Membrane materials enzyme sensors

In recent years the electrochemistry of the enzyme membrane has been a subject of great interest due to its significance in both theories and practical applications to biosensors (i-5). Since the enzyme electrode was first proposed and prepared by Clark et al. (6) and Updike et al. (7), enzyme-based biosensors have become a widely interested research field. Research efforts have been directed toward improved designs of the electrode and the necessary membrane materials required for the proper operation of sensors. Different methods have been developed for immobilizing the enzyme on the electrode surface, such as covalent and adsorptive couplings (8-12) of the enzymes to the electrode surface, entrapment of the enzymes in the carbon paste mixture (13 etc. The entrapment of the enzyme into a conducting polymer has become an attractive method (14-22) because of the conducting nature of the polymer matrix and of the easy preparation procedure of the enzyme electrode. The entrapment of enzymes in the polypyrrole film provides a simple way of enzyme immobilization for the construction of a biosensor. It is known that the PPy-... 

Time response. In most situations enzyme kinetics have very little effect on the response time of enzyme-based biosensors. From the analysis given above, it is clear that one should operate these devices under conditions where the analyte concentration within the sensor is much less than Km- For sensors which are in the membrane diffusion limiting regime (section 7.3.1.1 above), the response characteristics of the membrane material will be governing. These depend on the thickness of the membrane and the diffusivity of the analyte in the membrane material. An approximate estimate of the membrane lag time is... 

Beside these disposable systems enzyme-membrane-based devices (first-generation enzyme sensors) are working with a biological component that can be repeatedly used for thousands of measurements. The functional stability depends on the quality of the enzyme membrane material used. The autoanalyzer Stat-Profile 5 (Nova Biomedical, USA) and the lonometer (Fresenius, Germany) permit the analysis of metabolites, such as glucose and lactate, in addition to electrolytes and blood gases (see table 17.2). 

The technique of the immobilization of the biological elements has changed according with the different events, catalytic or affinity. The simplest way to retain enzymes on the tip of a transducer is to trap them behind a perm-selective membrane. This method has been mainly used in addition to embedding procedures in polyacrylamide gels. Then, mainly in the 80th, the trend shifted to use disposable membranes with bound bioactive material. Several companies put on the market preactivated membranes suitable for the immediate preparation of any bioactive membrane and this appeared as a real improvement at least for the easy use of enzyme sensors. 

It is important to realize that biocompatibility issues are not only relevant in respect of the well being of the host, but also in respect of the requirements of the sensor itself. More specifically, the required chemical interactions between the sensor and the body must not be interfere with by interactions—either chemical or physical— between the membrane material and contacting/adhering cells. For example, an encapsulating membrane of an electrochemically based sensor must maintain appropriate mass transport conditions for the analyte and electrolyte species, and must exclude species that could interfere with the electrochemistry, or denature or inhibit the activity of immobilized enzymes. Stability of mass transport conditions is especially critic, since any change in the permeability of the membrane or the surrounding tissues can affect the sensor calibration. Satisfactory stability can not be achieved without a biocompatible encapsulation material. 

By changing the enzyme and mediator, the amperometric sensor in Figure 11.39 is easily extended to the analysis of other substrates. Other bioselective materials may be incorporated into amperometric sensors. For example, a CO2 sensor has been developed using an amperometric O2 sensor with a two-layer membrane, one of which contains an immobilized preparation of autotrophic bacteria. As CO2 diffuses through the membranes, it is converted to O2 by the bacteria, increasing the concentration of O2 at the Pt cathode. 

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-... 

The design and implementation of a portable fiber-optic cholinesterase biosensor for the detection and determination of pesticides carbaryl and dichlorvos was presented by Andreou81. The sensing bioactive material was a three-layer sandwich. The enzyme cholinesterase was immobilized on the outer layer, consisting of hydrophilic modified polyvinylidenefluoride membrane. The membrane was in contact with an intermediate sol-gel layer that incorporated bromocresol purple, deposited on an inner disk. The sensor operated in a static mode at room temperature and the rate of the inhibited reaction served as an analytical signal. This method was successfully applied to the direct analysis of natural water samples (detection and determination of these pesticides), without sample pretreatment, and since the biosensor setup is fully portable (in a small case), it is suitable for in-field use. 

Biological Membranes Attractive supporting membranes for commercial exploitation of biological material, e.g. immobilization ot membrane bound enzymes in solid state sensors ISFET tvpe structures. 

CNTs and other nano-sized carbon structures are promising materials for bioapplications, which was predicted even previous to their discovery. These nanoparticles have been applied in bioimaging and drag delivery, as implant materials and scaffolds for tissue growth, to modulate neuronal development and for lipid bilayer membranes. Considerable research has been done in the field of biosensors. Novel optical properties of CNTs have made them potential quantum dot sensors, as well as light emitters. Electrical conductance of CNTs has been exploited for field transistor based biosensors. CNTs and other nano-sized carbon structures are considered third generation amperometric biosensors, where direct electron transfer between the enzyme active center and the transducer takes place. Nanoparticle functionalization is required to achieve their full potential in many fields, including bio-applications. 

Sensitive, selective detection of biochemically active compounds can be achieved by employing antigen-antibody, enzyme-substrate, and other receptor-protein pairs, several of which have been utilized in the development of piezoelectric immunoassay devices. The potential analytical uses of these materials has been reviewed, particularly with respect to the development of biochemical sensors [221-224], The receptor protein (e.g., enzyme, antibody) can be immobilized directly on the sensor surface, or it can be suspended in a suitable film or membrane. An example of the sensitivity and response range that can be... 

The potential applications of calixarmes range from molecular hosts for sensor techniques and medical diagnostics, use in decontamination of wastewater, construction of artificial enzymes, new materials for nonlinear optics to sieve membranes with molecular pores. 

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