Biochemical Engineering Fundamentals, Part 1 - Bailey J., Ollis D. 1989

Applications of Enzyme-Catalyzed Reactions
Technological Processes Involving Immobilized Enzymes
Applications of Immobilized Enzymes in Medicine and Chemical Analysis

Currently, more than 120 inherited Metabolic Disorders in humans are known; many of these defects are associated with a lack of activity of a single specific enzyme present in healthy individuals. For example, phenylketonuria (a condition leading to intellectual disability) is believed to be caused by a deficiency in the enzyme that converts phenylalanine into Tyrosine. At present, the therapeutic approach for treating phenylketonuria comes down to a special phenylalanine-free diet. An alternative Treatment could be the administration of the missing enzyme; however, an enzyme with the same activity isolated from animal sources triggers a severe human immunological response. This problem might be solved by enclosing the enzyme within microcapsules, fibers, or a gel. It is quite probable that an enzyme immobilized in this way will no longer elicit an immune defense reaction while still allowing small substrate molecules to contact the enzyme by diffusing through the walls of the microcapsule, fiber, or gel. Membrane-protected Enzymes are not subject to antibody attack; nonetheless, the concentration of biocatalysts On the surface of natural membranes reduces the efficiency of mass transfer and, consequently, The rate of substrate utilization.

Class="center">Table 4.13. Applications of immobilized enzymes in chemical processes and their potential areas of use^a

^a Reproduced with permission from: Immobilized Enzymes. Research and Development, Chibata I. (ed.), p. 164, Kodansha, Ltd., Tokyo, 1978.

A variation of the approach outlined above formed The basis of a proposed compact artificial Kidney design. According to this concept, urease and an ion-exchange resin or activated charcoal are co-encapsulated in a microcapsule; the ammonia generated during the decomposition of urea is adsorbed inside the microcapsule:

Among the attempts to apply immobilized enzymes in small-scale production, their use in steroid transformations is noteworthy (Section 2.1.2). For instance, cortisol, which is used in the treatment of Arthritis, can be produced from the cheap precursor 11-deoxycortisol using a Column packed with immobilized 11ß-hydroxylase; cortisol can then be converted into an even more valuable drug, prednisolone, in a packed-bed Reactor containing immobilized ∆1-dehydrogenase. Note the exceptionally high Specificity of these enzymes. Currently, the vast majority of industrial-scale steroid transformations are carried out microbiologically.

Immobilized enzymes are already widely used in analytical biochemistry, and their scope of application will undoubtedly expand further in the coming years. One example is enzyme electrodes, which enable continuous monitoring of low concentrations of biochemically important substances. Thus, in a urea-sensing electrode (Fig. 4.13), immobilized urease breaks down urea into ions that can be detected by conventional electrochemical Methods. Enzyme electrodes make it possible to automate standard biochemical assays, as shown schematically in Fig. 4.14. Such an automated system can be used, for example, to determine glucose or lactate concentrations using immobilized glucose oxidase or Lactate dehydrogenase.

The designs of enzyme electrodes intended for the determination of many other biologically important compounds are based on this same principle (Table 4.14). To study enzymatic reactions in membranes, a surface fluorimetry method has also been recently developed, making it possible to directly determine the concentrations of various enzymes, substrates, and Cofactors. Readers will find extensive supplementary material providing a more detailed Structure/133.html">Discussion of this problem in the literature listed at the end of the chapter.

The USE OF IMMOBILIZED biochemical compounds in Affinity Chromatography will be discussed in Chapter 11. This method, based on the exceptionally high affinity of certain substances in solution for an immobilized compound, makes it possible to isolate, purify, and analyze Enzyme Inhibitors, cofactors, Antigens, Antibodies, and other substances.

FIG. 4.13. Electrode for urea determination; in this electrode, urease is immobilized in a gel coated on The surface of a Glass electrode.

FIG. 4.14. Diagram of an automated system for determining glucose (using immobilized glucose oxidase) or lactate (using immobilized lactate dehydrogenase). [Reproduced with permission from: Hicks G. T., Updike S. J., The Preparation and Characterization of Lyophilized Polyacrylamide Enzyme Gels for Chemical Analysis, Anal. Chem., 38, 726 (1966). © American Chemical Society.]

Table 4.14. Some compounds that can be determined using immobilized enzyme electrodes.