Sensing microzone

Bio)chemical sensors can be active or passive according to whether they use a sensing microzone to accommodate a chemical or biochemical reaction and/or a biochemical e.g. immunological) or physico-chemical separation e.g. sorption). It should be noted that passive sensors e.g. a fibre-optic tip immersed in an industrial process stream) do not meet one of the essential requirements included in the definition of sensors as regeirds composition... 

The sensors discussed in this book are dealt with according to the classifications based on which the immobilized species is, the type of immobilization used and whether or not an additional separation is included. Thus, sensors have been included in three chapters according to the type of process that takes place in their sensing microzone, namely reaction-detection (Chapter 3), separation-detection (Chapter 4) and separation-reaction-detection (Chapter 5). 

No doubt, one of the most intuitive classifications of sensors is that based on the type of transducer used to reveal the physico-chemical changes that occur in the sensing microzone in the presence of the analyte. Figure 1.6 shows the principal types of transducing systems eire connected to or... 

Figure 1.14 — Classification of flow-through sensors according to external shape. SMZ sensing microzone D detector W waste. For details, see text.

Flow-through optical sensors bearing one or more immobilized enzymes at their sensing microzone can be classified according to the type of physical relationship between the microzone and the detection system or instrument used into those using fibre optics (photometric and luminemetric) and those integrating a biochemical reaction and detection (usually photometric). 

Mediated electrochemical sensors aside, there are few sensors involving a reaction at the sensing microzone by which the analyte is not retained to some extent during the time the analytical response is generated. Such is the case with sensors based on luminescence quenching and a few others. Although many of the reactions on which the analytical measurement rests in sensors based on acid—base reactions involve retention of protons, the sensors in question are dealt with in this Section. 

Saari and Seitz [250] used the tip of an optical fibre as the sensing microzone to immobilize a suitable reagent (fluoresceinamine) in order to construct a fluorescence sensor for pH measurements the design was inspired by previous work of Peterson et al., who used a dye immobilized at the tip of an optical fibre for pH absorptiometric measurements [251]. The... 

These sensors can be classified according to various criteria. One classification is based on space and time concepts in relation to the processes taking place at the sensing microzone (viz. whether they occur sequentially or simultaneously). As can be seen in Fig. 5.1, (a) mass transfer (of analytes... 

Figure 5.1 — Classification of (bio)chemical flow-through sensors based on integrated reaction, separation and detection according to whether the three processes take place sequentially (A,B) or simultaneously (C) at the sensing microzone. S sample R reagent. (Reproduced from [1] with permission of the Royal Society of Chemistry).

For a detailed description of the separation processes that may take place at the sensing microzone, the foundation of which is closely related to non-chromatographic continuous separation techniques based on mass transfer across a gas-liquid (gas diffusion), liquid-liquid (dialysis, ultrafiltration) or liquid-solid interface (sorption), interested readers are referred to specialized monographs e.g. [3]). 

The analytes typically determined by using this type of sensor are those usually addressed by gas-diffiision systems, viz. ammonia (or ammonium ion), carbon dioxide (or carbonates) and oxygen. The detection system used is most frequently photometric, fluorimetric or potentiometric, and can be integrated with or connected to the sensing microzone. The description below is based on the two choices shown in Fig. 5.4. 

In this type of sensor (Fig. 5.4.A), the sensing microzone accommodating the gas-diffiision membrane is traversed by a stream of (a) a carrier intended to transport the injected sample, condition it (e.g. adjusting its pH) and prepare/regenerate the sensor or b) the sample itself, introduced into the continuous manifold by aspiration or injection and subsequently mixed in a continuous fashion with another stream if needed. 

Figure 5.5 — Flow-through biosensor for the determination of L-glutamate. (A) Flow injection manifold. (B) Sensing microzone of the probe sensor (optrode), incorporated in the flow-cell (FTC). P pump IV injection valve MC mixing chamber AD air damper BFB bifurcated fibre bimdle LS light source PMT photomultiplier R recorder GLU L-glutamate 0-Glu 2-oxoglutarate E enzyme layer I optical insulator S sensing layer PS polyester support. For details, see text. (Adapted from [6] with permission of Elsevier Science Publishers).

The carrier used for this purpose consisted of a 0.1 M phosphate buffer of pH 7. The appearance of the sensing microzone is shown in Fig. 5.5.B. The oxygen optrode used was based on a 10-pm silicone rubber film containing dissolved decacyclene as indicator (S) that was fixed on a 110-pm thick polyester support (PS). A 9-pm black PTFE membrane (I) was used for optical insulation. The dye fluorescence was found to be markedly dependent on the concentration of oxygen, which exerted a quenching effect on it. The enzyme (glutamate oxidase) was immobilized on a 150-pm thick immunoaffmity membrane (E). The sensor was prepared similarly as reported by Trettnak et al. [7]. 

Recently, DeGrandpre [12] developed a probe-type sensor for the determination of PCO2 in sea water by direct immersion of the probe, which, however, has some connotations of flow-through sensor even though a pH indicator such as Phenol Red (piTj = 7.5) or Bromothymol Blue (pAn = 6.8) rather than the sample is circulated over the sensing microzone —the basic forms of these indicators have a high molar extinction coefficient at 560 and... 

This type of sensor is schematically depicted in Fig. 5.4.B. The sensing microzone is crossed by two streams rather than one. One such stream acts as the donor and contains the aspirated or injected sample, which is conditioned in order to provide a gaseous reaction product. The other stream acts as the acceptor for the volatile species transferred across the gas-diffusion membrane that isolates the two streams. 

Continuous circulation of the acceptor solution, where the sensing process takes place, makes them reusable and hence of practical interest. The acceptor solution may contain the ingredients needed to regenerate the reagent when this is immobilized on the sensing microzone. 

There are two possible configurations for this type of flow-through sensor integrating gas diffusion, reaction and detection that differ in whether the reagent is dissolved in the acceptor solution or immobilized on a sensing microzone located near the diffusion membrane. The descriptions below are based on such a difference. 

The sensing microzone of the flow-through sensor depicted in Fig. 5.9.B1 integrates gas-diffusion and detection with two analytical reactions [28], viz. (a) the urease-catalysed formation of ammonium ion by hydrolysis of urea (the analyte), which takes places on a hydrophilic enzyme membrane in contact with the sample-donor stream, which contains a gel where the enzyme is covalently bound and (b) an acid-b reaction that takes place at the microzone on the other side of the diffusion membrane and involves Bromothymol Blue as indicator. This is a sandwich-type sensor including a hydrophilic and a hydrophobic membrane across which the sample stream is circulated —whence it is formally similar to some enzyme electrodes. Since the enzymatic conversion of the analyte must be as efficient as possible, deteetion (based on fibre optics) is performed after the donor and acceptor streams have passed through the sensor. Unlike the previous sensor (Fig. 5.9.A), this does not rely on the wall-jet approach in addition, each stream has its own outlet and the system includes two sensing microzones... 

Sensors based on integrated dialysis, reaction and detection differ from those described in Section 4.3.1 in the fact that a (bio)chemical reaction takes place after separation (simultaneously with detection). They thus fit the generic configuration depicted in Fig. 5.1. A. Some of the ingredients of such a reaction may be immobilized at the sensing microzone, even though the reaction may also take place in the solution passed through it. 

The sorbent materials used to construct this type of sensor are widely varied (ion exchangers, adsorbent solids, polymers) and are employed as particles (larger than 30 pm in order to avoid overpressure in the flow system) or films. Most of these sensors are optical and rely on absorption, reflectance or molecular fluorescence measurements. In order to ensure that the sensing microzone is fully compatible with the detector, the sorbent material used must be as transparent as possible (photometry) or give rise to no appreciable light scatter (fluorimetry) so that the baseline (resulting from passage of the carrier) may be as low as possible. 

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