Fermentation technology

1. Fermentation technology
Presented by: Shashikala Metri

2. Fermentation
 Fermentation is the process of growing
microorganisms in a nutrient media by
maintaining physico- chemical
conditions and thereby converting feed
into a desired end product
 Fermentation technology is the use of
organisms to produce food,
pharmaceuticals and alcoholic
beverages on a large scale industrial
basis.

3. Fermentation
 The basic principle involved in the industrial
fermentation technology is that organisms are
grown under suitable conditions, by providing
raw materials meeting all the necessary
requirements such as carbon, nitrogen, salts,
trace elements and vitamins.
 The end products formed as a result of their
metabolism during their life span are released
into the media, which are extracted for use by
human being and that have a high commercial
value.

4. Major fermentation products
Group Product Organism
Industrial
chemicals
Ethanol
Lactic acid
Saccharomyces cerevisiae
Lactobacillus bulgaricus
Enzymes -amylase
Proteases
Lipases
Bacillus subtilis
Bacillus species
Saccharomyces lipolytica
Antibiotics Penicillin
Streptomycin
Chlorampenicol
Penicillium chrysogenum
Streptomyces griseus
Streptomyces venezuelae
Vitamins Riboflavin
Vitamin B12
Ashbya gossypi
Pseudomonas dentrificians

5. DESIGN OF FERMENTER
 A fermentation process requires a fermenter for successful production
.
 Fermentor is the large vessel containing considerable quantities of
nutrient media by maintaining favourable conditions.
 The design and nature of the fermentor varies depending upon the
type of fermentation carried out. Invariably all the fermentors provide
the following facilities for the process such as
 contamination free environment,
 specific temperature maintenance,
 maintenance of agitation and aeration, pH control,
 monitoring Dissolved Oxygen (DO),
 ports for nutrient and reagent feeding (antifoam agents, alkali or
acid),
 ports for inoculation and sampling,
 provide all aseptic conditions at the time of sample withdrawal
and addition of innoculum
 complete removal of broth from the tank and should be easy to
clean
 It should be designed in such away that it consumes less power,
have less evaporation, can be used for long periods of operation

6. FigI. An Ideal fermenter
DESIGN OF FERMENTER

7. Components of fermenter
 1. Basic component includes drive
motor, heaters, pump, etc.,
 2. Vessels and accessories
 3. Peripheral equipment (reagent
bottles)
 4. Instrumentation and sensor

8. Various components of an ideal
fermenter for batch process are:

9. Monitoring and controlling parts of
fermenter are:

10. Types of fermenter
Available in various sizes
According to the sizes classified as
 Small lab and research fermenter :1-50L
 Pilot plant fermenter: 50-1000 L
 Large size industrial production scale fermenter: more than 1000 L
 Broadly fermentes are also claified as
I. surface fermenters
 Tray fermenter
 Packed bed column fermenter
II. Submerged fermenters
 Simple fermenters (batch and continuous)
 Fed batch fermenter
 Air-lift
 Bubble fermenter
 Cyclone column fermenter
 Tower fermenter
 Other more advanced systems, etc

11. Types of fermenter
Surface fermenters
 Microbial cells cultured on surface layer of
the nutrient medium (solid/liquid) held in dish
or tray
 Used for production of citric acid from
Aspergillus niger and nicotinic acid from
Aspergillus terrus
 Microbial films can be developed on the
surfaces of suitable packing medium, may be
in the form of fixed bed, stones or plastic
sheets.

12. Tray fermenter
 TRAY FERMENTER
 one of the simplest and widely used fermenters.
 Its basic part is a wooden, metal, or plastic tray, often with
a perforated or wire mesh bottom to improve air circulation.
 A shallow layer of less than 0.15 m deep, pretreated
substrate is placed on the tray for fermentation.
 Temperature and humidity-controlled chambers are used
for keeping the individual trays or stacks.
 A spacing of at least one tray height is usually allowed
between stacked trays.
 Cheesecloth may be used to cover the trays to reduce
contamination.
 Inoculation and occasional mixing are done manually,
often by hand.

13. Tray fermenter
•Solid as well as liquid
medium are used
•If liquid medium, cells are
allowed to float easily and to
make a process continuous
•If solid medium is used the
micro-organisms are
allowed grow on moist solid
materials, process is called
Solid State Fermentation

14. Solid State Fermentation (SSF)
Solid State Fermentation Method (SSF)
 SSF defined as the growth of the micro-organisms on
(moist) solid material in the absence or near-absence
of free water
 Used for production of antibiotics, enzymes, alkaloids,
organic acids bio-pharmaceutical products
Advantages :
• Produce higher yields than submerged liquid
fermentation
• Possibilities of contamination by bacteria and yeast
is very less
• All natural habitats of fungi are easily maintained in
SSF
• culture media very simple , provides all nutrients for
growth of micro-organisms

15. SSF
Disadvantages:
•Causes problems in monitoring of the process parameters such
as pH, moisture content, and oxygen concentration
•Despite some automation, tray fermenters are
labor intensive
•Difficulties with processing hundreds of trays limit their
scalability
•Aeration may be difficult due to high level of solid content
•Substrates require pre treatment such as size reduction,
chemical or enzymatic hydrolyses

16. Packed bed fermenters
 This is type of surface culture bioreactor
 A bed of solid particles, with biocatalysts on
or within the matrix of solids, packed in a
column
 The solids used may be porous or non-
porous gels, and they may be compressible
or rigid in nature.
 A nutrient broth flows continuously over the
immobilised biocatalyst. The products
obtained in the packed bed bioreactor are
released into the fluid and removed.

17.  The concentration of
the nutrients can be
increased by
increasing the flow
rate of the nutrient
broth.
 Because of poor
mixing, difficult to
control the pH of
packed bed
bioreactors by the
addition of acid or
alkali.
Packed bed fermenter

18. Submerged fermenters
The microorganisms are dispersed in liquid
nutrient medium at maintained environmental
conditions.
on the mechanism of agitation Submerged
fermenters grouped as follows:
I. Mechanically stirred fermenter
○ batch operate fermenter
○ continuous stirred tank fermenter
II. Forced convection fermenters
○ Air –lift fermenter
○ Bubble column
○ Sparged tank fermenter
III. Pneumatic fermenter
○ Fluidized bed reactor

19.  These are equipped with a
mechanical agitator so as to maintain
homogencity and rapid dispersion
and mixing of materials
 Examples includes stirred tank
fermenter (batch or continuous
operated) , multistage fermenter,
paddle wheel reactor, and stirred loop
reactor
Mechanically stirred fermenter

20. Stirred tank fermenter (STF)
stirred tank fermenter
 batch operated
fermenter
 agitators consists of one
or more impellers
mounted on the shaft
 It is rotates with the help
of electric motor
 Advantage of this
fermenter flexibility in
design
 Used in the range of 1-
100 ton capacity sizes Stirred tank fermenter

21.  A continuous stirred
tank fermenter consists
of a cylindrical vessel
with motor driven
central shaft that
supports one or more
agitators (impellers).
 The shaft is fitted at
the top of the
bioreactor (ref. fig.).
The number of
impellers is variable
and depends on the
size of the fermenter
Continuous stirred tank fermenter
(CSTF)
Continuous stirred tank
fermenter

22. Continuous stirred tank
fermenter
 In this fresh medium is added continuously in
the fermenter vessel
 On the other end the medium is withdrawn for
the recovery of fermentation products
 As it is a continuous fermenter the Steady state
conditions can be achieved by
either Chemostatic or Turbidostatic principles.

23. Continuous stirred tank
fermenter(CSTF)
 Different types of continuous fermenter
are
a. Single stage: single fermenter is
inoculated and kept in continuous
operation by balancing the input and
output culture media
b. Recycle continuous fermentation: a
portion of the withdrawn culture or
residual unused substrate plus the
withdrawn culture is recycled

24. CSTF
c. Multistage
continuous
operation: involves
two
or more stages
with
the fermenter
being operated
in sequence
multistage

25. STF
Advantages of batch operated
 Less risk of contamination because of short
growth period
 Process is more economical and simple
 Raw material conversion level is high
Disadvantages:
 Low productivity due to time required fro
the sterilizing, filling, cooling, emptying and
cleaning
 More expenses are required for
subcultures for inoculation, labor and
process control

26. STF
Advantages of continuous operated
 Less labor expenses due to automation of
fermentation process
 Less toxicity risk to operator by toxins producing
microorganisms
 High yield and good quality product due invariable
operating parameters and automation of the
process
 Less stress on the fermenter as sterilization is not
frequent
Disadvantages:
 Higher investment costs in control and automation
equipment
 More risk of contamination and cell mutation

27. Bubble column fermenters
 In the bubble column
bioreactor, the air or gas is
introduced at the base of the
column through perforated
pipes or plates, or metal
micro porous spargers (ref
fig).
 The flow rate of the air/gas
influences the performance
factors —O2 transfer, mixing.
 May be fitted with perforated
plates to improve
performance. The vessel
used for bubble column
bioreactors is usually
cylindrical with an aspect ratio
of 4-6 (i.e., height to diameter
ratio). ..
Bubble column
fermenter

28. Air lift fermenter
 Airlift fermenter (ALF) is generally
classified as forced convection
fermenters without any mechanical
stirring arrangements for mixing.
 The turbulence caused by the fluid
(air/gas) flow ensures adequate
mixing of the liquid. The baffle or
draft tube is provided in the reactor.
 A baffle or draft tube divides the
fluid volume of the vessel into 2
inter-connected zones.
 Only one of the 2 zones is sparged
with air or other gas.
 The sparged zone is known as "
riser", the zone that receives no gas
is "downcomer“.
Air lift fermenter

29. Air lift fermenter
 Mainly 2 types
 Internal-loop airlift bioreactor
(ref Fig) has a single
container with a central draft
tube that creates interior
liquid circulation channels.
These bioreactors are simple
in design, with volume and
circulation at a fixed rate for
fermentation.
 External loop airlift
bioreactor (ref fig)
possesses an external loop
so that the liquid circulates
through separate
independent channels.
These reactors can be
suitably modified to suit the
requirements of different
fermentations.
Internal loop External loop

30. Air lift fermenter
Advantages
 The airlift bioreactors are more efficient than
bubble columns, particularly for more denser
suspensions of microorganisms as the mixing of
the contents is better compared to bubble
columns.
 Commonly employed for aerobic bioprocessing
technology.
 They ensure a controlled liquid flow in a recycle
system by pumping.
 Due to high efficiency, airlift bioreactors are
sometimes preferred e.g., methanol production,
waste water treatment, single-cell protein
production

31.  There are three different process of
fermentation viz.:
 (1) Batch fermentation
 (2) Feb-batch fermentation and
 (3) Continuous culture.
Batch fermentation:
 Nutrients are added in the fermentation for
the single time only and growth continues
until the particular nutrients are exhausted

32.  In the batch process when the microorganism is
added into a medium which supports its growth,
the culture passes through number of stages
known as ‘growth curve’
A typical growth curve consists of following stages
a) Lag phase
b) Acceleration phase
c) Log or exponential phase
d) Deceleration phase
e) Stationary phase
f) Death phase

33.  (a) Lag phase:
 Immediately after inoculation, there is no increase
in the numbers of the microbial cells for some time
and this period is called lag phase. In this is phase
the organisms adjust to the new environment in
which it is inoculated into.
 (b) Acceleration phase:
 The period when the cells just start increasing in
numbers is known as acceleration phase.
 (c) Log phase:
 This is the time period when the cell numbers
steadily increase.
 (d) Deceleration phase:
 The duration when the steady growth declines.

34.  (e) Stationary phase:
 The period where there is no change in the
microbial cell number is the stationary phase. This
phase is attained due to depletion of carbon source
or accumulation of the end products.
 (f) Death phase:
 The period in which the cell numbers decrease
steadily is the death phase. This is due to death of
the cells because of cessation of metabolic activity
and depletion of energy resources.
 Depending upon the product required the different
phases of the cell growth are maintained. For
microbial mass the log phase is preferred. For
production of secondary metabolites i.e. antibiotics,
the stationary phase is preferred.

35. Growth kinetics of batch
culture
The number of living cells (population of
growth rate dN/dt)varies with time in a batch
system as shown below:

36. where;
LAG Phase:
Number of bacteria does not change with time in lag phase.
LOG Phase:
Number of bacteria increases exponentially in log phase.

37. During log phase the number of
organisms in the reactor at any
time t can be calculated, by
using rate equation shown
below:

38. According to last equation, number of bacteria in the
reactor at any time t during log phase can be calculated,
as it is seen in the graph.
This rate equation can be integrated:

39. STATIONARY Phase:
There is no net change in number of bacteria with time
in stationary phase. Bacteria divide but also die at
equal rate. Most of the important biological products
(especially secondary metabolites like antibiotics) or
biomass are produced during this phase.
The biomass concentration at stationary phase is
determined by following equation
X = Y. SR
X=cell concentration
Y= yield factor for limiting nutrient
SR = original nutrient concentration in the medium

40.  Y’ measures the efficiency of a cell in
converting nutrients into biomass
 So the biomass at a particular time in the
during the fermentation is given by the
following equation.
X = Y (SR - s)
S= nutrient concentration at particular time
thus ‘Y’ is represented by the following equation
Y = X/ (SR - s)

41.  Feb-batch fermentation:
 In this type of fermentation, freshly prepared
culture media is added at regular intervals
without removing the culture fluid. This
increases the volume of the fermentation
culture. This type of fermentation is used for
production of proteins from recombinant
microorganisms.
 The total amount of the biomass in the
vessel increases but biomass concentration
is maintained constant

42. Continuous operations
Continuous fermentation:
 The growth rate and physiological conditions of
microorganisms can be maintained by using a
process of continuous culture (chemostat )
 In this the products are removed continuously
along with the cells and the same is
replenished with the cell girth and addition of
fresh culture media. This results in a steady or
constant volume of the contents of the
fermenter. This type of fermentation is used for
the production of single cell protein (S.S.P),
antibiotics and organic solvents.

43. CONTINUOUS fermentation
process
 The dilution rate is the ratio of inflowing
amount of medium to the volume of the
culture.
 Thus
 D = F / V
D= dilution rate (h-1)
F= flow rate (dm3 /h)
V= volume (dm3 )

44.  The change in cell concentration of cells at
perticular time period is expressed by the
following equation
dx/dt= growth rate – output
Or dx/dt = μx - Dx
In the process of continuous culture technique
the output is balanced by growth hence,
μx = Dx
μ – D
Dx / dt= D

45.  The biomass concentration in the
chemostat is determined by the
following equation
X = Y(SR - s)
X= steady state concentration
S= steady state residual concentration in
the medium

46. Advantages and disadvantages of batch and
continuous operations
BATCH SYSTEMS
 easy to operate and control
 genetic stability of organism
could be controlled if it is
genetically engineered
biocatalyst.
 lower contamination risk
 non-productive down time is a
disadvantage
 batch to batch variability is
problem
 accumulation of inhibitory
products is problem
CONTINUOUS SYSTEMS
 degeneration of
biocatalyst
 higher contamination risk
is a disadvantage
 efficient, higher
productivity
 product is obtained with
uniform characteristics;
quality of the product is
almost same from time to
time
 no accumulation of