Enzymatic sensor system

Tag K, Kwong AWK, Lehmann M, Chan C, Renneberg R, Riedel K, Kimze G (2000) Fast detection of high-molecular substances in wastewater based on an enzymatic hydrolysis combined with the Arxula BOD sensor system. J Appl Chem Biotechnol 75 1080-1082... 

Less wide-spread than the enzymatic determination of these amino acids is the application of enzyme sensor systems for precursors or other amino acids. Collins et al. [116] described a flow injection system for the determination of a-ketoglutarate during an industrial fermentation. The system was based on glutamate dehydrogenase (Eq. (11.5)) and glutamate oxidase (Eq. (11.4))... 

The product will diffuse out of the enzyme phase through the exterior membrane E and also diffuse toward the amperometric electrode detector (if one is part of the sensor system). The steady state product concentration within the enzyme phase will be determined by the balance of enzymatic production of product within the enzyme region, volume V, and its disappearance by diffusion or electrode reaction... 

The active site of organophosphorus hydrolase (OPH) contains two metal atoms (zinc in the wild-type enzyme) and catalyzes hydrolysis of numerous organophos-phate compounds including pesticides as well as chemical warfare agents such as sarin and soman. Rates of OPH catalyzed hydrolysis of organophosphates exceed those of chemical hydrolysis by NaOH at 4°C by factors of 40 to 2450 [41-43]. The enzyme has been described for use in sensor systems with exceptional detection limits reported for response times on the order of 10 seconds [44-48]. However, the presence of OP/CWA is detected by the inhibition of enzymatic rate determined by comparing rate measnrements in the presence and absence of the analyte. 

Aim of this worit was to demonstrate a particular example of a sensor system, which combines catalytic activity for urea and at the same time, enabling monitoring enzymatic reaction by optical recording. The proposed sensor system is based on multilayer polyelectrolyte microcapsules containing urease and a pH-sensitive fluorescent dye, which translates the enzymatic reaction into a fluorescendy registered signal. 

In this study we have demonstrated a particular example of a sensor system, which combines catalytic activity for the substrate (urea) and at the same time enabling to monitor the enzymatic reaction by co-encapsulated pH sensitive dye. Substrate sensitive enzyme urease was co-encapsulated together with SNARF-1 coupled to dextran in multilayer tnicrocapsules. Enzymatic activity was recorded by fluorescent changes caused by increasing of pH in course of enzymatic cleavage of urea as measured on... 

Overall, the implementation of lanthanide probes in chemical sensor technology is still in its initial stage. Up to now they have not found their way into commercialized sensor systems. Particularly, with respect to p02 and pH sensors, it cannot be foreseen that LLCs may displace established fluorescent indicators. Sensors for small molecules such as hydrogen peroxide, phosphate, or ATP can be useful in enzymatic assays in which the conversion of the substrate has to be monitored. In this case, the selectivity is provided by the enzyme involved. A concrete example is presented by means of a glucose sensor based on immobilized glucose oxidase and... 

Guivar JA, Fernandes EG, Zucolotto V. A peroxidase biomimetic system based on Fc304 nanoparticles in non-enzymatic sensors. Talanta 2015 141 307-14. http //dx.doi.org/ 10.1016/j.talanta.2015.03.017. 

In any case, when observing the evolution timeline, first works that are worth mentioning are potentiometric sensors, and only after first attempts with ion-selective electrodes and enzymatically converting systems are voltammetric types found. This progression also follows the timeline of ETs, as there were the ones using the potentiometric sensors the ones first proposed and developed, and only after a few years, the first voltammetric electronic tongue was reported [17]. 

Many complex systems have been spread on liquid interfaces for a variety of reasons. We begin this chapter with a discussion of the behavior of synthetic polymers at the liquid-air interface. Most of these systems are linear macromolecules however, rigid-rod polymers and more complex structures are of interest for potential optoelectronic applications. Biological macromolecules are spread at the liquid-vapor interface to fabricate sensors and other biomedical devices. In addition, the study of proteins at the air-water interface yields important information on enzymatic recognition, and membrane protein behavior. We touch on other biological systems, namely, phospholipids and cholesterol monolayers. These systems are so widely and routinely studied these days that they were also mentioned in some detail in Chapter IV. The closely related matter of bilayers and vesicles is also briefly addressed. 

A lot of analytical techniques have been proposed in recent decades and most of them are based on enzymes, called dehydrogenases, which are not sensitive to oxygen and need cofactors such as NAD". The key problems which seriously hamper a wide commercialization of biosensors and enzymatic kits based on NAD-dependent enzymes are necessity to add exogenous cofactor (NAD" ) into the samples to be analyzed to incorporate into the biologically active membrane of sensors covalently bounded NAD" to supply the analytical technique by NAD -regeneration systems. 

The high specificity required for the analysis of physiological fluids often necessitates the incorporation of permselective membranes between the sample and the sensor. A typical configuration is presented in Fig. 7, where the membrane system comprises three distinct layers. The outer membrane. A, which encounters the sample solution is indicated by the dashed lines. It most commonly serves to eliminate high molecular weight interferences, such as other enzymes and proteins. The substrate, S, and other small molecules are allowed to enter the enzyme layer, B, which typically consist of a gelatinous material or a porous solid support. The immobilized enzyme catalyzes the conversion of substrate, S, to product, P. The substrate, product or a cofactor may be the species detected electrochemically. In many cases the electrochemical sensor may be prone to interferences and a permselective membrane, C, is required. The response time and sensitivity of the enzyme electrode will depend on the rate of permeation through layers A, B and C the kinetics of enzymatic conversion as well as the charac-... 

Another possibility is to immobilise enzymes either on the sensor element itself or in the vicinity of the sensing element. The operation principle is in most cases a semi-continuous spectral difference measurement in combination with a kinetic data evaluation. A sample containing the analyte of interest is recorded by the sensor immediately after contact with the sample and again after a certain time. Provided that no other changes in the composition of the sample occur over time, the spectral differences between the two measurements are characteristic for the analyte (and the metabolic products of the enzymatic reaction) and can quantitatively evaluated. Provided that suitable enzymes are available that can be immobilised, this may be a viable option to build a sensor, in particular when the enzymatic reaction can not (easily) be monitored otherwise, e.g. by production or consumption of oxygen or a change of pH. In any case, the specific properties and stumbling blocks related to enzymatic systems must be observed (see chapter 16). 

Another approach, developed in our laboratory, consists of the compartmentalization of the sensing layer25"27. This concept, only applicable for multi-enzyme based sensors, consist in immobilizing the luminescence enzymes and the auxiliary enzymes on different membranes and then in stacking these membranes at the sensing tip of the optical fibre sensor. This configuration results in an enhancement of the sensor response, compared with the case where all the enzymes are co-immobilized on the same membrane. This was due to an hyperconcentration of the common intermediate, i.e. the final product of the auxiliary enzymatic system, which is also the substrate of the luminescence reaction, in the microcompartment existing between the two stacked membranes. 

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... 

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