Biosensors based on immobilization of biomolecules in sol-gel matrices - Electrochemical Sensors, Biosensors and Their Biomedical Applications - page 517

the construction of mediator-free electrodes that are highly sensitive, selective, and
stable [93, 94, 113]. Mediatorless amperometric biosensors have been prepared by
coupling GOD with HRP or soybean peroxidases, which mediate the reduction of
hydrogen peroxide [93, 94, 114]. The fundamental problem arising in the construc-
tion of an amperometric glucose biosensor is the selectivity of the substrate detection.
The often-used detection approach is electrochemical oxidation of the liberated hydro-
gen peroxide through the enzymatic reaction and requires a relatively high working
potential. At such a potential, endogenous or exogenous compounds present in bio-
logical samples (e.g. urate, ascorbate or paracetamol) can be electrochemically oxi-
dized, leading to a high level of interference and false results in the quantifi cation of
glucose concentration. To overcome this limitation Wang et al. [115] and Sampath and
Lev [93, 94] took the advantage of the catalytic properties of palladium-modifi ed car-
bon particles to detect glucose at lower potentials ( 0.5 V) via a screen-printing

process or by molding the porous organically modifi ed silica in a glass capillary.
One quick and stable biosensor performance was reported by the doping of grafting
copolymer poly(vinyl alcohol) grafting 4-vinylpyridine into TEOS sol-gel [116]. The
response of the glucose amperometric biosensor was less than 20 s with a linear range
up to 9 mM and a sensitivity of 405 nA/mM. This polymer retained well the activity of
entrapped enzyme and was able to fi rmly adhere to the electrode surface and enhanced
hydrophilic nature [2, 116]. Similarly the GOD entrapped in cellulose-doped titanium
oxide composite showed improved stability and lifetime of electrode, but the main
drawback of this biosensor was the long time (

0.3,

0.5 h) required to reach a steady state,
which limited its application in analysis of a practical sample online [117].
Organic phase enzyme biosensors have been received remarkable importance in
pharmaceutical and other sectors. Such organic phase biosensors provide some distinct
advantages, such as an extended analyte range due to increased analyte solubility of
certain reactants, monitoring of many hydrophobic substrates, prevention of undesir-
able side reactions and decreased microbial contamination. Enzymes usually lose their
protein conformation in pure organic solvents. To avoid this some of the studies added
trace amounts of water to the sides of the electrode [118]. Silica sol-gel-entrapped tyro-
sinase-based biosensors have also been reported for the phenol determination in non-
aqueous phase [119]. This sensor has taken about 18 seconds to reach steady-state
current. Recently Yu and Ju [120] reported a titania sol-gel-based tyrosinase biosen-
sor for the phenol determination in organic phase. At 100 mV vs SCE this biosen-

sor showed a good amperometric response to phenols in pure chloroform without any
mediator and rehydration of the enzyme. For catechol determination the sensor exhib-
ited a fast response of less than 5 s. The sensitivity of different phenols was as follows:
catechol

phenol p-cresol. The biosensor showed good reproducibility and stability

(70 days at 4 ), attributed to the titania sol-gel membrane which effectively retained the

essential water layer around the enzyme molecule needed for maintaining its activity in
organic phase.
The fouling and microbial contamination of the surface during operation are main
problems in the long-term use of enzyme biosensors. Wang et al. [121] reported for
the fi rst time a screen-printed electrode modifi ed with GOD/HRP that was stable up

β