Cell entrapment immobilized biocatalysts

In spite of usefulness of the simplification obtained by decreasing the experimental substrate concentration, many studies are aimed at the investigation of kinetic properties of immobilized biocatalysts within broader concentration ranges. In a previous paper [29], cells of Escherichia coli with penicillin acylase activity were immobilized by entrapment in calcium pectate gel and tested on the transformation of penicillin G to 6-amino penicillanic acid. Figure 9 shows experimental data from a microcalorimetric investigation of the penicillin G transformation in steady state. As appreciable particle-mass transfer was expected, the mathematical model that includes particle-mass balance was used. 

The decreased denaturating action of the precursor and procedure enables one to immobilize reduced amounts of biomaterial. It was demonstrated in Ref. [55] that biocatalysts prepared by entrapping endo-l,3-P-D-glucanase and a-D-galactosidasc in amounts comparable to that in living cells had a reasonable level of activity. When the TEOS is applied, the enzyme content in silica matrix can be up to 20-30 wt.% to counterbalance losses due to denaturation [50]. 

Mediators can exist free in solution physically entrapped behind a membrane - immobilized in a matrix along with the biocatalyst or covalently bound to a surface or polymer network, wherein the polymer can be conductive or insulating. - Detailed discussion of the various formats is outside scope of this review paper. However, selected immobilization chemistries reported in relation to enzymatic biofuel cells are reviewed in the sections below. 

Biological catalysts in the form of enzymes, cells, organelles, or synzymes that are tethered to a fixed bed, polymer, or other insoluble carrier or entrapped by a semi-impermeable membrane . Immobilization often confers added stability, permits reuse of the biocatalyst, and allows the development of flow reactors. The mode of immobilization may produce distinct populations of biocatalyst, each exhibiting different activities within the same sample. The study of immobilized enzymes can also provide insights into the chemical basis of enzyme latency, a well-known phenomenon characterized by the limited availability of active enzyme as a consequence of immobilization and/or encapsulization. 

Enzymes or cells, which are of relatively large size may be entrapped in a maze of polymeric molecules (a gel). This procedure is called immobilization by inclusion. When the biocatalyst is enclosed inside a semiper-... 

The term encapsulation has been used to distinguish entrapment preparations in which the biocatalyst environment is comparable to that of the bulk phase and where there is no covalent attachment of the protein to the containment medium (Fig. 6-1 D)[21J. Enzymes or whole cells may be encapsulated within the interior of a microscopic semi-permeable membranes (microencapsulation) or within the interior of macroscopic hollow-fiber membranes. Liposome encapsulation, a common microscopic encapsulation technique, involves the containment of an enzyme within the interior of a spherical surfactant bilayer, usually based on a phospholipid such as lecithin. The dimensions and shape of the liposome are variable and may consist of multiple amphiphile layers. Processes in which microscopic compart-mentalization (cf. living cells) such as multienzyme systems, charge transfer systems, or processes that require a gradient in concentration have employed liposome encapsulation. This method of immobilization is also commonly used for the delivery of therapeutic proteins. 

Opportunity for innovation and creativity still exists in the field of biocatalyst immobilization. Despite the tremendous volume of biocatalyst immobilization literature, there is no one technology that is universally applicable and no one technique that can be applied using a generic procedure. The limitations of individual immobilization techniques have been pointed out in each section. Operationally simple adsorption methods often are limited by the lack of stabilization and by protein leaching, especially under aqueous conditions. Restriction of diffusion can be severe for entrapped proteins and cells. Covalent methods often result in protein inactivation and a much higher carrier cost. The combined effects of... 

Transformations with immobilized enzymes or cells Often the stability of the biocatalyst can be increased by immobilization and many different enzymes and cells have been immobilized by a variety of different methods. The most popular method for the fixation of whole cells is entrapment or encapsulation with calcium alginate. Other natural gels e.g., carrageenan, collagen, chemically-modified natural polymers e.g., cellulose acetate and synthetic gels and polymers e.g., polyacrylamide or polyhydroxyethylmethacrylate can also be used for this type of immobilization. 

Immobilized enzymes are defined as enzymes physically confined or localized in a certain defined region of space with retention of their catalytic activities, which can be used repeatedly and continuously. This definition is applicable to the enzymes as well as aU types of biocatalysts such as cellular organelles, microbial cells, plant cells, and animal cells. In some cases, these biocatalysts are bound to or within insoluble supporting materials (carriers) by chemical or physical binding. In other cases, biocatalysts are free, but confined to limited domains or spaces of supporting materials (entrapment). 

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