Biologically Modified Carbon Paste Electrodes - Electroanalysis with Carbon Paste Electrodes - page 112

The. sketch. (Figure. 5.3A). shows. the. oxidation.of. a. reduced. substrate,. S Red ,. with.the. aid. of. the.
enzyme..The.enzyme,.in.fact.its.oxidized.cofactor.or.coenzyme,.is.reduced;.and.in.biological.sys-
tems.it.must.be.reconstituted.by.a.cosubstrate.in.the.oxidized.form,.which.is.reduced.as.a.conse-
quence..If.the.cosubstrate.is.oxygen,.the.enzymes.are.called.oxidases.with.the.possibilities.to.reduce.
O 2 . to. hydrogen. peroxide. or. water;. in. case. the. cofactor. is. a. biological. mediator,. such. as. nicotine.
adenine.dinucleotide,.NAD + ,.they.are.classiied.as.dehydrogenases.
On.the.other.hand,.hydrolases.are.enzymes.that.catalyze.the.hydrolytic.cleavage.of.a.substrate.
(Figure.5.3B)..To.some.extent.they.are.also.exploited.for.preparing.enzyme-modiied.carbon.paste.
sensors.

5.2.2.1 Oxidases
Oxidases.use.elemental.oxygen,.O 2 ,.as.an.electron.acceptor.for.the.oxidation.of.the.substrate..
The.simplest.way.to.regenerate.the.initial.state.of.the.enzyme.(Figure.5.3A).would.be.a.direct.
electrochemical. conversion. of. the. cofactor. or. coenzyme. that. is. located. at. the. active. center..
Practically,.it.is.very.dificult.in.most.cases.because.the.active.center.is.usually.buried.under-
neath.the.protein.shell.of.the.apoenzyme,.which.prevents.or.deteriorates.the.electron.transport.
inside.the.enzyme.
When. exploiting. oxidases. for. designing. biosensors,. there. are. quite. a. few. possibilities. to.
obtain.the.amperometric.signals.being.proportional.to.the.concentration.of.the.substrate.(see.
Figure.5.4).
Going.back.in.history,.it.is.found.that.the.irst.biosensor.was.designed.by.L..Clark.in.the.early.
1960s.[1908].and,.indeed,.it.was.based.on.the.detection.of.depletion.of.oxygen,.measured.by.the.
oxygen.sensor.(also.developed.by.him).and.which.can.be.estimated.as.the.very.irst.amperometric.
sensor.[1909-1911]..This.design.is.one.possibility.to.correlate.the.substrate.concentration.to.the.cur-
rent.signal.obtained.by.the.reduction.of.O 2 .(see.Figure.5.4,.pathway.“A”).
Many. oxidases. produce. hydrogen. peroxide. as. an. intermediate:. it. is. electroactive. and. can. be.
detected.either.by.reduction.to.water.(pathway.“B”).or.by.oxidation.to.oxygen.(pathway.“C”)..The.
possibilities.to.detect.the.intermediate.are.sketched.in.Figure.5.5.
Direct.electrochemical.conversion.of.H 2 O 2 .imposes.some.problems.because.both.electrochemi-
cal.reactions,.oxidation.(Figure.5.5,.pathway.“A”).and.reduction.(Figure.5.5,.pathway.“B”),.show.
very.high.overpotentials.with.all.the.risks.of.detecting.a.lot.of.other.compounds.

Artificial
electron
acceptor
replaces
oxygen

(ii)

E Ox

H 2 O

red.

2e -

S Red B

A red

H 2 O ox.

Direct
ox.

(i)

ox.

red.

or

ne -

ne - F

ne -

E

2e -

C D

O 2

A A Ox

S Ox

2/4

e - “Wire”

red.
ox.

E Red
Third

Used for
measure-
ment

red.

First
Sensor generation

Second

FIGURE 5.4 Way.of.action.of.oxidases,.electrochemical.detection,.and.sensor.generations.