Entrapping Enzymes

        The entrapment method of immobilization is based on the localization of an enzyme within the lattice of a polymer matrix or membrane. It is done in such a way as to retain protein while allowing penetration of substrate. It can be classified into lattice and micro capsule types.

        This method differs from the covalent binding and cross linking in that the enzyme itself does not bind to the gel matrix or membrane.  This results in a wide applicability. The conditions used in the chemical polymerization reaction are relatively severe and result in the loss of enzyme activity. Therefore, careful selection of the most suitable conditions for the immobilization of various enzymes is required.

        Lattice-Type entrapment involves entrapping enzymes within the interstitial spaces of a cross-linked water-insoluble polymer. Some synthetic polymers such as polyarylamide, polyvinylalcohol, etc... and natural polymer (starch) have been used to immobilize enzymes using this technique.

        Microcapsule-Type entrapping involves enclosing the enzymes within semi permeable polymer membranes. The preparation of enzyme micro capsules requires extremely well-controlled conditions and the procedures for micro capsulation of enzymes can be classified as:

• Interfacial Polymerization Method:  In this procedure, enzymes are enclosed in semi permeable membranes of polymers. An aqueous mixture of the enzyme and hydrophilic monomer are emulsified in a water-immiscible organic solvent. Then the same hydrophilic monomer is added to the organic solvent by stirring. Polymerization of the monomers then occurs at the interface between the aqueous and organic solvent phases in the emulsion. The result is that the enzyme in the aqueous phase is enclosed in a membrane of polymer.

• Liquid Drying:  In this process, a polymer is dissolved in a water-immiscible organic solvent which has a boiling point lower than that of water. An aqueous solution of enzyme is dispersed in the organic phase to form a first emulsion of water-in-oil type. The first emulsion containing aqueous micro droplets is then dispersed in an aqueous phase containing protective colloidal substances such as gelatin, and surfactants, and a secondary emulsion is prepared. The organic solvent in then removed by warming in vacuum. A polymer membrane is thus produced to give enzyme micro capsules.

• Phase Separation:  One purification method for polymers involves dissolving the polymer in an organic solvent and re-precipitating it.  This is accomplished by adding another organic solvent which is miscible with the first, but which does not dissolve the polymer.

• Particles - The particle form is described in the above section.
• Membranes - Enzyme membranes can be prepared by attaching enzymes to membrane-type carriers, or by molding into membrane form.  The molding is done after the enzymes have been enclosed within semi-permeate membranes of polymer by entrapment.
• Tubes - Enzyme tubes are produced using Nylon and polyacrylamide tubes as carriers. The polymer tube is first treated in a series of chemical reactions and the enzyme is bound by diazo coupling to give a tube in a final step.
• Fibers - Enzymes that have been immobilized by entrapment in fibers to form enzyme fibers.