(1)microbial growth and product formation

7/22/2019 (1)Microbial Growth and Product Formation

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MICROBIAL GROWTH

ANDPRODUCT FORMATION


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1.1 Phases of the Batch Growth Cycle

Lag phase

Exponential growth phase

Declining growth phaseStationary phase

Death phase


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Cell division occurs in the exponential phase.

The rate of increase of cell number (N) is proportional to the

no.of cells

Instead of cell no., we can use dry cell weight per volume X

(kgm-3) as a measure of cell concentration

During the exponential phase in a batch reactor we can write

)1(Xdt

dX

where is the specific growth rate of the cells

Eqn. (1) can be integrated from the end of the lag phase( , ) to any point in the exponential phase (X, t) to

give0XX lagtt

0

XX )( lagtte

or

0

lnX

X= )( lagtt (2)


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The time required for the cell numbers or dry weight to double,

ie the doubling time, is related to the specific growth rateby:

At low nutrient concentrations, depends on nutrient

concentration.

At high nutrient concentrations, reaches a maximum value.

The end of the exponential phase arises when some essential

nutrient is depleted, or when some toxic metabolite accumulates

to a sufficiently high level.

Following the exponential phase, the rate of exponential growthdecreases (declining growth phase) and is followed by the

stationary phase.

Following this is the death phase, when cell lysis occurs and the

population decreases.

dt

dt)2(ln

2lndt

or

(3)


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Monod assumes that only one substrate (the growth limiting

substrate, S) is important in determining the rate of cell growth.

For batch growth at constant volume:

where

At high substrate concentration, >> , eqn. (4) reduces to

zeroth order dependence on substrate concentration.

1.2 Relationship between growth rate and substrate concentration

)(

..max

SK

XS

dt

dX

s

(4)

max is the maximum specific growth rate

sK is the value of the limiting substrate

concentration which results in a growth

rate of half the maximum value

S is the concentration of the limiting substrate

S sK

ie at S sK max>> ; = (5)


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At low substrate concentration,


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Values of are generally quite small, implying that is near

for much of the period of batch growth. This apparent zeroth order

dependence on justifies the term exponential growth.

sK max

S


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The Arrhenius relationship generally holds for the temperature

dependence of ,

1.3 Temperature Effects

hence k=A. exp)(

RT

Ea

becomes )(

RT

Ea

=A. exp (7)

where A is Arrhenius constant

Ea is activation Energy

R is Universal Gas Constant

T is temperature

The temperature dependence of growth rate comes from the fact

that growth employs enzymes. The processes catalysed by enzymes

often have optima associated with the rise in k due to rise in

temperature, and the denaturation of enzymes at elevate

temperatures.


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The effect of pH on microbial growth parallels that observed for

enzymes.The effect of pH on enzymes is associated with the ionization of

-amino and carboxyl terminal groups in proteins, in addition to

many other ionizable groups on constituent amino acids, which

could give rise to complex kinetics when pH is varied.

1.4 pH Effects

Oxygen is consumed together with substrate and converted tobiomass and products. Varying the amount of dissolved oxygen

concentration has the same effect as varying the concentration of

the limiting substrate.

1.5 The Effect of Dissolved Oxygen Concentration


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A microbial cell consumes nutrients and generates products in

order to reproduce, through a set of reactions called cellular

metabolism.

Microorganisms are classified on the basis of their nutritionaland energy requirements for cellular metabolism.

1.6 Product Formation

1.6.1 Cellular Metabolism

Energy Source

Light

Chemical

(breaking of bonds)

Phototrophs

Chemotrophs

Organotrophs Litotrophs

(oxidation of organicmaterial) (oxidation ofinorganic material)


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Carbon Source

Autotroph

Heterotroph

CO2 as source of carbonorganic sources of carbon

Most organisms we deal with are chemoheterotrophs, ie requiring

organic carbon source and a chemical source of energy.

1.6.2 Growth-Associated Product Formation

End products of energy and carbon metabolisms are called primary

metabolites

These products are referred to as growth-associated products, as theirrate of production parallels the growth of the cell population.

Examples: - Ethanol produced by anaerobic fermentation of glucose by

yeast

- Production of gluconic acid from glucose by Gluconobacter


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1.6.3 Non Growth-Associated Product Formation

Most organisms we deal with are chemoheterotrophs, ie requiring

organic carbon source and a chemical source of energy.

Some products are produced in batch cultures at the end of the

exponential phase.They are formed from secondary metabolism, ie they are

secondary metabolites, and are non growth-associated products, and

their kinetics do not depend on rate of growth of cultureExamples :- Production of the antibiotic penicillin byP. chrysogenum


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1.6.4 The Intermediate Class of Products

An intermediate class of products, where product formation kinetics lie

between the two classes above, are called partially growth-associatedproducts.

Examples :- amino acids, lactic acid, citric acid, extracellular

polysaccharides (eg xantan), solvents (eg acetone).

1.7 Fermentation Profiles

An intermediate class of products, where product formation kinetics lie

between the two classes above, are called partially growth-associated

products.

Once a potentially useful fermentation process is identified, its time

course, or fermentation profile is plotted for use as a basis for

comparison in later scale-up and optimization of process parameters.


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1.8 Growth Yield

Growth yield is defined as YdS

dX

If defined asY is constant; then dXYdS

If the boundary conditions are:atat

00 ,,0 SSXXt SSXXtt ,,

then XXSSY 00 )(thus )( 00 SSYXX (8)


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Now take edn (4)

)(

..max

SK

XS

dt

dX

s

(4)Since )1(X

dt

dX

Thus)(

maxSK

S

s (9)


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Now take eqn. (8)

)( 00 SSYXX (8)

From eqn. (8) we get:XXSSY 00 )(

or00 YSXXYS

andY

XXS

Y

YSXXS

)()( 00

00


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Put this expression for S into eqn.(9) we get:

Y

XXSK

YXXS

s0

0

00

max

But from eqn.(1) we have:

)1(X

dt

dX

ie

dT

dX

X

1


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so

YXXSK

Y

XXS

dT

dX

X s 00

00

max

1

(10)

Integrating eqn.(10), we get:

0

00

0

0

00

0

0

0

max.

.lnln.

SY

XXSY

Y

XS

K

X

X

Y

XS

Y

XKS

t s

s

(11)


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1.9 Productivity and Production Rate

If we assume the Monod model holds, and if we define the

conversion from the limiting substrate S to microorganism biomass as

the yield , and if we assume that there is no lag phase, and

that there is no cell death, then what is the productivity of a constant

volume fermentation system?

dS

dXY

Biomass productivity is defined as how much biomass we can get perunit volume of fermenter, per unit time (e.g. kg/m/hr).For a batch system, time is needed between runs, to clean the

fermenter, load the medium, and sterilize it. This time is called the

Down Time .dt

A plot of biomass concentration vs time will look as follows:


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XMaximum productivity here

Point m where dT

dX

is maximum

0X

Maximum production rate is at point m.

Maximum productivity may be later since maximumproductivity is not calculated from time zero.

Productivity is overall production rate, ie involving down timedt


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Now recall Monodseqn (9) ie

(9)

)(

maxSK

S

s

The general plot of versus is as follows S


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The value of varies between substrates. If for a particular

substrate the value of is very small, then from eqn. (9) will

have a value close to max for most of the fermentation, untillsubstrate Sis almost exhausted, and until that point is reached, the

process has zero order kinetics.

sK

sK


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So a process with a small is less sensitive to substrate

concentration

And a process with a big is more sensitive to substrateconcentration

sK

sK


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In a process with a small the approximation to zero order

kinetics occur from high Sright down to very low values of S

The exponential phase will extend further before the microb will

begin to be affected by the depletion in S

sK


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Each student please pick a fermentation process for the

production of a different product, either from the list given

below, or from outside the list, and find and copy its

fermentation profiles on a piece of squared paper, make a

photocopy, keep the original, and submit the photocopy

during the next lecture:

Penicillin

Amino acid

Citric acid

Xantan

AcetoneEthanol

Biopolymer

Erythromycin

Yeast

ASSIGNMENT 1


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ASSIGNMENT 2

Use your original fermentation profiles

drawn on the squared paper and assuming

the down time of each fermentation is 4

hours, apply the method suggested in thelecture to estimate the productivity of

biomass production for your fermentation.

dt

(1)microbial growth and product formation

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