Batch Culture

Cell Growth Kinetics
- Introduction
- Growth patterns and kinetics in batch
culture
- growth phases
- effect of factors: oxygen supply
- heat generation
- Growth kinetics (Monod Equation)
- Growth in continuous culture (ideal
chemostat)
Growth Kinetics
Introduction
- Autocatalytic reaction: The rate of growth is directly related to ce
concentration
substrates + cells → extracellular products + more cells

∑S + X → ∑P + nX
S: substrate concentration (g/L); X: cell mass concentration (g/L);

P: product concentration (g/L); n: increased number of biomass.
Net specific growth rate (1/time):

1 dX
 net 
t: the time X dt
Growth Kinetics
Introduction
Net specific growth rate (1/time):
 net   g  k d
g : Gross specific growth rate (1/time)
kd : The rate of loss of cell mass due to cell death

or endogenous metabolism
Endogenous metabolism: during the stationary phase,

the cell catabolizes cellular reserves for new building blocks and
for energy-producing monomers.
Growth Kinetics
Introduction
Net specific replication rate (1/time):
1 dN
R 
N dt
R
R   '  kd
R
N : Cell number concentration (cell number /L)

 ' : Gross specific replication rate (1/time)
R
kd : The rate of cell death (1/time)

Growth Kinetics
Introduction
- Quantifying cell concentration:
- direct: no suspended solid and interference
compounds.
cell mass concentration – preferred

dry weight, optical density (turbidity) (600-700nm
Wave Length)
cell number density:

Petroff-Hausser slide (hemocytometer), plate counts,
etc.
Growth Kinetics
- Quantifying cell concentration:
- indirect: direct method is inapplicable. (mold solid
state fermentation)
Cell mass can be determined by measurement
of protein, DNA or ATP. e.g. 1mg ATP/g dry weight
bacterial cell.
If 100 mg ATP/L is measured, then the cell mass:
100 mg (ATP/L)/1 mg ATP/g dry cells=100 (g dry
weight cells/L)
Growth Kinetics
- Growth patterns and kinetics in batch culture
- growth phases
In batch culture:
- lag phase
- logrithmic or exponential growth phase
- deceleration phase
- stationary phase
- death phase
Typical growth curve for a bacterial population
Batch Growth Kinetics
Lag phase
A period of adaptation for the cells to their
new environment
• New enzymes are synthesized.
• A slight increase in cell mass and volume, but no
increase in cell number
• Prolonged by low inoculum volume, poor
inoculum condition (high % of dead cells), age of
inoculum, nutrient-poor medium
• Multiple lag phases: (diauxic growth) medium
contains more than one carbon source
Diauxic growth

Exponential growth phase
In this phase, the cells have adjusted to
their new environment and multiply rapidly
(exponentially)
• Balanced growth –all components of a cell
grow at the same rate.
• Growth rate is independent of nutrient
concentration, as nutrients are in excess.
The balance of cell mass in a batch culture
gives:
dX
  net X , X  X 0 at t  0
dt
Integration of the above equation yields :
X
ln  μnet t , or X  X 0e  net t
X0
X and X 0 are cell concentrations at time t and t  0
The slope  net is constant.

 net   R  μm
μm is the maximum specific growth rate (1/time)
Doubling time of cell mass: the time required to double

the microbial mass:
ln X / X 0 ln 2 0.693
d   
 net
 net
 net
Deceleration growth phase
Very short phase, during which growth decelerates

due to either:
• Depletion of one or more essential nutrients
• The accumulation of toxic by-products of growth
(e.g. Ethanol in yeast fermentations)
• Period of unbalanced growth: Cells undergo
internal restructuring to increase their chances of
survival
Stationary Phase:
With the exhaustion of nutrients (S≈0) and build-up
of waste and secondary metabolic products
- The growth rate equals the death rate.
- There is no net growth in the organism population.
- Cells may have active metabolism to produce secondary
metabolites.
Primary metabolites are growth-related: ethanol by S.
cerevisae.
Secondary metabolites are non-growth-related:
antibiotics, pigments.
Kinetic Pattern of Growth and
Product Formation
Growth-associated Mixed-growth-associated Non growth-associated

Stationary phase
- Cell lysis may occur and viable cell mass may drop.
A second growth phase may occur and cells may grow on
lysis products of lysed cells (cryptic growth)
- Endogenous metabolism occurs by catabolizing cellular
reserves for new building blocks and energy-producing
monomer (maintenance energy).
The rate describing the conversion of cell mass into
maintenance energy or the loss of cell mass due to cell
lysis:
dX
 kd X
dt
k d is the rate constant for endogenous metabolism.
Death Phase:
The living organism population decreases with time,
due to a lack of nutrients and toxic metabolic by-
products.
The rate of death usually follows:
dN '
 kd N
dt
'
k d is the first - order death rate constant.
Yield coefficients: defined based on the amount of
consumption of another material.
X
Growth yield : Y 
X /S S
P
Product yield : Y 
P/S S
Growth yield based on consumptio n of oxygen :
X
Y 
X / O2 O2
S  S assimilation  S assimilation  S growth energy S maintenance
into biomass into an energy
extracellular
product
Yield coefficients: defined based on the amount of
consumption of another material.
For most bacteria and yeast:

Yx/s =0.4-0.6 g/g glucose
Yx/O =0.9 – 1.4 g/g O2
2
At the end of the batch growth period, the measured yields are
apparent as of endogenous metabolism occurring, Kd > 0,
which changes the metabolic pathways of the substrate.
e.g. M App
YX / S  YX / S

Batch Culture

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Cell Growth Kinetics

substrates + cells → extracellular products + more cells

S: substrate concentration (g/L); X: cell mass concentration (g/L);

Net specific growth rate (1/time):

kd : The rate of loss of cell mass due to cell death

Endogenous metabolism: during the stationary phase,

N : Cell number concentration (cell number /L)

kd : The rate of cell death (1/time)

cell mass concentration – preferred

cell number density:

Typical growth curve for a bacterial population

Typical growth curve for a bacterial population

Doubling time of cell mass: the time required to double

Very short phase, during which growth decelerates

Growth-associated Mixed-growth-associated Non growth-associated

For most bacteria and yeast:

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