MICROBIAL GROWTH

KINETICS
Microbial Growth

Microorganisms grow in a wide spectrum of physical and chemical

environments

Growth and other physiological activities → response to their

physiochemical environment

Microbial growth is usually characterized by the time required to double

cell mass or cell number

Mass doubling time mau differ from cell doubling time → cell mass can
increase without an increase in cell number

Two processes associated with microbial growth: nutrient uptake from the
environment and release of metabolic end products to the environment
Bacterial Growth

Bacteria divide by binary

fission, cell doubles its mass
and the amount of all cell
constituents

Doubling time for some

bacteria are 15-20 minutes
under optimal growth
condition, however 45-60
minutes is more typical
Yeast Growth
Yeast commonly divide by
budding, exception include
yeast that grow by fission or by
forming hyphae

Sometimes, the buds don’t

separate from the mother cell
and long, branched chains of
cells (pseudomycelia) will form.

Under optimal condition, yeast

may divide in as little as 45
min, however, 90-120 min is
more typical
Mold Growth

Molds, characteristically grow by chain elongation and branching

Growth proceeds from the tip of mycelium (apical growth) by

forming septa between the cells

Depending on the physicochemical environment, the mycelium may

be long and diffuse, short and highly branched, or a mixture of the
two

In submerged culture the mycelia may exist as diffuse mycelia or

may form pellets with diameters ranging from 0.1 – 10 mm
Mold Growth (con’t)

Very difficult to follow growth (cell

number in molds) because the cells are
not easily separated

Increases in mass are used to monitor

growth

Typically molds have optimal mass

doubling times of 4-8 hours, some are
double in as little as 60-90 minutes.
Method for Measuring Microbial Growth

• Microscopic Count
Measurement of • Viable Plate Count
Cell Number • Slide Culture
• Coulter Counter

Measurement of
• Cell Dry Weight
Cell Mass: Direct • Turbidity
Methods

Estimation of Cell • Cell Components

• Nutrient Uptake
Mass: Indirect • Product Formation
Methods • Packed-Cell Volume
KINETICS OF MICROBIAL GROWTH
BATCH CULTURE
Closed system which contains an initial, limited
amount of nutrient.
BATCH CULTURE

Microbial Growth Phase

EXPONENTIAL
PHASE:
LAG PHASE: STATIONARY
growth rate PHASE:
a time off gradually
adaptation increases, cells growth rate
grow at a constant, decreases
maximum rate
ln biomass concentration
A = Lag phase
B = Log or exponential phase
C = Deceleration phase
D = Stationary phase

A B C D
Time

Growth of a typical microbial culture in batch condition

The stationary Microbial Microbial
phase in batch growth phase growth phase
culture (Borrow, 1961): (Bullock, 1965):
The growth rate has Balanced phase:
declined to zero early to middle
exponential phase Trophophase :
exponential phase
The population still
metabolically active Storage phase: late
exponential →
May produce products accumulation of lipid
called secondary and carbohydrate Idiophase :
metabolite stationary phase →
Maintenance phase: secondary
Bull (1974): maximum stationary phase metabolites are
population phase produced
 Material Balance on Cells

A material balance on the cell mass written around the

fermentor is given by equation 1.

Cells in – cells out + cell growth – cell death = cell accumulation

F F dX
Xo − X + X − X =
V V dt

Xo = the cell mass (g/l) in the feed

X = the cell mass (g/l) in the fermentor
F = the medium flow rate (l/hr)
V = the fermentor volume (l)
µ = the specific growth rate (hr-1)
α = the specific death rate (hr-1)
t = time (hr)
The Growth of Unicellular Microorganisms
Exponential phase: cells grow at a constant, maximum rate. The
exponential phase may be described by the equation:
dx
= x
dt …………………..1)

x = concentration of microbial biomass

t = time (h-1)
 = specific growtoh rate (h-1)

On integration and logarithmic equation gives :

xt = xo et …………….……2)
ln xt = ln xo + t ………………….3)
xo = original biomass concentration
xt = biomass concentration after the time interval, t hours

During the exponential phase, nutrients are in excess and the organisms is
growing at its maximum specific growth rate, max
The limitation of growth

The equations predict that growth will

continue indefinitely

However, after a certain time the growth

rate of the culture decreases until growth
ceases (substrate limitation, toxin
limitation, or a combination of the two)
 B C D

Biomass conce4ntration at
stationary phase

Initial substrate concentration

The effect of initial substrate conc. on the biomass conc.

at the onset of stationary phase, in batch culture
This figure show the nature of the limitation of growth
The limitation of growth (con’t)

•x = Y(SR –s) …………4)

•x = the concentration of biomass produced
A-B •Y
•SR
= the yield factor (g biomass produced g-1substrate consumed)
= the initial substrate concentration
•s = the residual substrate concentration

B-C • Utilization of the substrate is deleteriously affected by either the

accumulating toxins or the availability of another substrate

• An increase in the initial substrate concentration does not give a

C-D proportional increase in biomass.
• Exhaustion of another substrate or the accumulation of toxic product
The limitation of growth (con’t)

The decrease in growth rate and the cessation of growth,

due to the depletion of substrate → describe by the
relationship between  and s (Monod, 1942):

 max s
= …………………..5)
Ks+ s
s = the residual substrate concentration
Ks = the substrate utilization constant,
= equal to substrate concentration when  = ½ max
= a measure of the affinity of the organism for its
substrate
 C A B

Specific growth rate

Residual limiting substrate concentration

A-B : exponential phase where substrate conc. Is in excess and growth is at  =  max
C-A : the deceleration phase due to depletion of substrate which will not support  max

The effect of residual limiting substrate conc. on the

specific growth rate of a hypothetical bacterium
Low Ks value High Ks value
Organism has a very high Organism has a low affinity
affinity for the limiting for the substrate
substrate

Growth rate not affected until Growth rate deleteriously

the substrate concentration has affected at relatively high
decline to a very low level substrate concentration

Short deceleration phase Long deceleration phase

 Method 1
dx/dt = r = Y/X

Dry weight ( kg sel m-3)

x3 Method 2
dx/dt = r = (x3-x1) / 2

x2 Y
x1 µ = r/x
X

t1 t2 t3
waktu (jam)

Estimation of µ via curve fitting

 Slope = ks/µm
(kg glukosa h m-3)

1/µ (h)
1/µm

- 1/ks 1/S (m3kg glukosa -1)

Determination of µmax and ks by Lineweaver-Burke Plot

KINETICS OF PRODUCT FORMATION

Growth-linked products: primary metabolites

Non-growth-linked products: secondary metabolites

PEMBENTUKAN PRODUK PADA KULTUR CURAH

Batch fermentation may be used to produce:

Biomass Primary metabolite Secondary metabolite

production production
• Cultural condition • Condition to extend • Condition giving a
supporting the the exponential short exponential
maximum cell phase accompanied phase and an
population would be by product excretion extended stationary,
used or production phase
• Condition giving a
decreased growth
rate in the log phase
resulting in earlier
secondary
metabolite formation
Cellular Growth and Product Yields

Growth and product formation by microorganisms are bioconversion

processes

Nutrients fed to fermentation are converted to cell mass and metabolites

Each of these conversions may be quantized by a yield coefficient

expressed as the mass of cells or product formed per unit mass or nutrient
consumed, YX/S and YP/S for cells and product, respectively.

Yield coefficient represents the efficiency of conversion

Cellular growth and product yield

Yield can be calculating by measure the mass of

cells or product produced and substrate
consumed over some time period

X = Y
x/s
S

P = Y
P/s
S
Kinetic patterns or growth and product formation in
batch fermentation

X
or
P

Time Time Time

A B C

A. Growth-associated product formation

B. Mixed-growth-associated product formation
C. Non growth-associated product formation
Kinetics of Product formation

 Primary metabolites, synthesized by growing cell (growth linked products):

dp
=q px ………………..1)
dt

p = concentration of product
qp = specific rate of product formation

 Product formation is related to biomass production:

dp /dx = Yp/x ………………..2)

Yp/x = Yield of product in terms of substrate consumed

(g produk g-1 biomasa h-1)
Kinetics of Product Formation

 Multiply equation 2) by dx/dt:

dx/dt . dp/dx = Yp/x . dx/dt
dp/dt = Yp/x . dx/dt

but: dx / dt = x
therefore: dp / dt = Yp/x . x ……………….3)

equation 1): dp / dt = qp . x

combining equation 1) and 3)

qp . x = Yp/x . x
qp = Yp/x .  ……………….4)
Kinetics of Product Formation

From equation 4):

When product When product
formation is growth formation is non-growth
associated: associated:
• qp increases with  • qp may remain constant
over a wide range of 
or it may vary in a
complex manner
Kinetics of Product Formation

Relationship of specific growth rate and specific product synthesis rate

µ/
µmax

qp/
qpmax

time time
A B

A. Growth-associated product
formation
B. Mixed-growth-associated product
formation
C. Non growth-associated product
formation
time
C
Effect of temperature and pH on bacterial
growth – Secondary model
Effect of temperature and pH on bacterial
growth - Secondary model