Integrated detection-reaction

Flow-through sensors integrating detection and a chemical or biochemical reaction rely on immobilization in the probe proper or the flow-cell (or a special housing included in it) of a species intended to take part in or catalyse the reaction by which the analyte, viz. the catalyst or reagent, is measured, according to which the sensors described in this Chapter are divided into two broad categories. 

The equipment required to develop this type of sensor is very simple and resembles closely that used to implement ordinary liquid-solid separations in FI manifolds. The only difference lies in the replacement of the packed reactor located in the transport-reaction zone with a packed (usually photometric or fluorimetric) flow-cell accommodated in the detector. Whether the packing material is inert or active, it should meet the following requirements (a) its particle diameter should be large enough (< 80-100 fim) to avoid overpressure (b) it should be made of a material compatible with the nature of the integrated detection system e.g. almost transparent for absorbance measurements) and, (c) the retention/elution process should be fast enough to avoid kinetic problems. 

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. 

As with sensors based on a triply integrated process involving gas diffusion, there are few reported examples of sensors integrating dialysis, reaction and detection. There follows a description of die most salient examples based on the ingredient of the (bio)chemical reaction that is dialysed at the sensing micro2one. 

Though less commonplace, there are some electrochemical sensors based on integrated dialysis, reaction and detection. Most of them use a retained enzyme immobilized in a microcavity at the tip of the working electrode that... 

The descriptions of sensors integrating sorption, reaction and detection provided below are classified according to the type of immobilization... 

Equation (7.12) can be extended to more complex rate equations. Some of the advantages of the stirred flow method are (i) complicated rate expressions can be handled without integration, (ii) reaction can be carried out under constant conditions of solvent composition, ionic strength and transient intermediate can be built up in concentration, detected and measured. 

Gottschlich et al. [134] developed a microfluidic system that integrated enzymatic reactions, electrophoretic separation of the reactants from the products, and postseparation labeling of the proteins and peptides prior to fluorescence detection (see Fig. 12). Tryptic digestion of oxidized insulin p-chain was performed in 15 min under stopped flow conditions in a heated channel serving as the reactor, and the separation was completed in 60 s. Localized thermal control of the reaction channel was achieved using a resistive heating element. The separated reaction products were then labeled with naphthalene-2,3-dicarboxaldehyde (NDA) and detected by fluorescence detection. 

Pitts JR (2002) Detecting hydrogen with chemochromic thin films. Ind Physicist (AIP) Forum June/July 31 Polla DL, White RM, Muller RS (1985) Integrated chemical-reaction sensor. In Digest of technical papers of TRANSDUCERS 85. 1985 International conference on solid-state sensors and actuators, 11-14 June 1985, Philadelphia, pp 33-36... 

The main important details of these procedures for AA determination are contained in Table 18.8. As can be seen in Table 18.8, among these flow analysis techniques, stopped-flow procedures are the most applied to AA determination, being mostly kinetic enzymatic determinations. BI methodologies for AA determination use spectrophotometric detection and a commercial flow cell, which is filled with appropriate solid beads, works as a flow-through chemical sensor integrating online reaction, retention, and detection on the solid-phase disposable beads. 

Two distinctly different coulometric techniques are available (1) coulometric analysis with controlled potential of the working electrode, and (2) coulometric analysis with constant current. In the former method the substance being determined reacts with 100 per cent current efficiency at a working electrode, the potential of which is controlled. The completion of the reaction is indicated by the current decreasing to practically zero, and the quantity of the substance reacted is obtained from the reading of a coulometer in series with the cell or by means of a current-time integrating device. In method (2) a solution of the substance to be determined is electrolysed with constant current until the reaction is completed (as detected by a visual indicator in the solution or by amperometric, potentiometric, or spectrophotometric methods) and the circuit is then opened. The total quantity of electricity passed is derived from the product current (amperes) x time (seconds) the present practice is to include an electronic integrator in the circuit. 

HPAEC analyses were carried out to determine the oligomeric products released from various pectic substrates after depolymerization by the PL isoenzymes. Action pattern analyses for the concerted action of PL isoenzymes utilized 68% esterified pectin as substrate. One-ml reaction mixtures in a buffer system as detailed in section 2.2. comprising 0.5% (w/v) substrate and 5 U of enzyme were incubated for 30 s to 18 h, and then thermoinactivated. Samples of 750 pi were applied to a Carbopac PA-1 (Dionex) column before the carbohydrates were eluted over a period of 70 min using a gradient of 0.2 M KOH, 0.05 M K-acetate to 0.2 M KOH, 0.7 M K-acetate. Detection employed a Pulsed Electrochemical Detector (PED, Dionex) in the integrated amperometry mode according to the manufacturer s recommendations. 

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