Enzymatic immobilized enzymes

The second most important group of immobilized enzymes is stiU the penicillin G and V acylases. These are used in the pharmaceutical industry to make the intermediate 6-aminopenici11anic acid [551-16-6] (6-APA), which in turn is used to manufacture semisynthetic penicillins, in particular ampicilHn [69-53-4] and amoxicillin [26787-78-0]. This is a remarkable example of how a complex chemical synthesis can be replaced with a simple enzymatic one ... 

Since many years, pectolytic enzymes have been widely used in industrial beverage processing to improve either the quality and the yields in fruit juice extraction or the characteristics of the final product [1,2]. To this purpose, complex enzymatic mixtures, containing several pectolytic enzymes and often also cellulose, hemicellulose and ligninolytic activities, are usually employed in the free form. The interactions among enzymes, substrates and other components of fruit juice make the system very difficult to be investigated and only few publications are devoted to the study of enzymatic pools [3-5], An effective alternative way to carry out the depectinisation process is represented by the use of immobilized enzymes. This approach allows for a facile and efficient enzymatic reaction control to be achieved. In fact, it is possible to avoid or at least to reduce the level of extraneous substances originating from the raw pectinases in the final product. In addition, continuous processes can be set up. 

The high specificity required for the analysis of physiological fluids often necessitates the incorporation of permselective membranes between the sample and the sensor. A typical configuration is presented in Fig. 7, where the membrane system comprises three distinct layers. The outer membrane. A, which encounters the sample solution is indicated by the dashed lines. It most commonly serves to eliminate high molecular weight interferences, such as other enzymes and proteins. The substrate, S, and other small molecules are allowed to enter the enzyme layer, B, which typically consist of a gelatinous material or a porous solid support. The immobilized enzyme catalyzes the conversion of substrate, S, to product, P. The substrate, product or a cofactor may be the species detected electrochemically. In many cases the electrochemical sensor may be prone to interferences and a permselective membrane, C, is required. The response time and sensitivity of the enzyme electrode will depend on the rate of permeation through layers A, B and C the kinetics of enzymatic conversion as well as the charac-... 

In another approach, the alcohol moiety, formed by an enzymatic hydrolysis of an ester, can act as a nucleophile. In their synthesis of pityol (8-37a), a pheromone of the elm bark beetle, Faber and coworkers [17] used an enzyme-triggered reaction of the diastereomeric mixture of ( )-epoxy ester 8-35 employing an immobilized enzyme preparation (Novo SP 409) or whole lyophilized cells of Rhodococcus erythro-polis NCIMB 11540 (Scheme 8.9). As an intermediate, the enantiopure alcohol 8-36 is formed via kinetic resolution as a mixture ofdiastereomers, which leads to the diastereomeric THF derivatives pityol (8-37a) and 8-37b as a separable mixture with a... 

Membrane-Enclosed Enzymatic Catalysis (MEEC) has been developed as a useful, practical new method for the manipulation of enzymes in organic synthesis. The enzyme in soluble form is enclosed in commercially available dialysis membranes. It combines the simplicity of use of soluble enzymes with certain of the advantages of immobilized enzymes. Containment permits separation of the enzyme from the reaction medium, straightforward separation of the product, and recovery of the enzyme for reuse [53],... 

Osborne PC, Yamamoto K. 1998. Disposable, enzymatically modified printed film carbon electrodes for use in the high-performance liquid chromatographic-electrochemical detection of glucose or hydrogen peroxide from immobilized enzyme reactors. J Chrom B 707 3-8. 

Xu, S., Pan, C., Hu, L., Zhang, Y, Guo, Z., Li, X., Zou, H. Enzymatic reaction of the immobilized enzyme on porous silicon studied by matrix assisted laser desorption/ionizationtime of flight-mass spectrometry. Electrophoresis 2004, 25, 3669-3676. 

Enzymes can be used in several ways in chromatographic applications to improve selectivity or to enhance the detector response. Applications may involve enzymes with either a broad specificity toward a group of related compounds or a high specificity toward a particular compound. In the field of drug residue analysis, most current applications concern enzymatic reactions taking place in separate reactors incorporated in LC systems before or after the analytical column. Reactors with immobilized enzymes have proven to be suitable in such continuous flow systems. 

The wide variety of enzymes available gives for promise enzymatic derivatization to become a potent analytical tool in the future. Better understanding and theoretical formulations will lead to commercial availability of immobilized enzymes and consequently to more ready use of them. Since in such systems a low content of organic cosolvent in the mobile phase can only be tolerated (whereas a compromise has to be made as far as the optimum mobile phase pH is concerned), artificial enzymes, which are synthetic polymer chains having functional groups that mimic the biocatalytic activity of natural enzymes, are currently being synthesized and investigated as a means to overcome such limitations (276). 

Since all proteins contain both of these reactive groups, if there were a possibility of producing a polyurethane, and particularly a reticulated polyurethane, with excess isocyanate groups, it would be possible to produce an enzymatically active surface on a high-surface-area, high-void-volume reticulated struemre. This is possible and in fact is easier than the most common methods used currently to immobilize enzymes. 

A renewed interest in this research field may lead to the construction of functional immobilized biocatalysts that surpass the conventional definition, or usually credited advantages, of immobilized biocatalysts with regard to their capabilities as catalysts [22-24], i.e. immobilized enzyme systems in which, for example, an enzymatic process can be controlled by externally applied stimuli such as light, electric fields, pH, temperature, and mechanical force. In such cases, what is crucial in system construction is not to rely on a possible... 

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