Electrode, antimony enzyme

Vilker et al. were able to transfer the reducing equivalents to purified cytochrome CYPIOI through its natural redox partner putidaredoxin using an antimony-doped tin oxide working electrode, thus avoiding the reductase and NADH. The required oxygen was supplied at a platinum counter electrode by water electrolysis as shown in Fig. 31. A continuous catalytic cycle was sustained for more than 5h, and 2600 enzyme turnovers were obtained at a maximum production rate of 36 nmol S-C-vo-hydroxycamphor/nmol CYPlOl/min [151],... 

While the majority of enzyme electrodes fabricated have been rather large devices, there have been some recent reports concerning the development of miniaturized and even microsensors. For example, MeyerhoflF (M5) prepared an essentially disposable urea sensor (tip diameter 3 mm) by immobilizing urease at the surface of a new type of polymer-membrane electrode-based ammonia sensor (see Fig. 4). Alexander and Joseph (Al) have also prepared a new miniature urea sensor by immobilizing urease at the surface of pH-sensitive antimony wire. Similarly, lannello and Ycynych (II) immobilized urease on a pH-sensitive iridium dioxide electrode. In these latter investigations, ammonia liberated from the enzyme-catalyzed reaction alters the pH in the thin film of enzyme adjacent to the pH-sensitive wire. 

Other pH-sensing transducers used in biosensors are metal oxide electrodes. Beside the common antimony oxide electrode, palladium oxide and iridium oxide probes have been coupled with immobilized enzymes. These sensors may be miniaturized by using chemical vapor deposition technology. Moreover, they are mechanically more stable than glass electrodes. Unfortunately the measuring signal of metal oxide electrodes is affected by redox active substances. 

Kulys et al. (1986b) studied urea determination by difference measurement between two antimony electrodes covered with exchangable membranes (Fig. 68). Urease was attached in the pores of a macroporous membrane (thickness, 10 pm, pore diameter, 0.1 pm) by glutaraldehyde. This layer was covered with a monoacetylcellulose membrane. The membrane for the auxiliary electrode was prepared analogously, but using BSA instead of urease. The assay of urea was carried out with a differential amplifier which simultaneously differentiated the time course of the potential difference between enzyme and auxiliary electrode (kinetic method). Thus, a response time of only 20 s was possible. 

The pH electrode is another suitable transducer for the construction of a urea biosensor. The classical glass electrode is sensitive to H ions and urease is attached in a gel of either polyacrylamide [107] or methacrylamide-aoylamide cqxdymer [108]. me metallic electrodes are also sensitive to H ions (for example, the antimony electrode) and can also be used in conjunction with a urease membrane [109]. The enzyme pH electrode detects a very weak variation in proton concentration arising from an enzymatic reaction, and the signal amplitude is determined by the buffering capacity of the solution. Both the nature of the buffer solution and the working pH value of the biosensor can reduce its practical use. A differential measurement at different pH values can be used to correct for any variation. 

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