PRESENTED BY

SHIRJEEL AHMAD SIDDIQUI

B.Sc. Biotechnology
Semester VI
 Enzymes are biocatalyst synthesized by living cells.
 They are complex protein molecules that bring about
chemical reactions concerned with life.
 They can be produced in large amounts by
microorganisms for industrial applications.
 Enzymes in free state react with substrates to result in
products.
 Such use of enzymes is wasteful particularly for industrial
purposes ,since they are not stable and can’t be recovered
or reused.
 Immobilization of enzymes refers to the technique of
confining/anchoring the enzymes in or on an inert support
for their stability and functional reuse.
 By employing this technique, enzymes are made more
efficient and cost effective for their industrial use.
 Some workers regard immobilization as “a goose with a
golden egg” in enzyme technology
 It involves the physical binding of enzymes on the surface
of an inert support.
 The support materials may be inorganic (alumina, silica
gel, calcium phosphate gel) or organic (starch,
carboxymethyl cellulose,DEAE)
 Adsorption of enzymes involves weak forces like van der
Waals forces and hydrogen bonds.
 Therefore, adsorbed enzymes can be easily removed by
minor changes in pH or temperature.
 Sometimes this feature also serve as disadvantage for
industrial use.
 Enzymes can be immobilized by physical entrapment
inside a polymer or gel matrix.
 The size of the matrix pores is such that the enzyme is
retained while substrate and product molecule is passed
through.
 The matrices used for entrapping enzymes are made of
polyacrylamide gel ,collagen, gelatin, starch, cellulose,
silicone and rubber.
 The major limitation for entrapment of enzymes is their
leakage from matrix.
 It is used in industrial production of amino acids.
 It is also considered as a type of entrapment.
 It refers to the process of spherical particle formation
wherein a liquid or suspension is enclosed in semi-
permeable membrane .
 The membrane may be polymeric, lipoidal, or non-ionic in
nature.
 There are three distinct ways of encapsulation -
a) Building of special membrane reactors.
b) Formation of emulsions.
c) Stabilization of emulsions to form microcapsule.
 It involves the formation of covalent bonds between the
chemical group of enzymes and chemical group of
support.
 It can be achieved by forming the bonds commonly
specific as per the support and enzyme, it includes –
a) Cyanogen bromide activation - glycol groups of support
materials are activated by CNBr and binds to enzyme.
b) Diazotation – amino derivatives support is treated with
NaNO2 and HCl which in turn bind covalently to tyrosyl
or histidyl group of enzymes.
c) Peptide bond formation – The amino or carboxyl group
of support binds with carboxyl or amino groups of
enzyme respectively to form peptide bonds
 The enzyme molecules are immobilized by forming cross-
linking, through the involvement of polyfunctional
reagents.
 These reagents react with enzyme molecules and create
bridges which form the backbone to hold enzymes
molecules.
 Several reagents used in cross-linking like glutaraldehyde,
diazobenzidine, hexamethylene disisocyanate.
The selection of a particular method for immobilization of
enzymes is based on trial and error approach to choose the
ideal one.
The major factors to be considered are as follows –
 Enzyme catalytic activity
 Stability
 Regenerability
 Cost factor
The enzyme which have to be used in soluble form e.g.
enzymes in food additives, liquid detergents and diagnostic
reagents. They are stabilized using additives and chemical
methods, they include –
 Solvent stabilization
 Substrate stabilization
 Stabilization by polymers
 Stabilization by salts
 Stabilization by chemical modifications
 In food and beverage industry
 In biochemical analysis
 In affinity chromatography and purification
 Improved drug delivery
 Immobilization of artificial cells/organs
 Stable and more efficient in function.
 Can be reused many times.
 Products are enzyme free.
 Control of enzyme function is easy
 Suitable for industrial and medical use
 Minimize effluent disposal problems
Being so advantageous it has some disadvantages too –
 The possibility of loss of biological activity of an enzyme
during its immobilization.
 Not suitable for multiple enzymatic reactions.
 Expensive affair often requires sophisticated equipment.
 Pages 286-290, Enzyme Technology, Biotechnology,
U.Satyanaryana, Tenth edition,2016
 Image courtesy via internet –
 https://www.google.co.in/search?q=immobilized+enzymes
+and+their+products+table&source=lnms&tbm
 http://www.easybiologyclass.com/wp-
content/uploads/2015/04/Enzyme-whole-cell-
immobilization-methods-easybiologyclass.jpg
 https://www.google.co.in/search?q=images+of+adsorption
+in+immobilisation+of+enzymes&source=lnms&tbm=isc
h&sa=X&ved=