Enzyme reference materials application

The requirements for, preparation of, and application of enzyme reference materials have been discussed extensively (134, 135, 151, 152, 153, 154, 155, 156, 157, 158). Cooperation among clinical enzvmologists in Europe and the United States over the last several years has resulted in the availability of only a few enzyme reference materials. SRM 8430 from NIST is a preparation of human erythrocyte aspartate aminotransferase in a human albumin matrix (144) certified reference material (CRM) 319 from the Community Bureau of Reference (BCR) of the Commission of the European Communities is a preparation of porcine y-glutamvltransferase in a bovine serum matrix (159). The working group of the BCR is in the process of establishing protocols and evaluating... 

There are also RMs which are prepared for a specific application and are used for validation of relevant methods. Cobbaert et al. (1999) made use of Ion Selective Electrode (ISE)-protein-based materials when evaluating a procedure which used an electrode with an enzyme-linked biosensor to determine glucose and lactate in blood. Chance et al. (1999) are involved with the diagnosis of inherited disorders in newborn children and they prepared a series of reference materials consisting of blood spotted onto filter paper and dried, from which amino-acids can be eluted and... 

Use of the combination of reference methodology and materials to convert incompatible enzyme activity results for aspartate aminotransferase (146), alkaline phosphatase (147), and lactate dehydrogenase (148) to compatible values by use of a single scale, termed by the authors the International Clinical Enzyme Scale (ICES) has been suggested (149). Application of the ICES concept to the 1970 Scandinavian interlaboratory surv ey decreased the interlaboratory coefficient of variation from 38% to 16% for the enzymes tested. Similarly, in the 1971 New York State aspartate aminotransferase survev, the interlaboratory CV decreased from 41 %-44% to 2%-5%, a major improvement. The Scandinavian Committee on Enzymes has expressed serious concerns about the philosophy of the ICES approach, and they again endorsed the widely accepted approach of ongoing development of reference methodologies and proper use of reference materials (150). Since this flurry of activity in 1984 and 1985, there has not been any further application or acceptance of the ICES concept. 

Biopolymers are polymers formed in nature during the growth cycles of all organisms hence, they are also referred to as natural polymers. The biopolymers of interest in this review are those that serve in nature as either structural or reserve cellular materials. Their syntheses always involve enzyme-catalyzed, chain-growth polymerization reactions of activated monomers, which are generally formed within the cells by complex metabolic processes. The most prevalent structural and reserve biopolymers are the polysaccharides, of which many different types exist, but several other more limited types of polymers exist in nature which serve these roles and are of particular interest for materials applications. The latter include the polyesters and proteins produced by bacteria and the hydrocarbon elastomers produced by plants (e.g. natural rubber). In almost all cases (natural rubber is an exception), all of the repeating units of these biopolymers contain one or more chiral centers and the repeating units are always present in optically pure form that is, biopolymers with asymmetric centers are always 100% isotactic. 

Industrial applications represent more than 80% of the global market of enzymes. A distinction should be made between those cases in which the enzymatic conversion of the raw material into the product is the key operation and those in which the enzyme is used as an additive to modify certain functional property of the product. In the first case the enzymatic reaction is carried out in a controlled environment at optimized conditions with respect to the catalytic potential of the enzyme, while in the second case conditions for enzyme action are not specified to optimize its activity and sometimes not even controlled. Examples of the first case are the production of high-fructose syrups with immobilized glucose isomerase and the production of 6-aminopenicillanic acid from penicillin G with immobilized penicillin acylase examples of the second case are the use of fungal proteases in dough making and the use of pancreatin in leather bating. Most conventional uses of enzymes refer to... 

Enzyme Immunoassays. Liquid Chromatography Size-Exclusion Affinity Chromatography Clinical Applications. Peptides. Proteins Physiological Samples. Quality Assurance Reference Materials. 

The most fundamental classification of polymers is whether they are synthetic or naturally-occurring. Common synthetic polymers (Figure 5.1) are pervasive in commercial applications for example. Table 5.1 lists some synthetic polymers used for automotive applications. In contrast, natural polymers include macromolecules such as polysaccharides (e.g., starches, sugars, cellulose, gums, etc.), proteins e.g., enzymes), fibers (e.g., wool, silk, cotton), polyisoprenes (e.g., natural rubber), and nucleic acids (e.g., RNA, DNA). Accordingly, these polymer classes are often referred to as biopolymers, of which some recent materials applications will be discussed later in this chapter. 

Characterisation of the enzyme-responsive material prepared through the methods outlined above is essential not only to test the enzyme responsiveness of the polymer but also to characterise the material s overall performance under the conditions in which it will be used in its ultimate application. This section will provide a brief overview of standard and specialised techniques that have been employed for this purpose and covers mechanical, chemical, physical and biological properties as well as enzyme responsiveness. While we will discuss the reason for choosing particular techniques and explain their advantages and Umitations in the context of the analysis of enzyme-responsive materials, explanations of the working principles of the techniques will not be provided and the reader is referred to other specialised textbooks on this topic instead. 

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