Amperometric Biosensor for Pesticide Methamidophos Assay (original) (raw)
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Analytica Chimica Acta, 1994
and carbamic pesticides have been determined with an amperometric acetylcholinesterasebased 4-aminophenyl acetate biosensor. The glassy carbon enzyme membrane covered electrode poised at + 250 mV (vs. sodium chloride saturated calomel electrode) oxidizes the 4-aminophenol formed in the hydrolysis of 4aminophenyl acetate by acetylcholinesterase in the glutaraldehyde cross-linked layer. The activity of acetylcholinesterase is inhibited in the presence of pesticides. The decrease in activity of the enzyme is monitored by the 4-aminophenyl acetate sensor and is correlated to the concentration of pesticide present in solution. The influence of the acetylcholinesterase loading and the acetylcholinesterase to neutral protein (bovine serum albumin) ratio on the biosensor response was studied and the measuring conditions including pH, substrate concentration, and others were optimized. Detection liits of 4.0 and 13.0 nmol 1-t for paraoxon and carbaryl, respectively, were achieved with a 3-min preincubation time.
Biosensors and Bioelectronics, 2014
The work presented here describes a novel, easy and low-cost method of fabrication of a highly sensitive acetylcholinesterase biosensor and its application to detect organophosphate and organocarbamate pesticides. Acetylcholinesterase was electro-immobilized into a thick conducting layer of polypyrrole. Porcine skin gelatin and gluteraldehyde mixture was used for stabilizing the system. Acetylthiocholine chloride was used as the substrate. Polypyrrole catalyzed the electrochemical oxidation of thiocholine and promoted the electron transfer, thus lowering the oxidation potential and increasing the detection sensitivity. Electro oxidation of thiocholine in polypyrrole matrix occurred at 0.1 V under low potential scan rate. The thiocholine sensitivity of the electrode was found to be 143 mA/M. The sensor was applied to detect the sample organophosphate pesticide ethylparaoxon and organocarbamate pesticide carbofuran. The detection limit for paraoxon was found to be 1.1 ppb and that for carbofuran is 0.12 ppb. The sensor showed good intra and inter state precision with relative standard deviation (RSD) 0.742% and 6.56% respectively. Both dry and wet storage stability were studied. The sensor stored at 0 1C in dry condition had a good storage stability retaining 70% of its original activity for 4 months. During wet storage, the activity decrease followed the same trend, however, the operational stability at the end of the storage period was found to be less compared to the dry storage case. The developed biosensor is as a promising new tool for analysis of cholinesterase inhibitors.
Acetylcholinesterase inhibition-based biosensors for pesticide determination: A review
Analytical Biochemistry, 2012
Pesticides released intentionally into the environment and through various processes contaminate the environment. Although pesticides are associated with many health hazards, there is a lack of monitoring of these contaminants. Traditional chromatographic methods-high-performance liquid chromatography, capillary electrophoresis, and mass spectrometry-are effective for the analysis of pesticides in the environment but have certain limitations such as complexity, time-consuming sample preparation, and the requirement of expensive apparatus and trained persons to operate. Over the past decades, acetylcholinesterase (AChE) inhibition-based biosensors have emerged as simple, rapid, and ultra-sensitive tools for pesticide analysis in environmental monitoring, food safety, and quality control. These biosensors have the potential to complement or replace the classical analytical methods by simplifying or eliminating sample preparation and making field-testing easier and faster with significant decrease in cost per analysis. This article reviews the recent developments in AChE inhibition-based biosensors, which include various immobilization methods, different strategies for biosensor construction, the advantages and roles of various matrices used, analytical performance, and application methods for constructing AChE biosensors. These AChE biosensors exhibited detection limits and linearity in the ranges of 1.0Â10-11 to 42.19 lM (detection limits) and 1.0 Â 10 À11-1.0 Â 10 À2 to 74.5-9.9 Â 10 3 lM (linearity). These biosensors were stable for a period of 2 to 120 days. The future prospects for the development of better AChE biosensing systems are also discussed.
Amperometric detection of pesticides using polymer electrodes
… monitoring and assessment, 2006
The real time monitoring of some organophosphorus based pesticides is of great concern to environmentalists because the widespread use of pesticides is causing severe health hazards to all living beings and also hampering our ecological balance. The traditional methods of measurement of pesticide residues are time consuming, need sample pre-treatment, and lack desired specificity and accuracy. We have developed an amperometric biosensor for indirect measurement of the pesticide concentration precisely in ppb level. The method is based on the action of two enzymes namely acetylcholine esterase and choline oxidase which are uniquely immobilized in a polymeric porous network directly on the working electrode of a screen-printed sensor. Polyacrylamide matrix has been prepared by copolymerisation of acrylamide and N ,N -methylenebisacrylamide using Potassium peroxodisulphate (K 2 S 2 O 8 ) as initiator. A linear relationship was obtained between the range of 0 to 10 ppb.
Biotechnology & Biotechnological Equipment, 2012
A flow-injection system with integrated amperometric biosensor featuring an easily replaceable immobilzed acetylcholinesterase (AChE) membrane was studied. The amperometric biosensor was constructed on the basis of site-specific immobilization of AChE on a hybrid polymer membrane with integrated multi-walled carbon nanotubes. Multistage modification of the membrane and immobilization of the enzyme was proved by Fourier transform infrared spectroscopy. The optimum flow-rate of the flowinjection analysis (FIA) system was 0.5 mL/min. It gave a linear response to acetylthiocholine chloride from 2 μM to 100 μM, with an average RSD of 3.0% (n = 6). The sensitivity of the constructed biosensor was 0.093 µA/µM•cm 2. The K m app value of the immobilized AChE was 1.15 mM and the linear correlation coefficient R 2 , 0.9949. The method had a low detection limit for three organophosphorus pesticides (OPs) in model pesticide solutions-paraoxon ethyl (0.9×10-12 M), monocroptophos (1.8×10-12 M) and dichlorvos (2.0×10-12 M). This indicated that the action of multi-walled nanotubes and controlled site-specific enzyme immobilization ensured high electrocatalytic activity and selectivity of the biosensor towards pesticides. It was found that the biosensor can be reused 15 operation cycles. After storage for 30 days the enzyme membrane retained over 80% of its initial response. The FIA system was used for detection of anti-cholinesterase activity of two binary OP mixtures. The results for paraoxon + monocroptophos and paraoxon + dichlorvos showed that the total inhibition activity was not simply additive, but was lower than the sum of the individual inhibition values. Moreover, the difference between the sum of the individual inhibition values and the real results for the mixture was bigger for the binary system paraoxon and dichlorvos (7-10%) compared with that for paraoxon and monocroptophos (5-7%). The developed biosensor system is an ideal tool for monitoring of organophosphate pesticides.
Biosensors and Bioelectronics, 2003
A new highly sensitive amperometric method for the detection of organophosphorus compounds has been developed. The method is based on a ferophthalocyanine chemically modified carbon paste electrode coupled with acetylcholinesterase and choline oxidase co-immobilized onto the surface of a dialysis membrane. The activity of cholinesterase is non-competitively inhibited in the presence of pesticides. The highest sensitivity to inhibitors was found for a membrane containing low enzyme loading and this was subsequently used for the construction of an amperometric biosensor for pesticides. Analyses were done using acetylcholine as substrate; choline produced by hydrolysis in the enzymatic layer was oxidized by choline-oxidase and subsequently H 2 O 2 produced was electrochemically detected at '/0.35 V vs. Ag/AgCl. The decrease of substrate steady-state current caused by the addition of pesticide was used for evaluation. With this approach, up to 10 (10 M of paraoxon and carbofuran can be detected.
Sensors and Actuators B: Chemical, 2014
A novel amperometric biosensor based on a conducting polymer using multi walled carbon nanotube modified electrode was developed for detection of organophosphorus pesticides. Acetylcholinesterase (AChE) was successfully immobilized by covalent linkage on the modified graphite electrode. Carbon nanotubes were functionalized by electrochemical treatment. A conducting polymer; poly(4-(2,5-di(thiophen-2-yl)-1H-pyrrol-1-yl)benzenamine) (poly(SNS-NH 2)) was synthesized via electropolymerization to examine its matrix properties for biomolecule immobilization. This strategy enhanced electron transfer rate at a lower potential (+100 mV vs. Ag reference) and catalyzed electrochemical oxidation of acetylthiocholine effectively. Scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), contact angle measurements and Electrochemical Impedance Spectroscopy (EIS), Cyclic Voltammetry (CV) techniques were used to monitor changes in surface morphologies and electrochemical characterizations. The proposed biosensor design offered a fast response time (6 s), a wide linear range (0.05 mM and 8.00 mM) and a low detection limit (0.09 mM) with a high sensitivity (24.16 µA mM-1 cm-2) for acetylthiocholine. The inhibition responses of paraoxon, parathion and chlorfenvinphos on the enzymatic activity of AChE were detected. The fabricated biosensor was tested for the detection of pesticides in fortified tap water samples. The results were found to be in good agreement with the ones determined by HPLC/DAD technique.
Rapid Detection of Methomyl and Organophosphorous Pesticides with Portable Potentiometric Biosensor
Simple portable potentiometric biosensor has been assessed for rapid in-field detection of neurotoxic anticholineesterase pesticides. The proposed biosensor based on inhibition of the acetylcholinesterase enzyme (AChE) by carbamate, organophosphorous pesticides, namely; methomyl, prothiofos and chlorpyrifos. Comprehensive studies were performed for the optimum measuring parameters for each pesticide and its degradation products. Based on the relative inhibition action of pesticide on AChE activity, different sensitivities were recorded ranging between 0.25 ng to 37.5 μg mL -1 depending on the nature of pesticide tested and its detoxification affect (LD 50 ). From different degradation products of methomyl, inhibition by methomyl S-oxide and methomyl S,S-dioxide were more potent allowing determination of 0.20 and 0.088 ng mL -1 , respectively. Reasonable average recoveries ranging between 96.22±1.98 and 106.02±2.10 % were obtained. The applicability of the proposed method was tested by determination of pesticides in their commercial formulation and agricultural drainage water samples without pre-treatment procedures. Acceptable sensitivity and accuracy were obtained compared with the official chromatographic method indicated by good average recoveries between 94.73±2.74 and 108.3±0.80 %. The proposed sensors are promising and can be used as simple and direct screening devices for pesticide monitoring in field conditions available to unskilled end-users with significant decrease in cost per analysis to complement or replace the classical chromatographic methods.
Acetylcholine enzyme sensor for determining methamidophos insecticide
Analytica Chimica Acta, 2001
A sensitive screen-printed amperometric sensor suitable for rapid determination of the concentration of the insecticide methamidophos was developed. It was based on the principle of inhibition of acetylcholinesterase (AChE) activity. The first part of the study was focused on the screening of several genetically modified AChEs in order to select the most sensitive enzyme towards the methamidophos. Values for the bimolecular constant k i were also determined. In a second part of the study, we compared the lowest detectable methamidophos concentrations using different immobilised AChEs. The lowest detectable concentration in a standard solution was 1.4 ppb for the AChE(Dros)-B03 mutant compared to 4.8 ppb for the wild type AChE and 53 ppb for the electric eel source.
Talanta, 2002
A screen-printed biosensor for the detection of pesticides in water miscible organic solvents is described based on the use of p-aminophenyl acetate as acetylcholinesterase substrate. The oxidation of p-aminophenol, product of the enzymatic reaction was monitored at 100 mV vs. Ag/AgCl screen-printed reference electrode. Miscible organic solvents as ethanol and acetonitrile were tested. The acetylcholinesterase (AChE) was immobilised on a screen-printed electrode surface by entrapment in a PVA-SbQ polymer and the catalytic activity of immobilised AChE was studied in the presence of different percentages of organic solvents in buffer solution. The sensor shows good characteristics when experiments were performed in concentrations of organic solvents below 10%. No significant differences were observed when working with 1 and 5% acetonitrile in the reaction media. Detection limits as low as 1.91 × 10 − 8 M paraoxon and 1.24 × 10 − 9 M chlorpyrifos ethyl oxon were obtained when experiments are carried out in 5% acetonitrile.