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Freseuius Zeltsdlrift fiir
Fresenius Z. anal. Chern.2S7, 1-9(1977)
Anatytische Chemie © by Springer-Verlag 1977
Bio-Sensors Based on Ion-Selective Electrodes Karl Cammann Mineralog.-Petrograph. Institut der Universitat Miinchen, TheresienstraBe 41, D-8000 Munchen 2
Biiosensoren auf der GrundJage von ionenseJektiven EJektroden
Zusammeniassung. Ein Uberblick tiber Biosensoren wird gegeben. An Hand mehrerer BeispieJe von Enzymelektroden werden die entscheidenden Eigenschaften und die prinzipielle Arbeitsweise dieser neuen Sensoren dargesteHt. 72 Literaturzitate. Summary. A review is given on bio-sensors based on ion-selective electrodes. By means of several examples of enzyme electrodes the salient features and the operating principles of the new sensors are illustrated. 72 literature references. Key words: Biosensoren/Elektroden, ionenselektive; Eigenschaften, Arbeitsweise, Dberblick
1. Introduction Ion-selective potentiometry has become a well established analytical method in the last few years. This review will not deal with the basic principles of ion-selective electrodes, since this topic has already been covered in the three text books now available [7,39, 48J. No attempt is made here to present a complete summaIy of all electrode-based bio-sensors described in the literature; a few examples of enzyme electrodes have been chosen to illustrate the salient features and the operating principles of these new sensors. The term enzyme electrode is somewhat of a misnomer, as it is the substrate of an enzymic reaction that is measured. The term "substrate electrode" would seem to be preferable [47]. Nevertheless the former is more widely accepted and, in a provisional nomenclature suggestion of the IUPAC, an enzyme electrode is defined as a sensor "in which an ion-selective electrode is covered
with a coating that contains an enzyme which causes the reaction of an organic or inorganic substance (substrate) to produce a species to which the electrode responds" [49]. Apart from the measurement of substrate concentrations, enzyme electrodes are also used in the determination of related enzyme activities; by this, enzyme inhibitors and stimulating agents can also be determined. Enzymic reactions are extremely specific (sometimes even isomers with different optical activity are distinguished) and, because of this, enzyme electrodes can be highly selective sensors. In the literature major emphasis has, up till now, been restricted to macroscale enzyme electrodes, but research is now also being directed toward the microscale [28,57]. Recent progress in the field of miniaturized ion-selective electrodes [37,42,69J and glass fiber optics [35J have, for the first time, made possible the detection of changes in the composition of intracellular fluids. In situ and in vivo analysis without sample consumption is very important here as the complex steady state of different equilibria in living matter is not disturbed. Microelectrodes for monitoring pH, sodium, potassium, calcium and chloride ion levels have, up till now, been used successfully in physiological applications [36J. More or less selective macroelectrodes are now available for nearly two dozen different ions. Figure 1 shows that a number of other ions and even neutral compounds such as gases or enzymes and their substrates can be detected indirectly. One can expect that this large variety of compounds can then also be monitored with the corresponding microelectrodes now under development Biosensors (or bioprobes [58]) would then be an extremely valuable tool for researches in the field of medicine, biology or in general physiology. In order to understand fully the benefits of using ion-selective electrodes as sensors in biological systems, some features of these electrodes may be recalled.
2
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Fig. 1. Survey of the measurable substances with the aid of ion-selective electrodes [from 7]
2. Features of Iou-Selective Electrodes Ion-selective electrodes respond to the activity of the free uncomplexed species and not to the total concentration of the ion for which the electrode is selective. This is in no way a drawback since recent studies have shown that besides physiological applications, the activity of the free species is also of great importance in the field of pollution control. Often total concentration values without further ~nformation a~out the binding state of the corresponding Ion are meamngless if the toxic level problem is encountered [66]. However, if total concentrations are desired, only the measuring techniques have to be changed: direct-potentiometry via an activity calibration curve yields the activity of the free ion; standard addition gives the total concentration. Ion-selective electrodes can also differentiate between the various oxidation states of an element (e.g. NH 4+, NO" N0 3 - or S2-, S042- or Cl-, Cl0 4 -). One special advantage in the use of ion-selective electrodes as sensors for enzymic reactions is the fact that potentiometric measurements are not disturbed by turbid
samples. It is even possible to measure highly viscous samples (pastes, ointments etc.) or suspensions with high accuracy. Another advantage is their fast response time. This c~vers a s~an from milliseconds to a few minutes along With the dIrect and continuous delivery of an electrical signal ~roportion.al to the logarithm of the ion activity. Some lOn-selectIve electrodes have working ranges greater than 10 decades, in which the same relative accuracy can be achieved. A further important aspect lies in the simple and low-cost instrumentation; a pH meter is available in nearly all laboratories. The major disadvantage of ion-selective electrodes is lack of selectivity of many electrodes. This specificity of an ion-selective electrode can be expressed by the socalled selectivity coefficient which is related to an extended Nernst equation: (1)
3
K. Cammann: Bio-Sensors Based on Ion-Selective Electrodes
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Fig. 2A and B Gas sensors based on ion-selective electrodes. (A) Membrane-covered type (Orion, ElL); (B) Air-gap type [24]; M indicator electrode; B = reference electrode; K = salt bridge; L = air-gap; R = magnetic stirrer (from [7])
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