Prediction of wastewater quality using amperometric bioelectronic tongues

Wastewater samples from a Swedish chemi-thermo-mechanical pulp (CTMP) mill collected at different purification stages in a wastewater treatment plant (WWTP) were analyzed with an amperometric enzyme-based biosensor array in a flow-injection system. In order to resolve the complex composition of the wastewater, the array consists of several sensing elements which yield a multidimensional response. We used principal component analysis (PCA) to decompose the array's responses, and found that wastewater with different degrees of pollution can be differentiated. With the help of partial least squares regression (PLS-R), we could link the sensor responses to the Microtox (R) toxicity parameter, as well as to global organic pollution parameters (COD, BOD, and TOC). From investigating the influences of individual sensors in the array, it was found that the best models were in most cases obtained when all sensors in the array were included in the PLS-R model. We find that fast simultaneous determination of several global environmental parameters characterizing wastewaters is possible with this kind of biosensor array, in particular because of the link between the sensor responses and the biological effect onto the ecosystem into which the wastewater would be released. In conjunction with multivariate data analysis tools, there is strong potential to reduce the total time until a result is yielded from days to a few minutes. (C) 2015 Elsevier B.V. All rights reserved

Development of a Bioelectronic Tongue -Applications for Wastewater Analysis

A bioelectronic tongue has been developed for applications in wastewater analysis. The development of a biosensor array with complex signal analysis started from the idea of using group-selective phenol biosensors (tyrosinase and horseradish peroxidase) with chemometric analysis for signal processing. In a first step the ability to simultaneously determine each analyte in synthetic binary phenol mixtures was evaluated using multivariate data analysis on the responses from a single tyrosinase-modified solid graphite electrode. The next step was to construct a suitable device where different biosensors could be used in an array for multi-parameter detection of samples. Screen-printed electrodes of carbon and noble metals were first evaluated with the purpose of identifying electrodes that could be used as a basis for immobilisation of phenol- and pesticide-sensitive enzymes (horseradish peroxidase, soybean peroxidase, cellobiose dehydrogenase, acetylcholinesterase and butyrylcholinesterase). These enzymes were then immobilised on an array of eight radially distributed electrodes. To host the array and to provide equal hydrodynamics at each electrode in the array, a special electrochemical cell was constructed to enable flow-injection and steady-state measurements. Together with multivariate data analysis has this array system been successfully used for qualitative discrimination of wastewater samples as well as for quantitative determination of their toxicity and other pollution parameters such as chemical oxygen demand and biological oxygen demand. Pre-processing of data before multivariate analysis was shown to be necessary for reducing the noise that otherwise can hide the desired structural information. Several strategies to overcome noise problems due to drift in biosensors have been developed

Screen-printed carbon electrodes modified with cellobiose dehydrogenase: Amplification factor for catechol vs. reversibility of ferricyanide

A number of screen-printed carbon electrodes (SPCEs) have been electrochemically studied revealing strong correlation between the reversibility of the Fe(CN)/Fe(CN) couple and the sensitivity for catechol at the same electrodes modified with cellobiose dehydrogenase (CDH). Pretreatment of the electrode surfaces increased both the heterogeneous ferricyanide reaction rate and the catechol sensitivity. From cyclic voltammetric and chronoamperometric measurements of Fe(CN) it was concluded that the tested SPCEs behave as microelectrode arrays. Using the pinhole model the fraction of electroactive area was determined to directly correlate to a faster heterogeneous electron transfer for ferricyanide and a higher CDH-modified biosensor sensitivity for catechol. An electroactive area of 50% and higher is sufficient to create a sufficiently good biosensor for catechol

Direct heterogeneous electron transfer of theophylline oxidase

Direct electron transfer (DET) was shown between the heme containing enzyme theophylline oxidase (ThO) and the surface of both graphite and gold electrodes. As proof on graphite a steady state current for theophylline was recorded using the electrode modified with adsorbed ThO. The electrode showed a Michaelis–Menten-like response to theophylline with a detection limit of 0.2 mM and a Michaelis–Menten constant equal to 3.2 mM. These initial results open up a possibility for the development of reagentless third generation biosensor based on heterogeneous DET between ThO and an electrode. On gold DET between ThO and the surface of aldrithiol modified gold was studied with spectroelectrochemical measurements. DET was observed for soluble ThO as a change of its spectrum in a gold capillary responding to a change in the applied potential. It was shown that the redox conversion of the heme domain of the enzyme is directly (mediatorlessly) driven by the potential applied at the gold electrode. The measurements enabled an estimation of the formal potential (E°′) of the redox process equal to −275±50 mV versus Ag|AgClsat at pH 7.0. The experimentally determined number of the electrons involved in this heterogeneous electron transfer process was estimated to be equal to 0.53. The low precision in determination of the E°′ and the value of the number of electrons lower than one indicate that kinetic restrictions disturbed the evaluation of the true thermodynamic values from relatively fast spectroelectrochemical measurements. . . . This is the final, accepted and revised manuscript of this article. Use alternative location to go to the published article. Requires subscription

Amperometric sensors based on tyro sinase-modified screenprinted arrays

This paper describes the design, development and characteristics of a tyrosinase (polyphenol oxidase) modified amperometric screen-printed biosensor array, with the enzyme cross-linked in a redox-hydrogel namely the PVI13-dmeOs polymer. Two types of Au-screen-printed four-channel electrode arrays, differing in design and insulating layer, were compared and investigated. Au-, graphite-coated-Au- and Carbopack C-coated-Au-surfaces, serving as the basis for tyrosinase immobilisation, were investigated and the performances of the different arrays were evaluated and compared in terms of their electrocatalytic characteristics, as well as operational- and storage stability using catechol as model substrate. It was found that the Carbopack C-coated array was the best choice for tyrosinase immobilisation procedure mainly due to a higher mechanical stability of the deposited enzyme layer, combined with good sensitivity and stability for up to 6 months of use. In the batch mode the biosensors responded linearly to catechol up to 30 muM with limits of detection from 0.14 muM. Parameters from cyclic voltammograms indicated that the reversibility of the direct electrochemical reaction for catechol on the three types of electrode surfaces (no tyrosinase modification) was not the limiting factor for the construction and performance of tyrosinase biosensors. (C) 2003 Elsevier B.V. All rights reserved

Effect of interfering substances on current response of recombinant peroxidase and glucose oxidase-recombinant peroxidase modified graphite electrodes

Graphite electrodes have been modified with different forms of horseradish peroxidase (HRP). These included native HRP, wild-type recombinant HRP, and two single-point recombinant HRP mutants, N70V and N70D. The mediator-less response of these electrodes to H2O2 was studied indicating that electrodes modified with recombinant HRP forms are more stable than those modified with native HRP. Various interfering compounds were investigated for their effect on the current response to H2O2. It was found that interferences such as acetaminophen and dopamine affected the response by mediating the electron transfer (ET) between graphite and peroxidases. The mediating behaviour manifested itself as an increased current of the electrode to H2O2. The interfering effect was less pronounced for the electrodes modified with recombinant HRPs possessing better electronic coupling with the graphite surface. The interfering behaviour of acetaminophen on the response for glucose with the bienzyme electrode containing co-immobilised glucose oxidase and HRP was mainly ascribed to mediation of ET between graphite and HRP. It was experimentally proven that a high efficiency of direct ET between graphite and recombinant HRP substantially reduces the interfering effect of acetaminophen

Screen-printed multienzyme arrays for use in amperometric batch and flow systems

Screen-printing technology for electrode fabrication enables construction of amperometric devices suitable for combination of several enzyme electrodes. To develop a biosensor array for characterisation of wastewaters, tyrosinase and horseradish peroxidase (HRP) or cholinesterase-modified electrodes were combined on the same array. The behaviour of the tyrosinase-modified electrode in the presence of hydrogen peroxide (required co-substrate for the HRP-modified electrode) and acetylthiocholine chloride (required co-substrate for cholinesterase) was studied. Performance of bi-enzyme biosensor arrays in the batch mode and in the flow-injection system are discussed

A steady-state and flow-through cell for screen-printed eight-electrode arrays

An electrochemical cell has been developed enabling amperometric steady-state- and flow-injection measurements with screen-printed arrays consisting of eight working electrodes (circle divide = 1 mm) arranged radially around a printed Ag/AgCl reference electrode in the centre. The cell contained a rotator, providing similar hydrodynamics over all the working electrodes in the array, which was manually centered under the rotator. The reproducibility of steady-state measurements with eight-electrode platinum or gold arrays in this cell was studied by measuring and comparing currents from ferricyanide reduction at each electrode in the array. It was found that the relative standard deviation (R.S.D.) for the currents at different electrodes on one array was below 5%. Similar R.S.D. was found if measurements were compared between several arrays. This indicates that manual insertion/positioning of the eight-electrode array in the cell and hydrodynamics at the electrodes provided measurement reproducibility similar to the reproducibility of manufacturing eight-electrode platinum or gold arrays by screen-printing. A comparative study was performed between screen-printed and through mask sprayed carbon arrays. It was found that the reproducibility of the sprayed arrays was similar to that of the platinum or gold screen-printed arrays, with R.S.D. values below 6% regarding the variation between electrodes within the same array and the variation between different arrays. To enable flow-injection measurements, a tube (0.4 mm inner diameter) was inserted into a hole drilled through the centre of the steady-state cell rotator. This construction made it possible to inject the solution into the cell through the tube (not rotating), while the rotator was spinning over the eight-electrode array. It was found that this combination of flow-injection and mixing by a rotator provided a uniform current response over the array electrodes and that, at optimum conditions, the R.S.D. values between the eight electrodes in the array were nearly the same as in case of the steady-state measurements, i.e., below 5%. (C) 2004 Elsevier B.V. All rights reserved

Multivariate data analysis of dynamic amperometric biosensor responses from binary analyte mixtures - application of sensitivity correction algorithms

In this paper, it is demonstrated that a single-receptor biosensor can be used to quantitatively determine each analyte in binary Mixtures LIS in multivariate data analysis tools based on the dynamic responses received from flow injection peaks. Mixtures with different concentrations of two phenolic compounds, catechol and 4-chlorophenol, were measured with a graphite electrode modified with tyrosinase enzyme at an applied potential of -50 mV versus Ag/AgCl. A correction algorithm based on measurements of references in-between samples was applied to compensate for biosensor ageing as well as differences caused by deviations between biosensor preparations. After correction, the relative prediction errors with partial least squares regression (PLS-R) for catechol and 4-chlorophenol were 7.4 and 5.5%, respectively, using an analysis sequence measured on one biosensor. Additional validation mixtures of the two phenols were measured with a new biosensor, prepared with the same procedure but with a different batch of tyrosinase enzyme. Using the mixture responses for the first sensor as a calibration set in PLS-R. the relative prediction errors of the validation mixtures, after applying correction procedures. were 7.0% for catechol and 16.0% for 4-chlorophenol. These preliminary results indicate that by applying correction algorithms it could be possible to use less stable biosensors in continuous on-line measurements together with multivariate data analysis without time-consuming calibration procedures. (C) 2004 Elsevier B.V. All rights reserved