Fermentation

Fermentation
Fermentation is a metabolic process that
converts sugar to acids, gases or alcohol.
It occurs in yeast and bacteria, but also in
oxygen-starved muscle cells, as in the case of
lactic acid fermentation.
Fermentation is also used more broadly to refer
to the bulk growth of microorganisms on a
growth medium, often with the goal of producing
a specific chemical product.
The science of fermentation is known as
zymology
2

Range of Fermentation
Processes

To Produce Microbial cells or Biomass

To Produce Microbial Enzymes
To Produce Microbial Metabolites
To Produce Recombinant Products
To modify a compound which is
added to the fermentation
(Transformation)

Steps to carry out a

Fermentation
The formulation of media to be used in
culturing the process organism during
the development of the inoculum and
in the production fermenter.
The sterilization of the medium,
fermenters and ancillary equipment.
The production of an active, pure
culture in sufficient quantity to
inoculate the production vessel.
4

 The growth of the organism in the

production fermenter under optimum
conditions for product formation.
The extraction of the product and its
purification.
The disposal of effluents produced by
the process

Basic Design of a Fermenter

Various components of an ideal fermenter for batch process are:

Monitoring and controlling parts of fermenter are:

Basic Functions of a
Fermenter
The vessel should be capable of being operated
aseptically for a number of days and should be
reliable in long-term operation and meet the
requirements of containment regulations.
Adequate aeration and agitation should be
provided to meet the metabolic requirements of
the micro-organism. However, the mixing should
not cause damage to the organism.
Power consumption should be as low as possible.
A system of temperature control should be
provided.
9

 A system of pH control should be

provided.
Sampling facilities should be provided.
Evaporation losses from the fermenter
should not be excessive.
The vessel should be designed to require
the minimal use of labour in operation,
harvesting, cleaning and maintenance.
Ideally the vessel should be suitable for a
range of processes, but this may be
restricted because of containment
regulations.
10

 The vessel should be constructed to

ensure smooth internal surfaces, using
welds instead of flange joints whenever
possible.
The vessel should be of similar geometry
to both smaller and larger vessels in the
pilot plant or plant to facilitate scale-up.
The cheapest materials which enable
satisfactory results to be achieved
should be used.
There should be adequate service
provisions for individual plants
11

Materials for Body Construction of a

Fermenter
In fermentations with strict aseptic
requirements it is important to select
materials that can withstand repeated
steam sterilization cycles.
On a small scale (1 to 30 dm3) it is possible
to use glass and/or stainless steel.
Glass is useful because it gives smooth
surfaces, is non-toxic, corrosion proof and it
is usually easy to examine the interior of
the vessel.
12

Two basic types of Fermenters

A glass vessel with a round or flat
bottom and a top flanged carrying
plate.
All vessels of this type have to be
sterilized by autoclaving.

13

14

 A glass cylinder with stainless-steel top

and bottom plates.
Vessels with two stainless steel plates
cost approximately 50% more than
those with just a top plate.
At pilot and large scale, when all
fermenters are sterilized in situ, any
materials used will have to be
assessed on their ability to withstand
pressure sterilization and corrosion and
on their potential toxicity and cost.
15

16

 Pilot scale and Industrial scale vessels are

normally constructed of stainless steel or
at least have a stainless-steel cladding to
limit corrosion.
The American Iron and Steel Institute
(AISI) states that steels containing less
than 4% chromium are classified as steel
alloys and those containing more than 4%
are classified as stainless steels.
Mild steel coated with glass or phenolic
epoxy materials has occasionally been
used
17

 The corrosion resistance of stainless steel is

thought to depend on the existence of a thin
hydrous oxide film on the surface of the
metal.
The composition of this film varies with
different steel alloys and different
manufacturing process treatments such as
rolling, pickling or heat treatment.
The film is stabilized by chromium and is
considered to be continuous, non-porous,
insoluble and self healing.
If damaged, the film will repair itself when
exposed to air or an oxidizing agent.
18

 The minimum amount of chromium

needed to resist corrosion will
depend on the corroding agent in a
particular environment, such as
acids, alkalis, gases, soil, salt or fresh
water.
Increasing the chromium content
enhances resistance to corrosion, but
only grades of steel containing at
least 10 to 13% chromium develop
an effective film.
19

 The inclusion of nickel in high

percent chromium steels enhances
their resistance and improves their
engineering properties.
The presence of molybdenum
improves the resistance of stainless
steels to solutions of halogen salts
and pitting by chloride ions in brine
or sea water.
Corrosion resistance can also be
improved by tungsten, silicone and
other elements.

20

 AISI 316 grade steel 18% chromium, 10%

nickel and 2-1.5% molybdenum are now
commercially usd.
Tungsten, silicone
AISI 317 3-4% molybdenum- to prevent
leaching of metals in citric acid production
pH 1-2
AISI grade 304 18.5% chromium and 10%
nickeused for brewing industry.

21

 At this stage it is important to

consider the ways in which a reliable
aseptic seal is made between glass
and glass, glass and metal or metal
and metal joints such as between a
fermenter vessel and a detachable
top or base plate.

22

DESIGN OF A FERMENTOR
BAFFLES, IMPELLERS &
SPARGER

Standard geometry of a stirred

tank bioreactor
A stirred tank reactor will either be approximately
cylindrical or have a curved base. A curved base
assists in the mixing of the reactor contents.
Stirred tank bioreactors are generally constructed
to standard dimensions.
That is, they are constructed according to
recognised standards such as those published by
the International Standards Organisation and the
British Standards Institution.
These dimensions take into account both mixing
effectiveness and structural considerations.

Standard geometry of a stirred

tank bioreactor
A mechanically stirred tank bioreactor
fitted with
a sparger and
a rushton turbine
will typically have the following relative
dimensions:

Standard geometry of a stirred

tank bioreactor

3. Headspace volume
A bioreactor is divided in a working volume and a head-space volume.
The working volume is the fraction of the total volume taken up by the
medium, microbes, and gas bubbles.
The remaining volume is called the headspace.
Typically, the working volume will be 70-80% of the total fermenter
volume.
This value will however depend on the rate of foam formation during
the reactor. If the medium or the fermentation has a tendency to foam,
then a larger headspace and smaller working volume will need to be
used.

Headspace volume

4. Basic features of a stirred tank

bioreactor

A modern mechanically agitated bioreactor will contain:

Basic features
* An agitator system
* An oxygen delivery system
*A foam control system
* A temperature control system
* A pH control system
* Sampling ports
* A cleaning and sterilization system.
* A sump and dump line for emptying of the
reactor

4.1 Basic features of a stirred tank

bioreactor - Agitation system
The function of the agitation system is to
provide good mixing and thus increase mass transfer
rates through the bulk liquid and bubble boundary
layers.
provide the appropriate shear conditions required for
the breaking up of bubbles.
The agitation system consists of the agitator and the baffles.
The baffles are used to break the liquid flow to increase
turbulence and mixing efficiency.
The agitator consists of the components shown in the
following diagram:

Agitation system

The agitator consists of the components shown in the following

diagram:

Agitation system
The number of impellers will depend on the height
of the liquid in the reactor. Each impeller will have
between 2 and 6 blades. Most microbial
fermentations use a Rushton turbine impeller.
A single phase (ie. 240 V) agitator drive motor can
be used with small reactors. However for large
reactors, a 3 phase motor (ie 430 V) should be
used. The latter will tend to require less current and
therefore generate less heat.
Speed control or speed reduction devices are used
to control the agitation speed.

4.1.1 Basic features of a stirred tank bioreactor;

Agitation system - Top entry and bottom entry
impellers

The impeller shaft can enter from the bottom of the tank or from
the top. A top entry impeller ("overhung shaft") is more
expensive to install as the motor and the shaft will need to be
structurally supported:

Bottom entry impellers

A reactor with bottom entry impeller however will need higher maintenance due to damage
of the seal by particulates in the medium and by medium components that crystallize in the
seal when reactor is not in use:
Bottom entry agitators tend to require more maintenance than top entry impellers due to
the formation of crystals and other solids in the seals

4.1.2 Basic features of a STR

Agitation system - Mechanical seals
The mechanical seal is used prevent contaminants
from entering the reactor and to prevent organisms
from escaping through the shaft.
The seal uses vapours from the liquid for lubrication.
It is therefore important that you do not turn the shaft
when the tank is dry so as not to damage the seal.

5. Agitator design and

operation

Agitators are classified as having radial flow or axial flow

characteristics.
With radial flow mixing, the liquid flow from the impeller is initially
directed towards the wall of the reactor; ie. along the radius of the tank.
With axial flow mixing, the liquid flow from the impeller is directed
downwards towards the base of the reactor, ie. in the direction of the
axis of the tank.
Radial flow impellers are primarily used for gas-liquid contacting (such
as in the mixing of sparged bioreactors) and blending processes.
Axial flow impellers provide more gentle but efficient mixing and are
used for reactions involving shear sensitive cells and particles.

5.1. Agitator design and operation

- Radial flow impellers

Radial flow impellers contain two or more impeller blades

which are set at a vertical pitch:

Agitator design

The liquid flow from the blades is directed towards the walls of
the reactor; ie. along the radius of the tank.

Agitator design
Radial flow mixing is not as efficient as axial flow
mixing.
For radial flow impellers, a much higher input of
energy input is required to generate a given level
of flow.
Radial flow impellers do and are designed to,
generate high shear conditions. This is achieved by
the formation of vortices in the wake of the
impeller:

Agitator design

The high shear is effective at breaking up bubbles. For this

reason, radial flow impellers are used for the culture of
aerobic bacteria.
High shear can also damage shear sensitive materials such as
crystals and precipitates and shear sensitive cells such as
filamentous fungi and animal cells.

th radial flow impellers, vertical (or axial) mixing is achieved with the use of baffl

5.1.1 Agitator design and operation

Radial flow impellers - Rushton turbine
The most commonly used agitator in microbial
fermentations is the Rushton turbine.
Like all radial flow impellers, the Rushton turbine is
designed to provide the high shear conditions
required for breaking bubbles and thus increasing
the oxygen transfer rate.
The Rushton turbine has a 4 or 6 blades which are
fixed onto a disk.
The diameter of the Rushton turbine should be 1/3
of the tank diameter.

Radial flow impellers

A Rushton turbine is often referred to as a disk
turbine.
The disk design ensures that most of the motor
power is consumed at the tips of the agitator and
thus maximizing the energy used for bubble
shearing.

Radial flow impellers

5.2. Agitator design and

operation - Axial flow impellers
Axial flow impeller blades are pitched at an angle and thus direct
the liquid flow towards the base of the tank. Examples of axial
flow impellers are marine impellers and hydrofoil impellers.

Axial flow impellers

The resultant flow pattern is thus predominantly vertical; ie.

along the tank axis.

Axial flow impellers

Axial flow mixing is considerably more energy
efficient than radial flow mixing.
They are also more effective at lifting solids from
the base of the tank.
Axial flow impellers have low shear properties.
The angled pitch of the agitators coupled with
the thin trailing edges of the impeller blades
reduces formation of eddies in the wake of the
moving blades.

Axial flow impellers

Low shear conditions are achieved by pitching the

impeller blades at an angle and by making the edges
of the impeller blades thin and smooth.

Axial flow impellers

Axial flow impellers are used for mixing shear
sensitive processes such as crystallization and
precipitation reactions.
They are also used widely in the culture of animal
cells.
Their low shear characteristics generally makes
them ineffective at breaking up bubbles and thus
unsuitable for use in aeration of bacterial
fermentations

classification
Disc turbine
Vaned disc
Variable pitch open turbine
Marine propellers

Disc turbine
A disc with series of
rectangular vanes set in a
vertical plane around the
circumference.
Break up a fast air stream
without itself becoming
flooded in air bubbles

Vaned disc
A series of rectangular vanes
attached vertically to the
underside
Air from sparger hits its
underside & the air gets
displaced towards the vanes
Results in destruction of air

Varialble pitch open turbine

Vanes are attached directly to a
boss on the agitator shaft
Air bubbles hit any surface by
its action
Flood when superfial velocity
exceed 21m/h

Marine propeller
Blades are attached directly
to a boss on the agitator shaft
Air bubbles hit surface
A single low shear impeller
Mainly used in animla cell
culture vessel
Flood when superfial velocity
exceed 21m/h

Modern agitators
Rushton disc turbine
Scaba 6SRGT
Prochem maxflow T
Lightening A315
Ekato intermig

Modern Agitator
Develoments
Four other modern agitator
developments, the Scaba 6SRGT, the
Prochem Maxflo T, the Lightning
A315 and the Ekato Intermig are
derived from open turbines

65

67

 The Scaba 6SRGT agitator is one

which, at a given power input, can
handle a high air flow rate before
flooding.
This radial-flow agitator is also better
for bulk blending than a Rushton
turbine, but does not give good top
to bottom blending in a large
fermenter which leads to lower
concentrations of oxygen in broth
away from the agitators and higher
concentrations of nutrients, acid or

68

 Another is the Prochem Maxflo agitator. It

of four, five or six hydrofoil blades set at a
critical on a central hollow hub.
A high hydrodynamic thrust is created
during rotation, increasing the downwards
pumping capacity of the blades.
Good mixing and aeration in high viscosity
broths may also be achieved by a dual
impeller combination, where the lower
impeller acts as the gas disperser and the
upper impeller acts primarily as a device
for aiding circulation of vessel contents.
69

5.3. Agitator design and operation

Axial flow impellers - Intermig Impeller

Intermig impeller is a axial flow which is used for microbial

fermentations.

The impeller is shown below:

Intermig Impeller
The agitation system has two impellers. The bottom
impeller has a large axial flow section. The tips of the
impeller contain finger like extensions which create a
turbulent wake for breaking bubbles.
As the high shear region exists only at the tip, the
overall shear conditions in the reactor are lower than
would be generated by a radial flow impeller such as a
Rushton Turbine.
Intermig impellers are used widely for agitation and
aeration in fungal fermentations.

TYPES OF IMPELLERS

TYPES OF IMPELLERS

4.2 Basic features of a stirred tank

bioreactor - Oxygen delivery system.
The oxygen delivery system consists of
a compressor
inlet air sterilization system
an air sparger
exit air sterilization system

4.2.1 Basic features of a stirred tank bioreactor;

Oxygen delivery system - Compressor

Oxygen delivery system Compressor

A compressor forces the air into the reactor. The
compressor will need to generate sufficient pressure
to force the air through the filter, sparger holes and
into the liquid.
Air compressors used for large scale bioreactors
typically produce air at 250 kPa. The air should be
dry and oil free so as to not block the inlet air filter
or contaminate the medium.

4.2.2 Basic features of a stirred tank bioreactor;

Oxygen delivery system - Air sterilization system
Sterilization of the inlet air is undertaken to prevent
contaminating organisms from entering the reactor.
The exit air on the other hand is sterilized not only to
keep contaminants from entering but also to prevent
organisms in the reactor from contaminating the air.
A common method of sterilising the inlet and exit air
is filtration. For small reactors (with volumes less than
5 litres), disk shaped hydrophobic Teflon membranes
housed in a polypropylene housing is used. Teflon is
tough, reusable and does not readily block.

Sterilisation of the air

For larger laboratory scale fermenters (up to 1000 litres), pleated

membrane filters housed in polypropylene cartridges are used.

Sterilisation of the air

By pleating the membrane, it is possible to create a compact filter
with a very large surface area for air filtration. Increasing the filtration
area decreases the pressure required to pass a given volume of air
through the filter.
Sterilization of the inlet and exit air in large bioreactors (> 10,000
litres) can present a major design problem. Large scale membrane
filtration is a very expensive process. The filters are expensive as
they are difficult to make and the energy required to pass air through
a filter can be quite considerable.
Heat sterilization is alternative option. Steam can be used to sterilize
the air. With older style compressors, it was possible to use the heat
generated by the air compression process to sterilize the air.

Condenser
In small reactors, the exit air system will typically include a
condenser.

Condenser
The condenser is a simple heat exchanger
through which cool water is passed.
Volatile materials and water vapour
condense on the inner condenser surface.
This minimizes water evaporation and the
loss of volatiles.
Drying the air also prevents blocking of
the exit air filter with water

4.2.3 Basic features of a STR

Oxygen delivery system
Air sterilisation system - Positive pressure

During sterilisation the concept of "maintaining positive

pressure" will often be used.
Maintaining positive pressure means that during sterilisation,
cooling and filling and if appropriate, the fermentation process,
air must be pumped into the reactor.
In this way the reactor is always pressurised and thus aerial
contaminants will not be "sucked" into the reactor.
It is very important that positive pressure is maintained when
the bioreactor is cooled following sterilisation. Without air being
continuously pumped into the reactor, a vacuum will form and
contaminants will tend to be drawn into the reactor.

Air sterilisation system Positive pressure

Maintaining positive pressure at all stages of the fermentation

setup and operation is an important aspect of reducing the risk of
contamination

Without aeration, a
vacuum
forms as the reactor
cools.

With aeration, positive

pressure is always
maintained and
contaminants are pushed

Aeration system
Syn : sparger
A device that introduce air into medium
Has a pipe with minute holes (1/64 - 1/32 inch
or large)
Hole allows air under P to escape into medium
For mycelial growth inch holes
Impeller blades disperses air released through
sparger into medium

Sparger types

Porous
Orifice
Nozzle

4.2.4 Basic features of a stirred tank

bioreactor Oxygen delivery system - Sparger
The air sparger is used to break the incoming air
into small bubbles.
Although various designs can be used such as
porous materials made of glass or metal, the most
common type of filter used in modern bioreactors is
the sparge ring:

Porous sparger

Made of sintered glass, ceramics

or metal
Used mainly on a large scale
fermentors
Bubble size produced 10-100
times larger than pores
Throughput of air is low P drop
across it

Porous spargers

89

Oxygen delivery system - Sparger

A sparge ring consists of a hollow tube in which
small holes have been drilled. A sparge ring is
easier to clean than porous materials and is less
likely to block during a fermentation.
The sparge ring must located below the agitator
and will have approximately diameter as the
impeller.
Thus, the bubbles rise directly into the impeller
blades, facilitating bubble break up.

Orifice sparger
Those with drilled air holes on
their under surface of the tubes
making up ring or cross (6 mm)
Without agitation used to a
limited extend in yeast
manufacture & effluent
treatment

Oxygen delivery system - Sparger

During the emptying of a fermenter, it is important that the air feed valve
is closed. This will minimize the contamination of the inlet air line

4.2.5 Basic features of a STR

Oxygen delivery system - Effect of impeller speed

As discussed in another lecture, the shear forces that an

impeller generates play a major role in determining bubble size.
If the impeller speed is to slow then the bubbles will not be
broken down. In addition, if the impeller speed is too slow, then
the bubbles will tend to rise directly to the surface due to their
buoyancy.
Fast impeller speed
Slow impeller speed

The bubbles will not be

sheared into smaller
bubbles and will tend to rise
directly towards the surface

Smaller bubbles will be generated

and these bubbles will move with
throughout the reactor increasing
the gas hold up and bubble

Oxygen delivery system - Effect of impeller

speed
Another consequence of too slow an impeller speed is a
flooded impeller.
Under these conditions, the bubbles will accumulate and
coalesce under the impeller, leading to the formation of
large bubbles and poor oxygen transfer rates.
A similar phenomenon will happen when aeration rate is
too high.
In this case, the oxygen transfer efficiency will be low

FERMENTORS STRUCTURAL COMPONENTS

IN AERATION & AGITATION SYSTEM

The agitator
Stirrer glands & bearings
Baffles
The aeration system

Types of Seals

96

97

 With glass and metal, a seal can be made with

a compressible gasket, a lip seal or an '0' ring.
With metal to metal joints only an '0' ring is
suitable.
Nitryl or butyl rubbers are normally used for
these seals as they will withstand fermentation
process conditions.
A single '0' ring seal is adequate for GILSP and
levels 1 and B2, a double '0' ring seal is
required for levels 2 and B3 and a double '0'
ring seal with steam between the seals (steam
tracing) is necessary for levels 3 and B4

98

Stirrer Glands and Bearings

The satisfactory sealing of the stirrer
shaft assembly top plate has been one
of the most difficult problems to
overcome in the construction of
fermentation equipment which can be
operated aseptically for long periods.
The stirrer shaft can enter the vessel
from the top, side or bottom of the
vessel.
99

A simple Stirrer Seal

100

 A porous bronze bearing for a 13-mm shaft

was fitted in the centre of the fermenter
top and another in a yoke directly above it.
The bearings were pressed into steel
housings, which screwed into position in
the yoke and the fermenter top.
The lower bearing and housing were
covered with a skirt-like shield having a 6.5
mm overhang which rotated with the shaft
and prevented air-borne contaminants from
settling on the bearing and working their
way through it into the fermenter.
101

Four Main Types

102

The Stuffing Box (Packed Gland

Seal)
The shaft is sealed by several layers
of packing rings of asbestos or cotton
yarn, pressed against the shaft by a
gland follower.
Chain et al. (1954) used two stuffing
boxes on the agitator shaft with a
space in between kept filled with
steam.
These seals are sufficient for the
requirements of GILSP containment.

103

104

The Mechanical Seal

The seal is composed of two parts, one
part is stationary in the bearing housing,
the other rotates on the shaft, and the two
components are pressed together by
springs or expanding bellows.
The two meeting surfaces have to be
precision machined, the moving surface
normally consists of a carbon-faced unit
while the stationary unit is of stellite-faced
stainless steel.
105

106

Magnetic Drive
The problems of providing a
satisfactory seal when the impeller
shaft passes through the top or
bottom plate of the fermenter may be
solved by the use of a magnetic drive
in which the impeller shaft does not
pierce the vessel.
A magnetic drive consists of two
magnets: one driving and one driven.
107

 The driving magnet is held in

bearings in a housing on the outside
of the head plate and connected to a
drive shaft.
The internal driven magnet is placed
on one end of the impeller shaft and
held in bearings in a suitable housing
on the inner surface of the
headplate.

108

Aeration System (Spargers)

A sparger may be defined as a device
for introducing air into the liquid in a
fermenter.
Three basic types of sparger have
been used and may be described as
the Porous sparger, the Orifice
sparger (a perforated pipe) and the
Nozzle sparger (an open or partially
closed pipe).
109

Porous Sparger
The porous sparger of sintered glass,
ceramics or metal, has been used primarily
on a laboratory scale in non-agitated
vessels.
The bubble size produced from such
spargers is always 10 to 100 times larger
than the pore size of the aerator block.
There is also the problem of the fine holes
becoming blocked by growth of the
microbial culture.
110

Orifice Sparger
In small stirred fermenters the
perforated pipes were arranged below
the impeller in the form of crosses or
rings (ring sparger), approximately
three-quarters of the impeller
diameter.
In most designs the air holes were
drilled on the under surfaces of the
tubes making up the ring or cross.
111

 Sparger holes should be at least 6

mm (1/4 inch) diameter because of
the tendency of smaller holes to
block and to minimize the pressure
drop.

112

Nozzle Sparger
Single open or partially closed pipe
as a sparger to provide the stream of
air bubbles
Ideally the pipe should be positioned
centrally below the impeller and as
far away as possible from it to ensure
that the impeller is not flooded by
the air stream.
113

Valves and Steam Traps

Valves attached to fermenters and
ancillary equipment are used for
controlling the flow of liquids and
gases in a variety of ways.
There are four main types of valves,

114

 Simple ON/OFF valves which are

either fully open or fully closed.
Valves which provide coarse control
of flow rates.
Valves which may be adjusted very
precisely so that flow rates may be
accurately controlled.
Safety valves which are constructed
in such a way that liquids or gases
will flow in only one direction.
115

Gate Valves
In this valve, a sliding disc is moved in
or out of the flow path by turning the
stem of the valve.
It is suitable for general purposes on a
steam or a waterline for use when
fully open or fully closed and therefore
should not be used for regulating flow.
Not suitable for aseptic conditions
116

 there may be leakage round the

stem of the valve which is sealed by
a simple stuffing box.
This means that the nut around the
stem and the packing must be
checked regularly.

117

118

Globe Valves
In this valve, a horizontal disc or plug
is raised or lowered in its seating to
control the rate of flow.
It is not suitable for aseptic operation
because of potential leakage round
the valve stem which is similar in
design to that of the gate valve.
There is a high pressure drop across
the valve because of the flow path.
119

120

Piston Valves
The piston valve is similar to a globe
valve except that flow is controlled
by a piston passing between two
packing rings.
This design has proved in practice to
be very efficient under aseptic
operation.
There may be blockage problems
with mycelial culture and the
pressure drop is similar to a globe

121

122

Needle Valve
The needle valve is similar to the globe
valve, except that the disc is replaced by a
tapered plug or needle fitting into a
tapered valve seat.
The valve body can be used to give fine
control of steam or liquid flow.
Accurate control of flow is possible
because of the variable orifice formed
between the tapered plug and the tapered
seat.
123

124

Plug Valves
In this valve there is a parallel or tapered
plug sitting in a housing through which
an orifice, A, has been machined.
When the plug is turned through 90 the
valve is fully open and the flow path is
determined by the cross-sectional area of
A, which may not be as large as that of
the pipeline.

125

126

Ball Valve
This valve has been developed from the plug
valve.
The valve element is a stainless-steel ball
through which an orifice is machined.
The ball is sealed between two wiping
surfaces which wipe the surface and prevent
deposition of matter at this point.
The valve is suitable for aseptic operation, can
handle mycelial broths and can be operated
under high temperatures and pressures.
127

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Butterfly Valve
The butterfly valve consists of a disc
which rotates about a shaft in a housing.
The disc closes against a seal to stop the
flow of liquid.
This type of valve is normally used in
large diameter pipes operating under low
pressure where absolute closure is not
essential. It is not suitable for aseptic
operation.
129

130

Pinch Valve
In the pinch valve a flexible sleeve is closed
by a pair of pinch bars or some other
mechanism which can be operated by
compressed air remotely or automatically.
The valve is suitable for aseptic operation
with fermentation broths, even when
mycelial, as there are no dead spaces in
the valve structure, and the closing
mechanism is isolated from the contents of
the piping.
131

132

Diaphragm Valve
Like the pinch valve, the diaphragm
valve makes use of a flexible closure,
with or without a weir.
Suitable for aseptic operation.

133

Most Suitable Valve

Among these group of valves which
have just been described, globe and
butterfly valves are most commonly
used for ON/OFF applications, gate
valves for crude flow control, needle
valves for accurate flow control and
ball, pinch or diaphragm valves for
all sterile uses.
134

Check Valves
The purpose of the check valve is to prevent
accidental reversal of flow of liquid or gas in a pipe
due to breakdown in some part of the equipment.
There are three basic types of valve: swing check,
lift check and combined stop and check with a
number of variants.
The swing check valve is most commonly used in
fermenter designs.
The functional part is a hinged disc which closes
against a seat ring when the intended direction of
flow is accidentally reversed.
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4.3 Basic features of a STR

- foam control system

Foam control is an essential element of the operation of a

sparged bioreactor. The following photograph shows the
accumulation of foam in a 2 litre laboratory reactor.

Foam control system

Excessive foam formation can lead to blocked air exit filters
and to pressure build up in the reactor.
The latter can lead to a loss of medium, damage to the
reactor and even injury to operating personnel.
Foam is typically controlled with aid of antifoaming agents
based on silicone or on vegetable oils.
Excessive antifoam addition can however result in poor
oxygen transfer rates.

 The antifoam requirement will depend on

the nature of the medium.
Media rich in proteins will tend to foam more readily than simple media.
the products produced by the fermentation.
Secreted proteins or nucleic acids released as a result of cell death and
hydrolysis have detergent like properties.
the aeration rate and stirrer speed.
Increasing the aeration rate and stirrer speed increases foaming problems.
control
the use of mechanical foam control devices
Foam control devices such as mechanical and ultrasonic foam breakers help
to reduce the antifoam requirement.
The head space volume
The larger headspace volume, then the greater the tendency for the foam to
collapse under its own weight. For example, for fermentations in which high
levels of foam is produced, a 50% headspace volume may be required.
Condenser temperature
In laboratory scale reactors, a cold condenser temperature can help to
control the foam. The density of the foam increases when it moves from the
warm headspace volume to the cold condenser region. This causes the foam
to collapse.

Foam is typically detected using two

conductivity or "level" probes.

When the upper level probe is above the

foam level, no current will pass between
the level probes and the antifoam pump
remains turned off.

When the upper level probe is

immersed in the foam layer, a
current is carried in the foam. This
causes the antifoam to turn on.

Foam control system

One probe is immersed in the fermentation liquid
while the other placed above the liquid level.
When the foam reaches the upper upper probe, a
current is carried through the foam.
The detection of a current by the foam controller
results in the activation of a pump and the
antifoam is then added until the foam subsides.

4.4 Basic features of a stirred tank

bioreactor - Temperature control system
The temperature control system consists of
temperature probes
heat transfer system
Typically the heat transfer system will use a
"jacket" to transfer heat in or out of the reactor.
The jacket is a shell which surrounds part of the
reactor. The liquid in the jacket does not come in
direct contact with the fermentation fluid.

Temperature control system

The jacket will typically be "dimpled" to encourage turbulence in

the jacket and thus increase the heat transfer efficiency.
An alternative to using jackets are coils. Coils have a much
higher heat transfer efficiency than jackets. However coils take
up valuable reactor volume and can be difficult to clean and
sterilize.

Temperature control system

The heating/cooling requirements are provided by the following

methods:
Laboratory scale reactors
reactors
Heating
Electric heaters
requirements

Pilot and production scale

Steam generated in boilers

Cooling
Tap water or
Cooling water produced by
requirements refrigerated water baths
cooling towers or refrigerants
such as ammonia.

Temperature control system

In pilot and production scale reactors, heating is
typically only required during the initial stages and
final stages of the fermentation as most processes
which occur during a fermentation process,
including
the biological reactions (eg. growth)
chemical reactions
mixing
are exothermic.
Temperature is measured by : Mercury in glass thermometers,
bimetallic thermometers, pressure bulb thermometers,
thermocouples, metal resistence thermometers.

Type
Principle
Thermistors
Type of resistor measuring the
change in resistance. The output response from thermistors
is of a nonlinear
temperature versus resistance curve, with the resistance
decreasing as the temperature increases. Semiconductors
made of specific mixture of pure oxides of iron, nickel.

Resistance thermometer
Based on the changes in the
electrical resistance of metallic conductors (mostly platinum),
with changing temperature . A platinum wire of 100-resistance
is typically usedplatinum sensors are stable under both
sterilization and fermentation conditions.

Bimetallic thermometers Consist of a bimetallic helical

coil surrounded by a protective tube or wall. The coil winds or
unwinds with changes in temperature and causes movement
of a fixedpointer.

Metal resistant
thermometers

147

Bimetallic thermometers

148

4.5 Basic features of a stirred tank

bioreactor - pH control system

The pH control system

consists of
a pH probe
alkali delivery
system
acid delivery
The pH probe is typically steam sterilizable and glass
electrodes
system

(Silver electrodes, mercury electrodes.) For good insulation, both the

glass and reference electrodes were mounted in Teflon jackets and

4.5.1 Basic features of a stirred tank bioreactor

pH control system - Neutralizing agents
The neutralizing agents used to control pH should be noncorrosive. They should also be non-toxic to cells when diluted in
the medium.
Potassium hydroxide is preferred to NaOH, as potassium ions
tend to be less toxic to cells than sodium ions. However KOH is
more expensive than NaOH. Sodium carbonate is also commonly
used in small scale bioreactor systems.
Hydrochloric acid should never be used as it is corrosive even to
stainless steel.
Likewise sulphuric acid concentrations should not be between
10% and 80% as between this range, sulphuric acid is most
corrosive.

4.5.2. Basic features of a stirred tank bioreactor

pH control system - Setpoint and deadband

Setpoint and deadband

The pH control system (and indeed all other fermenter control

systems) are designed to have a deadband. A deadband is
used to prevent excessive alkali and acid addition.
The pH control deadband is shown in the following diagram:

Setpoint and deadband

The setpoint is the pH at which the fermenter is being attempted to be

controlled at. For example, if the fermentation is to be run at a constant
pH of 6.5, then the setpoint is set to 6.50.
If for example, a 5% deadband is used, then the upper deadband limit
will be
1.05 x 6.5 = 6.83
and the lower deadband limit will be
0.95 x 6.5 = 6.18
If the deadband is too small, then it is possible that pH will often
overshoot and undershoot the deadbands leading to excessive alkali and
acid addition. The trade off is that a wide deadband will lead to less
precise pH control.
As many fermentations tend to produce acids rather than substances
that increase the pH, acid addition is often not required. Indeed not all
fermentations need continuous pH control.

Pressure measurement:
Industrial and lab equipments are designed to withstand a
specified working pressure plus a factor of safety.
In a fermenter, pressure will influence the solubility of gases
and contribute to the maintenance of sterility when positive
pressure is present.
Pressure measuring sensors are: Bourdon tube pressure
gauge, Nested diaphragm type pressure sensor, piezoelectric
transducer.
The correct pressure is maintained by regulatory valves and
safety valves.
Gas/Liquid Flow rate:
Flow measurement and control of gases and liquid is
important in process management.
Rotameter, thermal mass flowmeters
Indirect measurement is by load cells and pumps
Agitator Shaft power:
sensors used to measure power consumption of a fermenter.
Watt meter, Torison dynamometer and strain gauge attached to
the agitator motor.

155

156

157

Aload cellis a transducer that is used to create an

electrical signal whose magnitude is directly proportional to
the force being measured. The various types ofload
cellsinclude hydraulicload cells, pneumatic load
cellsand strain gaugeload cells.

158

Rate of stirring:
to monitor rpm of the strirrer shaft.
Tachometers are used.
Microbial biomass:
Indirect methods dry weight , cell density
(spectrophotometer), cell number (Coulter counters)
Direct methods ( real time fuloremetric measurement of NAD
concentration).
Ion-specific Sensors
They developed to measure NH4+ Ca2+, K+, Mg2+, PO3-,
S02-, etc.
Response time of these electrodes ranges from 10 seconds to
several minutes depending on the concentration of the ion
species, the composition of the sample.
However, none of these probes is steam sterilizable.

160

Type
Principle
DO Probes
Based on galvanic or
polarographic principle. Measurement of partial oxygen
pressure.

Optodes (DO)
Optical sensors for oxygen
are constructed using an immobilized fluorophore that
undergoes dynamic quenching of the luminescence of a
ruthenium complex. The change in fluorescence signal
is measured by optical measurement device.

Exit gas analysis

There are a number of
methods available for determining the oxygen
concentration of exit gas from a bioreactor. Several of
these are based on exploiting the strong affinity shown
by oxygen to a magnetic field. Paramagnetic gases,
such as oxygen,
display a positive magnetic susceptibility.

Summary
Aware of standard geometry of a stirred tank
bioreactor
Know the basic features of a stirred tank
bioreactor
Understand working of the agitation system
Agitator design and operation
Components of the oxygen delivery system
Foam control
Temperature control system
pH control system
Cleaning and sterilization facilities

FERMENTORS STRUCTURAL COMPONENTS

IN AERATION & AGITATION SYSTEM

The agitator
Stirrer glands & bearings
Baffles
The aeration system

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