Enzymes are biocatalysts that facilitate biochemical reactions essential for the functioning of living organisms and have numerous applications in the pharmaceutical industry, including drug production and treatment of various diseases. Most industrial enzymes are derived from microorganisms through fermentation, with common strains including bacteria and fungi, which can be genetically engineered for higher yields. The production methods vary, with submerged fermentation being predominant, although solid-state fermentation presents advantages in certain contexts.

2. • Enzymes are the functional proteins or nucleic
acids (Ribozymes), also known as biocatalysts
that facilitate the execution of biochemical
reactions at the rates which are suitable for
the normal functioning, growth, and
proliferation of any living system, including
unicellular or multicellular plants as well as
animals

3. Unique properties
• The unique characteristics of enzymes makes
them highly applicable for a number of chemical
transformation reactions in pharmaceutical
industries, such as group protection and
deprotection
• selective acylation and deacylation,
• selective hydrolysis,
• deracemization,
• kinetic resolution of racemic mixtures,
• esterification

4. • The ability of enzymes to remain viable and perform catalytic activities
even outside their source organism, allows them to be exploited for
carrying out a number of industrial processes that rely on chemical
transformations of substrates to their corresponding products.
• [E] +[S] =[P] + [E]
• The reactions catalyzed by enzymes are highly efficient, that is, they occur
under ambient environmental conditions, that is, temperature, pH, and
pressure (the conditions depend on the physiological conditions of the
source organisms and its environmental conditions).
• For instance, an enzyme obtained from a mesophilic organism (optimal
growth temperature is 37◦C and growth temperature range from 20 to
40◦C), inhabiting neutral environments, shall work efficiently at mild
temperatures, neutral pH, and atmospheric pressure

5. Continued…………
Enzymes are applicable in pharmaceutical
industries, for
• group protection and deprotection, selective
acylation and
• deacylation, selective hydrolysis, deracemization,
kinetic
• resolution of racemic mixtures, esterification,
transesterification,

6. Name Use
Cystic Proteinase arthritis, osteoporosis, AIDS, immune-related diseases, atherosclerosis,
cancer, and for a wide variety of parasitic diseases such as malaria,
amebiasis, chagas disease, leishmaniasis, or African
Asparaginase Anticancer: treat acute lymphocytic leukemia (ALL).
deoxyribonuclease
I,
cystic fibrosis patients to improve mucociliary clearance and pulmonary
function.
streptokinase, dissolve blood clots that have formed in the blood vessels
urokinase, Urokinase is a thrombolytic (THROM-bo-LIT-ik) drug, sometimes called a
"clot-busting" drug
Hyaluronidase to increase absorption of fluids or medicines that are injected into your
skin.
Oegadenaswe Pegademase is used to treat a certain rare genetic immune system
problem (severe combined immune deficiency disease-SCID).
Pegademase replaces a certain natural substance (an enzyme called
adenosine deaminase-ADA) that is missing in people with SCID.
glucocerebrosidase. type 1 Gaucher disease. enlargement of the liver and spleen (
hepatosplenomegaly ),
Enzymes which are used for pharmaceutical applications are called Pharmaceutical
enzymes like

7. Production methods
• Majority of the industrial enzymes come from
microorganisms as they are the most convenient
sources, which gives the
• propensity of faster production,
• easy scale up,
• recovery and purification,
• strain manipulation for over-expression,
• enzyme activity,
• specificity modulations and so on

8. Ist Pharmaceutical enzyme
• The first enzyme produced industrially was the
fungal amylase Takadiastase which was
employed as a pharmaceutical agent for
digestive disorders.

9. • At present,around 200 types of microbial
enzymes from 4,000 known enzymes are used
commercially .Nearly 75% of the total
enzymes are produced by three top enzyme
companies,that is, Denmark-based
Novozymes, US-based DuPont, and
Switzerland-based Roche [10]

10. Source of enzymes
[Fermentation culture]
• Most of the industrial enzymes including those used in
pharmaceutical industries are produced through fermentation
of suitable microbial strains of bacteria and fungi due to their
easy handling,
• fast growth rates, and
• convenient scale up in large vessels (fermenters).
• Bacteria - Escherichia coli, Bacillus subtilis, lactic acid bacteria
• filamentous fungi such as Aspergillus oryzae, Aspergillus niger,
Trichoderma atroviride, and Yeasts ( Saccharomyces cerevisiae,
Pichia pastoris,)
• As the strains have been improved through genetic engineering,
they produce enzymes in very high yields.

11. Conditions for selection
• Selection of suitable microbial strains for the
production of various industrial enzymes is a very
important aspect for their successful industrial
applications.
• Ideally, the enzymes produced should be secreted
out in the fermentation medium by the producing
microbial strain as it makes the downstream
processing more convenient and economically
feasible; however, this may not be the case with
most of the industrial strains.

12. Production methods
Both the processes
have their own benefits
and limitations.
Most industries have
adopted SmF process
for enzyme production;
but there has been a
renewed interest in SSF
for certain specific
industries.

13. Typical bioreactor
A typical bioreactor consists of following
parts: • Agitator – used for the mixing of the
contents of the reactor which keeps the
“cells” in the perfect homogenous condition
for better transport of nutrients and oxygen
to the desired product(s).
• Baffle – used to break the vortex
formation in the vessel, which is usually
highly undesirable as it changes the center
of gravity of the system and consumes
additional power.
• Sparger – In aerobic cultivation process,
the purpose of the sparger is to supply
adequate oxygen to the growing cells
. • Jacket – The jacket provides the annular
area for circulation of constant temperature
of water which keeps the temperature of
the bioreactor at a constant value

14. Enzyme Production Through Solid-
State Fermentation
• Solid-state fermentation has a great potential for enzyme
production .
• This process offers several advantages over the SmF process, such
as high product titer, lesser effluent generation, use of simple
fermentation equipment, less trained labor, and so on .
• However, SSF is in general more suitable for those processes where
the crude fermented products themselves are used as the final
product rather than the isolated enzymes.
• Agroindustrial residues are commonly used substrates for the SSF
processes, including those used to produce enzymes. A variety of
substrates have been used for the cultivation of enzyme producing
microorganisms. Examples of the substrates used are wheat bran,
rice bran, sugar cane bagasse, wheat straw, rice straw, saw dust,
corncobs, banana waste, cassava waste, palm oil mill waste, oil
cakes, and so on .

15. Semisolid culture

17. Disadvantage of semisolid culture

18. Submerged fermentation involves the growth of the
microorganism as a suspension in a liquid medium in
which various nutrients are either dissolved or suspended
as particulate solids in many commercial media.
Submerged fermentation is a process involving the
development of microorganisms in a liquid broth.

19. Types
• Submerged fermentation for industrial production processes are manly
carried out in four ways, namely
• batch culture,
• continuous culture,
• perfusion culture, and
• fed batch culture. In batch culture,
• the production strains are inoculated in a fixed volume of medium and
thereafter no further addition of medium is carried out during the entire
fermentation process (except the addition of acid, alkali, and antifoam).
Bioreactor :A vessel which has provision of cell cultivation under sterile condition &
control of environmental conditions e.g., pH, Temperature, Dissolved oxygen etc. • It
can be used for the cultivation of microbial plant or animal cells.
• This process can either be aerobic or anaerombic.
• The bioreactors are commonly cylindrical, ranging in size from litres to cubic etres,
and are often made of stainless steel.

20. Advantage of submerged culture
• Generally, aerobic microorganisms are grown in submerged cultures in a stirred-tank
reactor for industrial production processes for extracellular enzymes and even for the
intracellularly over expressed enzymes as in the case of recombinant E. coli and other
bacterial and yeast strains..
• The fermenters for SmF applications have been developed successfully and are being
used for the production of enzymes and other biotechnological products .
• The main reason behind the success of SmF is feasibility of process scale up from pilot
scale fermenters (∼100 L) to production scale fermenters having capacities of millions of
liters (vessel volumes) .
there is no problem of mass transfer and heat dissipation

21. Disadvantage of Submerged
Culture
• • Initial investment cost is very high.
• mass transfer and heat dissipation generally
achieved required cost of higher energy
consumption

22. Down stream process: After
fermentation
• • Once fermentation is finished, the fermented
liquor is subjected to rapid cooling to about 5o C in
order to reduce deterioration.
• • Separation of micro-organisms is accomplished
either by filtration or by centrifugation of the
refrigerated broth with adjusted pH.
• • To obtain a higher purity of the enzyme, it is
precipitated with acetone, alcohols or inorganic salts
(ammonium or sodium sulfate).
• • In case of large scale operations, salts are preferred
to solvents because of explosion hazards

23. THANKS

24. Enzyme Production Through SmF
• It is carried out in a liquid medium, wherein
the nutrients and other medium components
are either dissolved or remain suspended in
an aqueous medium.
• The microorganisms also grow and proliferate
in the liquid fermentation medium in the
suspended state.
• .

25. • In continuous culture, the fermentation process is initiated in the batch
mode. However, after certain time period fresh medium or nutrient
concentrate is added into the fermenter at a rate which approximately
matches the growth rate of the microorganism used and simultaneously
the medium containing the product and biomass is continuously
withdrawn from the overflow line of the vessel .
• A perfusion culture is somehow similar to a continuous culture process, in
that it also involves the constant feeding of fresh media and removal of
spent media and product. However, it differs from the previous in
retention of high numbers of viable cells by using alternating tangential-
flow and standard tangential-flow filtration or by binding the cells to a
substrate (capillary fibers, membranes, microcarriers in fixed bed, and so
on) in the fermenter [26]. In the case of fed batch culture, the
concentrated components of the nutrients are added to the batch culture
in small lots at regular time points or based on the requirement of the
process [24, 25].