Biomedical Engineering Reference
In-Depth Information
17.3.2 Biosensors based on direct electron transfer of enzymes
17.3.2.1 Biosensors based on direct electron transfer of horseradish
peroxidase
Horseradish peroxidase (HRP) is a member of the large class of peroxidases, which are
enzymes defi ned as oxidoreductases using hydroperoxide as electron acceptor. HRP
has been widely used for the construction of amperometric biosensor for the determi-
nation of H
2
O
2
and small organic and inorganic substrates.
Poly(ester sulfonic acid), with the trade name of Eastman AQ, is a kind of anionic
ionomer. Enzymes can be added directly to aqueous dispersions of AQ ionomers and
deposited onto solid surfaces to form fi lms without denaturation or signifi cant loss of
activity. The embedded HRP in AQ fi lms retained the electrocatalytic activity for oxy-
gen, nitrite, and hydrogen peroxide [246]. A certain volume of air can increase the
reduction peak of HRP-AQ fi lms in CVs. This increase in reduction peak was accom-
panied by the disappearance of the oxidation peak for HRP-Fe (II) because HRP-Fe (II)
reacted with oxygen. An increase in the amount of oxygen in solution increased the
reduction peak current. NO
2
can also be reduced by the HRP-AQ/PG electrode.
When NO
2
was added into a pH 6.0 buffer, a catalytic peak was observed at about
0.84 V. The peak current increased with the concentration of NO
2
. Electrochemical
catalytic response of hydrogen peroxide was also observed by HRP-AQ fi lms. When
H
2
O
2
was added to a pH 7.0 buffer, an increase in reduction peak at about
0.36 V
was seen with the disappearance of oxidation peak for HRP-Fe(II). The reduction
peak current increased with the concentration of H
2
O
2
in solution. However, there was
no reduction current observed at AQ fi lm electrodes in the presence of H
2
O
2
.
Biomembranes in living organisms are composed of lipid, protein, and carbohydrate.
The biomembranes can be prepared by casting aqueous vesicle of lipid onto the surface
of electrodes. The lipids were arranged in multiple bilayers by self-assembling and retain
their bilayer properties similar to biomembrane after drying. When we immobilize HRP
in the DDAB fi lms by casting the mixture of the aqueous vesicle of DDAB and HRP
on GC electrodes, the enzyme electrode showed catalytic activity toward H
2
O
2
[204].
The electrocatalytic reduction of H
2
O
2
by HRP immobilized in the DDAB fi lms was
tested by CVs. When H
2
O
2
was added to the buffer solution, the cathodic peak increases
dramatically and the anodic peak almost disappears for the HRP-DDAB/GC electrode.
These results indicate that HRP incorporated into the DDAB fi lms can catalyze the
reduction of H
2
O
2
effi ciently and still retains its biological activity. The amperometric
response of the enzyme electrode resulted from increasing concentrations of H
2
O
2
was
investigated in the stirred buffer solution (pH 5.5). When an aliquot of H
2
O
2
is added
into the buffer solution, the reductive current rises steeply to reach a stable value. There
is a linear relationship between the current response and H
2
O
2
concentration in the range
from 1
10
3
to 4
10
3
M. The regression equation is
y
0.04007
0.05186
x
,
with a correlation coeffi cient of 0.998.
Biopolymers such as agarose hydrogel can also be used to immobilize HRP and to
reduce oxygen, hydrogen peroxide and nitric oxide [231]. For an HRP-agarose/EPG
in a pH 7.0 PBS, a reduction peak at
0.30 V was observed after addition of H
2
O
2
.
Search WWH ::
Custom Search