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Enzymes may be immobilized by a variety of methods, which may be broadly classified as physical, where weak interactions between support and enzyme exist, and chemical, where covalent bonds are formed with the enzyme. To the physical methods belong: (i) containment of an enzyme within a membrane reactor, (ii) adsorption (physical, ionic) on a water-insoluble matrix, (iii) inclusion (or gel entrapment), (iv) microencapsulation with a solid membrane, (v) microencapsulation with a liquid membrane, and (vi) formation of enzymatic Langmuir-Blodgett films. The chemical immobilization methods include: (i) covalent attachment to a water-insoluble matrix, (ii) crosslinking with use of a multifunctional, low molecular weight reagent, and (iii) co-crosslinking with other neutral substances, e.g. proteins. Numerous other methods which are combinations of the ones listed or original and specific of a given support or enzyme have been devised. However, no single method and support is best for all enzymes and their applications. This is because of the widely different chemical characteristics and composition of enzymes, the different properties of substrates and products, and the different uses to which the product can be applied. Besides, all of the methods present advantages and drawbacks. Adsorption is simple, cheap and effective but frequently reversible, covalent attachment and crosslinking are effective and durable, but expensive and easily worsening the enzyme performance, and in membrane reactor-confinment, entrapment and microencapsulations diffusional problems are inherent. Consequently, as a rule the optimal immobilization conditions for a chosen enzyme and its application are found empirically by a process of trial and error in a way to ensure the highest possible retention of activity of the enzyme, its operational stability and durability. |
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