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Amperometric Biosensors
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Introduction Enzyme Catalyzed redox reactions The function of the enzyme is to generate or consume an electroactive species in a stoichiometric relationship with its substrate or target analyte Amperometric transducer allows for the electrometrical reactions to proceed at the electrode surface, thus giving rise to a current
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Introduction - Amperometric Biosensor Schematic
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Introduction S+C enzyme P + C’ S = Substrate C = Cofactor / Coreactant P = Product C’ = Cofactor Product
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Introduction Three generations 1 st generation: the normal product of the reaction diffuses to the transducer and causes electrical response 2 nd generation: involves specific mediators between reaction and transducer to generate improved response
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Introduction 3 rd generation: reaction itself causes the response
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Amperometric Glucose Biosensor Developed by Updike and Hicks First reported use of enzyme electrode Glucose in blood-diagnosis of diabetes O 2 detection H 2 O 2 detection Enzyme mediation
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Glucose Biosensor – O 2 Central – Pt cathode, surrounding Ag/AgCl anode, KCl electrolyte solution Teflon membrane Cathode (0.6V) O 2 + 4H + +4e - 2H 2 O Anode4Ag + 4Cl - 4AgCl + 4e - Glucose +O 2 glucoseoxidase Gluconic acid + H 2 O
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Glucose Biosensor – O 2
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Glucose Biosensor – H 2 O 2 Reverse polarity of electrodes for the O 2 set up Replace Teflon membrane with cellulose acetate membrane Cathode: 2Ag + 2Cl - 2AgCl + 2e - Pt Anode (0.6V): H 2 O 2 O 2 +2H + + 2e - Counter electrode: 4H + +O 2 2H 2 O – 4e -
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Glucose Biosensor – H 2 O 2
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Glucose Biosensor – redox mediators Replace O 2 with synthetic e- acceptor to shuttle e- from the flavin redox center of the enzyme to the electrode Measurements become insensitive to O 2 fluctuations, and can be carried out at lower potentials Enzyme mediators – dimethyl ferrocene, medola blue
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Glucose Biosensor – Redox mediators Glucose + Gox(FAD)gluconic acid + Gox(FADH 2 ) Gox(FADH 2 )+2M ox Gox(FAD)+2M red +2H + 2Mred2M ox + 2e-
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Common Enzyme electrodes Measured Species EnzymeDetected species LactateLactate dehydrgenase NADH LactateLactate oxidase H2O2H2O2 PhenolTyrosinaseQuinone CholesterolCholesterol oxidase O2O2
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Some Commercial applications YSI : Glucose/Lactate Prostate specific antigen (PSA) NOVA Bioprofile analyzer (PO 2, Gluc, Gln, Glu, Lactate)
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Other Applications Glucose: meat freshness, fermentation Rapid cell number monitor Monitor of herbicides in surface waters Amperometric and novel fluorescent DNA probes Rapid electrochemical sensor for Iron
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References A.G. Elie,Principles of Potentiometric and Amperometric Biosensors,University of Virginia,(2002) P.V. Climent, M.L.M.Serralheiro, and M.J.F. Rebelo, Pure and Applied Chemistry 73, pp.1993-1999, 2001 http://www.lsbu.ac.uk.html
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