Amperometric Biosensors

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

Introduction - Amperometric Biosensor Schematic

Introduction

S+C enzyme P + C’

S = Substrate

C = Cofactor / Coreactant

P = Product

C’ = Cofactor Product

Introduction

Three generations 1st generation: the normal product of the

reaction diffuses to the transducer and causes electrical response

2nd generation: involves specific mediators between reaction and transducer to generate improved response

Introduction

3rd generation: reaction itself causes the response

Amperometric Glucose Biosensor

Developed by Updike and Hicks First reported use of enzyme electrode Glucose in blood-diagnosis of diabetes O2 detection

H2O2 detection

Enzyme mediation

Glucose Biosensor – O2

Central – Pt cathode, surrounding Ag/AgCl anode, KCl electrolyte solution

Teflon membrane Cathode (0.6V) O2 + 4H+ +4e- 2H2O Anode 4Ag + 4Cl- 4AgCl +

4e-

Glucose +O2 glucoseoxidase Gluconic acid + H2O

Glucose Biosensor – O2

Glucose Biosensor – H2O2

Reverse polarity of electrodes for the O2 set up

Replace Teflon membrane with cellulose acetate membrane

Cathode: 2Ag + 2Cl-2AgCl + 2e-

Pt Anode (0.6V): H2O2 O2+2H+ + 2e-

Counter electrode: 4H++O2 2H2O – 4e-

Glucose Biosensor – H2O2

Glucose Biosensor – redox mediators

Replace O2 with synthetic e- acceptor to shuttle e- from the flavin redox center of the enzyme to the electrode

Measurements become insensitive to O2 fluctuations, and can be carried out at lower potentials

Enzyme mediators – dimethyl ferrocene, medola blue

Glucose Biosensor – Redox mediators

Glucose + Gox(FAD) gluconic acid + Gox(FADH2)

Gox(FADH2)+2Mox Gox(FAD)+2Mred+2H+

2Mred 2Mox + 2e-

Common Enzyme electrodesMeasured Species

Enzyme Detected species

Lactate Lactate dehydrgenase

NADH

Lactate Lactate oxidase

H2O2

Phenol Tyrosinase Quinone

Cholesterol Cholesterol oxidase

O2

Some Commercial applications

YSI : Glucose/Lactate Prostate specific antigen (PSA) NOVA Bioprofile analyzer (PO2, Gluc,

Gln, Glu, Lactate)

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

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

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