Enzyme potentiometric

A.P. Soldatkin, D.V. Gorchkoh, C. Martelet, and N. Jaffrezic-Renault, New enzyme potentiometric sensor for hypochlorite species detection. Sens. Actuat. B 43, 99-104 (1997). 

Caras, S. D., Petelenz, D., Janata, J., pH-based Enzyme Potentiometric Sensors. Part 2. Glucose-Sensitive Field Effect Tansistor , J. Anal. Chem. 57 (1985) 1920-1923. 

It has been reported [48] that the detection limit can be lowered significantly by the addition of approximately 5% of doped PPA into the enzyme containing layer. This effect was demonstrated on enzyme potentiometric sensors for urea and creatinine based on field-effect transistors and on ion-selective electrodes. The origin of this effect is not obvious and no explanation has been offered in the original paper. 

The catalytic hydrolysis of each molecule of these compounds releases two protons, the measurement and correlation of which to the OP concentration forms the basis of a potentiometric enzyme electrode. The basic element of this very simple enzyme potentiometric electrode is a pH electrode modified with an immobilized purified organophosphorus hydrolase (OPH) layer formed by cross-linking OPH with bovine serum albumin and glutaraldehyde. Thus, potentiometric OP biosensors were prepared by coupling OPH and a glass pH electrode. The sensors were constructed by immobilizing OPH on the surface of a pH-sensitive... 

Potcntiomctric Biosensors Potentiometric electrodes for the analysis of molecules of biochemical importance can be constructed in a fashion similar to that used for gas-sensing electrodes. The most common class of potentiometric biosensors are the so-called enzyme electrodes, in which an enzyme is trapped or immobilized at the surface of an ion-selective electrode. Reaction of the analyte with the enzyme produces a product whose concentration is monitored by the ion-selective electrode. Potentiometric biosensors have also been designed around other biologically active species, including antibodies, bacterial particles, tissue, and hormone receptors. 

Schematic diagram of an enzyme-based potentiometric biosensor for urea in which urease is trapped between two membranes.

Potentiometric electrodes also can be designed to respond to molecules by incorporating a reaction producing an ion whose concentration can be determined using a traditional ion-selective electrode. Gas-sensing electrodes, for example, include a gas-permeable membrane that isolates the ion-selective electrode from the solution containing the analyte. Diffusion of a dissolved gas across the membrane alters the composition of the inner solution in a manner that can be followed with an ion-selective electrode. Enzyme electrodes operate in the same way. 

Directions for preparing a potentiometric biosensor for penicillin are provided in this experiment. The enzyme penicillinase is immobilized in a polyacrylamide polymer formed on the surface of a glass pH electrode. The electrode shows a linear response to penicillin G over a concentration range of 10 M to 10 M. 

The sensor is an ammonium ion-selective electrode surrounded by a gel impregnated with the enzyme mease (Figme 6-11) (22). The generated ammonium ions are detected after 30-60 s to reach a steady-state potential. Alternately, the changes in the proton concentration can be probed with glass pH or other pH-sensitive electrodes. As expected for potentiometric probes, the potential is a linear function of the logarithm of the urea concentration in the sample solution. 

Instead of immobilizing the antibody onto the transducer, it is possible to use a bare (amperometric or potentiometric) electrode for probing enzyme immunoassay reactions (42). In this case, the content of the immunoassay reaction vessel is injected to an appropriate flow system containing an electrochemical detector, or the electrode can be inserted into the reaction vessel. Remarkably low (femtomolar) detection limits have been reported in connection with the use of the alkaline phosphatase label (43,44). This enzyme catalyzes the hydrolysis of phosphate esters to liberate easily oxidizable phenolic products. 

An entirely different breed of instruments has been introduced by Beckman for measuring substrate concentrations using enzymes as the reagent and specific potentiometric electrodes... 

Enzyme sensors are based primarily on the immobilization of an enzyme onto an electrode, either a metallic electrode used in amperometry (e.g., detection of the enzyme-catalyzed oxidation of glucose) or an ISE employed in potentiometry (e.g., detection of the enzyme-catalyzed liberation of hydronium or ammonium ions). The first potentiometric enzyme electrode, which appeared in 1969 due to Guilbault and Montalvo [140], was a probe for urea with immobilized urease on a glass electrode. Hill and co-workers [141] described in 1986 the second-generation biosensor using ferrocene as a mediator. This device was later marketed as the glucose pen . The development of enzyme-based sensors for the detection of glucose in blood represents a major area of biosensor research. 

The final method of coupling enzyme reactions to electrochemistry is to immobilize an enzyme directly at the electrode surface. Enzyme electrodes provide the advantages already discussed for immobilization of enzymes. In addition, the transport of enzyme product from the enzyme active site to the electrode surface is greatly enhanced when the enzyme is very near to the electrode. The concept of combining an enzyme reaction with an amperometric probe should offer all of the advantages discussed earlier for ion-selective (potentiometric) electrodes with a much higher sensitivity. In addition, since the response of amperometric electrodes is linear, background can be selected. 

Kobos, R. K. Potentiometric Enzyme Methods, in Ion-Selective Electrodes in Analytical Chemistry, Vol. 2 (Freiser, H., ed.) New York Plenum Press, 1980, p. I... 

Fig. 7. Schematic diagram of an enzyme electrode. A, B and C comprise the membrane system, while D is the detector (either amperometric or potentiometric)...

A fairly sensitive ( 10 A/) homogeneous ECIA technique for human IgG using chloroperoxidase catalyzed COj production and subsequent potentiometric detection has recently been reported A more complex scheme using enzymes and amperometric determination of H2O2 has demonstrated micromolar sensitivity... 

Uithoven K.A., Schmidt J.C., Ballman M.E., Rapid identification of biological warfare agents using an instrument employing a light addressable potentiometric sensor and a flow-through immunofiltration-enzyme assay system, Biosens. Bioelectron. 2000, 14 761-770. 

In AChE-based biosensors acetylthiocholine is commonly used as a substrate. The thiocholine produced during the catalytic reaction can be monitored using spectromet-ric, amperometric [44] (Fig. 2.2) or potentiometric methods. The enzyme activity is indirectly proportional to the pesticide concentration. La Rosa et al. [45] used 4-ami-nophenyl acetate as the enzyme substrate for a cholinesterase sensor for pesticide determination. This system allowed the determination of esterase activities via oxidation of the enzymatic product 4-aminophenol rather than the typical thiocholine. Sulfonylureas are reversible inhibitors of acetolactate synthase (ALS). By taking advantage of this inhibition mechanism ALS has been entrapped in photo cured polymer of polyvinyl alcohol bearing styrylpyridinium groups (PVA-SbQ) to prepare an amperometric biosensor for... 

P. Mulchandani, A. Mulchandani, I. Kaneva, and W. Chen, Biosensor for direct determination of orga-nophosphate nerve agents. 1. Potentiometric enzyme electrode. Biosens. Bioelectron. 14, 77-85 (1999). 

M.Y. Keating and G.A. Rechnitz, Potentiometric enzyme immunoassay for digoxin using polystyrene beads. Anal. Lett. 18, 1-10 (1985). 

T. Fonong and G.A. Rechnitz, Homogeneous potentiometric enzyme immunoassay for human immunoglobulin G. Anal. Chem. 56, 2586-2590 (1984). 

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