Enzyme-immobilized capillary

Keywords MonolithPorous polymer, Separation, HPLC, Capillary electrochromatography, Enzyme immobilization, Modification... 

Smirnova et al. demonstrated the determination of the insecticide, carbaryl, using a two-chip system. The first chip (for the hydrolysis of carbaryl) had a simple Y-shaped channel while the second chip (for the diazo couphng reaction between hydrolyzed products and 2,4,6-trimethylaniline)—the extraction required special channel shapes with a partial surface— modification obtained by using capillary-restricted modification (CARM) (Figure 35.11). " Determination of carbaryl pesticide in water with sufficient sensitivity was carried out with an analysis time of 8 min. In a similar manner, Honda et al. developed a combination of a tube-type enzyme-immobilized microreactor and a microextractor with partial surface modification to produce optically pure amino acids. 

See also Capillary Electrophoresis Food Chemistry Applications. Chiroptical Analysis. Enzymes Immobilized Enzymes Enzyme-Based Electrodes. Extraction Solvent Extraction Principles. Flow Injection Analysis Principles. lon-SelectIve Electrodes Food Applications. Liquid Chromatography Food Applications. Spectrophotometry Organic Compounds. 

Whole cells or enzymes immobilized in capillary membrane reactors... 

Edwards et al. (1999a) reported that during removal of polyphenols in an enzyme-immobilized membrane reactor, in order to remove the colored quinone-type by-products, which passed through the membrane, a chitosan-packed post-adsorption column was required to be integrated into the system. In another study Edwards et al. (1999b) utilized chito-san gel as an immobilization matrix for polyphenol oxidase on polysul-fone capillary membranes. They reported that during the treatment of... 

To fulfill such requirements, attempts have been made in the past decade by researchers working on peptide mapping and proteomics through development of immobilized microfluidic enzymatic reactors. Microfluidic enzymatic microreactors are an alternative to in-solution method employing immobilization of proteases on microchaimels of chip-based reactors or surfaces of capillaries. The microreactors that enable proteolytic digestion by enzymes immobilized on solid supports are also referred to as immobilized enzyme reactors, IMERs. The great potential of IMERS for proteomic applications comprise rapid and enhance... 

Immobilization of Enzyme in Capillary Microreactor IMER systems in capillary microreactors are now routinely being used in protein digestion for peptide mapping and proteomics. Similar to microfluidic charmels on chips, enzyme immobilization in capillary can be classified into two categories based on the immobilization location. 

Scheme 10.12 Scheme of the surface modification and enzyme immobilization in fused silica capillaries [119], Source Copyright 2008 Elsevier. 

Immobilization of Enzyme in Capillary Microreactor by Entrapment Entrapment technique involves the entrapment of enzymes in gel matrix. The enzyme is mixed with gel formation ingredients and upon gel formation, the enzyme remains trapped in the matrix. Another form of entrapment is the formation of membrane around the droplet of enzyme, which is typically in solution. Immobilization by entrapment differs from adsorption and covalent bonding in that enzymes are free in solution but restricted in movement by the lattice structure of a gel. The membrane must be permeable to diffusion of substrate and product molecules... 

Another category of enzymatic transformations in multiphase systems is enzymes immobilized on the reactor wall as presented in Table 10.4. Enzymes are advantageously used in immobilized form because this strategy allows for increased volumetric productivity and improves stability. Continuous mode of operation is employed in these systems. The approaches commonly used for immobilization in conventional multiphase biocatalysis can also be employed in microreactors such as covalent methods, cross-linked enzyme aggregates (CLEA), and adsorption methods. The experimental setups can either be chip-type reactors with activated charmel surface walls where enzyme binds, or enzyme immobilized monolith reactors, where a support is packed inside a capillary tube. 

Biocatalytic reactions performed using immobilized enzyme microreactors under continuous flow mode have been found effective for hydrolysis reactions [121,158-161], with the enzyme either trapped in the matrix [159], covalently linked to modified surface wall [160,121], enzymes entrapped in hydrogels [162], or enzymes immobilized on monolith [179]. The experimental setup consists of either chip-type microreactors with activated chaimel walls where enzymes bind, enzymes that bind to beads, enzymes entrapped in the matrix, enzymes adsorbed in nanoporous materials, and most recently, nanosprings as supports for immobilized enzymes in chip-based reactors, or enzyme immobilized monolith reactors, where support is packed inside a capillary tube (Table 10.4). 

HOLLOW-FIBER MEMBRANES. A hollow-fiher membrane is a capillary having an inside diameter of - inn and an outside diameter < I mm and whose wall functions as a semipermeahlc membrane. The fibers can he employed singly or grouped into a bundle which may contain tens of thousands of fibers and up to several million libers as in reverse osmosis (Fig. 11. In most eases, hollow fibers are used as cylindrical membranes that permit selective exchange of materials across (heir walls. However, they can also he used as containers to effect the controlled release of a specific material, or as reactors to chemically modify a permeate as il diffuses through a chemically activated hollow-liher wall. e g., loaded with immobilized enzyme. 

Several techniques have been developed for the determination of purine and pyrimidine derivatives in food sample and in particular for hypoxanthine quantification biosensors (220-223) and electrochemical methods making use of immobilized enzyme electrode (224 -227), electrochemical enzymatic-based HA methods (228,229), enzyme reaction with fluorimetric detection (230), radioimmunoassay (231), colorimetric methods (232), capillary electrophoresis (233), and TLC (234). Many HPLC methods have also been developed and are reported in Table 4 (235-247) the most recent ones are described next. 

Alternative protocols for covalent coupling were described by Wu et al. [48], Sims et al. [49], and McKie et al. [50]. In the latter approach, single-stranded DNA was linked in a multistep procedure with a short primer covalently coupled to the antibody. The single-stranded DNA-marker also included a Hind III restriction site. By adding the restriction enzyme previous to the PCR-step, the DNA-marker sequence was released from the immobilized immuno-complex to the supernatant liquid phase and subsequently transferred to capillary vessels (see Section 2.2.3). 

Henares et al. [210] developed a multiple enzyme linked immunosorbent assay (ELISA) chip by using capillary-assembled microchip (CAs-CHIP), which involved simple embedding of a 2 to 3 mm length of square capillaries possessing valving and immunoreaction functions into the microchannels fabricated on a PDMS substrate. The authors immobilized different anti-IgGs... 

The basic hydrodynamic equations are the Navier-Stokes equations [51]. These equations are listed in their general form in Appendix C. The combination of these equations, for example, with Darcy s law, the fluid flow in crossflow filtration in tubular or capillary membranes can be described [52]. In most cases of enzyme or microbial membrane reactors where enzymes are immobilized within the membrane matrix or in a thin layer at the matrix/shell interface or the live cells are inoculated into the shell, a cake layer is not formed on the membrane surface. The concentration-polarization layer can exist but this layer does not alter the value of the convective velocity. Several studies have modeled the convective-flow profiles in a hollow-fiber and/or flat-sheet membranes [11, 35, 44, 53-56]. Bruining [44] gives a general description of flows and pressures for enzyme membrane reactor. Three main modes... 

Kawakami et al. [430] covalently immobilized protease P (from Aspergillus mdleus) onto a capillary (PEEK) monolith with an inner diameter varying between 0.1 and 2.0 mm using an in situ sol-gel method. The enzyme protease P can be used for transesterification. To determine the reaction rate of the system, (S)-glyddol and vinyl n-butyrate (Scheme 4.105) were used as substrates to be converted by the enzyme into... 

Acetylcholineesterase and choline oxidase Carbon-fiber micro-electrode for determination of ACh and choline, mounted in a glass capillary tube. Enzyme was immobilized on the surface of the carbon fiber with albumin. Electrode was dip-coated with Nafion. Evaluation of selectivity and dynamic range, at a fixed potential of 1.2 V versus Ag-AgCl. Calibration graph for ACh and Ch was rectilinear between 0.1 and 3mM. Interference from ascorbic acid was not observed in the range 0.1 to 0.3 mM. [79]... 

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