Enzyme Immobilization on Chitosan-Based Supports - Chitosan Based Hydrogels: Functions and Applications - page 384

nanoparticles. Different from laponite, layered double hydroxides (ldhs), Zn 3 Al(OH) 8 Cl,
are anionic clays and display a layered structure built on a stacking of positive layers. The
positively charged layer may be an attractive point to immobilize biomolecules depending
on their IEP.
Fan et al. explore two phenol biosensors for phenol determination based on chitosan-
laponite composite matrix and chitosan-ldhs composite film, respectively [148,149].
Chitosan was utilized to improve the analytical performance of the pure clay-modified
bioelectrode. PPO was simply entrapped into this novel composite film. GA was avoided
in making the biosensor. These biosensors exhibited good affinity to the substrate; the
apparent Michaelis-Menten constant ( K m ) for the chitosan-laponite sensor and the chito-
san-ldh sensor was 0.16 and 0.13 mM, respectively. The enzyme electrode provided a
linear response to catechol over a concentration range of 3.6 × 10 −9 to 4 × 10 −5 M with a
sensitivity of 2750 ± 52 mA M −1 cm −2 for chitosan-laponite sensors and a concentration
range of 5.3 × 10 −9 to 4 × 10 −5 M with a sensitivity of 674 ± 4 mA M −1 cm −2 for chitosan-ldhs
sensors. It seems that substrate affinity and sensitivity of chitosan-ldhs sensors are rela-
tively higher due to highly improved adhesive ability.
8.5.4.6.3 MMT-Chitosan-Gold Nanocomposite Film
Gold nanoparticles stabilized by chitosan (AuCS) were hybridized with exfoliated clay
nanoplates through electrostatic interaction. The resulting clay (MMT)-chitosan-GNP
nanocomposite (Clay-AuCS) was used to modify GCE. HRP, as a model peroxidase, was
entrapped between the Clay-AuCS film and another clay layer. The results demonstrated
that the quasireversible, surface-controlled electron transfer kinetics for HRP was realized
on Clay-AuCS-modified GCE. The nanocomposite showed advantages over clay and AuCS
alone in fabricating biosensors. Moreover, HRP retained its native secondary structure in
the Clay-AuCS film and bioelectrocatalytic activity with good sensitivity and fast response
toward hydrogen peroxide [150].
8.5.4.7 Chitosan-Inorganic Salt Composite Material
8.5.4.7.1 Chitosan-Nano-Calcium Carbonate Composite
A great challenge for the fabrication of biosensors comes from the effective immobilization
of enzyme to the solid electrode surface. In recent years, calcium carbonate has been
proved to intensify enzyme performance and has been widely used in industry, medicine,
microcapsule fabrication, and many other bio-related fields. In particular, because of the
large surface-to-volume ratio, the high hydrophilicity, and biocompatible characteristics,
nanosized calcium carbonate (nano-CaCO 3 ) has been demonstrated to be a promising
enzyme immobilization matrix.
Gong et al. present a facile approach for constructing a novel functional hybrid film
composed of nano-CaCO 3 -chitosan by a one-step coelectrodeposition method (Figure 8.14) .
The generated nano-CaCO 3 -based matrix possessed a 3D porous, network-like structure,
providing a favorable and biocompatible microenvironment to immobilize enzyme. By
using such a composite film as an enzyme immobilization matrix, a highly sensitive and
stable AChE sensor was achieved for the determination of methyl parathion as a model of
organophosphate pesticide (OP) compounds. The detection limit was found to be as low as
1 ng mL −1 ( S / N = 3). The designed biosensor exhibited good reproducibility and acceptable
stability [151].