Dr. Gul Shahnaz Immobilization of enzymes Immobilization of enzymes (or cells) refers to the technique of confining/anchoring the enzymes (or cells) in or on an inert support for their stability and functional reuse. Immobilized enzymes possess higher resistance to environmental changes and can be recovered/recycled easily when compared to the free forms. Problems and Limitations Associated with Industrial Enzymes Enzyme immobilization methods Immobilization strategies entail fixing or entrapping enzymes within solid support materials. Researchers have suggested several support materials, besides many beneficial approaches for the immobilization of enzymes. The main function of the support is to stabilize the structures of enzymes and accordingly preserve their efficacy to a great extent by rendering them more resistant to the surrounding environments.
Enzyme immobilization permits an easy recovery of both the used enzymes and their support materials, and this is particularly beneficial in the food, medical, and pharmaceutical applications.
Enzymes in their immobilized forms possess much higher stability and are also easier to handle when compared to their free forms.
Additionally, the enzymatic reaction can occur in a nonaqueous medium where the solid supports preserve the enzyme’s constituents and make them stronger, which enhances their catalytic activity and renders them reusable for several times.
Another benefit of the immobilization process is that the catalysts can alter from homogeneous to heterogeneous forms after the enzymatic binding, which assists in separating the enzymes, producing products with high purity Adsorption • The enzyme here are adhere to carrier matrix due to hydrophobic effects or by forming salt links. • Binding is strong but may be weakened by substrate, pH etc.
• Commonly used matrices :-
ion exchange matrices, porous carbon,clays, hydrous metal oxides, polymeric aromatic resins etc. Covalent binding • The enzyme here are attached to certain matrix by forming covalent bonds • Binding is very strong. It generally occurs with side chain of the enzyme. Lysine is very useful, as they are very reactive, usually exposed to surface and very rarely occur at active site of enzyme. • Commonly used matrices :- agarose, cellulose, poly acrylamides etc. • Activation by CNBr Cont…. • Activation by Ethyl chloroformate Cont…. • Immobilization of enzyme using Glutaraldehyde Methods for immobilization
c) Entrapment d) Membrane confinement
Entrapment
• The enzyme here are entrapped with in suitable gels or fibers,
with or with out covalent bonds.
• Commonly used matrices :-
Cellulose acetate fibers, calcium alginate etc. Membrane confinement
• The enzyme molecule here are confined to semipermiable
membrane. Thus in a aqueous phase beg of membrane is free to move with out allowing enzyme to come out. Advantages • Enzymes are costly, Immobilization permits their repeated use. • The product can be easily separated without any additional cost. • Immobilized enzymes can be used in nonaqueous system. • Continuous production system can be used. • Thermo stability of some enzyme can be increased by immobilization. • Enzyme can be used at much higher concentration then free enzyme. Disadvantages
• Additional cost. • It sometime affects stability and activity of enzyme. • This approach can not be used when one of the substrates is insoluble. • Some immobilization methods restricts the diffusion of substrate. Applications of immobilized enzymes
