Enzyme Immobilization systems

5 Application of Nanocomposites as Nanosensor for Smart/Intelligent Packaging 

Nanotechnology-based sensors have the potential to bring about a paradigm shift in revolutionizing the speed and accuracy with which industries or regulatory agencies detect the presence of molecular contaminants or adulterants in complex food matrices with color changes that occur to metal nanoparticle solutions in the presence of analytes 

---

Immobilized system the air circulates over a film of microorganisms that grows on a solid surface. In an immobilized bioreactor, particulate biocatalysts for enzyme production and conversion of penicillin to 6-aminopenicillanic acid are used. 

Fig. 12. On-line enzyme reactor system designs, merging stream system (Top) and immobilized-enzyme reactor system (Bottom). A = mobile phase, B = enzyme solution...

Enzyme electrodes belong to the family of biosensors. These also include systems with tissue sections or immobilized microorganism suspensions playing an analogous role as immobilized enzyme layers in enzyme electrodes. While the stability of enzyme electrode systems is the most difficult problem connected with their practical application, this is still more true with the bacteria and tissue electrodes. 

Although the catalysts do not interfere with each other, the immobilization process resulted in higher yields and ees than with the raw enzyme the initial activity was over five times greater. Additionally, the biocomposite with enzyme was able to be recycled and maintained its activity, which demonstrates the utility of such an immobilization system for potential cascades with mutually interfering catalysts. 

Figure 2 Effect of enzyme immobilization on luminescent image spatial resolution evaluated using coupled enzymatic reactions on nylon net as a model system, (a) Immobilized 3a-hydroxysteroid dehydrogenase (b) immobilized 3a-hydroxysteroid dehydrogenase and FMN-NADH oxidoreductase (c) immobilized 3a-hydroxysteroid dehydrogenase, FMN-NADH oxidoreductase, and bacterial luciferase. (From Ref. 47. Copyright John Wiley Sons Ltd. Reproduced with permission.)...

The method of enzyme immobilization constitutes a key factor in the construction of these systems as it is the biocatalytic membrane that largely determines sensitivity, stability and response-time characteristics of the biosensor. 

It was reported that PEGylated lipase entrapped in PVA cryogel could be conveniently used in organic solvent biocatalysis [279], This method for enzyme immobilization is more convenient in comparison to other types of immobilization that take advantage of enzyme covalent linkage to insoluble matrix, since the chemical step which is time consuming and harmful to enzyme activity is avoided. The application of this catalytic system to the hydrolysis of acetoxycoumarins demonstrated the feasibility of proposed method in the hydrolysis products of pharmaceutical interest and to obtain regioselective enrichment of one of the two monodeacetylated derivatives. 

The general method utilized to prepare E5-Ab solutions obviates the need for stocking large numbers of reagents which would be necessary if different activation methods were used for each antibody. A number of specific antibodies immobilized by this process have shown response similar to that of the same antibodies when adsorbed as immune complexes in the Stratus system. In addition, the dendrimer-coupled antibodies have shown dramatic improvements in sensitivity, flexibility and precision for the enzyme immunoassay system. Feasibility demonstration of an assay for DNA probes is a prelude to what can possibly be achieved with these dendrimer-based reagents. 

A. Aoki, M. Ueda, H. Nakajima, and A. Tanaka, Construction of a photo controllable enzyme reaction system by co-immobilization of an enzyme and a semiconductor. Biocatalysis, 2, 89-95 (1989). 

In an early application, an enzyme electrode system was reported for the determination of creatinine and creatine, using a combination of creatinine amidohy-drolase, creatine amidinohydrolase and sarcosine oxidase, co-immobilized on an asymmetric cellulose acetate membrane. Thus, the hydrogen peroxide produced was detected to give a quantitative measure of creatine and creatinine in biological fluids [70]. 

In vitro enzymatic polymerizations have the potential for processes that are more regio-selective and stereoselective, proceed under more moderate conditions, and are more benign toward the environment than the traditional chemical processes. However, little of this potential has been realized. A major problem is that the reaction rates are slow compared to non-enzymatic processes. Enzymatic polymerizations are limited to moderate temperatures (often no higher than 50-75°C) because enzymes are denaturated and deactivated at higher temperatures. Also, the effective concentrations of enzymes in many systems are low because the enzymes are not soluble. Research efforts to address these factors include enzyme immobilization to increase enzyme stability and activity, solubilization of enzymes by association with a surfactant or covalent bonding with an appropriate compound, and genetic engineering of enzymes to tailor their catalytic activity to specific applications. 

The use of enzymes as biocatalysts for the synthesis of water-soluble conducting polymers is simple, environmentally benign, and gives yields of over 90% due to the high efficiency of the enzyme catalyst. Since the use of an enzyme solution does not allow the recovery and reuse of the expensive enzyme, well-established strategies of enzyme immobilization onto solid supports have been applied to HRP [22-30]. A recent work reported an alternative method that allows the recycle and reuse of HRP in the biocatalytic synthesis of ICPs. The method is based on the use of a biphasic catalytic system in which the enzyme is encapsulated by simple solubilization into an IL. The main strategy consisted of encapsulating the HRP in room-temperature IPs insoluble in water, and the other components of the reaction... 

Complete desialylation of erythrocytes is not necessary for erythrocyte sequestration liberation of 15-20% of membrane sialic acids was found to be sufficient for a significant diminution of the life-time of erythrocytes.481,482 For an in vitro system with rat-peritoneal macrophages (see later), this threshold value was found to be only 10% when using soluble V. cholerae sialidase, but it was —30% when the erythrocytes were treated with the same enzyme immobilized on Sepharose.488... 

The model active transport system described by Dr. Thomas is based on an asymmetric arrangement of two enzymes. A model active transport system was also described by Blumenthal et al. several years ago based on a single enzyme immobilized between asymmetric boundaries [Blumenthal, Caplan, and Kedem, Biophys. J., 7, 735 (1967)]. In the latter case the phenomenological coefficients were measured, and it was possible to demonstrate Onsager symmetry and the correlation between the thermodynamic coefficients and the kinetic constants. 

---