Entrapment immobilization

The aim of this chapter is to give a detailed overview of the characterization of biocatalysts and the development of membrane bioreactors, in particular, the main aspects of biocatalyst kinetics and their immobilization/ entrapment, either within the porous membrane structure, or on its surface. Thansport models that can help to predict the behaviour of membrane bioreactors, as well as the most relevant theoretical models and operating parameters, are presented below. This data is then analysed in order to ascertain how to improve effectiveness and productivity of the membrane bioreactors. Some relevant fields of application are also discussed in order to show the potential of such systems. 

The entrapment method, initially invented by Desai and Hubbell, appears promising to modify biomaterial surfaces to improve their hydrophilicity [61,62]. With this method, the biomaterial surface can be non-covalently but stably modified with water-soluble polymers. The sketch map of this method is presented in Fig. 7.6. A PLA sample is immersed in an ECM solution in a solvent mixture (e.g., a good solvent of PLA and a non-solvent of PLA). Then PLA is moved to a non-solvent of the scaffold material. The ECM is successfully immobilized (entrapped) on the surfaces of PLA. The immersion of PLA in the ECM solution in the solvent mixture makes the PLA surfaces swelled to a certain extent, allowing the ECM molecules to at least partially penetrate into the swelled pore surfaces. The immersion in the non-solvent afterwards makes the porous PLA surfaces solidified, resulting in the entrapment of the ECM molecules... 

Because enzymes can be intraceUularly associated with cell membranes, whole microbial cells, viable or nonviable, can be used to exploit the activity of one or more types of enzyme and cofactor regeneration, eg, alcohol production from sugar with yeast cells. Viable cells may be further stabilized by entrapment in aqueous gel beads or attached to the surface of spherical particles. Otherwise cells are usually homogenized and cross-linked with glutaraldehyde [111-30-8] to form an insoluble yet penetrable matrix. This is the method upon which the principal industrial appHcations of immobilized enzymes is based. 

Other immobilization methods are based on chemical and physical binding to soHd supports, eg, polysaccharides, polymers, glass, and other chemically and physically stable materials, which are usually modified with functional groups such as amine, carboxy, epoxy, phenyl, or alkane to enable covalent coupling to amino acid side chains on the enzyme surface. These supports may be macroporous, with pore diameters in the range 30—300 nm, to facihtate accommodation of enzyme within a support particle. Ionic and nonionic adsorption to macroporous supports is a gentle, simple, and often efficient method. Use of powdered enzyme, or enzyme precipitated on inert supports, may be adequate for use in nonaqueous media. Entrapment in polysaccharide/polymer gels is used for both cells and isolated enzymes. 

The success of the enzyme electrode depends, in part, on the immobilization of the enzyme layer. The objective is to provide intimate contact between the enzyme and the sensing surface while maintaining (and even improving) the enzyme stability. Several physical and chemical schemes can thus be used to immobilize the enzyme onto the electrode. The simplest approach is to entrap a solution of the... 

Adsorption on solid matrices, which improves (at optimal protein/support ratios) enzyme dispersion, reduces diffusion limitations and favors substrate access to individual enzyme molecules. Immobilized lipases with excellent activity and stability were obtained by entrapping the enzymes in hydrophobic sol-gel materials [20]. Finally, in order to minimize substrate diffusion limitations and maximize enzyme dispersion, various approaches have been attempted to solubilize the biocatalysts in organic solvents. The most widespread method is the one based on the covalent linking of the amphiphilic polymer polyethylene glycol (PEG) to enzyme molecules [21]. 

All these immobilization techniques run the risk of altering activity compared with the native enzyme. Improved activity is occasionally reported, but this is the exception. The immobilization techniques listed above are in approximate order of loss in activity. Physical entrapment normally causes no change. Adsorption will distort the shape of the molecule compared with the native... 

Like enzymes, whole cells are sometime immobilized by attachment to a surface or by entrapment within a carrier material. One motivation for this is similar to the motivation for using biomass recycle in a continuous process. The cells are grown under optimal conditions for cell growth but are used at conditions optimized for transformation of substrate. A great variety of reactor types have been proposed including packed beds, fluidized and spouted beds, and air-lift reactors. A semicommercial process for beer used an air-lift reactor to achieve reaction times of 1 day compared with 5-7 days for the normal batch process. Unfortunately, the beer suffered from a mismatched flavour profile that was attributed to mass transfer limitations. 

FIG. 18 Schematic drawing of a liposome with entrapped functional molecules, coated with an S-layer lattice, that can be used as immobilization matrix for functional molecules. Alternatively, liposomes can be coated with genetically modified S-layer subunits incorporating functional domains. (Modified from Ref. 59.) (b) Electron micrograph of a freeze-etched preparation of an S-layer-coated liposome (bar, 100 nm). 

Dynamic light-scattering experiments or the analysis of some physicochemical properties have shown that finite amounts of formamide, A-methylformamide, AA-dimethyl-formamide, ethylene glycol, glycerol, acetonitrile, methanol, and 1,2 propanediol can be entrapped within the micellar core of AOT-reversed micelles [33-36], The encapsulation of formamide and A-methylformamide nanoclusters in AOT-reversed micelles involves a significant breakage of the H-bond network characterizing their structure in the pure state. Moreover, from solvation dynamics measurements it was deduced that the intramicellar formamide is nearly completely immobilized [34,35],... 

Solutions of surfactant-stabilized nanogels share both the advantage of gels (drastic reduction of molecular diffusion and of internal dynamics of solubilizates entrapped in the micellar aggregates) and of nonviscous liquids (nanogel-containing reversed micelles diffuse and are dispersed in a macroscopicaUy nonviscous medium). Effects on the lifetime of excited species and on the catalytic activity and stability of immobilized enzymes can be expected. 

Bioencapsulation is a technology aimed toward the immobilization and incorporation (entrapping) of a biologically active compound on or inside solid particles (microspheres) or liquid vesicles in order to stabilize, stracture, and protect the active compound and allow control of its release. 

Immobilization is the technique of choice in many food industry processes and especially in beverage production. Many immobilization technologies have already been tested and some are applied in the production of beer, wine, vinegar, and other food products using a traditional approach with cultme adhesion (i.e., Acetobacter in vinegar production) or more modem approaches with entrapment of yeast biomass (i.e., sparkling wines, cheeses, and yogurts). 

Prepolymer method for entrapment of biocatalyst developed by Fukui and Tanaka is considered to be one of the best immobilization methods. Characteristic features and advantages of the prepolymer method can be summarized as follows ... 

Immobilized spores Entrapment in Caalginate/ Methyl ketone from Isoparaffin-water Improvement in catalyst 20... 

Solidification and stabilization processes. These immobilize the contaminants through physical or chemical processes. Solidification involves the entrapment of contaminants into a consolidated mass and stabilization is the conversion of contaminants to a chemical form that is less available. 

Vorlop et al. described a novel cross-linked and subsequently poly(vinyl alcohol-entrapped PaHNL for synthesis of (//(-cyanohydrins. These immobilized lens-shaped biocatalysts have a well-defined macroscopic size in the millimeter range, show no catalyst leaching, and can be recycled efficiently. Furthermore, this immobilization method is cheap and the entrapped (/ )-oxynitrilases gave similar good results compared with those of free enzymes. The (//(-cyanohydrin was obtained in good yields and with high enantioselectivity of up to >99% ee [55],... 

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