Amperometric biosensor

High sensitivity, selectivity, and ability to operate in turbid solutions are advantages of electrochemical biosensors. Amperometric detection is based on measuring the oxidation or reduction of an electroactive compound at a working electrode (sensor). A potentiostat is used to apply a constant potential to the working electrode with respect to a second electrode (reference electrode). A potentiostat is a simple electronic circuit that can be constructed using a battery, two operational amplifiers, and several resistors. The applied potential is an electrochemical driving force that causes the oxidation or reduction reaction. 

Thus electrochemical transducers dominate in the construction of microbial biosensors. Amperometric sensors offer greater sensitivity, since their response is linear as opposed to the logarithmic response of potentiometric sensors. This was described in detail by Mascini and Memoli [260] who compared microbial sensors based on amperometric and potentiometric electrodes with the yeast Saccharomyces for the determination of glucose. 

Amperometric or voltammetric biosensors typically rely on an enzyme system that catalyt-ically converts electrochemically non-active analytes into products that can be oxidized or reduced at a working electrode. Although these devices are the most commonly reported class of biosensors, they tend to have a small dynamic range due to saturation kinetics of the enzyme, and a large overpotential is required for oxidation of the analyte this may lead to oxidation of interfering compounds as well (e.g., ascorbate in the detection of hydrogen peroxide). In addition to the use in enzyme-based biosensors, amperometric transducers have also been used to measure enzyme-labelled tracers for affinity-based biosensor (mainly immunosensors and genosensors). Enzymes which are commonly used for this purpose include horseradish peroxidase (HRP) [17] and alkaline phosphatase (AP) [18,19,21]. 

The other type of electrochemical biosensors, amperometric biosensors, measure the current under the influence of a constant potential that is applied between the reference and working electrodes. Many types of electrodes have been used in amperometric biosensors. However, considering the glucose biosensor, the Clark electrode has... 

Bucur B, Radu GL, Toader CN (2008) Analysis of methanol-ethanol mixmres from falsified beverages using a dual biosensors amperometric systean based on alcohol dehydrogenase and alcohol oxidase. EurFood Res Technol 226(6) 1335-1342... 

Enzymes have also been used as selective biorecognition elements to determine a specific substrate in enviromnental microfiuidic biosensors. Amperometric biosensors, which have proven to have great potential since the 1980s, can be incorporated into microfiuidic devices transferring their benefits to lower dimension devices. 

Schematic showing the reactions by which an amperometric biosensor responds to glucose.

This experiment describes the use of a commercially available amperometric biosensor for glucose that utilizes the enzyme glucose oxidase. The concentration of glucose in artificial... 

Commercially available kits for monitoring blood-glucose use an amperometric biosensor incorporating the enzyme glucose oxidase. This experiment describes how such monitors can be adapted to the quantitative analysis of glucose in beverages. 

Abass and colleagues developed an amperometric biosensor for NHA that uses the enzyme glutamate dehydrogenase to catalyze the following reaction. 

Describe the major problems encountered in the detection of the NADH product of dehydrogenase-based amperometric biosensors. Discuss a common approach to circumvent these problems. 

Wang, B.Q. and Dong, S.J. (2000) Sol-gel-derived amperometric biosensor for hydrogen peroxide based on methylene green incorporated in Nafion film. Talanta, 51, 565—572. 

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... 

FIGURE 2.5 (a) Amperometric response of CNT modified screen-printed biosensor for methyl parathion in 0.1 M PBS containing 0.1 M NaCl (pH 7.4) at applied potential of 0.50 V and (b) die relative inhibition of CNT AChE/CHO biosensor as a function of methyl parathion concentration (adapted from [40]). 

MWNTs favored the detection of insecticide from 1.5 to 80 nM with a detection limit of InM at an inhibition of 10% (Fig. 2.7). Bucur et al. [58] employed two kinds of AChE, wild type Drosophila melanogaster and a mutant E69W, for the pesticide detection using flow injection analysis. Mutant AChE showed lower detection limit (1 X 10-7 M) than the wild type (1 X 10 6 M) for omethoate. An amperometric FIA biosensor was reported by immobilizing OPH on aminopropyl control pore glass beads [27], The amperometric response of the biosensor was linear up to 120 and 140 pM for paraoxon and methyl-parathion, respectively, with a detection limit of 20 nM (for both the pesticides). Neufeld et al. [59] reported a sensitive, rapid, small, and inexpensive amperometric microflow injection electrochemical biosensor for the identification and quantification of dimethyl 2,2 -dichlorovinyl phosphate (DDVP) on the spot. The electrochemical cell was made up of a screen-printed electrode covered with an enzymatic membrane and combined with a flow cell and computer-controlled potentiostat. Potassium hexacyanoferrate (III) was used as mediator to generate very sharp, rapid, and reproducible electric signals. Other reports on pesticide biosensors could be found in review [17],... 

K. Rekha, M.D. Gouda, M.S. Thakur, and N.G. Karanth, Ascorbate oxidase based amperometric biosensor for organophosphorous pesticide monitoring. Biosens. Bioelectron. 15, 499-520 (2000). 

P. Mulchandani, W. Chen, and A. Mulchandani, Flow injection amperometric enzyme biosensor for direct determination of organophosphate nerve agents. Environ. Sci. Technol. 35, 2562-2565 (2001). 

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