Barna Kovács

Optical biosensor for urea with improved response time

An optical biosensor for urea measurements was developed. The operation of the sensor is based on the well-known urease enzyme-catalyzed hydrolysis of urea. The ammonium ions liberated in the reaction are detected with an ion selective optode membrane containing nonactin as ion selective ionophore and ETH 5294 chromoionophore in a thin (1 microm) plasticized poly(vinylchloride) film. The basic sensing element was home made of a microscope glass slide, a HeNe laser light source, photodiode light detector and light in coupling, de-coupling elements. The transducer membrane and the enzyme containing reaction layer were sandwich-cast with spin coating onto the surface of the sensing slide. The attenuation of the laser light propagating inside the glass wave-guide was used as signal for urea measurements. With this arrangement membranes provided good sensitivity (0.05 absorption unit when going from 0.1 to 1 mM urea) and short (16-20 s) response time. Taking advantage on the improved response time, flow injection urea measurements were made in the 0.01-2 mM concentration range. Thirty sample/hour analysis-rate, good peak-to-peak reproducibility (RSD=0.02) and recovery (95-104%) was achieved with buffer diluted urea solutions. Applications for the analysis of real samples are planned to do in the future.

Investigation of concentration profiles inside operating biocatalytic sensors with scanning electrochemical microscopy (SECM)

Scanning electrochemical microscopy (SECM) with amperometric or potentiometric measuring tips was used to investigate biocatalytic reactions inside the enzyme layer of a biosensor during its operation. The well known glucose oxidase catalyzed oxidation of glucose has been selected for the studies. Local, instantaneous concentration of dissolved oxygen and hydrogen peroxide was studied observing the amperometric current while miniaturized potentiometric tip served for local pH measurements. Liquid enzyme layer immobilized with Cellophane membrane or cross linked polyacrilamide gel membrane containing entrapped enzyme served for biocatalytic media in the SECM imaging. Local maximum of H(2)O(2) and minimum of O(2) profiles were found at approximately 200 microm far from the substrate/enzyme layer boundary. From the experimental findings guidelines to design well functioning biocatalytic sensors could be concluded. The concentration profiles obtained with SECM techniques were compared with the results of simple model calculations carried out with the method of finite changes. Most of earlier made SECM studies dealing with enzyme reactions imaged the electrolyte being in contact with the immobilized enzyme. The data in our investigation, however, were collected inside the working catalytic layer.

A new potentiometric sensor for the determination of α-amylase activity

A platinum redox sensor for the direct potentiometric determination of α-amylase concentration has been described. The sensor measured the amount of triiodide released from a starch-triiodide complex, which was correlated with the α-amylase activity after biocatalytic starch degradation. The composition and stability of the potassium triiodide solution was optimized. The starch-triiodide complex was characterized potentiometrically at variable starch and triiodide concentrations. The response mechanism of the platinum redox sensor towards α-amylase was proposed and the appropriate theoretical model was elaborated. The results obtained using the redox sensor exhibited satisfactory accuracy and precision and good agreement with a standard spectrophotometric method and high-sensitive fully automated descret analyser method. The sensor was tested on pure α-amylase (EC 3.2.1.1, Fluka, Switzerland), industrial granulated α-amylase Duramyl 120 T and an industrial cogranulate of protease and α-amylase Everlase/Duramyl 8.0 T/60 T. The detection limit was found to be 1.944 mU for α-amylase in the range of 0-0.54 U (0-15 μg), 0.030 mKNU for Duramyl 120 T in the range of 0-9.6 mKNU (0-80 μg) and 0.032 mKNU for Everlase/Duramyl 8.0 T/60 T in the range of 0-9.24 mKNU (0-140 μg).

Fabrication of optical chemical ammonia sensors using anodized alumina supports and sol-gel method.

In this comparative study, the fabrication and the sensing properties of various reflectometric optical ammonia gas sensors are described. In the first set of experiments the role of the support material was investigated on four different sensor membranes. Two of them were prepared by the adsorption of bromocresol green indicator on anodized aluminum plates. The applied anodizing voltages were 12 V and 24 V, which resulted in different dynamic ranges and response times for gaseous ammonia. The sol-gel method was used for the preparation of the other batch of sensors. These layers were coated on anodized aluminum plates (24 V) and on standard microscope cover glasses. In spite of the identical sensing chemistry, slightly different response times were measured merely because of the aluminum surface porosity. Gas molecules can remain entrapped in the pores, which results in delayed recovery time. On the other hand, the porous oxide film provides excellent adhesion, making the anodized aluminum an attractive support for the sol-gel layer.

Opto-electrochemical planar wave-guide sensor for copper (II) ion.

An opto-electrochemical sensor for copper (II) ion was developed. The sensor consists of a planar conductive indium-tin-oxide (ITO) glass support, which was coated by a polymeric copper (II) sensitive membrane. The sensing membrane is made of plasticized polyurethane matrix containing Zincon colorimetric reagent immobilized as ion pair with tetraoctylammonium ion. The instrumentation used commonly for wave-guide sensor development was extended with a potentiostat that made possible the control of the electric potential of the conductive surface. In this way the transport rate of ionic species through the membrane-sample interface could be influenced. In the presence of copper (II) ions the color of the membrane turned from red to blue, which was monitored optically. By applying positive potential to the conductive surface the direction of the ion diffusion at the membrane/sample interface was changed. As a result the sensing layer was regenerated within 2 minutes and was ready for further measurement. The sensor measured Cu(2+) ion in a concentration range of 1-200 microM.

Optical Ammonia Sensors for Environmental Applications

Triphenyl-methane based indicators bromophenol blue and bromocresol green were immobilized in ormosil sol-gel films prepared from phenyl-triethoxysilane in order to prepare colorimetric ammonia sensors. The sensing layers contained different molar ratios of the indicators, in this way—depending on the pKa values of the indicators—sensors with tailored dynamic ranges could be fabricated. Adsorption based immobilization was also used for the fabrication of chemical layers immersing anodized aluminum films in aqueous solutions of the indicators. Simultaneous immobilization of the indicators was considerably effortful on the anodized aluminum film, compared with the sol-gel film; therefore, the significant temperature dependency of the sensor response was used to shift the dynamic range. The effect of an ionic liquid additive was also investigated to increase the lifetime of these sensors. All sensors were tested spectrophotometrically, although a simple reflection based read-out device was also constructed using two light emitting diodes as light sources and two photodiodes as detectors. The device did not produce a concentration dependent signal but gave a digital yes or no (1 or 0) indicating that ammonia concentrations had reached a preset level.

Voltammetric concentration measurements in diffusion-hindered media

In voltammetric analysis the signal—in most cases—depends on transport processes. When getting concentration values of analytes from voltammetric calibrating curves, it is expected that the mass transport characteristics in sample solutions and in calibrating standards are identical. Standard addition methods are used in analytical practice when making calibrating standard with transport properties matching that of the samples would be difficult. Voltammetric measurements can also be carried out in soil—in sediment—or in gel samples. The mass transport conditions in these media, however, can considerably differ from those existing in aqueous solutions. The application of standard addition technique, however, is not an option there. In this work, a glassy carbon electrode was applied with a built-in diffusion layer on its measuring surface and chronoamperometric measurements were carried out. The current–time transients taken in aqueous solution and in tortuous, diffusion-hindered media were compared. Ascorbic acid and iodine as analytes as well as silica sand sediments and green pepper pulps as tortuous matrices were used. It was proved that if the modified electrode is used for analysis, then short time chronoamperometric transients taken in tortuous media can be evaluated by calibration data taken in aqueous standard solutions.

Characterization of a novel dissolved CO2 sensor for utilization in environmental monitoring and aquaculture industry

A novel optical fiber sensor is presented for measuring dissolved CO2 for water quality monitoring applications, where the optical signal is based either on refractive index changes or on color change. The sensing chemistry is based on the acid-basic equilibrium of 4-nitrophenol, that is converted into the anionic form by addition quaternary ammonium hydroxide. The CO2 sensitive layer was characterized and tested by using simple absorbance/reflectance measurement setups where the sensor was connected to a fiber optic CCD spectrometer. A prototype simulating a real shallow raceway aquaculture system was developed and its hydraulic behavior characterized. A commercially available partial-pressure- NDIR sensor was used as a reference for dissolved CO2 tests with the new optical fiber sensor under development. Preliminary tests allowed verifying the suitability of the new optical sensor for accurately tracking the dissolved carbon dioxide concentration in a suitable operation range. Direct comparison of the new sensor and the reference sensor system allowed to demonstrate the suitability of the new technology but also to identify some fragilities there are presently being addressed.

LPG based fiber optic sensor for carbon dioxide

In this work a novel optical-fiber sensor for carbon dioxide measurement is presented. A polymeric sensitive layer based on the acid-base equilibrium of phenol and of its derivative 4-nitro-phenol is used for carbon dioxide determination. The sensitive material presents changes in color and in its refractive index. Colorimetric and refractometric measurements were performed. The results show the sensor is more sensitive for lower concentrations and a saturation effect occurs for higher levels. For the colorimetric response, a resolution of ±0.15% was estimated and a response time of 30s was measured. For the refractometric measurements, a resolution of ±0.50% could be estimated and a response time of 12s was measured. Reversibility and reproducibility were also demonstrated.

Colorimetric and refractometric measurements of carbon dioxide

In this work, a polymeric sensitive layer based on the acid-base equilibrium of phenol and of its derivative p-nitro-phenol is presented for carbon dioxide measurements. Thin films casted on glass slides were tested, using a LED source (λc at 410 nm) and an Ocean Optics USB4000 spectrometer, in the 0% to 15.25% CO2 concentrations range, showing a 40% maximum transmittance variation with a 51s response time and a 0.15% resolution. Preliminary results indicate that CO2also induces refractive index changes in the sensitive layer. Using a fiber based interferometric setup, a CO2 dependent refractive index change of ~0.045 RIU was observed, in the 0%-90% CO2 concentration range.