Choline esterase, immobilization

In 1989, to the best of our knowledge, the first report appeared on the use of an avidin-biotin system for the immobilization of enzymes in the preparation of biosensors. Walt et al. have immobilized biotin-modified enzymes (urease, esterase, and penicillinase) on the surface of biotin-modified optical fiber using avidin as a binder (Figure 5a). They have demonstrated the general use of this procedure in immobilizing several types of enzymes. At nearly the same time, Gunaratna and Wilson used an enzyme column in which choline esterase and choline oxidase were immobilized through avidin-biotin complexation for the determination of acetyl-... 

Figure 27.18 Common configuration for postcolumn reactors with electrochemical analysis. (A) LC-chemical reaction-EC. Postcolumn addition of a chemical reagent (for example, Cu2+ or an enzyme). (B) LC-enzyme-LC. Electrochemical detection following postcolumn reaction with an immobilized enzyme or other catalyst (for example, dehydrogenase or choline esterase). (C) LC-EC-EC. Electrochemical generation of a derivatizing reagent. The response at the second electrode is proportional to analyte concentration (for example, production of Br2 for detection of thioethers). (D) LC-EC-EC. Electrochemical derivatization of an analyte. In this case a compound of a more favorable redox potential is produced and detected at the second electrode (for example, detection of reduced disulfides by the catalytic oxidation of Hg). (E) LC-hv-EC. Photochemical reaction of an analyte to produce a species that is electrochemically active (for example, detection of nitro compounds and phenylalanine). Various combinations of these five arrangements have also been used. [Reprinted with permission from Bioanalytical Systems, Inc.]...

Assay of enzyme inhibitors. In 1965, Guilbault and co-workers [252] proposed an electrochemical apparatus for monitoring enzyme activity in urethane pads. The immobilized choline esterase hydrolyzed butyryl (or acetyl) thiocholine iodide (Eq, (57)) ... 

If an inhibitor is present in air or water then the activity of choline esterase decreases and the degree of activity loss is related to the concentration of the inhibitor (e.g., pesticides). If the inhibition is reversible then the inhibitor is washed out, if it is irreversible then the pad with the immobilized enzyme must be exchanged. An improved system was later commercialized by Midwest Research, Kansas City, Missouri, USA [253]. 

Some 1-carbonyl-3-methyl-2-pyrazoline-4,5-dione-4-arylhydrazones are active against Trichinella spiral is. 3 Against microfilarial infections, 2-ethyl-6-methyl-2,3,4,4a,5,6,7,8-octahydro-lH-pyrazino(l,2-c)pyrimidine-l-one seems to be very promising.0,0-Dimethyl-2,2,2-trichloro-l-hydroxyethylphosphonate, a choline esterase inhibitor, immobilizes the adult Onchocerca volvulus and consequently is useful in treating onchocerciasis. 1 8... 

The same group reported in 1986 a sensitive and selective HPLC method employing CL detection utilizing immobilized enzymes for simultaneous determination of acetylcholine and choline [187], Both compounds were separated on a reversed-phase column, passed through an immobilized enzyme column (acetylcholine esterase and choline oxidase), and converted to hydrogen peroxide, which was subsequently detected by the PO-CL reaction. In this period, other advances in this area were carried out such as the combination of solid-state PO CL detection and postcolumn chemical reaction systems in LC [188] or the development of a new low-dispersion system for narrow-bore LC [189],... 

Multienzyme systems have been used in carbon paste electrodes, providing bio-catalytic cascades that result in an analytical amperometric signal. For example, acetylcholine esterase (AChE) and choline oxidase (ChOx) have been co-immobilized in carbon pastes, either with monomeric TTF [145] or flexible ferrocene-containing polymers [146] as electron mediators. The primary reaction includes the hydrolysis of acetylcholine biocatalyzed by AChE, then the choline produced is oxidized by the electrically contacted ChOx giving an analytical amperometric signal corresponding to the acetylcholine concentration. 

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