Polyacrylamide matrix

The entrapment method is based on confining the enzyme within the lattice of a polymeric matrix. Polyacrylamide gels have successfully yielded stable enzyme films with a high retention of activity... 

Crosslinking or entrapment Entrapment in dextran matrix, polyacrylamide or DNA crosslinked by using some agents such as glutar aldehyde [43] ... 

Entrapment in a polymer matrix (polyacrylamide, alginate, fc-carrageenan, polyurethane). 

The bacteria cells are suspended in a matrix polyacrylamide gel, and the reacting species must diffuse in and out of the matrix. The diffusivities of the substrate (fumaric acid) and product (L-aspartic acid) in the gel decrease as their concentrations increase due to the tendency of the gel to shrink at low pH. 

Both the ease of use of this method for characterization of proteins and nucleic acids, and the abiHty to analyze many samples simultaneously for comparative purposes, have led to the prevalence of this technique. The drawbacks of a polyacrylamide matrix is that acrylamide is a neurotoxin, the reagents must be combined extremely carefiiUy, and the gels are not as pHable as most agarose gels. 

Blotting techniques may be used in a variety and combination of electrophoretic systems which makes their use widespread and convenient when protein concentrations are minimal and agarose or polyacrylamide is the matrix choice. 

Generally, electrophoresis is carried out not in free solution but in a porous support matrix such as polyacrylamide or agarose, which retards the movement of molecules according to their dimensions relative to the size of the pores in the matrix. 

Polymetric matrix Polydiallyldimethylammonium bromide [9] Polypyrrole [10[ Poly (MA -dimethyb-S -pyrrolidinium bromide [11J Styrene-divinyl benzene copolymers [4] Polyacrylamide [12]... 

Widhalm et al. (1991) reported the use of noncrosslinked polyacrylamide for protein separation in fused silica capillaries. This matrix has low viscosity and can be replaced between separations, greatly facilitating automation of the separation. A wide range of noncrosslinked polymers has been used for size-based protein separations. Noncrosslinked polymers do not form a gel, and it is inappropriate to refer to this separation as gel electrophoresis. A number of names have been used for the method. In an effort to standardize nomenclature, IUPAC has used the term capillary sieving electrophoresis. 

This process involves the suspension of the biocatalyst in a monomer solution which is polymerized, and the enzymes are entrapped within the polymer lattice during the crosslinking process. This method differs from the covalent binding that the enzyme itself does not bind to the gel matrix. Due to the size of the biomolecule it will not diffuse out of the polymer network but small substrate or product molecules can transfer across or within it to ensure the continuous transformation. For sensing purposes, the polymer matrix can be formed directly on the surface of the fiber, or polymerized onto a transparent support (for instance, glass) that is then coupled to the fiber. The most popular matrices include polyacrylamide (Figure 5), silicone rubber, poly(vinyl alcohol), starch and polyurethane. 

Enzymes can be immobilized by matrix entrapment, by microencapsulation, by physical or ionic adsorption, by covalent binding to organic or inorganic polymer-carriers, or by whole cell immobilization (5 ). Particularly impressive is the great number of chemical reactions developed for the covalent binding of enzymes to inorganic carriers such as glass, to natural polymers such as cellulose or Sepharose, and to synthetic polymers such as nylon, polyacrylamide, and other vinyl polymers and... 

One of the extensively used synthetic polymers used as a support for immobilization of biocatalysts is polyacrylamide (PAAm) [287,288], The major advantage is that it can be polymerized either chemically or by using radiation. Advantages of y-ray polymerization against chemical polymerization is that the polymerization can be carried out even under frozen conditions thus allowing the matrix to be molded to any form such as beads or membranes [289-291], However one of the major drawbacks of this polymer especially in a membranous form is its brittleness. 

A variety of techniques has been used varying from simple adsorption to entrapment within a polymer matrix but the commonest method is the covalent linking of the ligand to a suitable polymer, such as dextran, agarose or polyacrylamide. 

Figure 8.2 Polyacrylamide gel formation and hydrolysis of acrylamide to acrylate. (A) Acrylamide and A,A-methylenebisacrylamide (bis) are copolymerized in a reaction catalyzed by ammonium persulfate [(NH4)2S208] and TEMED. (B) A very short stretch of cross-linked polyacrylamide is represented. Cross-linking between similar structures leads to the formation of ropelike bundles of polyacrylamide that are themselves cross-linked together forming the gel matrix. In the lower portion of (B) is shown how pendant, neutral carboxamide groups can become hydrolyzed to charged carboxyls.

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