Glutaraldehyde immobilization

Polyethylenimine/glutaraldehyde immobilization Prussian blue 60a 1 5 0 - Lupu et al. [25]... 

Glutaraldehyde Immobilization of Trypsin. A sample of hydro-gel-p-amlnobenzenesulfonyl carbamate (0.1-0.2 g) was suspended in 5.0 mL pH 7 phosphate buffer solution in a 10 mL vial equipped with a stirring bar and Teflon lined screw cap. To the suspension was added an excess of 50% glutaraldehyde (0.5 mL), and the... 

Immobilization. The fixing property of PEIs has previously been discussed. Another appHcation of this property is enzyme immobilization (419). Enzymes can be bound by reactive compounds, eg, isothiocyanate (420) to the PEI skeleton, or immobilized on soHd supports, eg, cotton by adhesion with the aid of PEIs. In every case, fixing considerably simplifies the performance of enzyme-catalyzed reactions, thus faciHtating preparative work. This technique has been appHed to glutaraldehyde-sensitive enzymes (421), a-glucose transferase (422), and pectin lyase, pectin esterase, and endopolygalacturonase (423). 

Two types of immobilization are used for immobilizing glucose isomerase. The intracellular enzyme is either immobilized within the bacterial cells to produce a whole-ceU product, or the enzyme is released from the cells, recovered, and immobilized onto an inert carrier. An example of the whole-ceU process is one in which cells are dismpted by homogenization, cross-linked with glutaraldehyde, flocculated using a cationic flocculent, and extmded (42). 

In a second example, a cell—gelatin mixture is cross-linked with glutaraldehyde (43). When soluble enzyme is used for binding, the enzyme is first released from the cell, then recovered and concentrated. Examples of this type of immobilization include binding enzyme to a DEAE-ceUulose—titanium dioxide—polystyrene carrier (44) or absorbing enzyme onto alumina followed by cross-linking with glutaraldehyde (45,46). 

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. 

Fig. 17 (a) Elastin-based stimulus-responsive gold nanoparticles. Reproduced from [131] by permission of The Royal Society of Chemistry (b) Functionalization of a glass surface with ELP. In the first step, the glass surface is aminosilylated with N-2-(aminoethyl)-3-aminopropyl-trimethoxysilane, then modified with glutaraldehyde. Subsequently, the stimulus-responsive biopolymer is covalently immobilized using reductive amination. Reproduced from [132] by permission of The Royal Society of Chemistry... 

In another investigation, ELP[V5L2G3-90] with three lysines in the N-terminal region was immobihzed on a glass surface in a microreactor to enable temperature-controlled positioning of ELP fusion proteins. For this purpose, the glass surface was first functionalized with A -2-(aminoethyl)-3-aminopropyltrimethoxysilane, followed by glutaraldehyde treatment and reductive amination to immobilize the biopolymer on the surface (Fig. 17b) [132]. 

Affinity microparticles (AMPs) were obtained by cross-linking the S-layer lattice on S-layer-carrying cell wall fragments with glutaraldehyde, reducing Schiff bases with sodium borohydride, and immobilizing protein A as an IgG-specific ligand [92]. Thus, AMPs rep-... 

A semi-interpenetrated network was obtained by bulk polymerization of 2-hydroxye-thyl methacrylate incorporated in DMF treated PET films by solvent-exchange technique, followed by treatment of films in e-lectrical discharges. Heparinization was accomplished by reacting glutaraldehyde with heparin and poly(2-hydroxyethyl methacrylate) present on the surface of modified polyester films. The immobilization of heparin was indirectly evidenced by chromatographying the silylated hydrolyza-tes of heparinized PET films and heparin, respectively. In vitro experiments demonstrated the enhanced thromboresistance of heparinized films. 

Purified MeHNL was crystallized by the sitting-drop vapor-diffusion method. The 10-20 mm bipyramidal crystals formed were cross-linked with glutaraldehyde and used as biocatalyst for the synthesis of optically active cyanohydrins. The cross-linked crystals were more stable than Celite-immobilized enzymes when incubated in organic solvents, especially in polar solvents. After six consecutive batch reactions in dibutyl ether, the remaining activity of the cross-linked crystals was more than 70 times higher than for the immobilized enzymes. Nevertheless, the specific activity of the cross-linked crystals per milligram protein was reduced compared with the activity of Celite-immobilized enzymes [53],... 

Corynebacterium glutamicum (CGMCC No. 1464) cells immobilized in calcium alginate beads cross-linked with polyethenimine and glutaraldehyde have been employed for the production of nicotinamide from 3-cyanopyridine [21], The reaction was mn at 10-15 °C,... 

A thermally stable NHase from Comamonas testosteroni 5-MGAM-4D (ATCC 55 744) [22] was recombinantly expressed in Escherichia coli, and the resulting transformant cells immobilized in alginate beads that were subsequently chemically cross-linked with glutaraldehyde and polyethylenimine. This immobilized cell catalyst (at 0.5 % dew per reaction volume) was added to an aqueous reaction mixture containing 32wt% 3-cyanopyridine at 25 °C, and a quantitative conversion to nicotinamide was obtained. The versatility of this catalyst system was further illustrated by a systematic study of substrates, which included... 

Scheme 1 Covalent enzyme immobilization Aminopropyl-modified silica reacts with glutaraldehyde and the N-terminus of the enzyme.

In view of the conductive and electrocatalytic features of carbon nanotubes (CNTs), AChE and choline oxidases (COx) have been covalently coimmobilized on multiwall carbon nanotubes (MWNTs) for the preparation of an organophosphorus pesticide (OP) biosensor [40, 41], Another OP biosensor has also been constructed by adsorption of AChE on MWNTs modified thick film [8], More recently AChE has been covalently linked with MWNTs doped glutaraldehyde cross-linked chitosan composite film [11], in which biopolymer chitosan provides biocompatible nature to the enzyme and MWNTs improve the conductive nature of chitosan. Even though these enzyme immobilization techniques have been reported in the last three decades, no method can be commonly used for all the enzymes by retaining their complete activity. 

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