This document summarizes various techniques for immobilizing enzymes and their therapeutic applications. It defines enzyme immobilization as restricting an enzyme's movement by trapping it within an inert support material. The main techniques discussed are adsorption, entrapment, encapsulation, covalent bonding, and cross-linking. Adsorption physically binds enzymes to carrier surfaces, while entrapment traps them within porous matrices. Encapsulation encloses enzymes within semi-permeable membranes. Covalent bonding and cross-linking form strong chemical bonds between enzymes and supports. Therapeutic uses of immobilized enzymes include producing antibiotics, diagnosing and treating diseases, and applications in food processing, research, biodiesel production, and waste treatment.

1. SUBMITTED BY:
ANANYA ROY
(Y23254003)
M.PHARM I SEM
Immobilizati on: Various Techniques and Its
Therapeutic Applications
Subject :- Pharmaceutical Biotechnology
PHS CC-1203
DEPARTMENT OF PHARMACEUTICAL SCIENCES
DR. HARISINGH GOUR VISHWAVIDYALAYA, SAGAR
(M.P.)
SUBMITTED TO:
Prof. Umesh K. Patil
Dr. Udita Agarwal
Dr. Priyanka Jain
Mr. Satyamshyam Vishwakarma

2. Immobilization:
Various techniques
and its therapeutic
applications

3. CONTENTS
1) Brief Introduction on Enzymes
2) Enzyme Immobilization
- Definition
- Advantages
- Disadvantages
3) Carriers
- Properties of an Ideal Carrier Matrix For Enzyme Immobilization
- Classification of Carriers
4) Methods of Enzyme Immobilization
- Adsorption
- Entrapment
- Encapsulation
- Covalent Bonding
- Cross Linking
5) Therapeutic Application of Enzyme Immobilization
6) References

4. Brief Introduction On Enzymes
❖ What are Enzymes?
❖ How to identify Enzymes?
❖ How does Enzyme work?
❖ Enzyme Biotechnology

5. WHAT ARE ENZYMES?
Enzymes are proteins that act as biocatalyst
to speed up the chemical reactions.

6. HOW TO IDENTIFY ENZYMES?
The suffix “ ASE ” is used for the
identification of Enzymes.
Example :
• Oxidoreductase
• Transferase
• Hydrolase
• Isomerase

7. HOW DOES ENZYME WORK?
- Enzymes work by speeding up the chemical
reaction.
- They act as catalyst - means they make reaction
happen faster.
- Enzyme have active sites (the most important part
of Enzyme) on which the substrate binds.
- When the substrate binds to the enzyme, it
undergoes a reaction and is transformed into a
"PRODUCT"

8. EXAMPLE :
Suppose we have 2 Substrate A and B
Enzyme will do some necessary changes in the substrate,
by combining both of the substrates, it is transformed
into PRODUCT and is separate out.

9. In Enzyme Biotechnology, we
modify enzyme with the help of
biotechnology sо, we can use this
enzymes widely as per human
requirements.
ENZYME BIOTECHNOLOGY

10. ENZYME IMMOBILIZATION
It means the movement of
enzyme is limited upto a certain
area space in which the enzyme
is entrapped in.
NO MOVEMENT

11. DEFINATION :
- “Enzyme Immobilization" is a process where
enzymes аrе trapped in a matrix or inert solid
support material, restricting the movement of
the enzyme either completely or to a small
limited region.
- This allows the enzyme to remain in a fixed
position while still being active maintaining their
catalytic activity.

12. ADVANTAGES OF ENZYME
IMMOBILIZATION :
• Protection from degradation and deactivation.
• Cost effective.
• Reusability for many reactions.
• Ability to control the reactions.
• Enhanced stability.
• Easy separation of enzyme from the product.

13. • The possibility of loss of biological activity of an enzyme.
• It is an expensive process often requiring sophisticated
equipment.
• Some time native structure of enzyme is disrupted due
to immobilization.
• All the enzymes are not immobilized by immobilization.
• Cost of carriers.
DISADVANTAGES OF ENZYME
IMMOBILIZATION :

14. • Inert (does not harm the enzyme or react with it)
• Cost effective.
• Physically strong and stable.
• Regenerability after use.
• Reduction in product inhibition.
• Enhancement of enzyme specificity.
CARRIER PROPERTIES

15. 1) Inorganic Carriers
CLASSIFICATION OF CARRIERS :
Ex : Glass silica, Metals like - Aluminum oxide
or Titanium oxide.
➢ These carriers are made of inorganic material.
➢ Provide high pressure stability.

16. 2) Organic Natural Carriers
Ex : Cellulose derivatives, Agarose, Dextran,
Carboxy methyl cellulase, Diethylaminoethyl
cellulose (DEAE).
➢ These carriers are made of organic materials.
➢ Provide biocompatible environment for
enzyme enhancing its stability & activity.

17. 3) Organic Synthetic Carriers
➢ These carriers are made up of synthetic
polymers.
➢ Provide high chemical and mechanical stability.
Ex : Polyvinyl acetate, Polystyrene,
Acrylic polymers.

18. Methods of Enzyme Immobilization
Covalent Bonding
Cross Linking
Adsorption
Entrapment
Encapsulation
Physical Chemical

19. ADSORPTION
• Involves the physical binding of the enzyme on the surface
of carrier matrix.
• In this process the enzyme molecules is adhere (bind) to
the surface of carrier matrix with the help of weak
interaction forces such as –
Van der wall forces, Electrostatic forces, Hydrophobic
bond, Hydrogen bond.
Ex :
α-amylase Calcium phosphate
Catalase Activated charcoal
Invertase DEAE

20. Advantages Disadvantages
1. Low surface area for
binding.
2. Risk of microbial attack.
3. Weak forces - chances
of desorption.
1. Simple and economical.
2. Limited loss of activity.
3. Can be recycled,
regenerated, reused.

21. ENTRAPMENT
- In this process, Enzymes are entrapped within the
carrier matrix/gels/fibers.
- Porous matrix is used.
- In this, the size of the matrix pore is
such that the enzyme is retained in
it, while the substrate and product
molecule pass through.
- Water insoluble polymer are used
here,
Ex : Polyacrylamide, Starch.

22. Entrapment of enzyme inside the gels.
Ex : Polyacrylamide gels, Polyvinyl alcohol gels etc.
1) Enzyme inclusion in gels
Enzyme are entrapped inside fibers.
Ex : Cellulose etc.
2) Enzyme inclusion in fibers
Enzyme are entrapped inside a microcapsule matrix.
Ex : Polyamines.
3) Enzyme inclusion in microcapsule
SOME OTHER TYPE OF ENTRAPMENT

23. ENCAPSULATION
It is a type of entrapment in which the enzyme
molecules are regularly taken up on aqueous medium
which further enclosed in a semi-permeable membrane.
- The membrane should be polymeric
lipoidal or non-ionic in nature.
- Mostly the semi-permeable
membrane is made up of nylon or
cellulose.

24. COVALENT BONDING
(STRONGEST IMMOBILIZATION)
- In this method, the enzyme molecules adhere
to the carrier matrix by forming covalent
bonds.
- The functional group that may take part in
bonding are amino group, carboxyl group,
hydroxyl group, phenolic group, thiol group.
- Most widely used method.
Ex : Cellulose, Sepharose, Aminobenzyl etc.

25. Advantages Disadvantages
Changes in structure
of enzyme may
cause loss of activity.
The binding force
between enzyme and
carrier is so strong
that no leakage of the
enzyme occurs.

26. CROSS LINKING
- In this method, cross links are
formed between the enzyme
molecule
(solid support is not required)
- Used polyfunctional reagent such as
glutaraldehyde diazobenzidine and
which react with enzyme molecules
and form this cross linking between
enzyme.

27. Advantages Disadvantages
1. May cause some
changes.
2. Denaturation of
enzymes.
1. Enzyme strongly
bounds.
2. Higher stable
(ph temp)
3. Simple and cost
effective.

29. THERAPEUTIC APPLICATIONS OF ENZYME
IMMOBILIZATION
1. Industrialproduction:Industrial productionof antibiotics,beverages, amino acids etc.
uses immobilizedenzymes.
2.Immobilized enzymes are widely used for diagnosisand treatment of many diseases
such as inborn disorder.
3. Food industry: Enzymes like pectinases and cellulasesimmobilizedon suitable carriers
are successfully used in the productionof jams, jellys and syrups from fruits and
vegetables.
4. Research: The use of immobilized enzyme allow researcher to increase the efficiency
of different enzymes such as different proteases for cell and organelle lysis.
5. Biodieselproduction from vegetable oils.
6. Textile industry: Scouring, bio polishing and desizing of fabrics.
7. Waste water management: Treatment of sewage and industrialeffluents.
8. Detergent industry: Immobilizationof lipase enzyme for effective dirt removal from
cloths.

30. REFERENCES :
• Vyas S. P. and Dixit V.K., Pharmaceutical Biotechnology, First Edition, 2005
(reprint), C.B.S. Publishers and Distributors, New Delhi.
• Patil A.S. et al., A Text Book of Pharmaceutical Biotechnology, First
Edition, 2019, S. Vikas and Company, Jalandhar.
• Rajesh Gollapudi and Sujitha Paladugu, Concise course in Pharmaceutical
Biotechnology, First Edition, 2020, S. Vikas and Company, Jalandhar.
• Sikander Ali et. al., "Enzymes Immobilization:An Overview of Techniques,
Support Materials and its Applications", International Journal of Scientific
& Technology Research, 6 (9), 2017.
• Viet T.Q. et al., "Immobilizationof Cellulase Enzyme in Calcium Alginate
Gel and its ImmobilizedStability", American Journal of Research
Communication, 1(12), 2013.

31. THANK YOU