Enzymes can be immobilized to support structures using various techniques. This confines and anchors the enzymes, making them stable and reusable. The main immobilization techniques are adsorption, entrapment, covalent binding, and cross-linking. Adsorption uses weak bonds to attach enzymes to inert supports, but the bonding is weak. Covalent binding and cross-linking form stronger covalent bonds or cross-links between enzymes and supports or between enzyme molecules. Immobilization can impact enzyme properties and kinetics but allows for reuse of enzymes in industrial, analytical, and therapeutic applications.

1. ENZYME IMMOBLIZATION
• INTRODUCTION
• Enzyme are large biological molecules responsible for the thousands of metabolic processes that
sustain life.
• Since enzyme are not stable and they can not be recovered for reuse.
• DEFINATION :
• “Immobilization of enzyme (or cells) refer to the technique of confining anchoring the enzyme
in on an inert support for their stability and function reuse”

2. PROPERTIES
• The material used for enzyme immobilization called carrier matrices .
• The ideal carrier matrix has the following properties are:
• Low costs
• Inert less
• Physical strength
• Regenerability after use

3. IMMOBILIZATION TECHNIQUES
• Immobilization techniques can be classified by basically two methods, the
chemical and the physical method. The former is covalent bond formation
dependent .
• The two Method consist of :
• Chemical method consist of Covalent binding, Cross linking
• Physical method consist of Adsorption, Entrapment:

4. METHODS OR TECHNIQUES
• ADSORPTION
• ENTRAPMENT
• COVALENT BINDING
• CROSS LINKING

5. ADSORPTION
• Adsorption involves the physical binding of enzyme (or cells) on the surface of an inert support.
• The support material may be inorganic (e.g. alumina ,silica gel ,calcium) or organic ( starch,
carboxymethyl,cellulose ).
• Adsorption of enzyme molecules involves weak forces such as Vander Waals forces and
hydrogen bonds.
• Therefore ,the adsorbed enzyme can be easily removed by minor changes in pH, ionic strength
or Temperature.

6. ADVANTAGES & DISADVANTAGES
• Advantages
• 1.The procedure of immobilization is simple.
• 2. It is possible to separate and purify the enzymes while being immobilized.
• 3. The enzymes are not usually deactivated by adsorption.
• 4. The adsorption is a reversible process.
• Disadvantages
• 1. The bonding strength is weak.
• 2. The state of immobilization is very sensitive to solution pH, ionic strength, and temperature.
• 3. The amount of enzymes loaded on a unit amount of support is usually low.

8. ENTRAPMENT
• Enzymes can be entrapped within cross-linked polymers by forming a highly cross-linked network of polymer in
the presence of an enzyme.
• This method has a major advantage in the fact that there is no chemical modification of the enzyme, therefore, the
intrinsic properties of an enzyme are not altered.
• However, the enzyme may be deactivated during the gel formation. Enzyme leakage is also a problem. The most
commonly employed crosslinked polymer is the polyacrylamide gel system.
• This has been used to immobilize alcohol dehydrogenase, glucose oxidase, amino acid oxidase, hexokmase,
glucose isomerase, urease, and many other enzymes.

9. ENZYME CAN BE ENTRAPPED
BY SEVERAL WAYS :
• 1.Enzyme inclusion in gels.
• 2.Enzyme inclusion in fibers.
• 3.Enzyme inclusion in microcapsules

10. COVALENT BINDING
• Immobilization of the enzyme can be achieved by creation covalent bonds between the chemical
groups of enzyme and chemical groups of the support.
• The following are the common methods of covalent binding:
• 1.cyanogen bromide activation
• 2.Diazotation
• 3.Peptide bonds formation
• 4.Activation by Bi or polyfunctional regents

11. CROSS –LINKING
• The absence of a solid support is a characteristics feature of immobilization of enzymes by
cross-linking.
• The enzyme molecules are immobilized by creating cross-links between them , through the
involvement of poly-functional reagent.
• These reagent in fact react with the enzyme molecules and create brides which form the
backbone to hold enzyme molecules.
• Glutaraldehyde is the most extensively used cross-linking reagent

12. EFFECT OF IMMOBILIZATION ON ENZYME PROPERTIES
• Enzyme immobilization is frequently associated with alterations in enzyme properties
,particularly the kinetic properties of enzyme some are listed below:
• There is substantial decrease in the enzyme specificity . This may be due to conformational
changes that occur when the enzyme gets immobilized.
• The kinetic constants km and V max of an immobilization enzyme differ from that of the native
enzyme .This is because the conformational changes of the enzyme will affect the affinity
between enzyme and substrate.

13. • APPLICAION OF IMMOBILIZED ENZYMES AND
CELLS
• INDUSTRIAL USES
• ANALYTICAL
• THERAPEUTIC