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Antifoaming Agents
Inducers
Precursors
Inhibitors in Fermentation Technology
Dr. Pavan J Kundur
Assistant Professor
Department of Biotechnology and Microbiology
P C Jabin Science College
Hubballi
What is the Foam ?
A foam is a substance that is formed by
trapping pockets of gas in a liquid or solid.
A bath sponge and the head on a glass of
beer are examples of foams.
In most foams, the volume of gas is large,
with thin films of liquid or solid separating
the regions of gas.
An important division of solid foams is into
closed-cell foams and open-cell foams.
In a closed-cell foam, the gas forms
discrete pockets, each completely
surrounded by the solid material.
Cappuccinos are topped with a layer of steamed-milk
foam.
Antifoams
Antifoams are necessary to reduce foam formation during
fermentation.
Foaming is largely due to media proteins that become
attached to the air-broth interface where they denature to form
a stable foam “skin” that is not easily disrupted.
If uncontrolled the foam may block air filters, resulting in the
loss of aseptic conditions; the fermenter becomes
contaminated and microorganisms are released into the
environment.
Of possibly the most importance is the need to allow
“freeboard” in fermenters to provide space for the foam
generated. If foaming is minimized, then throughputs can be
increased.
There are three possible approaches to controlling foam
production:
the use of a defined medium and a modification of some of
the physical parameters, e.g. pH, temperature, aeration and
agitation (if the foam is due to media components), use of
chemical foam breakers and addition of chemical antifoams.
Antifoams are surface active agents that reduce the surface
tension in the foams and destabilize the protein films by
(i) hydrophobic bridges between two surfaces;
(ii) displacement of the adsorbed protein; and
(iii) rapid spreading on the surface of the film.
The ideal antifoam should have the following properties:
1. readily and rapidly dispersed with rapid action
2. high activity at low concentrations
3. prolonged action
4. non-toxic to fermentation microorganisms, humans or
animals
5. low cost
6. thermostability
7. compatibility with other media components and the
process, i.e., having no effect on oxygen transfer rates or
downstream processing operations
8. be heat sterilisable
Natural antifoams include plant oils (e.g., from soya, sunflower
and rapeseed), deodorized fish oil, mineral oils
The synthetic antifoams are mostly silicon oils, poly alcohols
and alkylated glycols.
Since antifoams are of low solubility, they need a carrier,
e.g., lard oil, liquid paraffin or castor oil, which may be
metabolized and therefore affect the fermentation process.
Many of the surface-active agents, particularly the oils, are
added as emulsions of suspended oil droplets which can
destabilise the foams by acting as hydrophobic bridges
between the two film surfaces or by displacing the stabilising
adsorbed material, e.g. protein, at the bubble–liquid
interface.
The concentrations of many antifoams which are
necessary to control foaming may reduce the oxygen
transfer rate by as much as 50%.
Thus, antifoam addition should be kept to an absolute
minimum.
Some antifoams may reduce the oxygen transfer rate as
well as adversely affect downstream processing steps,
especially membrane filtration.
If the oxygen transfer rate is too severely affected
mechanical foam breakers may have to be considered.
Precursors
Precursors are defined as “substances added prior to or
simultaneously with the fermentation which are incorporated
without any major change into the molecule of the
fermentation product and which generally serve to increase
the yield or improve the quality of the product”.
They are required in certain industrial fermentations and are
provided through crude nutritive constituents, e.g., corn steep
liquor or by direct addition of more pure compounds.
Some fermentations must be supplemented with specific
precursors, notably for secondary metabolite production.
When required, they are often added in controlled
quantities and in a relatively pure form, examples include,
D-threonine is used as a precursor in L-isoleucine
production by Serratia marcesans, and anthranillic acid
additions are made to fermentations of the yeast
Hansenula anomola during L-tryptophan production.
The use of corn steep liquor as side-chain precursors in
penicillin fermentations results in six different penicillins as
opposed to the use of phenylacetic acid which results in
mainly Penicillin G formation.
Inducers and Elicitors
•If product formation is dependent upon the presence of a
specific inducer compound or a structural analogue, it must
be incorporated into the culture medium or added at a
specific point during the fermentation.
•The majority of enzymes of industrial interest are inducible.
Inducers are often substrates such as starches or dextrins
for amylase.
•In plant cell culture the production of secondary
metabolites, such as flavanoids and terpenoids can be
triggered by adding elicitors.
These may be isolated from various microorganisms,
particularly plant pathogens. Inducers are often necessary in
fermentations of genetically modified microorganisms.
This is because the growth of genetically modified
microorganisms (GMMs) can be impaired when the cloned
genes are “switched on”, due to the very high levels of their
transcription and translation.
Consequently, inducible systems for the cloned genes are
incorporated that allow initial maximization of growth to
establish high biomass density, whereupon the cloned gene
can then be “switched on” by the addition of the specific
chemical inducer,
e.g., a commercial system developed based on the alcA
promoter in Aspergillus nidulans to express human interferon
α2.
The system is induced by volatile chemicals like ethyl methyl
ketone which are added when the biomass has increased to
an adequate level and growth medium contains non-
repressing carbon source.
Inhibitors
Inhibitors are used to redirect metabolism towards the target
product and reduce formation of other metabolic intermediates;
others halt a pathway at a certain point to prevent further
metabolism of the target product.
An example of an inhibitor specifically employed to redirect
metabolism is sodium bisulphite, which is used in the
production of glycerol by S. cerevisiae.
Some GMMs contain plasmids bearing an antibiotic resistance
gene, as well as the heterologous gene(s).
The incorporation of this antibiotic into the medium used for
the production of the heterologous product selectively inhibits
any plasmid-free cells that may arise.
Thank You

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Anti-foaming agents, inducers, precursors and inhibitors in Fermentation technology

  • 1. Antifoaming Agents Inducers Precursors Inhibitors in Fermentation Technology Dr. Pavan J Kundur Assistant Professor Department of Biotechnology and Microbiology P C Jabin Science College Hubballi
  • 2. What is the Foam ? A foam is a substance that is formed by trapping pockets of gas in a liquid or solid. A bath sponge and the head on a glass of beer are examples of foams. In most foams, the volume of gas is large, with thin films of liquid or solid separating the regions of gas. An important division of solid foams is into closed-cell foams and open-cell foams. In a closed-cell foam, the gas forms discrete pockets, each completely surrounded by the solid material. Cappuccinos are topped with a layer of steamed-milk foam.
  • 3. Antifoams Antifoams are necessary to reduce foam formation during fermentation. Foaming is largely due to media proteins that become attached to the air-broth interface where they denature to form a stable foam “skin” that is not easily disrupted. If uncontrolled the foam may block air filters, resulting in the loss of aseptic conditions; the fermenter becomes contaminated and microorganisms are released into the environment. Of possibly the most importance is the need to allow “freeboard” in fermenters to provide space for the foam generated. If foaming is minimized, then throughputs can be increased.
  • 4. There are three possible approaches to controlling foam production: the use of a defined medium and a modification of some of the physical parameters, e.g. pH, temperature, aeration and agitation (if the foam is due to media components), use of chemical foam breakers and addition of chemical antifoams. Antifoams are surface active agents that reduce the surface tension in the foams and destabilize the protein films by (i) hydrophobic bridges between two surfaces; (ii) displacement of the adsorbed protein; and (iii) rapid spreading on the surface of the film.
  • 5. The ideal antifoam should have the following properties: 1. readily and rapidly dispersed with rapid action 2. high activity at low concentrations 3. prolonged action 4. non-toxic to fermentation microorganisms, humans or animals 5. low cost 6. thermostability 7. compatibility with other media components and the process, i.e., having no effect on oxygen transfer rates or downstream processing operations 8. be heat sterilisable Natural antifoams include plant oils (e.g., from soya, sunflower and rapeseed), deodorized fish oil, mineral oils
  • 6. The synthetic antifoams are mostly silicon oils, poly alcohols and alkylated glycols. Since antifoams are of low solubility, they need a carrier, e.g., lard oil, liquid paraffin or castor oil, which may be metabolized and therefore affect the fermentation process. Many of the surface-active agents, particularly the oils, are added as emulsions of suspended oil droplets which can destabilise the foams by acting as hydrophobic bridges between the two film surfaces or by displacing the stabilising adsorbed material, e.g. protein, at the bubble–liquid interface.
  • 7. The concentrations of many antifoams which are necessary to control foaming may reduce the oxygen transfer rate by as much as 50%. Thus, antifoam addition should be kept to an absolute minimum. Some antifoams may reduce the oxygen transfer rate as well as adversely affect downstream processing steps, especially membrane filtration. If the oxygen transfer rate is too severely affected mechanical foam breakers may have to be considered.
  • 8. Precursors Precursors are defined as “substances added prior to or simultaneously with the fermentation which are incorporated without any major change into the molecule of the fermentation product and which generally serve to increase the yield or improve the quality of the product”. They are required in certain industrial fermentations and are provided through crude nutritive constituents, e.g., corn steep liquor or by direct addition of more pure compounds.
  • 9. Some fermentations must be supplemented with specific precursors, notably for secondary metabolite production. When required, they are often added in controlled quantities and in a relatively pure form, examples include, D-threonine is used as a precursor in L-isoleucine production by Serratia marcesans, and anthranillic acid additions are made to fermentations of the yeast Hansenula anomola during L-tryptophan production. The use of corn steep liquor as side-chain precursors in penicillin fermentations results in six different penicillins as opposed to the use of phenylacetic acid which results in mainly Penicillin G formation.
  • 10. Inducers and Elicitors •If product formation is dependent upon the presence of a specific inducer compound or a structural analogue, it must be incorporated into the culture medium or added at a specific point during the fermentation. •The majority of enzymes of industrial interest are inducible. Inducers are often substrates such as starches or dextrins for amylase. •In plant cell culture the production of secondary metabolites, such as flavanoids and terpenoids can be triggered by adding elicitors.
  • 11. These may be isolated from various microorganisms, particularly plant pathogens. Inducers are often necessary in fermentations of genetically modified microorganisms. This is because the growth of genetically modified microorganisms (GMMs) can be impaired when the cloned genes are “switched on”, due to the very high levels of their transcription and translation. Consequently, inducible systems for the cloned genes are incorporated that allow initial maximization of growth to establish high biomass density, whereupon the cloned gene can then be “switched on” by the addition of the specific chemical inducer, e.g., a commercial system developed based on the alcA promoter in Aspergillus nidulans to express human interferon α2.
  • 12. The system is induced by volatile chemicals like ethyl methyl ketone which are added when the biomass has increased to an adequate level and growth medium contains non- repressing carbon source.
  • 13. Inhibitors Inhibitors are used to redirect metabolism towards the target product and reduce formation of other metabolic intermediates; others halt a pathway at a certain point to prevent further metabolism of the target product. An example of an inhibitor specifically employed to redirect metabolism is sodium bisulphite, which is used in the production of glycerol by S. cerevisiae. Some GMMs contain plasmids bearing an antibiotic resistance gene, as well as the heterologous gene(s). The incorporation of this antibiotic into the medium used for the production of the heterologous product selectively inhibits any plasmid-free cells that may arise.