is
responsible for the uptake of
substrate
– Amount of enzyme is
sufficiently low to be growth
limiting
– Low population density
• Most commonly used
expression for growth

S K
S
S
m
+
=
µ
µ
µ
m
- maximum growth rate when
S >> K
s
• K
s
- saturation constant -
concentration of the rate-limiting
substrate when the specific rate
of growth is equal to one half of
the maximum.
• K
s
= S when µ = 1/2µ
m

• In general µ = µ
m
for S >>
K
s
and for S << K
s

S
m
K
S µ
µ =
Cell Growth - Monod
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0 5 10 15 20 25
Time (hr)
C
e
l
l

C
o
n
c
e
n
t
r
a
t
i
o
n

(
g
/
L
)
Does not
account for
death phase
Does not
account for
lag phase
Unstructured, Nonsegregated Models
• Other models - Section 6.3.2.1 in text
assuming one limiting substrate
– Blackman
– Tessier
– Moser
– Contois
• Multiple Substrates are growth limiting
– Usually do not use unstructured, nonsegregated
model
Inhibition Models
(Very similar to enzyme models)
• Substrate Inhibition
– High substrate concentration inhibits growth
– If a single-substrate enzyme catalyzed reaction is the
rate-limiting step then inhibition of enzyme activity
results in inhibition of microbial growth.
Noncompetitive Substrate Competitive Substrate


|
|
.
|

\
|
+
|
.
|

\
|
+
=
I
S
m
K
S
S
K
1 1
µ
µ
S
K
S
K
S
I
S
m
+
|
|
.
|

\
|
+
=
1
µ
µ
Inhibition Models - cont.
• Product Inhibition
– High concentrations of product can be inhibitory
– Underlying mechanism of product inhibition is unknown
– Approximated as exponential or linear decay functions
Noncompetitive Product Competitive Product

|
|
.
|

\
|
+
|
.
|

\
|
+
=
P
S
m
K
P
S
K
1 1
µ
µ
S
K
P
K
S
P
S
m
+
|
|
.
|

\
|
+
=
1
µ
µ
Inhibition Models - cont.
• Inhibition by Toxic Compounds
– inhibition of growth is analogous to enzyme inhibition
Noncompetitive Competitive



Uncompetitive Cell Death
|
|
.
|

\
|
+
|
.
|

\
|
+
=
I
S
m
K
I
S
K
1 1
µ
µ
S
K
I
K
S
I
S
m
+
|
|
.
|

\
|
+
=
1
µ
µ
|
|
.
|

\
|
+
|
|
.
|

\
|
+
+
=
I I
S
m
K
I
S
K I
K
S
1
) / 1 (
µ
µ
'
d
S
m
k
S K
S
÷
+
=
µ
µ
Batch Reactors
• Cell Growth


• Substrate Utilization


• Product (cometabolic contaminants use negative sign)

X
S K
S
X
dt
dX
r
S
m
X
+
= = =
µ
µ
S X S
m
S X
S
Y
X
S K
S
Y
X
dt
dS
r
/ /
+
÷ = ÷ = =
µ µ
|
µ
o
| oµ µ
+
+
= =
+ = = =
S K
S
dt
dP
X
q
Y
dt
dP
X
q
S
m
p
g g X P p
1
1
/
Logistic Equation
• Batch Growth
Equation
• Combines Batch
growth, Monod and
Yield Coefficients
• No maintenance
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0 5 10 15 20
Time (hr)
C
e
l
l

C
o
n
c
e
n
t
r
a
t
i
o
n

(
g
/
L
)
Rate Expression for Growth
(1)

Yield Expression
(2)
Substituting into Eq. 1 for S from Eq 2:


Integrating

Logistic Equation
X
S K
S
X
dt
dX
r
S
m
X
+
= = =
µ
µ
) (
0 / 0
S S Y X X
S X
÷ = ÷
X
X X S Y Y K
X X S Y
dt
dX
S X S X S
S X m
) (
) (
0 0 / /
0 0 /
÷ + +
÷ +
=
µ
| | t S Y X X S Y
X S Y
Y K
X
X
X S Y
X S Y Y K
m S X S X
S X
S X S
S X
S X S X S
µ = ÷ +
+
÷
|
|
.
|

\
|
+
+ +
0 / 0 0 /
0 0 /
/
0 0 0 /
0 0 / /
/ ) ( ln
) (
ln
) (
) (
Describes
Sigmoidal
shape batch
growth curve
Unstructured, Nonsegregated Models
• Disadvantage of Unstructured, Nonsegregated Models
– No attempt to utilize or recognize knowledge about cellular
metabolism and regulation
– Show no lag phase
– give no insight to the variables that influence growth
– assume a black box
– assume dynamic response of a cell is dominated by an
internal process with a time delay on the order of the
response time
– most processes are assumed to be too fast (psuedo ss) or too
slow to influence the observed response.

Filamentous Organisms
• Types of Organisms
– mold
– bacteria or yeast entrapped in a spherical gel particle
– formation of microbial pettlets in suspension
• Model - no mass transfer limitations

– R - radius of the cell floc or pellet or mold colony
Then the growth of the biomass (M)can be written as



const k
dt
dR
= =
3 / 2
2 2
4 4
M
dt
dM
or
R k
dt
dR
R
dt
dM
p
¸
µ t t µ
=
= =
3 / 1
) 36 ( tµ ¸
p
k =
Where:
Filamentous Organisms - cont.
• Integrating the equation:



• M
0
is usually very small then
• Model is supported by experimental data.


3 3
3 / 1
0
3 3
|
.
|

\
|
~
|
.
|

\
|
+ =
t t
M M
¸ ¸
3
t M ·
Chemically Structured Models

• Improvement over nonstructured, nonsegregated models
• Need less fudge factors, inhibitors, substrate inhibition, high
concentration different rates etc.
• Model the kinetic interactions amoung cellular
subcomponents
• Try to use Intrinsic variables - concentration per unit cell
mass- Not extrinsic variables - concentration per reactor
volume
• More predictive
• Incorporate our knowledge of cell biology

In Class Exercise
• The results shown below correspond to typical
batch culture dynamics. Calculate the following:
• A) µ
max
assuming Monod growth
• B)

Y
X/S

• C) o and | assuming mixed growth and
• non-growth associated product formation

Quantifying Growth Kinetics • Structured vs Unstructured – S .exponential growth phase of batch or in continuous • Segregated vs Nonsegregated – S .model divides the cell mass into components – U .assumes fixed cell composition .all cells are the same type .assumes different types of cells exist – N .

Unstructured.saturation constant concentration of the rate-limiting substrate when the specific rate of growth is equal to one half of the maximum. • Ks = S when  = 1/2m • In general  = m for S >> Ks and  m S for S << Ks  KS . Nonsegregated Models (Monod) mS • Assumptions  – One limiting substrate – Semi empirical relationship – Single enzyme system with Michaelis-Menten kinetics is responsible for the uptake of substrate – Amount of enzyme is sufficiently low to be growth limiting – Low population density KS  S • Most commonly used expression for growth  m .maximum growth rate when S >> Ks • Ks .

2 0.3 0.Cell Growth .6 0.5 0.1 0 0 5 10 15 20 Does not account for death phase Does not account for lag phase 25 Time (hr) .7 Cell Concentration (g/L) 0.4 0.Monod 0.

2.Unstructured. Nonsegregated Models • Other models .1 in text assuming one limiting substrate – – – – Blackman Tessier Moser Contois • Multiple Substrates are growth limiting – Usually do not use unstructured.Section 6. nonsegregated model .3.

Inhibition Models (Very similar to enzyme models) • Substrate Inhibition – High substrate concentration inhibits growth – If a single-substrate enzyme catalyzed reaction is the rate-limiting step then inhibition of enzyme activity results in inhibition of microbial growth. Noncompetitive Substrate m   KS 1  S  S  1   K  I     Competitive Substrate m S   S K S 1   K I   S   .

Inhibition Models .cont. • Product Inhibition – High concentrations of product can be inhibitory – Underlying mechanism of product inhibition is unknown – Approximated as exponential or linear decay functions Noncompetitive Product  m  KS 1  S  P   1   K    P  Competitive Product  m S  P  K S 1   K S  P   .

cont.Inhibition Models . • Inhibition by Toxic Compounds – inhibition of growth is analogous to enzyme inhibition Noncompetitive m   KS 1  S  I  1   K  I     Competitive m S   I K S 1   K I   S   Uncompetitive m S  Cell Death       KS I   (1  I / K )  S 1  K  I I    mS KS  S '  kd .

Batch Reactors • Cell Growth rX   S dX  X  m X dt KS  S • Substrate Utilization m S X dS X rS    dt YX / S K S  S YX / S • Product (cometabolic contaminants use negative sign) 1 qp  X 1 qp  X dP  YP / X  g   g   dt m S dP   dt KS  S .

2 0.3 0.4 0.Logistic Equation 0.1 0 0 5 10 15 20 • Batch Growth Equation • Combines Batch growth.7 0. Monod and Yield Coefficients • No maintenance Time (hr) Cell Concentration (g/L) .6 0.5 0.

Logistic Equation Rate Expression for Growth mS dX rX   X  X dt KS  S (1) Yield Expression X  X 0  YX / S ( S 0  S ) Substituting into Eq. 1 for S from Eq 2: (2)  m (YX / S S 0  X 0  X ) dX  X dt ( K S YX / S  YX / S S 0  X 0  X ) Integrating Describes Sigmoidal shape batch growth curve ( K S YX / S  YX / S S0  X 0 )  X  K S YX / S   ln   (Y S  X ) ln (YX / S S0  X 0  X ) / YX / S S0    mt (YX / S S0  X 0 ) X /S 0 0  X0  .

Unstructured. . Nonsegregated Models – No attempt to utilize or recognize knowledge about cellular metabolism and regulation – Show no lag phase – give no insight to the variables that influence growth – assume a black box – assume dynamic response of a cell is dominated by an internal process with a time delay on the order of the response time – most processes are assumed to be too fast (psuedo ss) or too slow to influence the observed response. Nonsegregated Models • Disadvantage of Unstructured.

no mass transfer limitations dR  k  const dt – R .radius of the cell floc or pellet or mold colony dM dR   4R 2  k p 4R 2  dt dt or dM  M 2 / 3 dt Then the growth of the biomass (M)can be written as Where:   k p (36 ) 1/ 3 .Filamentous Organisms • Types of Organisms – mold – bacteria or yeast entrapped in a spherical gel particle – formation of microbial pettlets in suspension • Model .

3 . • Integrating the equation:  1/ 3 t   t  M   M0      3 3  3 3 • M0 is usually very small then M  t • Model is supported by experimental data.Filamentous Organisms .cont.

Not extrinsic variables . high concentration different rates etc. nonsegregated models • Need less fudge factors.Chemically Structured Models • Improvement over nonstructured. inhibitors.concentration per reactor volume • More predictive • Incorporate our knowledge of cell biology . substrate inhibition. • Model the kinetic interactions amoung cellular subcomponents • Try to use Intrinsic variables .concentration per unit cell mass.

Calculate the following: • A) max assuming Monod growth • B) YX/S • C)  and  assuming mixed growth and • non-growth associated product formation .In Class Exercise • The results shown below correspond to typical batch culture dynamics.

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