CHAPTER 20 FLOW ANALYSIS IN THE PROTECTION OF THE

ENVIRONMENT

Krystyna Pyrzyńska, Ewa Poboży, Marek Trojanowicz The Faculty of Chemistry of Warsaw University

ABSTRACT The short time available for determination, the mechanization of the procedure, as advanced as possible, or automation of the measuring system that allows operation without any control by personnel are among the most important requirements of contemporary environmental analysis – carried out both in the laboratory and in direct process control. Carrying out the measurements of the changes of the controlled chemical and physico-chemical environmental parameters – including their recording in the time span close to the real time regime – is, to a large extent, made possible thanks to flow analysis methods. The essence of theses methods is to carry out analytical measurements during the time of the sample’s flow through the detector. In industrial monitoring, most often in the measuring systems where sample processing is not employed, many methods have been applied for dozens of years – both to control the course of a process and to reduced the environmental risks involved. Progress in this field, observed in recent years, comes from wider and wider application in continuous monitoring such methods and devices which perform various transformations of the analysed sample such as buffering, transforming the analyte into a product which enables its detection, or transfer of the analyte into another phase. Development of laboratory flow analysis methods was launched in the 1950s by the determinations utilizing stream segmentation in order to limit sample dispersion in the measuring system. These methods, at first applied mainly in clinical laboratories, have recently become the standard methods of the determination of many waste components and in water quality control with the use of devices provided by many manufacturers. Making the laboratory flow analysis methods more efficient has been achieved by moving away from utilizing the equilibrium signal and basing the analytical determination upon measurements of the instantaneous signal in the detector. Such determinations are carried out mainly in systems without stream segmentation, but by injecting low volumes of the sample (microlitres up to fraction of millilitre). Procedures utilizing FIA (Flow Injection Analysis) and SIA (Sequential Injection Analysis) and many commercially available instruments are more and more frequently employed in environmental analysis and introduced to the legal acts by which environmental analysis is regulated. Such systems, when properly connected (interfaced) to the spectrophotometric devices allowing multicomponent determination or high performance chromatographs, can be very useful in speciation determinations and in the analysis of environmental samples characterized by complex matrices.

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1 INTRODUCTION Chemical analysis, like any sphere of human activity which makes use of scientific achievements and the development of various technologies, undergoes continuous development and improvement. This is brought about mostly by the growing need for taking advantage of making the analyses useful, but also by discoveries of new phenomena and new materials, stimulated by the development of the new technologies and by the intellectual competitiveness, which facilitates the scientific and technological progress. Development of flow analysis methods reflects many tendencies concerning the whole of analytical chemistry on which the contemporary chemical analysis is based. The need for mechanizing and automatizing the analytical procedure, the necessity of the constant improvement in efficiency, the growing requirements of precision and accuracy are the tendencies seen on every field where chemical analysis is employed. More elaborate approaches include: employing microelectronics and micromechanics to miniaturize analytical instrumentation, succouring known physico-chemical measuring methods by applying biochemical interactions observed in living organisms or employing equally advanced informatics methods and artificial intelligence in transforming and directing the data sets in the chemical analysis.

Flow analysis is one of the most frequently used methods of mechanizing various individual stages of the analytical procedure. Here, our attention should be drawn to the word mechanization – instead of the commonly overused automation. The latter concerns complex measuring systems, which not only mechanically use some activities (dilution, solution transferring, sample injecting, etc.), but also equipped with the control systems, functioning independently in the feedback regime and participating in decision making, e.g., concerning the change of the parameters of the determination carried out [1,2].

Flow analysis is one of many ideas for mechanizing analytical procedure – parallelly to making very complicated and sophisticated analysers: discrete, packet, centrifugal, and parallel [2]. The analysis is supposed to help carry out the stage of analyte detection under the conditions of the flow of the analysed sample through the detector. This is conductive to a number of crucial consequences, such as the proper design of the detector, the possibility of using kinetic effects and of moduling the speed of the analyte transporting to the sensitive element of the detector, the necessity of the effective data collection and processing, or the necessity of the particularly careful control of the transfer error, and also some advantages and limitations due to the use of the instantaneous signal – not the equilibrium signal - as the source of the analytical information.

It is difficult to indicate exactly the beginnings of flow analysis. It seems that these were the process electrochemical measurements of the conductance, the redox potential and pH, carried out in industrial installations and water and waste purification plants as early as in the 1940s. Habitually, the notion of the flow analysis does not include chromatographic methods, despite the fact that detection, which – in the column version – is already a hundred years old, has been carried out in the flow detectors. The beginning of the development of the laboratory flow analysis methods was 1957 [3], when Skeggs, in order to limit sample dispersion in the system, worked out the idea of flow measurements with stream segmentation, to limit sample dispersion in the system. The studies of Ruzicka and Hansen [4] in the mid 1970s initiated the development of injection methods in flow analysis, however, a few similar researches may be found in the subject’s bibliography. Each of these methodologies was undergoing different phases of development, it was found to be more or less interesting, depending on the particular fields of chemical analysis, and – with variable acceptance, it was gaining the

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status of the routine analytical method, equipped with the commercially available instrumentation, the enacted legislation, and the description of its employment. Even the commercial availability of the proper instruments does not mean that the methods can be ranked among the routine ones -- this can be explained by their absence in the legal norms and regulations in many countries. At present, the environmental analysis and process analysis are the largest sphere of the employment of flow analysis methods. 2 FLOW ANALYSIS METHODS The existing and utilizeded methods of analysis can be classified differently with reference to: the design of the measuring instrument, the way the measurement is carried out, the sphere of method’s employment, the applied instrumental detection methods, the state of matter of the introduced samples, and the kind of phase in which the analytical determination is carried out. From the perspective of the practical applicatios, one should distinguish between the methods (and instrumentation) where the analysed liquid or gas sample is introduced into the system in the continuous mode (without sampling) and the methods where the samples with the specified volume or mass are introduced into the measuring system in various ways in time intervals. The first of these methods, with the continuous collection of the analysed sample, are applied primarily in the process analyses of the installations where constant monitoring of the contents of an component or all components is necessary. This is less often applied on the laboratory scale. Such determinations are, to a different degree, carried out both with the use of the detection methods that do not require the transformation of the analyte and the methods where the chemical reaction is applied in order to make a product whose content is, in turn, measured in the detector. The methods with periodic sampling are primarily the domain of the laboratory systems – however this is not their exclusive feature. We can distinguish here three groups of methods. In the continuous CFA (continuous flow analysis) methods with stream segmentation, the air bubbles of the same size are introduced into the channel in order to limit the dispersion of the analyte under the flow conditions, which, in turn, is supposed to lead to the best possible effectiveness of determination (shortening of the determination time span). The samples are sucked into the system in turns with the carrier solution [5]. In a number of the injection methods, the microlitre volumes of samples are introduced into the system without stream segmentation. In the flow injection analysis (FIA) a sample is injected into the carrier that flows continuously in the system [6 –8]. In the method of sequential injection analysis (SIA), adequate design of the apparatus permits introduction a few samples and reagent zones into the system, which, mingling on the way to the detector, enable the detection of the analyte [9]. In these methods, on the way from the point of sample injection to the point it reaches the detector, a number of varied sample transformation methods may be carried out. In turn, in the method of the direct injection to the detector – BIA (batch injection analysis) – which may be defined as the tubingless flow injection analysis – one doses small volume of the sample directly onto the sensitive surface of the detecting element [10]. In yet another kind of the injection method, that is BFIA (bead flow injection analysis), the transformation of the analyte together with the detection takes place on the suspended particles of the properly selected solid materials introduced to the flow system [11]. The examples of the diagrams of FIA and SIA flow systems are presented in Figure1.

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Flow injection analysis (FIA)

Sequential injection analysis (SIA)

Figure 1. The diagrams of the FIA and SIA flow systems.

3 THE FLOW METHODS IN THE ENVIRONMENTAL PROCESS ANALYSIS Process analysis constitutes a huge section of contemporary analytical chemistry, developed for more than 50 years on the borderland of chemistry and chemical technology. However, it is treated as marginal in the university education of analysts, it already has a vast monographic bibliography and a huge diversity of the measuring devices constructed specially for the process analysis applications [12 – 16]. Together with the most commonly used spectrophotometric (UV/VIS) and electrochemical analysers, in the process analysis one also uses gas and liquid chromatographs, the IR and Raman spectrophotometers, mass spectrometers and nuclear resonance spectrometers. The way the analytical instruments are used gives rise to the situation when, in most cases, either the devices themselves serve as flow analysers, where detection takes place during the flow of the liquid or gas sample through the detector, or the properly made probe (sensor) is introduced directly into the analyzed stream. In a number of cases, depending on the measuring process itself, the periodic instruments, e.g., process chromatographs, mass and NMR spectrometers may also serve as the analyzers. Generally speaking, the fundamental aims of process analysis are: control of the course of the process and its safety, control of the raw materials, reagents and products, control

Waste Carrier solution

Reagent

Pump

Sample

Detector

Mixing spiral

Waste

Pump

Carrier solution

Sample

Detector

Reagent Standard

Selection valve Mixing

spiral

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of the energy consumption in the process, and control of the pollution of the environment. The efficiency of employing the analyser with certain functional parameters necessary for controlling the process is expressed by the parameter defined as measurability which takes into account the precision of determinations, the sampling frequency, and the delay in receiving the results of determinations [16]. The place of installing the analyser that monitors the process may be crucial, too [17]. The profitability of installing the analyser may be basically influenced by its servicing and maintenance requirements [18]. The process analytical measurements may be carried out both in the laboratory – far from the installations – or directly at the technological stream, using the submersion probes, analysers with either the continuous or periodic sample collection [19], as well as employing noninvasive detection methods. Usually, the analysers installed at the technological stream differ substantially from their laboratory conterparts in the required resistance to the aggressive chemical environment and the disturbances caused by the electric field, in the necessity of equipping them with data transmission and the remote control systems, and – particularly – in the durability and operational reliability. Process environmental analysers are a very wide range of devices, used to control the level of pollutants in waste and in the close environment of various industrial installations, the processes of the waste treatment, purification of the exhaust gases or conditioning the natural waters for the industrial or municipal purposes. Spectrophotometric and electrochemical analysers are most commonly used, as far for monitoring the level of various components in solutions (waters, liquid wastes) is concerned; although, UV and fluorimetric analysers are used as well. A typical process analyser, manufactured and installed nowadays, is a device controlled by microprocessors, equipped with a user-programmed systems, autocalibration systems, malfunctioning warning systems, systems warning against exceeding given concentration limits, systems of assistance and self-starting in the case of failures in power supply or data transmission. Environmental spectrophotometric analysers are in most cases based on the principle of absorption of light by the colored products of the reaction between the analyte and the appropriate reagent, and either on acid-base or complexometric titrations with the photometric end-point detection. They are manufactured by many companies in numerous countries, such as Seres in France [20], Hatch in the USA [21], Skalar in Holland [22], Lange in Germany [23], Polymetron in Switzerland [24] and many more. For example, in the wide offer of Seres, apart from the analysers of many inorganic anions and cations one can find, for the need of water and waste analysis, flow analysers for detecting hydrazine and phenols, analysers with near IR detection for the determination of Total Organic Carbon (TOC) and – also IR based – analyser for the determination of the content of oils in the waters and wastes. OPAL analyser (organic pollution alarm), manufactured by GLI International [25], enables the detection of volatile organic compounds with the photoionization detector. The probe used for the process measurements of nitrates, based on the UV radiation absorption, is offered by Dr Lange company [23]. Electrochemical analysers equipped with the ion-selective electrodes are a large group of the process flow analysers used broadly in the environmental analysis. Among others, these are the devices of such firms as the aforementioned Polymetron, GLI International, Contronic from Sweden and Environment from France [26]. The process analysers used for the simultaneous determinations of, e.g., ammonia, fluorides, and nitrates in waters and wastes are manufactured as well, and the brand new Blue Box system manufactured by Bran and Luebbe [27] may serve for the simultaneous control and data transmission from 100 different sensors and detectors. Amperometric detection

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is used in many different process analysers, e.g., to determine the dissolved oxygen, hydrazine, free chlorine, ozone, and nitrates – this being offered by Polymetron [24]. This very same manufacturer also delivers the flow analyser used for determining methanol for the control of the processes of the biological denitrification on the biofilters and in the wastes where a biosensor has been employed. Amperometric detection with the bacterial biosensor is, in turn, used in the flow analyser employed for the continuous measuring of the biological oxygen demand (BOD) – this is manufactured by a Japanese firm Nissan Electric. 4 LABORATORY CONTINUOUS FLOW ANALYSERS USED IN THE ENVIRONMENT PROTECTION The idea of laboratory continuous flow analysers and the stream segmentation with the use of the air bubbles, which had appeared at the end of the 1950s, mechanized, for the first time so efficiently, carrying out wet analyses in all kinds of the laboratory analysers. It was the result of performing simultaneously – under the flow conditions and with limited sample dispersion –many operations of transforming the sample and, additionally, many various detections. The standard AutoAnalyser instruments manufactured by Technicon became available shortly after the publication of the first research and patenting the invention. In the first place they were used in the clinical laboratories, but gradually it was also introduced to the environmental, agricultural, and industrial laboratories [5, 28, 29]. The analysers in the clinical laboratories evolved during the next twenty years, reaching the stage of complex combines for the rapid multicomponent determinations, such as SMAC by Technicon. Despite this development, they were relatively early ousted by varied discrete analysers manufactured by many other companies. This trend could not be reversed by modifying the analysers into the efficient, single-stream devices, employing the capsule chemistry technology, e.g., clinical analyser CHEM 1 by Technicon. Flow analysers with stream segmentation, recording the equilibrium signal, were much slower accepted and improved – with the improvements taking place mainly in the manufacturers’ laboratories. They are manufactured exclusively for the environmental and industrial laboratories by few specializing companies in various countries (e.g. Bran and Luebbe in Germany [27], Lachat Instruments in the USA [30], Skalar in Holland [22], Alliance Instruments in France [31], and Burkard in Great Britain [32]). For many years, they have not been the object of any scientific research, the results of which would be published in any scientific journal of international circulation. Apart from bibliography and application notes provided by the manufacturers, many methods are included to the standard determination methods, e.g., colorimetric determination of ammonium nitrogen [33] or nitrates (III) and (V) in the waters. Laboratory flow analysers, built in recent years for the need of the environmental laboratories, are mostly modular devices used for the simultaneous determination of from a couple to a dozen different components with two kinds of detection: either spectrophotometric or potentiometric with ion-selective membrane electrode. Usually they allow analysis of several samples per hour, and the number of applications developed and next provided to the users usually reaches a few hundreds – which includes analytical procedures for, e.g., food products, fertilizers, soil extracts or pharmaceutical preparations. These are mostly determinations of various inorganic components in waters and in wastes. There are also organic analytes, e.g., detergents, phenols, urea, metalorganic lead compounds, and a number of various parameters: chemical oxygen demand, water hardness, total nitrogen by Kjeldahl method, total

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acidity and alkalinity. For many analysers, e.g., TRAACS by Bran and Luebbe and Lachat analysers, various local and international organizations (ISO, EPA, AOAC, etc.) suggest certain regulations which concern the methods developed by these organizations. In many cases a flow analyser for a specific purposes can be equipped with other detectors: for example SAN analyser by Skalar can cooperate with a flame photometer in order to determine sodium and potassium in soil extracts and fertilizers, and with an IR detector in order to determine the total organic carbon using the persulphate method. Several Lachat analysers are connected to the high-performance ion chromatographs. One of the most inviting features of flow analysers is their mechanized carrying out many operations of transforming the sample, such as dilution or reactions with various reagents. These are much more complicated and labour-consuming operations when performed manually or in the discrete systems such as: mineralization in determinations of total nitrogen or phosphorus, or the flow distillation in determinations of phenol, fluorides and cyanides. In the case of complex determinations which require many operations for very complex samples (e.g., heavily loaded wastes), single component laboratory analysers are employed, such as single channel TRAAC analyser from Bran and Luebbe, which is equipped with a flow dialysis unit, SINGLE from Alliance Instruments which is used for phenol, cyanide, and total organic carbon determination, as well as ammonia analyser from Timberline Instruments [35]. 5 THE EMPLOYMENT OF THE FLOW INJECTION METHODS IN THE PROTECTION OF THE ENVIRONMENT As opposed to the standard methods and to the stream segmentation analysers, applying injection analysis enables carrying out the measurements before reaching the state of equilibrium. As long as the configuration of the system remains unchanged, the equilibrium signal is not necessary for measuring. In FIA and SIA methods, it is possible then to carry out – in mechanized mode – various environmental analyses with much higher frequency. The design of the flow system depends both on the chemical aspect of determination and on the detector employed. The systems which are characterised by small dispersion of the sample zone are used for determinations where the modification of the sample is not required because of employing selective detection method. It is often the case that in order to attain the proper sensitivity and selectivity of the measurement, the composition of the sample in the systems where the dispersion is bigger needs to be modified. The reaction part of the flow systems may include separation modules where the processes of diffusion, dialysis, and extraction are used – together with employing the reactors where the redox processes, sorption, ion exchange, and enzymatic or immunochemical reaction take place. Flow injection techniques can be also applied to multicomponent determinations through the change of configuration of the system (parallel or serial arrangement of suitable detectors) or through taking advantage of the difference in kinetics of the reactions of individual analytes.

5.1 The employment of FIA and SIA with the spectrophotometric and

electrochemical detections The greatest number of the studies described in literature is devoted to employing the FIA and SIA methods to the analysis of the surface, ground, drinking, and sea water. Other kinds of the analysed environmental samples are waste, sediments, air and aerosols. The two things that are determined in water samples are macro- and microcomponents – the former being Na, K, Ca, Mg, chlorides, nitrates, sulphates,

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phosphates, and the ammonium ions, which are found at mg 1-1 concentration level, while the latter are some inorganic ions (fluorides, nitrites, cyanides), metals, and the organic and inorganic compounds present at trace concentration level. Because of their qualities, flow methods have been used to monitor the selected components of water for years. In such analyses, the pretreatment and filtering the sample is frequently required. As far as monitoring is concerned, it is important to carry out these two stages in the flow system as well. Wang and others [36] presented the automatic FIA system enabling, with the use of ultrasounds, the continuous filtration of the samples of river water with 90% efficiency. In such a system PO4

3-, NO2-, NO3

- and NH4+ were

determined. In the protection of the environment, frequently controlling the concentration level of such organic components as pesticides, phenol-derivatives, or surfactants becomes important. The most recent studies concerning the employment of FIA and SIA methods in the environmental analysis are discussed below.

In environmental analysis, equally important is elaborating the methods that permit to carry out multicomponent and speciation measurements. One of the ways of conducting these measurements is the simultaneous use of several detectors, e.g., potentiometric detectors with integrated ion-selective electrodes [37, 38]. The simultaneous determination of fluorides and phenolic compounds was made possible with employing fluoride ion-selective electrode and the amperometric detector [39]. The automated SIA systems, whose design enables multicomponent measurements of NO2

-, NO3

-, NH4+, PO4

3-, the total nitrogen and phosphorus can be applied to monitor their concentration in the wastes [40].

The FIA and SIA systems are also applied for determination of the elements on different oxidation levels. Most often, one of the forms is determined directly and, after the reaction of either oxidation or reduction, a total concentration of the analyte is measured. The concentration of the second form is calculated as the difference between the two results. This is the way the nitrates (III)/(V), Cr (III)/Cr(VI) [42], Fe(II)/Fe(III) [43,44] and Se(IV)/(VI) [45] may be determined. In the iron speciation determinations the systems with two detectors are used [46]. The content of Fe (II) was determined with the use of the spectrophotometric detector, and the total concentration of iron with FAAS method. Introducing the sample between two reagent zones – and this is possible in SIA systems – enabled to determine simultaneously NO2

-, NO3-, Fe (II) and Fe (III)

[47]. The selectivity and sensitivity of Cr (VI) and total aluminum determination was improved by introducing an additional stage of extraction to the organic phase. A new automatic FIA system, serving for the nitrogen compounds speciation in sea water, was presented by Tovar and others [49]. The method is based on determining the absorbance of azo dye formed in the reaction between nitrates (III) with N-(-1-naphthylo)ethylenediamine and sulphanilamide. Ammonium ions were determined after oxidation to nitrates (III), and nitrates (V) after reduction to nitrates (III). The list of FIA methods used in aluminum speciation in environmental samples is presented in articles [50, 51].

Frequently, in analyses of environmental samples, determination of a given compound – present at low concentration in a complex matrix – is required. In determinations of pesticides, this requirement is met by the FIA method linked with the biosensors and the immunochemical systems. The pesticides were determined using the potentiometric detection based on inhibition of acetylcholinesterase immobilized in the enzymatic flow reactor [52, 54]. The enzyme may be immobilized in the layer of polymer on the surface of a platinum electrode [54] or silk-screen printed electrodes[56, 57]. Employing the system with three enzymes permitted the simultaneous determination of several compounds [54]. Application of the FIA system with a graphite

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electrode and fixed tyrozinase leads to the results remaining in agreement with those obtained by the traditionally accepted method of determination of the phenolic compounds in the environmental samples [58].

To determine the herbicide of 2,4-dichlorophenoxyacetic acid (2,4–D), the FIA system equipped with various immunochemical reactors and amperometric detection was employed. The column reactor with immunoglobuline G deposited on the porous bed was found the best solution to the problem. The same herbicide was also determined after the employment of the amperometric immunosensor made of a gold electrode modified with cystamine, to which 2,4–D was attached, while the antibody was conjugated with peroxidase [60]. Other kinds of detection, different from the amperometric detection can be employed to the flow systems with the reactors. To determine the 2,4–D electrochemiluminescence was used where, as the result of the electrode reactions, a luminescent compound is formed [61]. To determine herbicides, the fluorescence detection was also used [62 – 64]. Aaron and Coly [65 – 66] presented the review of FIA methods with both the luminescence and fluorescence detection earlier applied to pesticide determination in the environmental samples.

Ramanathan and Danielson [67] described the use of the thermic biosensors to detect the pesticides in FIA systems. The estimation of the analytes’ concentration was based on measuring the changes in temperature during the time of the enzymatic reaction (the reaction of the pesticide hydrolysis by the enzyme or the enzyme inhibition). The biosensor based on tyrozinase was used to monitor the process of biodegradion of the phenolic compounds found in waters, wastes and sediments [68].

In the FIA system, the anionic surfactants in the surface waters and in the wastes were determined by employing potentiometric detection with the cylindrical ion-selective electrode [69]. Coupling this approach with the continuous phase extraction, the determinations on the level 0.03 mg 1-1 were made possible. Determination of the cation surfactants was carried out using the spectrophotometric detection, based on measuring the changes in the absorbance of Fe(III)–SCN complex in presence of these compounds [70].

An important parameter, defining water quality, is the presence of harmful bacterias and toxins. For their analysis, FIA systems with immunochemical reactors and amperometric [71,72] or fluorescence [73] detection are used. A columnar reactor containing microorganisms immobilized on the surface of porous glass has been used for determining trichloroethylene in water [74]. As a result of biodegradation, chloride anions have been formed and their concentration has been measured with ion-selective chloride electrode.

Anodic inversion voltammetry (anodic stripping) is a sensitive and cheap method of detection of trace amounts of metals in the FIA/SIA systems. The employment of this technique in flow systems allows simultaneous measurements of Cd(II) and Pb(II) [75-77], and the pulse voltammetry of Cd(II), Pb(II), Cu(II), and Zn(II) [78]. Such systems were used in analyses of Cu(II) in drinking water [79] and Cu(II), Pb(II), Cd(II), and Zn(II) in river sediments [80]. A mercury film electrode was used as a working electrode. For determination of phosphates in drinking water and wastes, amperometric detection was used together with the process of the reduction of molibdenophosphoric acid to molibden blue on a glassy carbon electrode [81]. Potentiometric detection with ion-selective electrodes was used in measuring the concentration of chlorides and fluorides in waters [82,83]. Spectrophotometric detection is one of the most frequently used in the flow systems. It is the result of the improvement of detectors in which photodiodes and the optical wave-guides were used. This allows performing measurements in a wide range of wavelengths, assuring good sensitivity of the

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measurements and low internal noise of the instrument. For the determination of metal cations, reactions yielding colored complexes are used. Reaction of complex formation with aluminon and chromazulon S was used to determine Al(III) in the soils [84]; with diethyldithiocarbamate – to determine Cu(II) in the water samples [85]; with o-cresolopthaleine – to determine Ca(II) in drinking water [86]. Trace concentrations of Co(II) and Ni(II) in waters and soils were determined spectrophotometrically after the reaction with PAR [87]. Employing fluorimetric detection allowed to determine Mg(II) after its reaction with 8-hydroxychinoline-5 sulfonic acid in the brands of the mineral water available on the market [88]. Employing two selective reagents, that is malachite green and ammonium thiocyanide enabled to determine simultaneously Cd(II) and Zn(II) [89]. Improving the sensitivity and selectivity of the spectrophotometric detection is possible thanks to employing the optodes [90]. The optode with Nafion membrane and with the immobilized organic ligand (PAN) was used for determinations of copper in the river water. This method’s detection limit was 15 µg/l. The agreement between the results obtained, as compared to those by AAS methods, was good.

Inorganic anions in the environmental samples were determined using color yielding reactions, specific for a given ion, linked with spectrophotometric detection, are most frequently utilized. In this way nitrates(III) [91, 92], phosphates [93-95], sulphates(VI) [96], thiocyanides [97, 98], and iodides were analyzed. Very often, the preconcentration is necessary because of the low concentration of nitrates(III) in the environmental samples. To attain this, the microcolumns C18 were used and the preconcentration was being conducted after the reaction between the sulphanilamide and N-(-1-naphthyl)ethylenediamine [100]. Ammonium ion in the SIA systems were determined in waters and industrial wastes [101] and in aerosols [102, 103]. These systems had also the additional diffusion modules, in order to liberate ammonium ion in the form of gaseous ammonia which would pass trough a semi-permeable membrane. The employment of both the flow system and the spectrophotometric detector (in its measuring cell the Sephadex QAE A-25 bed was fixed), allowed to determine various phenolic derivatives: phenol, 2-naphthol, 3,4-dimethylophenol, 1-naphthol, and 2,4-dichlorophenol – after their preconcentration [104]. Ten times higher sensitivity (as compared with conventional FIA systems) was attained in the system where the desorption of the analyzed compounds took place directly in the detector. SIA method with the spectrophotometric detection was used for determination of the phenolic compounds in the waste, employing the color yielding reaction with 4-aminoantipirine [105], and the UV detection served for determinating the aromatic hydrocarbons [106]. In the determinations of many organic compounds, it is necessary to use, as far as the flow system is concerned, the extraction to the organic phase. Teflon tubing with the an organic phase forming a film on its internal surface [107] were applied for these purposes. Such systems were used to determine the nitro-phenolic compounds in the wastes. 5.2 The application of the BIA methods in environmental analysis In the tubingless method of BIA analysis, the low volume sample (of a few microlitres) is directly introduced on the surface of either the working electrode or another detector, e.g., the optical one, allowing determination of the analytes during the flow of the solution at the detector’s surface. The examples of the possible designs used in the electrochemical detectors for BIA are shown in Figure 2.

Seeking new, selective and sensitive electrode materials is the main trend in the development of this method. In the environmental determinations, BIA is used primarily to determine trace metals Pb(II), Cd(II), Cu(II), Zn(II) with the anodic stripping

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detection. Working electrodes are most often thin layer detectors with the mercury electrode, carbon electrodes [109-111], carbon fibre electrodes or mercury covered graphite electrodes [112-114]. In order to improve the selectivity and to lessen the influence of the sample matrix, the modified electrodes, covered with polymers [109, 110] are used. In BIA method, for the determination of metals, the electrodes covered either with Nafion [115-117] or with a polymer containing the sulphonic groups were most frequently used. Employing the modified electrodes enables to determine the metals in the presence of interfering surfactants. Despite the fact that the currents registered for these electrodes are lower than for the unmodified electrodes, the measurements on the nM level [119] are possible. Turyan et al. [113] proposed application of a rotating disc microelectrode in the BIA technique, made of mercury covered graphite, for the determination of Pb(II), Zn(II), and Cu(II). With 5 minutes concentration time, the detection limit was established at the level of 30 µg/l. BIA method was used to determine metals in the natural waters [109, 115 – 118] and the industrial wastes [111, 114, 115].

Figure.2. Diagrams showing cross sections of electrochemical detectors for the BIA

method. (A) a large-volume detector with solution inlet perpendicular to the surface (wall-jet) [120], (B) a capillary detector with a paste working electrode [121], (C) thin layer detector with a rotating disc electrode [113]. In (A): A-disc electrode, B-ring electrode, C-auxilliary electrode, D-reference electrode. In (B): 1-paste working electrode, 2-Teflon capillary, 3-sample solution, 4-channel for sample injection, 5- auxilliary electrode, 6-reference electrode. In (C): 1-inner cell, 2-Teflon capillary, 3-sample, 4-outer large-volume cell, 5-base electrolyte, 6-sample channel, 11-ceramic junction, 12-rotating working electrode, 13- auxilliary electrode, 14-reference electrode.

5.3 Detection by atomic spectrometry and mass spectrometry. Coupling the flow-injection methods with the methods of absorption and emission atomic spectrometry, as well as mass spectrometry with ionization in inductively coupled plasma (ICP-MS), can improve the parameters of the latter ones, mainly through introducing into the flow system various techniques of sample processing. Direct introduction of the sample with dilution in the flow system, analyte concentration, or generation of volatile compounds are examples of operations illustrating the advantages of joining FIA or SIA with flame or electrothermal absorption atomic spectrometry (FAAS, ETAAS), atomic emission spectrometry or

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mass spectrometry with excitation in inductively coupled plasma (ICP-AES, ICP-MS). Apart from the techniques of isolation and preconcentration, much interest lies upon mineralization of a sample in the flow conditions [122], especially when it is the slowest stage of the analytical procedure. The processing of the sample in the flow conditions is also helpful in determining the forms of the element on different oxidation levels, which is used in speciation [123, 124]. The basic aim of the analytical procedure is the exact determination of the analyte in samples with various matrices. It is often a very difficult task in complex matrices, due to the fact that spectral and non-spectral interferences may appear [125, 126]. In many cases, especially when using the FAAS or ICP-MS methods, detectability may be insufficient. Although the ETAAS and ICP-MS methods make it possible to attain lower detection limits, but it is often accompanied by much stronger matrix effects. The measurement in the ETAAS method is disturbed by high content of salt in the sample, and is susceptible to changes in the matrix composition [127]. In many cases, strong disturbances of matrix origin cannot be adequately eliminated with the accessible background correction methods. Using organic solvents should also be avoided, for they lower the precision of measurements and sensitivity of the detection. Both, the high salinity of the sample and the presence of considerable amount of organic solvents, cause strong disturbances in ICP-MS. Non-spectral matrix interferences can be eliminated by diluting the sample; yet, it worsens the detectability [128]. The internal standards can only be used when they are well chosen for the analytes in regard to their mass and ionization energy. Therefore, in order to avoid these difficulties, it is advisable to process the sample in a way that allows the improvement of the detection limit, both by elimination of the inteferences and concentrating the analytes [129]. Such operations can be successfully used in FIA and SIA systems, with good repeatability and limited risk of contamination. Various procedures of the in-flow separation and concentration, based on ion exchange and adsorption, precipitation and co-precipitation, solvent extraction and volatile hydride generation, have already been developed. Miniature columns with various sorbents and open hydrophobic (e.g. Teflon) tubing are most frequently used for analytes sorption. Many research papers, as well as review articles [130–134] and books [8,135,136], have been devoted to these issues. Many scientific publications concerning these matters can be found in novelties from the atomic spectrometry field published in Journal of Analytical Atomic Spectrometry [137 – 139]. Columnar sorption in the FIA and SIA systems. Systems using the analyte sorption on suitably chosen packings of microcolumns belong to the most often used methods of sample processing and modification of its composition in the flow systems. These techniques are used mostly to improve the sensitivity of determination with the FAAS and ICP-AES methods; and to separate the determined components from the matrix components, which cause interferences in ETAAS and ICP-MS. Scientific publications offer various configurations of the flow systems using microcolumns. The differences may be found mainly in the way of concentrating/separating the analyte, the kind of the packing of the microcolumn, its position in the system, and the way the eluent solution is injected into the electrothermal atomizer when using the AAS detection.

Two ways of the sample processing and modifying its composition using microcolumns can be distinguished. In the first one, the constant and known volume of solution dosed through the loop of the injection valve, is pumped through the microcolumn bed. In this case, usually used to isolate the analyte, the volume of the

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sample is limited (usually to 1 ml) due to its dispersion; yet, the influence of the pulsation of the used peristaltic pump is eliminated (repeatable sample volume). In the second method, with the constant time of the analysed solution flow through the microcolumn, the analyte can be concentrated from a larger volume of solution; at the same time reaching higher enrichment factors (EF). Repeatability of the process carried out in this way depends, to a great extent, on the stability of the solution flow through the bed. The efficiency of the analyte concentration method and, consequently, the attained EF values, depend on the time of the solution flow through the microcolumn. Conducting the stage of the concentration for a longer time at the same flow rate, causes the EF increase, but the number of samples analysed in a given time span decreases. In order to compare the efficiency of the used flow systems with concentration, the concept of concentration efficiency (CE) or concentration factor (CF) is introduced. CF is defined as the product of the EF and the number of samples handled per a unit of time, e.g. one hour. For both methods of injecting the sample onto the microcolumn (at constant volume or at constant time), the increase of the analysed solution flow rate, which assures the high efficiency of this method, is, however, limited by the kinetics of the analyte sorption process.

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Pic.1. Diagram of the flow system with the ETAAS detection with a microcolumn

Figure 3. Diagram of a flow system with the ETAAS detection and the microcolumn placed at the end of the autosampler arm. P1, P2-peristaltic pumps, V-valve.

The selectivity of the sorption process depends mainly on the choice of the microcolumn packing. The packing should have appropriate physical and chemical properties, good kinetics of analyte sorption and desorption, and its volume should not change when the acidity of the flowing solution changes. In the flow systems with columnar sorption, cation and anion exchangers, chelate resins containing functional groups bound with polymeric matrix with covalent bond, and non-ionic sorbents used for extraction of neutral metal complexes, are employed. Yet, normal cation- and anion-exchange resins have rather low selectivity because the differences in their affinity to, e.g., metal ions, are related to their physical properties, such as charge or size of the solvated ions. However, changing metal ions into negatively charged complexes, e.g.,

Concentration stage Autosampler arm

Sample

Reagent

Waste

Microcolumn

Eluent Washing solution

P2

P1

Elution stage

Sample

Reagent

WasteP1

P2

V

V

Microcolumn

Graphite cuvette

Eluent Washing solution

Waste

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chlorides, makes it possible to separate them from other, positively charged, matrix components on the anion-exchange resins. Chelate resins show ion-selective properties, and their affinity to metal ions is basically related with complexing properties of the functional group of these exchangers. Among many chelate exchangers, Chelex 100 and Muromac A-1 resins with iminodiacetic groups were used for metal ion concentration and isolation [129,131,132]. Ion exchanger with fibrous matrix (fibrous materials), e.g., of cellulose type, despite their worse mechanic properties, turned out to be very useful in flow systems, due to their good kinetics of analyte sorption and desorption [140-143]. The properties of other exchangers used in the FIA and SIA methods are described in Fang’s works [129,135,136].

Despite many new syntheses of solid sorbents, another method of obtaining chelate stationary phases is used. They are obtained through immobilization of complexing reagents on anion-exchange resins or non-ionic sorbents as the result of the ionic exchange and/or physical adsorption [144 – 147]. This method allows the control of their ion exchange capacity and selectivity through the choice of a selective organic reagent, appropriate for specified metal ions. Also hydrophobic stationary phases, such as C18 silica gel or polymeric sorbents of the XAD type, are used in flow systems for the metal complexes sorption, mainly with dithiocarbamates (DDTC, APDC) and ammonium diethyldithiophosphate (DDPA). DDPA is more selective than dithiocarbamates and its solution is more stable in acid environment. Among other complexing reagents, 1-nitroso-2-naphthol [148], 1,10-fenantroline [149], and porphyrins [150] are worth mentioning. The structure of the flow systems with the columnar sorption with the FAAS and CP-AES detection is very similar and both methods of introducing the sample (at the constant volume or constant time of the solution flow through the column) are used. The eluent stream, most frequently it is a mineral acid solution or, in the case of non-ionic sorbents, organic solvent, after the elution of concentrated analytes, is directed into the detector’s nebulizer. Using ETAAS as a detection method demands different setup of the system. The stage of the sample concentration on the microcolumn can in this case be conducted simultaneously with the on-going processes of pyrolysis and atomization in the graphite cuvette or the flow of the solution into the cuvette and during the measuring of the absorbance. Contrary to the measuring in the flame AAS, with the employment of the electrothermal atomization the sorbent deposit has to be washed after the concentration stage due to the possibility of interference from the remains of the matrix components. The volume of the solution injected into the graphite cuvette is limited to 50-70 µl, but the volume of the used eluent used in the process of analyte elution is often greater. Therefore various suggestions are put forward, such as slow dosing of the solution from the microcolumn into an initially heated atomizer, repeated dosing of the eluate with evaporation of the solvent at each dose, and injecting only this part of the eluate that contains the highest concentration of the analyte. In order to lower dispersion, individual solutions are separated by short air bubbles. In Figure 3, a diagram of a flow system with ETAAS detection is shown; in this system, the microcolumn is placed at the end of the autosampler arm. Sorption on a column can also be employed in concentrating the analyte at the site of natural water sampling [151,152]. Later, the microcolumns are taken to a laboratory and installed into the FI or SI system in order to perform the process of elution and final detection. Such procedure in trace analysis and speciation studies, apart from the advantages of using a closed flow system, is also efficient and precise.

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The employment of hydrophobic channels to concentration Open Teflon hydrophobic tubing, arranged in proper patterns – therefore their name: knotted reactors – were initially used as filterless precipitate collectors in the methods of the analyte isolation and concentration based on precipitation. These methods have been proven to be very useful in collecting large quantities of precipitates with low resistance to the solution flowing in such systems. Because of the configuration of the channels, the solution inside the tubing continuously changes the direction of the flow, and the aroused centrifugal force directs the particles of the precipitate towards the inner walls, where they accumulate easily. The precipitate is subsequently rinsed and dissolved in a small volume of a properly chosen eluent, which is immediately directed to the detectors (Figure.2). If the ETAAS detection is used, the solution is separated by bubbles of air; if the ICP-MS method is used – the solution is separated by small volumes of the carrier solution [153]. Burguera et al. [154] used the high volume of knotted reactors to determine iron in geothermic samples containing considerable amount of sulphur compounds. Their presence in the analysed sample causes significant decrease of the absorbance value in the ETAAS method. They were isolated from the analyte in the form of free sulphur precipitated by using hydrogen peroxide stream and irradiating this fragment of the system with microwave radiation. After dissolving in CCl4, sulphur was directed to the waste, and the solution containing iron was introduced into the autosampler arm of the graphite cuvette. In order to remove the interference from the sulphates remaining in the sample, the solution of lutetium salt was used as a chemical modifier. In recent years Teflon tubing hydrophobic reactors have been most frequently used for sorption of neutral metal ion complexes with organic ligands, as an alternative method of concentration on octadecylsilanol C18 column packings. In this method, the interactions between the inner wall of the channel and the metal complexes present in the solution are utilized [155]. The solution of the sample can be pumped through such reactors with high velocity because of much lower resistance in comparison to microcolumns, and so the analyte can be concentrated within a given time span from a much larger volume. However, the efficiency of the sorption is lower, only 40-50 %. A different system of a channel reactor was also examined, the so called serpent reactor in the shape of the figure “eight” [156], but its geometrical shape decreases the centrifugal force that directs the solution towards the channel walls and therefore the sorption is also abated.

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Figure 4. Precipitate concentration method using a knotted reactor

DDTC, APDC, and DDPA are used both as organic ligands forming neutral complexes with metal ions [157,158] and in methods of column sorption with non-polar sorbents. Selective formation of complexes of As(III) and Fe(III) with APDC was used in speciation studies of inorganic arsenic and iron forms in natural water samples [159]. The mechanism of creating ionic pairs can also be used for the concentration and isolation of the metal ions present in the solution as charged complexes. To determine cadmium with the ETAAS detection, the sorption of ionic pair was used. The pair was formed from the negative Cd(II) complex with nitroso-R-salt and tetrabutylammonium ion [160]. In order to avoid analyte loss during the washing out the residual amounts of the sample matrix remains from the inside of the tubing reactor, complexing reagent should be added to the washing solution (it is usually de-ionised water or diluted acid solution). It inhibits dissociation of the complexes formed, especially for lead, tin, and bismuth ions [157]. Solutions of methanol, ethanol, and methyl-isobutyl ketone are used for the desorption of concentrated complexes. For plasma detectors, however, diluted nitric acid solutions or organic solvents, injected through suitable membranes, are recommended as eluents [161,162].

SI-LOV systems In the flow systems of separation and concentration, based on ionic exchange and adsorption, sorbents are constant and indispensable elements of the system. Yet, their long-term exploitation can cause changes in the volume of the microcolumn packing and, as the result, the resistance of the flow increases considerably.

The properties of the employed sorbents can be changed because of deactivation or functional group loss. In order to avoid these disadvantageous phenomena, the packing of microcolumns should be replaced after each measurement, in a repeatable way. It is possible with the employment of a special valve (LOV – lab-on-valve) [163], the functioning of which is based on connecting a sequence of flow microchannels. The valve cooperates with the standard multi-positional injection valve used in SIA. The

Concentration stage

Elution

Waste

Analyte

Matrix Sample

Reagent

Air

ETAAS ICP-MS

Carrier solution

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employment of the SI-LOV system makes it possible to take various (low) sample volumes with large repeatability and to add reagents in appropriate points of the system, as well as and carrying out the separation/concentration processes using column sorption [164].

Figure 5. SI-LOV system with a reproducible column reactor The flow system with the LOV valve and reproducible microcolumn packing was

used for the determination of trace amounts of nickel and bismuth in environmental samples with the ETAAS [165] and ICP-MS detection [166]. The diagram of this system is shown in Figure 5. Cationic ion exchanger SP Sephadex C-25 with sulfonic groups constituted the packing of the microcolumn. Specified amount of exchanger was introduced into the system from ionite suspension (position 6 in the valve); this exchanger could freely move between position C1 and C2 of the microcolumn during the stage of the analysed sample concentration. The rinsing of the ionite bed was followed by elution of the analyte with nitric acid solution. The eluate was then directed to graphite cuvette of the AAS spectrophotometer or the nebulizer of the ICP-MS detector. After the whole measurement cycle was finished, the grains of the exchanger were removed from the microcolumn channel by altering the carrier solution flow rate. The grains (40–125 µm in diameter) of SP Sephadex C-25 cation exchanger are held in the microcolumn C1 when the flow rate is low (< 1.2 ml/min.). However, when the flow rate exceeds 6 ml/min, the size of the grains decreases – they are crushed and can easily be removed through the opening at the end of the microcolumn.

Carrier solution

Pump

Mixing spiral

Sample

Eluent

Air

Detector

Sorbent suspension

Microcolumns C1 and C2

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Figure 6. Diagrams of procedures used for analyte concentration/isolation in the SI-

LOV system with reproducible microcolumn packing. (A) after the concentration stage the analyte is eluted from the bed and determined with the ETAAS or ICP-MS method. (B) the microcolumn packing together with the concentrated analyte is directly dosed into the atomiser in the ETAAS method.

With the employment of the SI-LOV system a different procedure can be used

with the ETAAS detection. After the concentration stage, the analyte is not eluted from the microcolumn with the eluent solution, and the ionite grains still containing the analyte are directly dosed into the spectrophotometer atomizer, where they undergo the thermal decomposition (pyrolysis and atomisation). This method, however, cannot be used to determine metals, for which the temperature of the pyrolysis stage is lower than 1000oC, such as cadmium, bismuth, and lead. The diagrams of these two procedures using reproducible microcolumn packing in the LOV system are shown in Figure 6. Validation of this method was performed by determining nickel in certified reference materials of biological and environmental origin [167]. In both cases, the same magnitude of the detection limit and concentration factor were achieved (Table 1). However, the precision of the determination, expressed as relative standard deviation (RSD), was lower when the eluent was used for the analyte recovery. It is worth adding that the detection limit for nickel determination (with the ETAAS detection) in a standard SIA system with a multiple usage of the same microcolumn packing was respectively 42 and 24 ng/l for the same and reverse directions of the solution flow at the concentration and elution stages, with the precision (RSD) ca. 5%. Lower concentration factor values that were attained for the ICP-MS detection were caused by

Analyte

Sample

Matrix

Waste

Eluent Air

Carrier

ETAAS

ETAAS

ICP-MS

Air

Carrier

A) Elution B) Ionite pyrolysis

Waste

H2O

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higher dispersion of the solution in the channels connecting the employed flow system and the nebulizer. TABLE 1 The comparison of nickel determination methods using reproducible microcolumn.

Elution Ionite pyrolysis

ETAAS ICP-MS ETAAS

Precision, % RSD

Detection limit, ng/l

Concentration factor

1,5 2,9 3,4

10,2 13 9

71,1 35,2 72,1

The possibility of using hydrophobic column packings for the concentration/isolation of the analyte in the SI-LOV system was also examined. Two types of stationary phases of different structure and physical properties were examined: co-polymer of styrene-and divinylbenzene with octadecylsilanol groups (C18–PS/DVB) and politetrafluoroethylene (PTFE) and used for sorption of cadmium complexes with DDPA [169]. The grains of C18–PS/DVB sorbent can easily be moved in the LOV system between the positions C1 and C2 of the microcolumn. Preparation of the packing in the form of suspension was similar to the already described preparation of cationic hydrophilic exchanger. Because of its greater density, the Teflon sorbent had to be constantly kept in a suspension state with an extra stirrer in the container. Due to the fact that both examined hydrophobic sorbents do not show the property of decreasing their volume at greater solution flow rate, as it was observed for SP Sephadex C-25 cation exchangter, only the procedure using elution of the concentrated analyte with ethanol solution could be examined. The results of using the ETAAS detection for determination of cadmium are shown in Table 2. Working with C18-PS/DVB sorbent is easier and therefore the precision of determination was better than for PTFE sorbent. Yet, from the analytic point of view, Teflon sorbent is better because a broader linear range, greater sensitivity and the value of concentration factor were gained. It is important, however, to inject the suspension of this packing into the microcolumn at very low flow rate – 0.24 ml/min. TABLE 2 Comparison of the analytical methods of cadmium determination in the SI-LOV system using C18-PS/DVB and PTFE sorbents [169].

PTFE C18-PS/DVB

Precision, %RSD

Detection limit, ng/l

Linearity range, ng/l

Concentration factor

4,3 3,4

15 126

0,05 - 1,0 0,2 - 1,5

17,2 7,4

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6 MINIATURIZED SYSTEMS FOR FLOW ANALYSIS The miniaturization of analytical instrumentation, observed in recent years, is the result of the technological possibilities as well as of the growing demand for such equipment. The high scale integration electronics, optotechnology, microtechnology, and modern material engineering are used more and more widely in constructing the analytical detectors themselves, as well as whole analysers with system of transporting and processing the sample, controlling the measurement and data processing. The construction of miniaturized flow systems (microfluidics) is an important field for employing miniaturization in analytical chemistry; it is also connected with the needs of environmental analysis (e.g., carrying out field determinations), protection of work places, and employment of such systems in large, complex measuring systems controlling whole industrial processes, and production of medicine and food.

The development of these instruments started at the beginning of the 1980s with development of integrated microflow systems (microconduits) to be used in flow-injection analysis with potentiometric detection (membrane electrodes) and photometric with optical wave-guide light transmission. These miniaturized devices include also integrated Chemifold modules used in Tecator commercial instruments with numerous applications in environmental analysis, chiefly for the inorganic analytes [170]. The channels with the diameter of 0.5 – 1.0 mm and external pumping equipment were used in these systems. As the development progressed, the pumping equipment was substituted with miniature piezoelectric pumps [171] or electroosmotic flow, which, in turn, called for using channels of much smaller diameters [172]. Another idea for miniaturization of flow systems, mostly in their sequential SIA version, was implementation of miniaturized both detecting and sample-processing elements, into a multipositional selecting injection valve. These systems are often called “lab-on-valve” (LOV), which is a much exaggerated term as far as their functional potential is concerned. Their application to trace analysis with atomic spectroscopy and mass spectrometry methods have been described above in more detail. In the attempts to construct miniaturized total chemical analysis systems, much attention is devoted not only to elaborating various methods of solution transportation or construction of miniature detectors directly on the so called analytic chip, but also to developing the methods of processing the sample in these integrated devices [173]. For environmental use, a system for concentrating gas analytes before chromatographic determination on the chip was elaborated and used for determination of dimethyl phosphonate [174]. Microsystems for determination of metal fluorescent ions with derivatization and concentration through accumulation on the chip with the capillary electrophoresis system were also described [175]. Miniaturized FIA systems for the use in environmental analysis, adopting various detection methods, were developed. For detection of phosphates with absorption spectrophotometric detection, a microsystem using silicon micropumps [176], as well as a miniature FIA system with electrokinetic transportation [177] were elaborated. Spectrometric determination of ammonia in waste and drinking water with indophenol blue was also designed for a silicon chip with an etched optic cuvette and an external syringe type pumps [178]. Amperometric enzymatic detection of phosphororganic pesticides with the employment of phosphororganic hydrolase was used in the FIA system with a flow detector having a biosensor on a silicon chip [179]. Flow systems, miniaturized on chips, provide special analytical opportunities; in such systems, apart from performing derivatization or concentration, electroosmotic flow with electromigration separation is used. Such systems permit carrying out complex analyses in short time and with little consumption of reagents. Similar systems,

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with separate thin-walled amperometric electrodes, were elaborated to determine various environmentally important compounds. To determine phenols, that are most common in rivers, silk-screen printed gold electrodes were employed [180]; to determine phosphororganic pesticides – printed carbon electrodes [181]. A flow chip electroforetic system with a gold disc electrode detector was created for environmental determination of organic peroxides [182]. BIBLIOGRAPHY [1] Stockwel P.B., Talanta, 27, 835 (1980). [2] Trojanowicz M., Automatyzacja w analizie chemicznej. Warszawa: WNT, 1992. [3] Skeggs Jr. L.T., Am. J. Clin. Pathol., 28, 311 (1957). [4] Ruzicka J., and Hansen E.H., Anal. Chim. Acta, 78, 145 (1975). [5] Furman W.B., Continuous Flow Analysis. Theory and Practice. New York:

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