Sensitive bi-enzymatic biosensor based on polyphenoloxidases-gold nanoparticles-chitosan hybrid film-graphene doped carbon paste electrode for carbamates detection.

A bi-enzymatic biosensor (LACC-TYR-AuNPs-CS/GPE) for carbamates was prepared in a single step by electrodeposition of a hybrid film onto a graphene doped carbon paste electrode (GPE). Graphene and the gold nanoparticles (AuNPs) were morphologically characterized by transmission electron microscopy, X-ray photoelectron spectroscopy, dynamic light scattering and laser Doppler velocimetry. The electrodeposited hybrid film was composed of laccase (LACC), tyrosinase (TYR) and AuNPs entrapped in a chitosan (CS) polymeric matrix. Experimental parameters, namely graphene redox state, AuNPs:CS ratio, enzymes concentration, pH and inhibition time were evaluated. LACC-TYR-AuNPs-CS/GPE exhibited an improved Michaelis-Menten kinetic constant (26.9±0.5M) when compared with LACC-AuNPs-CS/GPE (37.8±0.2M) and TYR-AuNPs-CS/GPE (52.3±0.4M). Using 4-aminophenol as substrate at pH5.5, the device presented wide linear ranges, low detection limits (1.68×10(-9)±1.18×10(-10)-2.15×10(-7)±3.41×10(-9)M), high accuracy, sensitivity (1.13×10(6)±8.11×10(4)-2.19×10(8)±2.51×10(7)%inhibitionM(-1)), repeatability (1.2-5.8% RSD), reproducibility (3.2-6.5% RSD) and stability (ca. twenty days) to determine carbaryl, formetanate hydrochloride, propoxur and ziram in citrus fruits based on their inhibitory capacity on the polyphenoloxidases activity. Recoveries at two fortified levels ranged from 93.8±0.3% (lemon) to 97.8±0.3% (orange). Glucose, citric acid and ascorbic acid do not interfere significantly in the electroanalysis. The proposed electroanalytical procedure can be a promising tool for food safety control.

[1]  Damià Barceló,et al.  Evaluation of a highly sensitive amperometric biosensor with low cholinesterase charge immobilized on a chemically modified carbon paste electrode for trace determination of carbamates in fruit, vegetable and water samples , 1999 .

[2]  Armando C. Duarte,et al.  Advances in point-of-care technologies with biosensors based on carbon nanotubes , 2013 .

[3]  Ajaya K. Singh,et al.  Polymer thin films embedded with metal nanoparticles for electrochemical biosensors applications. , 2013, Biosensors & bioelectronics.

[4]  Simone Morais,et al.  Simple laccase-based biosensor for formetanate hydrochloride quantification in fruits. , 2014, Bioelectrochemistry.

[5]  T. Ng,et al.  A laccase with inhibitory activity against HIV-1 reverse transcriptase from the mycorrhizal fungus Lepiota ventriosospora , 2013 .

[6]  Jean-Louis Marty,et al.  Biosensors for Pesticide Detection: New Trends , 2012 .

[7]  Mirela Diaconu,et al.  Bienzymatic sensor based on the use of redox enzymes and chitosan–MWCNT nanocomposite. Evaluation of total phenolic content in plant extracts , 2011 .

[8]  Claude Durrieu,et al.  A bi-enzymatic whole cell conductometric biosensor for heavy metal ions and pesticides detection in water samples. , 2005, Biosensors & bioelectronics.

[9]  J. Kochana,et al.  Tyrosinase/laccase bienzyme biosensor for amperometric determination of phenolic compounds , 2008 .

[10]  C. Alonso,et al.  Nanostructured rough gold electrodes for the development of lactate oxidase-based biosensors. , 2010, Biosensors & bioelectronics.

[11]  Antonio V. Herrera-Herrera,et al.  Dispersive Solid‐Phase Extraction , 2015 .

[12]  J. Kimling,et al.  Turkevich method for gold nanoparticle synthesis revisited. , 2006, The journal of physical chemistry. B.

[13]  Ke-Jing Huang,et al.  Amperometric immunobiosensor for α-fetoprotein using Au nanoparticles/chitosan/TiO(2)-graphene composite based platform. , 2013, Bioelectrochemistry.

[14]  S. Morais,et al.  Subacute Effects of the Thiodicarb Pesticide on Target Organs of Male Wistar Rats: Biochemical, Histological, and Flow Cytometry Studies , 2013, Journal of toxicology and environmental health. Part A.

[15]  John B. Shoven,et al.  I , Edinburgh Medical and Surgical Journal.

[16]  R. Yu,et al.  Bienzymatic amperometric biosensor for choline based on mediator thionine in situ electropolymerized within a carbon paste electrode. , 2004, Analytical biochemistry.

[17]  R. K. Shervedani,et al.  Direct electrochemistry of dopamine on gold-Agaricus bisporus laccase enzyme electrode: characterization and quantitative detection. , 2012, Bioelectrochemistry.

[18]  B. Pletschke,et al.  Review on the use of enzymes for the detection of organochlorine, organophosphate and carbamate pesticides in the environment. , 2011, Chemosphere.

[19]  H. J. Salavagionea,et al.  Spectroelectrochemical study of the oxidation of diaminophenols on platinum electrodes in acidic medium , 2005 .

[20]  K. Vårum,et al.  Influence of chitosan structure on the formation and stability of DNA-chitosan polyelectrolyte complexes. , 2005, Biomacromolecules.

[21]  C. Tortolini,et al.  Laccase-based biosensor for the determination of polyphenol index in wine. , 2010, Talanta.

[22]  V. Zucolotto,et al.  The use of mixed self-assembled monolayers as a strategy to improve the efficiency of carbamate detection in environmental monitoring , 2013 .

[23]  Cristina Freire,et al.  Laccase-Prussian blue film-graphene doped carbon paste modified electrode for carbamate pesticides quantification. , 2013, Biosensors & bioelectronics.

[24]  Eric R. Ziegel,et al.  Statistics and Chemometrics for Analytical Chemistry , 2004, Technometrics.

[25]  Zhen Yang,et al.  Catalytic Properties of Tyrosinase from Potato and Edible Fungi , 2006 .

[26]  O. Fatibello‐Filho,et al.  Synergic effect studies of the bi-enzymatic system laccase-peroxidase in a voltammetric biosensor for catecholamines. , 2003, Talanta.

[27]  Qin Xu,et al.  In situ immobilization of glucose oxidase in chitosan-gold nanoparticle hybrid film on Prussian Blue modified electrode for high-sensitivity glucose detection , 2006 .

[28]  S. Bose,et al.  Recent advances in graphene-based biosensors. , 2011, Biosensors & bioelectronics.

[29]  F. Pariente,et al.  Laccase biosensors based on different enzyme immobilization strategies for phenolic compounds determination. , 2013, Talanta.

[30]  H. Salavagione,et al.  Spectroelectrochemical study of the oxidation of diaminophenols on platinum electrodes in acidic medium , 2004 .

[31]  José M. Pingarrón,et al.  Amperometric selective biosensing of dimethyl- and diethyldithiocarbamates based on inhibition processes in a medium of reversed micelles , 1997 .

[32]  Mark E. Orazem,et al.  Electrochemical Impedance Spectroscopy: Orazem/Electrochemical , 2008 .

[33]  J. Caetano,et al.  Determination of carbaryl in tomato in natura using an amperometric biosensor based on the inhibition of acetylcholinesterase activity , 2008 .

[34]  M. Yazdani-Pedram,et al.  Hybrid ternary organic-inorganic films based on interpolymer complexes and silica , 2004 .

[35]  Damià Barceló,et al.  Sensors and biosensors in support of EU Directives , 2009 .

[36]  C. Delerue-Matos,et al.  Extraction of ochratoxin A in bread samples by the QuEChERS methodology. , 2012, Food chemistry.

[37]  Jean-Louis Marty,et al.  Biosensors based on highly sensitive acetylcholinesterases for enhanced carbamate insecticides detection , 2006 .

[38]  J. Oehlmann,et al.  Before the curtain falls: endocrine-active pesticides--a German contamination legacy. , 2011, Reviews of environmental contamination and toxicology.

[39]  Wenping Zhao,et al.  Acetylcholinesterase biosensor based on chitosan/prussian blue/multiwall carbon nanotubes/hollow gold nanospheres nanocomposite film by one-step electrodeposition. , 2013, Biosensors & bioelectronics.

[40]  Tapas Kuila,et al.  Efficient synthesis of graphene sheets using pyrrole as a reducing agent , 2011 .

[41]  J. Dupont,et al.  Bioelectroanalytical Determination of Rutin Based on bi‐Enzymatic Sensor Containing Iridium Nanoparticles in Ionic Liquid Phase Supported in Clay , 2010 .

[42]  Ruo Yuan,et al.  Bi-enzyme synergetic catalysis to in situ generate coreactant of peroxydisulfate solution for ultrasensitive electrochemiluminescence immunoassay. , 2012, Biosensors & bioelectronics.

[43]  S. Andreescu,et al.  Studies of the binding and signaling of surface-immobilized periplasmic glucose receptors on gold nanoparticles: a glucose biosensor application. , 2008, Analytical biochemistry.

[44]  Steven J Lehotay,et al.  Fast and easy multiresidue method employing acetonitrile extraction/partitioning and "dispersive solid-phase extraction" for the determination of pesticide residues in produce. , 2003, Journal of AOAC International.

[45]  Simone Morais,et al.  Biosensor based on multi-walled carbon nanotubes paste electrode modified with laccase for pirimicarb pesticide quantification. , 2013, Talanta.

[46]  Jonathan N. Coleman,et al.  The preparation of hybrid films of carbon nanotubes and nano-graphite/graphene with excellent mechanical and electrical properties , 2010 .

[47]  Xiuhua Zhang,et al.  A novel tyrosinase biosensor based on chitosan-carbon-coated nickel nanocomposite film. , 2012, Bioelectrochemistry.

[48]  R. Kaner,et al.  Honeycomb carbon: a review of graphene. , 2010, Chemical reviews.

[49]  K. Temsamani,et al.  Investigation of biosensor signal bioamplification: comparison of direct electrochemistry phenomena of individual Laccase, and dual Laccase-Tyrosinase copper enzymes, at a Sonogel-Carbon electrode. , 2008, Talanta.

[50]  A. Telefoncu,et al.  Maltose biosensing based on co-immobilization of alpha-glucosidase and pyranose oxidase. , 2010, Bioelectrochemistry.

[51]  Alessandra Bonanni,et al.  Graphene for electrochemical sensing and biosensing , 2010 .

[52]  K. Kit,et al.  Production and characterization of thick, thin and ultra-thin chitosan/PEO films , 2011 .

[53]  Åke Bergman,et al.  State of the Science of Endocrine Disrupting Chemicals - 2012 , 2012 .

[54]  G. Zeng,et al.  A tyrosinase biosensor based on ordered mesoporous carbon-Au/L-lysine/Au nanoparticles for simultaneous determination of hydroquinone and catechol. , 2013, The Analyst.

[55]  T. Dantas,et al.  Effect of molecular weight and ionic strength on the formation of polyelectrolyte complexes based on poly(methacrylic acid) and chitosan. , 2006, Biomacromolecules.

[56]  Xiaogang Qu,et al.  Graphene Oxide: Intrinsic Peroxidase Catalytic Activity and Its Application to Glucose Detection , 2010, Advanced materials.