MICROBIAL GROWTH KINETICS

Batch Growth Kinetics

Batch grows refers to culturing cell in a vessel with an initial change of medium that is not altered
by further nutrient addition or removal. This form of cultivation is simple and widely used both
in laboratory and industrially. It occurs in a closed system that contains an initial limited amount
of substrate. Characteristics of batch fermentation system
 Simplest fermentor operation
 Sterilisation can be performed in the reactor
 All nutrients are added before inoculation
 Maximum levels of C and N are limited by inhibition of cell growth
 Biomass production limited by C/N load and production of toxic waste products
 Cells are harvested when biomass levels or product levels start to decline
The inoculated microorganism will pass through a number of growth phases. These are:

i. Lag phase
The first phase observed under batch conditions is the lag phase in which the growth rate is
essentially zero. When an inoculum is placed into fresh medium, growth begins after a period of
time called the lag phase. It is the transition period before the log phase. The lag phase is thought
to be due to the physiological adaptation of the cell to the culture conditions. This may involve a
time requirement for induction of specific messenger RNA (mRNA) and protein synthesis to meet
new culture requirements. The lag phase may also be due to low initial densities of organisms
that result in dilution of exoenzymes (enzymes released from the cell) and of nutrients that leak
from growing cells. Normally, such materials are shared by cells in close proximity. But when cell
density is low, these materials are diluted and not as easily taken up. As a result, initiation of cell
growth and division and the transition to exponential phase may be slowed.

ii. The log phase

The second phase of growth observed in a batch system is the log phase. The log phase also
called the exponential phase is characterized by a period of the exponential growth—the most
rapid growth possible under the conditions present in the batch system.
During exponential growth, the rate of increase of cells in the culture is proportional to the
number of cells present at any particular time. The cell numbers increase exponentially at a
constant maximum rate. In mathematical terms, we can write:
dx/dt = µX ………………………….. (1)
Where:
X is concentration of biomass
t is time in hours
µ is specific growth rate in g/g/h or h-1

Integrating equation (1) gives

Xt = Xo eµt ………………………….… (2)

Where:
Xt biomass concentration at time t
Xo is the original biomass concentration
e is the natural logarithm

By taking the natural logarithm on both side of equation 2

ln Xt = ln Xo + µt ………………………… (3)
(y = c + mX)

Plot of natural logarithm of biomass concentration vs time (t) should produce a straight line
with slope equal to µ.
Doubling time
Doubling time is the amount of time to do one binary fission.
When all condition essential for cell growth are present, the rate of cell increase is proportional
to the number of viable cell present

dx/dt  X … First order autocatalytic ………...1

or dx/dt = X ……………………………………….. ..2

dx/x = dt………………………………………. …3

Integrating equation 3 gives;

ln X2/X1 = µ (t2-t1) ……………………………… 3
For the case where X2 = 2X1 (doubling in cell mass), the doubling time;
td = t2 –t1 = ln 2 ………………………………. 4
td = 0.693/2 ……………………………5

iii. Deceleration phase

Growth rate slowly decreases due to:

 Consumption of nutrient or essential nutrient become depleted (substrate limitation)

 Accumulation of toxic product
The Monod Kinetic try to describe how substrate change with growth rate. This is described by
Monod equation:

 = max S
(Ks + S)
Where:
S is residual of substrate concentration
Ks is an affinity constant which equal to substrate concentration S at µ= µmax
2
iv. The Stationary Phase
The stationary phase in a batch culture can be defined as a state of no net growth, which can be
expressed by the following equation: dX/dt = 0. Although there is no net growth in stationary
phase, cells still grow and divide. Growth is simply balanced by an equal number of cells dying.
There are several reasons why a batch culture may reach stationary phase. One common reason
is that the carbon and energy source or an essential nutrient becomes completely used up. When
a carbon source is used up it does not necessarily mean that all growth stops. This is because
dying cells can lyse and provide a source of nutrients. Growth on dead cells is called endogenous
metabolism. Endogenous metabolism occurs throughout the growth cycle, but it can be best
observed during stationary phase when growth is measured in terms of oxygen uptake or
evolution of carbon dioxide. Thus, in many growth curves the stationary phase actually shows a
small amount of growth. A second reason that stationary phase may be observed is that waste
products build up to a point where they begin to inhibit cell growth or are toxic to cells. This
generally occurs only in cultures with high cell density.

v. The Death Phase

The final phase of the growth curve is the death phase, which is characterized by a net loss of
culturable cells. Even in the death phase there may be individual cells that are metabolizing and
dividing, but more viable cells are lost than are gained so there is a net loss of viable cells. The
death phase can be described by the following equation: dX/dt = -kd X, where k d is the specific
death rate.

Yield coefficient (Y)

A measure of the overall efficiency of the conversion of substrate to cell mass or specific product
is given by a yield coefficient (Y) where it varies depending on organism, pH, temperature and
substrate.
The yield coefficient of cell at different substrate concentrations can be determined by plotting
biomass concentration at stationary phase against initial substrate concentration. The slope gives
an estimate of yield coefficient. The following equations are used in determine the yield
coefficient.

Parameter equation unit

Cell (Yx/S) X/S g/g
Product (Yp/s) P/S g/g
P/X  P/X g/g
Example 1
During the production of ethanol in a 5 L batch bioreactor by Aspergillus glaucus, approximately
10% (V/V) of spore suspension containing 12X 10−12 spores/mL was used as inoculum. The
following profile of the batch culture fermentation of ethanol was obtained.

Time (h) X (g/L) S(g/L) P (g/L)

0 0.02 80 0
1 0.04 65 0.01
2 0.1 43 0.05
3 7.8 40 0.1
4 10.5 37 0.3
5 15.3 30 0.5
6 16.4 24 7.0
7 16.0 20 12.0
8 17.0 15 20.5
9 18.2 5 22.0
10 18.3 2 21.8
11 17.0 0.2 21.0

Determine the following:

i. Specific growth rate (µ), maximum specific growth rate (µmax) and doubling time (td) of
Aspergillus glaucus.
ii. Yield, Yx/s, Yp/s and Yp/x.
iii. Overall and maximum productivity

Solution

I. (a) Specific Growth rate.

In the log phase
ln Xt = ln Xo + t This is equivalent to (y = c + mX).
Where:
Xt = Biomass concentration at time t.
Xo = Biomass concentration at initial state.
 = Specific growth rate.

A plot of lnX versus Time should give a straight line with gradient equals to 
 Time (h) X (g/L) ln X
3 7.8 2.054
4 10.5 2.351
5 15.3 2.728
6 16.4 2.797

So the specific growth rate (µ) = 0.260 h-1

b) The doubling time (Td) of Aspergillus glaucus

Doubling time (td) is given by the following equation

𝑙𝑛2
𝑇𝑑 =
𝜇

0.693
𝑇𝑑 =
0,260

Td = 2.665 h
c) The maximum specific growth rate (µmax) of Aspergillus glaucus

In the log phase, specific growth rate (µ) is at maximum that is the µmax (the maximum specific
growth rate). Therefore, the maximum specific rate = specific growth rate.

So, maximum specific rate = 0.260 h-1

II. Determine the yields Yx/s, Yp/s and Yp/x

a) Yield coefficient of biomass

𝑋𝑚𝑎𝑥−𝑋0
(Yx/s) = ΔX/Δs = 𝑆0−𝑆𝑥𝑚𝑎𝑥

18.3 − 0.02
𝑌(𝑥⁄𝑠) =
80 − 2
Therefore, Yx/s = 0.234g/g

b) Yield coefficient of products

𝑃𝑚𝑎𝑥−𝑃0
(Yp/s) = ΔP/ΔS= 𝑆0−𝑆𝑝𝑚𝑎𝑥

22.0 − 0
𝑌 (𝑝⁄𝑠) =
80 − 5
Therefore, Yp/s = 0.293g/g

c) Yield coefficient of products

𝑃𝑚𝑎𝑥−𝑃0
(Yp/x) = ΔP/Δx = 𝑋𝑝𝑚𝑎𝑥−𝑋0

22.0 − 0
𝑌(𝑝⁄𝑥) =
18.2 − 0.02

Therefore, Yp/x = 1.210g/g

 iii. Overall and maximum productivities of ethanol produced by Aspergillus glaucus

a. The overall productivity of ethanol produced by Aspergillus glaucusis as thus:

𝑃𝑓𝑖𝑛𝑎𝑙 − 𝑃0
𝑂𝑣𝑒𝑟𝑎𝑙𝑙 𝑝𝑟𝑜𝑑𝑢𝑐𝑡𝑖𝑣𝑖𝑡𝑦 =
𝑇𝑓𝑖𝑛𝑎𝑙 − 𝑇0

21.0 − 0
=
11.0 − 0

Therefore, overall productivity = 1.909g/L/h

b. Maximum productivity

𝑃𝑚𝑎𝑥 − 𝑃0
=
𝑇𝑝𝑚𝑎𝑥 − 𝑇0

22 − 0
=
9−0

Therefore, maximum productivity = 2.444g/L/h