Electrochemical interferences

Biosensors fabricated on the Nafion and polyion-modified palladium strips are reported by C.-J. Yuan [193], They found that Nafion membrane is capable of eliminating the electrochemical interferences of oxidative species (ascorbic acid and uric acid) on the enzyme electrode. Furthermore, it can restricting the oxidized anionic interferent to adhere on its surface, thereby the fouling of the electrode was avoided. Notably, the stability of the proposed PVA-SbQ/GOD planar electrode is superior to the most commercially available membrane-covered electrodes which have a use life of about ten days only. Compared to the conventional three-dimensional electrodes the proposed planar electrode exhibits a similar... 

Figure 28.15, curve a, displays the emission spectrum. Anthracene (An) can be added to this system without electrochemical interference, yet it alters the spectrum to the anthracene-like distribution of Figure 28.15, curve b. Apparently, the exothermic energy transfer... 

For these reasons, the drawbacks of electrochemical interferences are the primary subject of many research groups involved in the biosensor field, and strategies to overcome them have become a major goal. 

For this reason, in the last decade, inorganic electrochemical mediators, which catalyse the oxidation or reduction of H202 have been preferred to HRP and have been used for the assembling of oxidase-based biosensors [18-20]. This results in a decrease of the applied potential and the consequent avoidance of electrochemical interferences. Many electrochemical mediators have been used and many of them have found broad application, especially in glucose biosensors for diabetes control. However, due to the solubility of the mediator, they are generally employed in a single-use sensor and present some problems due to the low operative stability. 

Our research in this field, which is summarised in this chapter, has been directed at obtaining a sensor modified with PB as electrochemical mediator which could avoid electrochemical interferences and could also couple the advantages of the screen-printed electrodes. For this purpose, an in-depth study of the modification procedure for PB deposition on the electrode surface was first conducted and then when an optimised procedure capable of providing an efficient and stable PB layer was obtained, it was applied with screen-printed electrodes in real analytical systems. Thus, our main goal has been not only to obtain a PB modification procedure suitable for a mass production of modified screen-printed electrodes, as already pointed out above, but also to achieve a stable PB layer in terms of operative and storage stability. 

It should also be pointed out that in the case of an in vivo measurement, the microdialysis probe will be able to recover not only glucose but also many other biological compounds with low molecular weight from the subcutaneous tissue. The electrochemical interferents are greatly reduced by the use of PB at a low applied potential. However, other biological compounds could negatively affect the stability of the enzymatic membrane. Also, it is possible to have a sort of passivation or fouling of the electrode surface due to the absorption of... 

In this application, the use of wild-type electric eel AChE and a recombinant AChE, specifically selected as very sensitive to dichlorvos, was compared. The effect of the matrix extract was determined by using various sample solvent ratios, 1 2.5, 1 5, 1 10, and 1 20. The optimal extraction ratio, considering the electrochemical interferences and the effect on enzyme activity and bioavailability of the pesticide, was 1 10. 

To perform electrochemical analysis with the choline oxidase biosensor, dichlorvos needed to be transferred to PBS solution, thus avoiding any electrochemical interference by organic solvents. 

The problem of interfering substances which are also oxidized at this potential can be overcome by differential methods using an enzyme covered electrode and a blank electrode and measuring the differential signal. Such systems are able to compensate for electrochemical interference but cannot hinder fouling of the electrodes [71]. 

For long term operation other approaches have to be used. To protect the Pt working electrodes against fouling and to prevent erroneous reading due to electrochemical interference, an electropolymerised semi-permeable membrane can be utilized [76]. 

The major problem that has plagued these kinds of implantable biosensors is the gradual decrease in sensitivity and in some cases a complete loss of function within just hours of implantation. Biofouling, oxygen limitation, electrochemical interference and GOD inactivation have been considered as explanations of this behaviour. For instance, a tissue reaction to the sensor implantation may result in a limitation in the blood supply to the tissue surrounding the probe and thus in a lower availability of glucose and oxygen. 

Their biosensor consisted in a Pt wire on which GOD was immobilized by the electropolymerization of m-phenylenediamine. The advantage of this type of immobilization consists in creating an effective barrier against electrochemical interference due to the polymer formed onto the electrode. Moreover, an extended linearity for the glucose sensor was also obtained, and this was a requisite for the direct measurement of the subcutaneous glucose at diabetic levels, since in practice an extremely low dilution of the subcutaneous fluid was realized. 

Palmisano et al. [34] developed a lactate biosensor where the LOD enzyme was electrochemically immobilized in a bilayer membrane formed from 1,2-diaminobenzene and an overoxidized polypyrrole, thus achieving an effective exclusion of electrochemical interferents. This system allowed the measurement of lactate content in untreated, undiluted milk and yoghurt samples. 

For the measurement a moderate reduction potential between — 100 and + 100 mV vs. Ag/AgCl is appUed (Fig. 2.12). In this region the potential for electrochemical interferences is very low. However, the biggest problems arise from the high reactivity of compormds I and II with reducing substrates (electron donors), which compete with the electrode for the reduction of peroxidase. Ascorbic acid, naturally occurring phenolics and aromatic amines are among those compounds. The competitive reaction of reductants should be... 

Elimination of electrochemical interferences in glucose biosensors. Trends in Analytical Chemistry TRAC, 29 (4), 306-318. 

Jtinsson and Gorton (1985) adsorbed N-methylphenazinium (NMP+) onto a spectral carbon electrode containing covalently bound GOD. Whereas the immobilized enzyme was stable for 2 months, fresh NMP+ had to be adsorbed daily. An important advantage of this sensor is the low required potential, +50 mV vs. SCE, which keeps electrochemical interferences to a minimum. 

Reduced dimethylferrocene is nearly insoluble in water and has a half wave potential of 100 mV vs SCE, which diminishes electrochemical interferences in glucose determination. The adsorbed mediator becomes active as an electron acceptor only by anodic oxidation at +160 mV to the ferricinium ion (FecpR+) ... 

A glucose microsensor based on difference measurement between two gold microelectrodes, one covered by native and one by denatured GOD, has been described by Takatsu and Morizuma (1987). This combination permits the elimination of electrochemical interferences without resorting the use of permselective membranes. The gold electrodes were... 

For anodic NADH oxidation a potential of more than +0.4 V is necessary. Since this high overpotential favors electrochemical interferences, various investigations have been conducted to decrease the oxidation potential by using mediators or pretreating the electrode. Cenas et al. (1984) found that after electrochemical pretreatment of glassy carbon electrodes in the range of -0.8-1.8 V, NADH oxidation occurs at 6-0.2 V vs Ag/AgCl. LDH was entrapped on top of the electrode by means of a dialysis membrane. The oxidation current was proportional to lactate concentration up to 10 mmol/1. Presumably because of adsorption of NADH oxidation products the half life of the sensor was less than 3 days. 

Mullen et al. (1986) decreased the permeability of LOD membranes in order to extend the linear range to higher lactate concentration. A polycarbonate membrane was treated with methyltrichlorosilane prior to enzyme immobilization by crosslinking with glutaraldehyde and BSA. In this way the upper limit of linearity was shifted from 0.2 to 18 mmolA. The permeation of electrochemical interferents was diminished concomitantly. On the other hand, the silanization increased the response time from 0.5-1 min to 1-3 min and reduced the sensitivity by 98-99%. 

Laccase also catalyzes the 02-dependent oxidation of ascorbic acid, ferrocyanide, iodide, and uric acid. These reactions have been utilized to eliminate electrochemical interferences in amperometric hydrogen peroxide detection at membrane-covered enzyme electrodes (Wollen-berger et al., 1986). The capacity of the laccase membrane to oxidize ferrocyanide has been characterized by anodic oxidation of ferrocyanide at +0.4 V (Fig. 62). When a fresh enzyme membrane is used, a current signal appears only at substrate concentrations above 5 mmolA the current increases linearly with increasing concentration. This threshold concentration decreases with increasing membrane age until the remaining enzyme activity is too low for complete substrate oxidation. 

In view of the low physiological uric acid concentrations, in the assay of uric acid electrochemical interferences by other anodically oxidizable substances become particularly troublesome. Kulys et al. (1983) coimmobilized HRP with uricase in order to eliminate these interferences. This approach permits uric acid measurement to be performed at a potential of 0 V vs SCE. However, the autoxidation of ferrocyanide used as HRP substrate is a serious drawback of this method. 

With a similar lactose sensor, Pilloton et al. (1987) assayed lactose in milk samples and obtained a good correlation with the reduction method described by Fehling. The enzyme was fixed to a nylon membrane. A cellulose acetate membrane (molecular cutoff 100 Dalton) was included in order to eliminate electrochemical interferences. The lifetime of the sensor was 1 month. 

Matsumoto et al. (1985) developed a sensor with the same enzyme sequence by immobilizing the enzymes covalently to silanized glassy carbon by glutaraldehyde. The sensor had a half-life of 7 weeks. Electrochemical interferences were compensated for by use of an additional, enzyme-free electrode. 

The above measuring principle has been modified for the measurement of HRP substrates which are themselves electrochemically active. To avoid electrochemical interference a Clark-type oxygen probe was used together with catalase coimmobilized with GOD. The hydrogen peroxide not consumed by HRP is cleaved by catalase to oxygen which is indicated at the electrode. Consequently, the biochemical basis of the sensor is the competition of HRP and catalase for the common substrate, H202. 

Enzyme electrodes comprising coimmobilized COD and CEH can be used for the determination of total cholesterol. On the other hand, coupling of COD with HRP enables cholesterol measurement at low electrode overvoltage, which avoids electrochemical interferences. Fig. 89 shows the diversity of the potential sequences of enzymatic and electrochemical reactions in cholesterol electrodes. 

In the optimized conditions, the dynamic range for Carbo-firran detection was 10 10-107 M with a detection limit of 4.9 x 10 10 M, for an analysis time of 15 min. This is an important feature, considering that the immobilization can determine a loss of the activity of the enzyme that influence the sensitivity as well as dynamic range of the pesticide detection (31). Moreover, the proposed method was less prone to electrochemical interferences since the incubation and measurement were performed in two separate steps. 

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