Glucose determination with enzyme electrodes

F. Ricci, A. Amine, C.S. Tuta, A.A. Ciucu, F. Lucarelli, G. Palleschi and D. Moscone, Prussian Blue and enzyme bulk-modified screen-printed electrodes for hydrogen peroxide and glucose determination with improved storage and operational stability, Anal. Chim. Acta, 485(1) (2003) 111-120. 

Figure SJ Activity of the various states of the [NiFe] hydrogenase from A. vinosum as determined with a Pt electrode at 30°C.The reaction was performed in SOmM Tris/HCI (pH 8.0) in a volume of 2 ml. Oxygen was scavenged by adding glucose (90 mM) and glucose oxidase (2.5 mg/ml). Hydrogen peroxide was removed by catalase. When the system was anaerobic, an aliquot of H2-saturated water was added, and a little later enzyme (S-IOnM) was injected. Benzyl viologen (4.2mM) was used as electron acceptor.

All electrodes react with their environment via the surfaces in ways which will determine their electrochemical performance. Properly selected surface modification can effectively enhance the electrode heterogeneous catalysis property, especially selectivity and activity. The bulk materials can be chosen to provide mechanical, chemical, electrical, and structural integrity. In this part, several surface modification methods will be introduced in terms of metal film deposition, metal ion implantation, electrochemical activation, organic surface coating, nanoparticle deposition, glucose oxidase (GOx) enzyme-modified electrode, and DNA-modified electrode. 

A great number of enzymes have already been combined with electrodes. In some cases, several possible enzymatic routes may be selected, depending on the substrate to be analyzed. One such example is that used to determine aspartame (Fig. 3) (18-20). Enzyme electrodes use, ideally, only one enzyme and monitor the main substrate-enzyme reaction, for example, glucose oxidase-glucose. If the main substrate-enzyme reaction is not electrochemically detectable or if signal amplification is required, bi- or trienzymatic sequences may be applied. 

In another advance in the use of immobilized enzymes in analysis, glucose oxidase was combined with an oxygen electrode to give an enzyme electrode for determining glucose concentration (15, 16). The... 

An electrode for measuring urea has been described (Gll), consisting of a thin film of urease, immobilized in acrylamide gel, on the surface of a glass electrode responsive to NH. Conditions are carefully selected to ensure stability of the enzyme, and the potential developed is proportional to the logarithm of the urea concentration. Blood glucose and lactate have been determined with a membrane electrode in which the enzyme (glucose oxidase or lactate dehydrogenase) is trapped in a porous or jellied layer at the membrane surface (W20). 

Di Gloria et al. [54] proposed the use of electrode-enzyme coupling for the amplified readout of a redox label, using a lidocaine-ferrocene conjugate in combination with GOx. The use of ferrocene as a mediator for glucose determination within GOx enzyme electrodes provided the... 

Techniques such as potentiometry, polarography, and microcalorimetry have been chosen in exploiting the benefits of immobilized enzymes (see Chapter 4). Enzymes incorporated into membranes form part of enzyme electrodes. The surface of an ion-sensitive electrode is coated with a layer of porous gel in which an enzyme has been polymerized. When the electrode is immersed in a solution of the appropriate substrate, the action of the enzyme produces ions to which the electrode is sensitive. For example, an oxygen electrode coated with a layer containing glucose oxidase can be used to determine glucose by the amount of oxygen consumed m the reaction, and urea can be estimated by the combination of a selective ammonium ion-sensitive electrode and a urease membrane. 

Gorton et al. (1985) employed GOD adsorbed on spectral carbon for glucose determination in a wall jet flow-through cell of an FIA system. Up to 120 samples per hour have been measured with good precision. The enzyme electrode was stable for 3-7 days. Coating of the spectral carbon by a Pd/Au layer of 20 pm thickness permits H2O2 to be anodically indicated at potentials as low as +350 mV vs SCE. The enzyme activity is not affected by this modification. 

Renneberg et al. (1983a) described an enzyme electrode-based assay of factor VIII, which is important for blood coagulation diagnostics. AP was used as the marker enzyme and the hydrolysis of glucose-6-phosphate was measured with a glucose electrode. This combination allowed the determination of 1.6-16 ng of factor VIII in human plasma. 

There are over 300 enzymes available commercially, and many of these can be used in one way or another for analytical purposes. One of methods of use involves the determination of an analyte or substrate by means of the enzyme which reacts specifically with that substrate. Examples are 1. Glucose and glucose oxidase, O2 released, measurement by O2-ISE 2. Urea and urease, NHs and CO2 released, measurement by an NH4 - or CO2-ISE 3. Pectin and pectin-esterase, titration of H+ released. In chnical work, the opposing approach is often used, and the amount of an enzyme determined by adding the proper substrate to a solution of the enzyme and measuring with an ion-selective electrode a product of the reaction. Early work in this area, and in that of immobihsed enzyme electrodes, was carried out by Katz and Rechnitz [15,16], Guilbault and his co-workers [17] and Guilhault and Montalvo [18]. 

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