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Afasfa Kinetics
Afasfa Kinetics
CHNG 3804
Biological Systems
Kinetics and Modelling
Background
A catalyst is a substance that affect the rate of reaction
without altering the reaction equilibrium.
Enzymes, enzyme complexes, cell organelles and whole
cells are catalysts in bioprocesses.
The bio-catalyst are either viable or non-viable, growing
or non-growing.
In Most bioprocesses the reaction is homogenous (i.e.
there is not temperature and concentration gradients in
the reactor.
Reaction Yield
The extent to which reactants are converted to products
is expressed as the yield of reaction.
The theoretical yield that can be obtained from
equilibrium reaction can be different from actual yield.
Yield =
Reaction Thermodynamics
A + bB yY + zZ
C y Cz
K = Y bZ
C A CB
0
Grxn
RT
0
0
Hrxn
Srxn
ln K =
+
RT
R
LnK =
Reaction Rate
Volumetric rate. The total mass of
reactant converted in a system depends
on the size of the reactor. It is more
common to specify the rate of reaction as
the rate per unit volume (rA). The
volumetric rate of reaction is calculated in
a constant volume.
aA + bB yY + zZ
dMA
=Mi Mo R A
dt
dMA
RA =
dt
R A 1 dMA
rA =
=
V
V dt
dC A
rA =
dt
dMA
dt
1 1 dMA
rA = ( or )
X E
dt
1 1 dC A
rA = ( or )
x e dt
RA =
X: cell concentration,
e: enzyme concentration
Kinetics of a Reaction
rA = kCaA CBb
k = Ae
E
RT
ln k = ln A
The kinetics of many biological reactions are either zeroorder, first order or a combination of these called MichealisMenten kinetics.
Bacterial Growth
Growth of population of bacteria
Factor affecting growth
Nutritional requirement
E
RT
Temperature
Major factors affecting growth rate of bacteria
Upper limit determined by
Protein termolability
lysis
Collapse of cell membrane
GR = b( T Tm )
Min.
Optimum
Max.
Thermophiles
40-50
55-75
60-80
Mesophiles
10-15
30-45
35-47
Psychrotrophs
obligate
Facultative
-5-5
15-18
19-22
-5-5
23-30
30-35
Nutrient Concentration
Growth rate depends on the type and amount of
nutrients supplied.
Growth of E-Coli in minimal medium + glucose is less
than that obtained by growth in complete medium
(e.g. nutrient broth) containing amino acids and
growth factors.
Medium Populations
For each species of microorganisms, growth is
most rapid at certain degree of alkalinity or
acidity.
Bacteria usually grow in the pH range of 4-8.
Yeasts prefer a pH range of 3-6.
Molds grow well in a pH range of 3-7.
Higher eukaryotic cells (human and animal)
grow best in a range of 6.5-7.5.
H2O
Water Availability
Not all water in and around cell is available for use in
cellular reactions.
Water bound by molecules is unavailable. The higher the
solute concentration, the less available the water in
solution.
In order to use water bound up by solutes, energy must
be expended by the cell.
Reduction of water availability by addition of sugars, salt,
drying is basis of food preservation.
Most bacteria grow only in aw range of 0.85-1.
aw = mole of pure water/(mole of pure water + moles of
solute)
H2O2 + O2
pH
Most bacteria can tolerate range of 1.5 to 2 pH unit either side of their
optimum.
Bacteria are well buffered internally to cope with small changes in pH.
Larger change in pH require energy to pump H+ ions into or out of the cell.
pH affects:
Growth of Populations
Given a supply of nutrients, bacteria will grow at
a logarithmic rate
N = N02kt
K: growth rate (generation per hour)
Mean generation time (MGT) is a time taken for
a population to double (1/K)
Phases of Growth
Lag phase: time required for the cell to repair
sub lethal damage and adapt to new
environment
Exponential phase: Grow at a max. rate for that
environment.
Stationary phase: cell division balanced by cell
death
Death phase: death rate greater than growth
rate
Microbial Growth
Typical Simplified Batch growth given in
microbiology text books.
Stationary
Death
Log Cell
Number
Exponential
Lag
Time
Modeling/Data Analysis
Noise
Exacerbated the need to take derivatives
Need to consider experimental errors and
the uncertainty of parameters.
Mathematical Models
A mathematical model is a set of relationships
between inputs (manipulated variables) and outputs
(the fermenter states).
In the case of fermentations, the inputs are typically
the conditions in the fermenter, such as temperature,
media composition and substrate concentration.
The outputs are typically biomass, products and byproducts.
In more complex models intracellular compounds
may also be included.
Uses of Models
The motivations for modelling fermentations are varied,
reasons including;
improving understanding,
process control,
fermenter simulation,
optimisation and
estimation of conditions within the fermenter.
Types of Models
Types of Models
Unstructured
Structured
Single component
Heterogeneous individual
cells
Microbial Growth
Many but not all microorganisms grow
exponentially
Dependent on type of microorganism
Dependent on sugar/substrate
concentration
Microbial Growth
Growth Rate
Stationary
Death
Log Cell
Number
Exponential
Lag
S
Time
Growth Equations
Biomass formation is most simply described by
assuming that the rate of growth is proportional to the
number or mass of cells present.
dX
= X
dt
This proportionality is termed the specific growth rate
and is generally symbolised by and has units of hr-1.
This proportionality is not however constant, and is
affected by many factors such as the type of substrate
and its concentration.
Mycelial fermentations generally dont fit this model well
Growth Equations
In the following equations, a number of parameters and
variables are common;
max S
km + S
Monod Kinetics
Typical value of Km quite small
Specific growth rate during lag phase is constant.
Saturation constant (affinity constant, Km) inversely
proportional to affinity of the organism for its substrate.
Substance
Glucose
Km
(mg/L)
1.0
Organism
Enterobacter Aerogenes
Glucose
2.0-4.0
Escherichia coli
Glucose
25.0
Saccharomyces cerevisiae
Ribose
3.0
Hansenula polymorphia
Ammonia
0.1
Eni aerogenes
Ka
max
1 + K s S
max S
Bx + S
Growth Equations
All of these equations predict that the growth
rate decreases as the substrate concentration
decreases.
A number of authors have looked at these
different types of models and found few
differences between them.
Some equations have the advantage that they
can be integrated analytically.
However as the growth equation is generally
part of a much larger model, this is of little use.
Also necessary to include by-product inhibition
terms.
max S
km + S + S 2 / K I
Method 3
Construct model and use a minimisation routine to
determine parameters
Requires a lot of noise free data for complex models
Enzyme kinetics
[E ][S ] = K
[ES ] S
-d[S]0/dt
k2
E + S ES E + P
zero order
kinetics
/2
dP
dt
v = k 2 [ES ]
v=
first order
kinetics
K
[E0 ] = [E ] + [ES ]
[E ] = [E0 ] [ES ]
[S]0
Same form of
equation as Monod
Michaelis Menten
kinetics assumes that
the combination of the
Enzyme and the
substrate is rapid and
reversible.
1
v
Km
vmax
Simple
But, tends to crowd
data at high substrate
concentrations
Where
vmax = k2 [E0 ]
and
KM = KS
1
Km
1
S
v = vmax
Km
Hanes Plot
K mv
S
v max
Km
v
S
vmax
S
S
K
=
+ m
v vmax vmax
Km
vmax
S
v
Substrate Uptake
Biomass
Substrate for growth
CO2+H2O
CO2+H2O
Cell
Substrate for production
Substrate Uptake
Equation below typical for batch system
dS
1 dX
1 dP
=
mX
dt
YX / S dt
YP / S dt
Substrate (sugar) uptake is generally assumed to be
linearly related to
Cell numbers Maintenance Energy (m)
Cell growth Yield Coefficient (YX/S)
Product formation - Yield Coefficient (YP/S)
For simulations need to ensure that sugar
concentration is not < 0
CO2+H2O
Excreted product
It is a function of temperature
More complex form equation is used for spores
and as the above profile is not linear in such
systems.
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Parameter Determination
Product Formation
By Product Inhibition
By Product Inhibition
Inhibitory By-Products
Substrate concentrations
Dissolved Oxygen concentrations
Acetate (E.coli)
Ethanol (Yeast)
Inhibition term = 1
Inhibitory By-Product
by product tolerance
uninhibited
production rate
By Product Inhibition
Red Xs data
Blue Line Model n=1
Green line Model
n~2
Without extra data
hard to tell if n=1
over predicts both
P
Pmax
Running Models
Need to solve several ODEs
Biomass
Substrate
Product
By-product
Oxygen
Software
Matlab, Simulink
Excel (Eulers method, need small step size, simple
models)
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Review
References
Pauline M. Doran, Bioprocess Engineering
Principles, Elsevier Academic Press, 2004.
M. L. Shuler and F. kargi, Bioprocess Engineering:
Basic concepts, Prentice-Hall (1992).
Bailey, J. E., Ollis, D. F., Biochemical Engineering
fundamentals, McGraw Hill (1986)
Wang, D I C, Cooney, C H, Demain, A L, Humphery A
E, Lilly M D, Fermentation and enzyme technology, J.
Wiley, 1979.
Aiba, S, Humphery A E and Millis, N F, Biochemical
engineering, Academic press 1973.
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