Preparation and characterization of platinum coatings for long life-time BOD biosensor

Sensors and Actuators B 47 (1998) 21 – 29

Preparation and characterization of platinum coatings for long life-time BOD biosensor Kaido Tammeveski a, Timo Kikas a, Toomas Tenno a,*, Lauri Niinisto¨ b a

b

Institute of Physical Chemistry, Uni6ersity of Tartu, Jakobi 2, EE-2400 Tartu, Estonia Laboratory of Inorganic and Analytical Chemistry, Helsinki Uni6ersity of Technology, P.O. Box 6100, FIN-02015 Espoo, Finland Accepted 10 December 1997

Abstract An easy method to prepare platinum coatings is proposed. Platinum coatings were prepared on glass substrate by applying a thermal decomposition method. Electrochemical and surface properties of the Pt coatings obtained were studied. A polycrystalline Pt layer is formed which shows electrocatalytic activity towards oxygen reduction similar to that of bulk platinum. An amperometric oxygen sensor was constructed by using Pt-coated glass substrate as a cathode. The oxygen sensor exhibited good performance characteristics. A biosensor for the determination of biochemical oxygen demand (BOD) was developed using the oxygen sensor as a base transducer and by immobilizing bacteria Bacillus subtilis in agarose gel film on the top of the sensor’s membrane. The BOD biosensor was tested in various media. © 1998 Elsevier Science S.A. All rights reserved. Keywords: Atomic force microscopy; Biochemical oxygen demand (BOD) biosensor; Oxygen reduction; Oxygen sensor; Platinum coating; X-ray photoelectron spectroscopy

1. Introduction Amperometric oxygen sensors based on thin-film electrodes have found an increasing application in the field of biosensing [1,2]. Thin-film platinum is a suitable material in the fabrication of electrochemical biosensors [3,4]. This is mainly due to its high electrocatalytic activity and chemical inertness. Platinum is also one of the best electrocatalysts towards oxygen reduction [5], and has therefore been widely used as cathode material in amperometric oxygen sensors [6]. The major pathway of oxygen reduction on platinum proceeds through the four-electron reaction. The amount of hydrogen peroxide produced at the surface of platinum is relatively small. In recent years, there has been a growing need for miniaturized oxygen sensors and biosensors [7,8]. Those are mainly produced in a planar form on various substrates. Thermal evaporation and sputtering techniques are widely used for the formation of Pt films. However, miniaturization is not the main goal in the development of BOD sensors because of large sam* Corresponding author. Tel.: + 372 7 465180; fax: + 372 7 465160; e-mail: [email protected] 0925-4005/98/$19.00 © 1998 Elsevier Science S.A. All rights reserved. PII S0925-4005(98)00004-5

ple volumes. Special care should be taken to achieve a stable signal for a long period. This, in turn, requires that optimum sensor construction and immobilization procedure be used [9]. The removal of biodegradable organic substances is a major concern in biological wastewater treatment plants. The biochemical oxygen demand (BOD) test is a widely employed method for the estimation of organic pollution in wastewater. A conventional method for the estimation of BOD requires 5 days for sample incubation [10]. This period is too long for effective bioprocess control. A more rapid method for the measurement of the BOD value is needed. A BOD biosensor based on immobilized microorganisms has been proposed for that purpose [11]. The microbial BOD sensor consists of immobilized microorganisms in combination with an oxygen electrode, and it measures the respiration activity of cells. The decrease in oxygen concentration in the microbial layer depends on the substrate concentration and this relationship provides the basis for the BOD determination. Various microorganisms, Trichosporon cutaneum more frequently than others, have been used in the preparation of BOD sensors [12–15]. Obviously, microorganisms showing bio-oxidation activity towards

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a wider range of substrates are preferred. Basically, there are two possibilities of performing the BOD test with microbial sensors: (1) steady-state measurement with a typical response time of 15 – 20 min; (2) kinetic measurement where the signal is recorded within 15–30 s [12]. In the work reported in this paper, a thermal decomposition method for the fabrication of Pt coatings was proposed. The BOD biosensor was developed using an improved oxygen sensor with Pt film cathode and was further characterized in the test media.

tron spectroscopy (XPS) and atomic force microscopy (AFM). XRD images were obtained by a Philips MPD 1880 X-ray diffractometer using Cu Ka X-ray source. XPS analysis was performed by the KRATOS Axis 165 instrument using monochromated Al Ka radiation and two measuring points for each sample. The surface morphology was examined by using a Park Scientific Instruments AutoProbe CP microscope in contact mode. SEM/EDX was used to confirm the AFM results.

2.4. Sensor design 2. Experimental

2.1. Preparation of Pt coatings A thermal decomposition method was used for the preparation of platinum coatings on glass substrate. An ethanolic solution of hexachloroplatinic acid containing organic additives (colophony, turpentine) and boric acid was prepared. The final solution contained 3% H2PtCl6. Two procedures were used to prepare Pt films on glass: the dipping of the substrate in coating solution or the application of a certain amount of the solution onto the substrate surface. The substrate was dried in air for 5 min followed by firing in flame for 1 – 2 min. The coating procedure was repeated up to 15 times. The method used in this study for the formation of Pt coatings is different from that of screen-printing where an ink containing Pt was used [16].

A conventional two-electrode Clark-type oxygen sensor was constructed as a base transducer for the BOD biosensor. Thermally prepared Pt-coated glass was used as the sensor’s cathode. Usually 10 coating cycles were made for the application of Pt coating in the sensor fabrication. An Ag/AgCl reference electrode was formed by the anodization of a silver wire in 0.1 M HCl. The electrodes were mounted in a Perspex body which was filled with the 1.0 M KCl internal electrolyte. The top of the sensor was covered with a 25 mm thick Teflon membrane. The construction of the oxygen sensor was similar to the one described earlier [17]. A specially designed holder was used to attach an agarose gel film with immobilized bacteria Bacillus subtilis. Fig. 1 depicts schematically the BOD sensor used.

2.2. Electrochemical measurements The electrochemical properties of Pt films were investigated by cyclic voltammetry (CV) and the rotating disk electrode technique (RDE). The solutions of 0.5 M H2SO4, 0.1 M KOH and 1.0 M KCl were prepared from p.a. quality reagents (Merck) and Milli-Q water (Millipore). Electrochemical experiments were performed in a three-electrode cell. A saturated calomel electrode (SCE) was used as a reference and Pt-wire served as a counter electrode. Pt-coated glass with an exposed area of 0.2 cm2 was embedded in a Teflon holder. The oxygen reduction data were collected in O2-saturated 0.1 M KOH and 1.0 M KCl by using an EDI 101 rotating disk electrode and a CTV 101 speed control unit (Radiometer). The electrode rotation rate (v) was varied from 360 to 3100 rpm. The potential was applied by Polarographic Analyzer PA-2 connected with an IBM PC.

2.3. Surface characterization The surface characterization of Pt-coatings was carried out by X-ray diffraction (XRD), X-ray photoelec-

Fig. 1. Schematic drawing of BOD sensor: (1) epoxy; (2) end cap; (3) Ag/AgCl electrode; (4) glass tube with planar end surface; (5) Perspex body; (6) threaded cover; (7) Teflon membrane holder; (8) bacteria– agarose membrane holder; (9) Pt coating; (10) electrolyte (1 M KCl); (11) Teflon membrane; (12) agarose gel membrane with immobilized bacterial cells; (13) test media.

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2.5. Microorganism immobilization

2.6. Measurements with the biosensor

The bacteria Bacillus subtilis were produced in the Institute of Molecular and Cellular Biology of the University of Tartu. The bacterial cells grown on congealed nutrient substratum were planted into 100 ml of nutrient substratum solution, which contained 0.5 ml of the solution of microelements. The process of growing lasted for 24 h on the shaker at the temperature of 30°C in aerobic conditions. The growth was stopped when the cells reached the stationary phase of growth. The cells were gathered by centrifuging them for 10 min at a speed of 1500 rpm. The solution was decanted and the sediment of bacteria was suspended into 0.1 M phosphate buffer (pH 6.86). For the purpose of cleaning, the suspension of cells was washed twice in the phosphate buffer and then suspended in 3 ml of the same buffering solution. The biomembrane was made of the 2% solution of agarose in the phosphate buffer and of the suspension of the bacteria. The agarose solution heated to the boil was cooled down to 36°C and 2 ml of the bacterial suspension was added to it. The resulting mixture was spread on the polymer net of a particular thickness, which, in order to gain a certain and even thickness of the bacterial layer on the net, was then placed between two glass plates until the formation of a persistent layer of gel. In most experiments a bacterial membrane thickness of 0.3 mm was used. The net with the immobilized bacteria was put into the phosphate buffer for storage at room temperature. Subsequently, a disk of a particular size was cut from the net and attached to the oxygen sensor.

The measurements were carried out at 25°C in the measuring cell, the volume of which was 150 ml. The measuring system was thermostated and the solutions were mixed with a magnetic stirrer. The solutions were saturated by air oxygen with the aid of a microcompressor. The BOD biosensor was calibrated with a BOD standard solution that contained 150 mg of glucose and 150 mg glutamic acid in 1 l water. The BOD biosensor was tested in a medium containing lower alcohols ranging from ethanol to hexanol. The test solutions were prepared by the dilution of stock solutions of alcohols (0.8 g l − 1) in water. Measurements were also conducted in phenol solutions and phenolcontaining wastewater solutions. A classical BOD7 test was performed to compare the BOD values obtained by the BOD sensor.

Fig. 2. AFM images of Pt coating on glass substrate: (a) three coating cycles; (b) six coating cycles.

3. Results and discussion

3.1. Surface characterization A series of platinum coatings on glass was produced using the thermal decomposition method. The thermogravimetric data show that the temperature of the decomposition of H2PtCl6 is about 450°C [18]. For that reason the Pt-coated glass was fired up to 500°C. The repetitive coating increased the amount of platinum on glass. The adhesion of platinum to glass substrate was fairly good to meet the needs for practical application in electrochemical sensors. AFM is a useful tool to examine the surface morphology of thin films and coatings. An increase in the number of coating cycles resulted in a change of morphology. There was a clear tendency towards platinum hillock formation. Relatively large Pt hillocks were formed already at small Pt loadings. The number of hillocks per surface area increased with increase of platinum loading. It is interesting to note that the height of a hillock did not increase noticeably at higher Pt loadings. Fig. 2 presents the AFM images of threeand six-times-coated substrate. For a six-times-coated sample the number of hillocks is high enough to cover the substrate surface completely. For the given coating the average height of Pt hillocks was about 60 nm and the width was within 100–200 nm. The morphological development of Pt coatings can vary in different fabrication series, but the main tendencies remain the same. The crystallinity of the Pt deposits was studied by the XRD method. XRD patterns revealed clear peaks even for a single-time-coated substrate. This indicates that the firing temperature is sufficient to form a crystalline

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deposit of platinum. The intensity of peaks increased with the increase of the Pt loading. The peaks of Pt planes (111), (200), (220), (311) and (222) were observed within the range of angles 30°B2UB 90°. The Pt(111) reflection showed the highest intensity. The XPS analysis was carried out with the Pt/glass samples in order to examine the chemical composition and state of the Pt coating surface. Samples made by one, two, three and five coating cycles were investigated. Survey spectra of the Pt/glass samples consisted mainly of platinum, oxygen and carbon. The level of residual chloride, sodium and silicon was relatively low (less than 2 wt.%). The mass concentration of Pt in the surface layer was nearly constant for all samples under study. The surface was contaminated by hydrocarbons due to exposure to atmospheric conditions. The spectra of Pt 4f electrons were used to examine the state of the valency of platinum in the coating (see Fig. 3). The binding energy (BE) values for Pt 4f5/2 and 4f7/2 peaks were 74.6 and 71.3, respectively, and were nearly constant for different samples. The BE values are very close to those typical of metallic platinum [19]. The other valence state Pt atoms were present in a negligible amount.

3.2. Electrochemical measurements The cyclic voltammetric (CV) experiments were performed with Pt coatings in argon-saturated 0.5 M H2SO4 and 0.1 M KOH. It is interesting to note that already the second scan revealed the presence of typical features of the CV curves of polycrystalline platinum. This indicates that the surface of platinum is rather clean after the firing. By repetitive potential cycling the hydrogen adsorption – desorption peaks started to be more sharp and the peak of Pt oxide reduction in-

Fig. 3. XPS spectrum of Pt 4f region for five-times coated Pt/glass sample.

Fig. 4. Cyclic voltammograms for Pt coatings in Ar-saturated 0.5 M H2SO4: (1) five coating cycles; (2) 10 coating cycles. Sweep rate 50 mV s − 1, 15 repetitive potential scans.

creased. The platinum oxide formation peaks gradually improved and finally two peaks were clearly separated. The shape of cyclovoltammograms did not change considerably with the increase of the number of coating cycles (see Fig. 4). The roughness factor, fr (the ratio of real and geometrical surface area), was determined from the area under the hydrogen desorption curve, assuming a charge of 210 mC consumed per cm2 of polycrystalline Pt. It was found that the fr value increased with the increase of the number of coating cycles. For a 10-times-coated sample the average value of fr was close to 5. It is noteworthy that fr is not reproducible for different coating series. The reason for that seems to be connected with the difficulty of controlling the firing procedure. The cyclic voltammograms of Pt coatings were also recorded in 0.1 M KOH. The main tendencies were similar as compared with the behaviour in acid solution. The CV curves remained stable after about 15 potential scans. Just after the CV experiments the solution of 0.1 M KOH was saturated with oxygen. The oxygen reduction curves were recorded in the potential range from 0.1 to − 0.9 V. Well-defined oxygen reduction waves with a broad diffusion-limited current plateau were observed at all rotation rates. Tafel plots for oxygen reduction on Pt coatings were constructed from the RDE data. Two slope regions were obtained for the reduction of oxygen in 0.1 M KOH. At low current densities the slope was close to − 70 mV dec − 1 and did not change noticeably with the number of coating cycles. The slope was slightly higher than that found for bulk Pt and thin Pt films on glassy carbon substrate in alkaline solution [5]. The apparent electrocatalytic activity of Pt coatings towards oxygen reduction (estimated on the bases of half-wave potential values) was even higher as com-

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pared with the bulk platinum electrode. This is mainly due to a higher roughness factor. The exchange current density of Pt coatings in the reduction of oxygen was in the order of 10 − 9 A cm − 2, which agrees well with that of bulk Pt. The reduction of oxygen was also studied in the solution of 1.0 M KCl as this was an internal electrolyte of our oxygen sensor. The voltammetry curves for oxygen reduction are presented in Fig. 5. The reduction reaction is limited by diffusion in a wide region of potentials. This was confirmed by the Koutecky–Levich analysis (plotting i − 1 versus v − 1/2). The experimental Levich slope value 0.32 mA cm − 2 rad − 1/2 s1/2 was close to the theoretically calculated value for the four-electron transfer per oxygen molecule. At higher rotation rates the current slightly increased at higher cathodic polarization and therefore the plateau was not ideal. The oxygen reduction behaviour of Pt coatings studied in this work was similar to that of thick-film Pt electrodes obtained by screenprinting with platinum ink [16]. The presence of a current plateau on i,E-curves is favourable for the development of amperometric sensors with good performance characteristics.

3.3. Oxygen sensor The operating characteristics of the oxygen sensor were evaluated in order to be sure that it meets the requirements for use as a transducer for BOD biosensors. The following sensor properties were investigated: voltammetric behaviour, linearity of response, response time and stability. The current – voltage curve for the oxygen sensor was recorded by changing the potential from 0 to −1.0 V in 50 mV increments (see Fig. 6). Up

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Fig. 6. Current – voltage curve for oxygen sensor in air-saturated aqueous solution. The inset shows a calibration curve for oxygen sensor in O2 – N2 gas mixtures.

to − 0.1 V the current is negligible. At more negative potentials the reduction of oxygen starts to occur and the current rises to its limiting value. The diffusion-limited plateau region ranges from −0.5 to − 0.9 V. The extent of the plateau region can vary for different sensors, decreasing slightly with increasing the sensor’s age. At − 0.85 V the hydrogen evolution reaction commences. For further work the sensor’s cathode was polarized at − 0.7 V with respect to the internal Ag/ AgCl electrode. Oxygen–nitrogen gas mixtures of different ratios (overall pressure 1 atm) were used for electrode calibration. The sensor current was a linear function of the oxygen concentration with a correlation coefficient of 0.9996 (see inset of Fig. 6). The sensor signal was in the order of microamperes and therefore no amplification was needed. The signal-to-noise ratio was high enough to perform reliable measurements. The response of the oxygen sensor towards a step change in oxygen concentration was also studied. The response time of the sensor depends mainly on the thickness of the polymer membrane. The 95% response time was within 15–20 s. The output signal of the oxygen sensor was measured in air-saturated water at 25°C after every week. The sensor was fairly stable (the current variation was less than 9 2%) during a 6month period. The long-term stability of the given oxygen sensor seems to be connected with the relatively high surface roughness of the working electrode. This reduces the effect of the poisoning of the electrode surface.

3.4. Biosensor model Fig. 5. Voltammetry curves for oxygen reduction on Pt coating electrode in O2-saturated 1 M KCl at various rotation rates (v). Sweep rate 10 mV s − 1.

An attempt was made to develop a model for the microbial BOD sensor. This is rather similar to the model reported in a previous study [20]. The main

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difference of the models is that in the present case we have a sensor whose outer membrane is absent and this simplifies the model. In the model effective parameters are used which makes it possible to introduce the concentrational distribution of the substrate and oxygen as a continuous curve. The sensor response (I) is related to the concentration of substrate (cS) according to the following equation: dI = R

j cS j M + cS

(1)

where dI is the normalized sensor current I, R =(6ox/ ce) × (lb/2Pb) is the coefficient of diffusion resistance of bacterial layer to oxygen, j =kbcblb is the limiting value of the substrate flow density, M =(lb/2P %b)+ (1/ kblb)(1+ (KM/cb)) is the coefficient of complex (diffusion –kinetic) resistance to the substrate, 6ox is the stoichiometric coefficient of oxygen in biochemical reaction, ce is the effective concentration of oxygen in the test solution, lb is the thickness of the bacterial layer of the sensor, Pb is the permeability of the bacterial layer to oxygen, P %b is the permeability of the bacterial layer to the substrate, kb is the rate constant of biochemical reaction, cb is the concentration of bacteria in the bacterial layer and KM is the Michaelis – Menten constant. For a given construction of biosensor it can be assumed that the diffusional component of complex resistance to the substrate is relatively low as compared to the kinetic component:





lb 1 K 1+ M  2P %b kblb cb

(2)

It is also possible to assume that the concentration of bacteria in the bacterial membrane (cb) is low in comparison to the substrate concentration (cS). In that case we can obtain: j M $ KM

dI = R

j c¯Sb KM + c¯Sb

(5)

where: c¯Sb = − (p/2)+ (p/2)2 − q is an average value of substrate concentration in the bacterial layer, and p= j M− cS and q= −KMcS are the coefficients of the equation.

3.5. Biosensor performance The Bacillus subtilis-based BOD sensor was tested in the solutions of various alcohols. The sensor response depended on the amount of the substrate added. The values of the stabilized output signal were registered in the substrate solutions of different concentrations. The resulting BOD sensor was calibrated with the solution of glucose and glutamic acid (each 150 mg per 1 l solution), which is the standard solution of BOD. It took 15–30 min to achieve a stabilized output. Fig. 7 shows the calibration graph obtained from the experimental data of the stabilized output signal versus the BOD value of the substrate solution. For this purpose the average output signal was calculated on the basis of 12 experiments. In the present study the BOD value of standard solution was also tested by the standard method of BOD7 determination. The results remained within the limits of the allowed experimentation error (15–20%). The parallel measurement results coincided in the range 1–5%. Particular attention was paid to the biosensor’s response to different substrates. The first experiment was carried out by using solutions of straight-chained alcohols (see Fig. 8). The representatives of one homological line were taken for the experiment to follow the metabolic activity of the bacteria. Hydroxy compounds were found to be suitable for this purpose for the following reasons: (1) because of the hydroxy substi-

(3)

Eq. (1) for the studied sensor in the given conditions can be transferred to the type of Michaelis – Menten as follows: dI =

dImaxcS KM + cS

(4)

where dImax = R×j is the limiting value of the sensor output current. It follows that in the absence of diffusion resistance to the substrate and at a low concentration of the bacteria, the dependence of the sensor current on substrate concentration is in accordance with the Michaelis– Menten scheme (i.e., the sensor response is under kinetic control). The influence of the diffusional component of complex resistance to the substrate can be taken into account using the following expression:

Fig. 7. Calibration curve for BOD sensor in glucose – glutamic acid standard solution.

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Table 1 The BOD values of phenol solutions obtained by BOD7 test and BOD sensor

Fig. 8. The dependence of normalized response on the concentration of various alcohols.

tute, they have relatively good solubility in water; (2) the wastewater used in the experiment contained several different hydroxy compounds (phenols). Biodegradability of alcohols by Bacillus subtilis increases with the growth of the carbon chain. As is seen in Fig. 8, the shape of the curve changes with change of the length of carbon chain. The experimental data were analyzed using the sensor models given above. The least-squares method was used to fit the data. The standard deviation of the curves of different substrates was obtained and compared for both models. It can be concluded that for ethanol the kinetic model fits quite well. The higher the chain length, the lower the concentration at which diffusional control starts to play a role. Similar experiments were carried out with different isoalcohol solutions. Three first members were used from the isoalcohol line (isopropanol, isobutanol, isopentanol). Results showed a trend similar to the previous one: biodegradability of isoalcohols by Bacillus subtilis increases with the growth of the carbon chain. By comparing the results mentioned above it was found that isoalcohols have lower biodegradability in comparison with linear alcohols of similar length of the carbon chain. It was also important to inspect the usability of the biosensor in the phenolic test environment. For this purpose measurements were carried out in the solution of phenol whose BOD7 had been measured using the conventional method for the BOD determination. The results were compared to the calibration curve (CC) and are given in Table 1. As can be seen, there is a good correlation between the BOD values measured by biosensor and the BOD7 method. Identical experiments were carried out by using the solutions of hydroxy compounds with a similar length of the carbon chain (hexanol, cyclohexanol, phenol).

BOD7 (mg l−1)

BOD value from CC (mg l−1)

1.7 3.5 5.2 7.0 8.7 10.4 12.2

2.0 3.9 5.8 7.7 9.5 11.3 13.0

Phenol with its aromatic cycle was found to be less biodegradable. At the same time, cyclohexanol appears to be more easily degradable than its linear analogue. In order to evaluate the suitability of the BOD biosensor for testing wastewaters containing phenolic compounds, measurements were carried out with actual wastewaters from the Kohtla-Ja¨rve oil-shale industry. The wastewater of the dephenolization plant contained a relatively high amount of phenolic compounds. The sensor was adapted to the measurements in solutions containing phenols. Several wastewater samples were studied and the BOD value obtained by the sensor was higher than that measured by BOD7. However, on the basis on several experimental series, it was obtained that the ratio of BOD values measured by both methods did not change considerably. Hence, we can conclude that the sensor can be utilized for the BOD estimation of wastewater samples of certain origin. The main advantage of the BOD sensor of the given construction was the ease of the bacterial membrane replacement. It took only 1 min to change a membrane, whereas the sensor response changed only slightly. This was achieved due to optimum bacterial membrane preparation. Scores of membrane disks of uniform

Fig. 9. Long-term stability of oxygen sensor and BOD sensor.

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properties were obtained from one net of immobilized bacteria. Fig. 9 presents the data of long-term stability of the oxygen sensor and the BOD sensor. The bacterial membrane was stable up to 5 months. As the oxygen sensor was stable for an even longer period, the lifetime of a biosensor is determined by the stability of the bacterial membrane.

[4]

[5]

[6]

4. Conclusions A simple thermal decomposition method was used for the fabrication of platinum coatings on glass substrates. The major advantage of the given method over those of physical evaporation is that no sophisticated equipment is needed in the preparation procedure. The surface analysis revealed that the coating was composed of metallic platinum. The Pt-coating electrodes showed high electrocatalytic activity towards oxygen reduction. A two-electrode Clark-type oxygen sensor was constructed using a thin-film Pt cathode. A biosensor for the determination of biochemical oxygen demand was designed using the oxygen sensor as a basic device. The BOD sensor can be used as a monitoring device.

[7] [8] [9]

[10]

[11]

[12]

[13]

[14]

Acknowledgements The authors would like to thank Mrs K. Orupo˜ld, Mr J. Raudsepp, Mr A. Mashirin and Dr K. Kukli for their valuable comments. K.T. wishes to thank the CIMO, Finland, for awarding a scholarship to study at Helsinki University of Technology (HUT). Professor K. Heiskanen and Ms K. Ilmonen (Laboratory of Mechanical Process and Recycling Technology, HUT) are gratefully acknowledged for their help in AFM investigations. We wish to thank Dr L.-S. Johansson (Centre for Chemical Analysis, HUT) for performing XPS analysis. This work was supported by the Estonian Science Foundation (grant No. 2346). Partial financial support was provided by the Phare HESR Programme (ES 9502.02).

[15]

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Biographies Kaido Tamme6eski received his diploma in chemistry and M.Sc. degree in chemistry from the University of Tartu in 1989 and 1993, respectively. He is currently studying for his Ph.D. degree at the University of Tartu. His research interests include electrochemical kinetics and electrochemical sensors.

K. Tamme6eski et al. / Sensors and Actuators B 47 (1998) 21–29

Timo Kikas received his B.Sc. and M.Sc. degrees in chemistry from the University of Tartu in 1994 and 1997, respectively. In 1997 he started his postgraduate studies at doctoral level at the University of Tartu. His research interests are in environmental analysis and biosensors. Toomas Tenno received his diploma in chemistry and his Ph.D. in chemistry from the University of Tartu in 1965 and 1973, respectively. Since 1992 he has been Professor of Colloid and Environmental Chemistry and since 1993 Head of the Institute of Physical Chemistry at the University of Tartu. He is a member of the Royal Society of Chemistry and a member of board of the Estonian Chemical Society. His research interests in-

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clude environmental chemistry and electroanalytical chemistry. Lauri Niinisto¨ was born in Helsinki, Finland, in 1941. He received his M.Sc. and D.Techn. degrees from Helsinki University of Technology (HUT) in 1968 and 1973, respectively. His doctoral research was partly conducted at the University of Stockholm in 1971– 1972. Since 1977 he has been Professor of Inorganic Chemistry at HUT but also worked in France, the USA and Austria. His current research is focused on preparation and characterization of thin films for optoelectronic and electronic devices, including sensors. He is the author or co-author of more than 200 original publications and reviews.