Photonic crystals in biology

Poster Session, Tuesday, June 15
Theme A1 - B702
Tyrosinase-Fe 3 O 4 -Chitosan Nanobiocomposite Film for Biosensor Application
1 and -Ozkan 1 *
1 Izmir Institute of Technology, Chemical Engineering Department, Gulbahce Koyu, 35430-
Abstract- Tyrosinase-Fe 3 O 4 -chitosan nanobiocomposite film was prepared in order to determine the pesticide residue as a biosensor
application in the following research. The immobilized tyrosinase enzyme on the Fe 3 O 4 -chitosan nanocomposite prepared by coating of
chitosan onto the Fe 3 O 4 nanoparticles was also characterized by the measurements of atomic force microscopy (AFM), Fourier transform
infrared (FTIR) spectra, thermogravimetric analysis (TGA) and differential scanning calorimetric analysis (DSC).
Contaminated pesticide residues for ground water, raw food
materials and processed foods results form the application of
organophosphate pesticides (OP) to increase the amount and
quality of the product [1]. Reliable analytical procedures are
required in order to determine the pesticide residue at low
level. Spectrophotometric and chromatographic techniques
are analytical methods commonly used [2]. These methods
have a number of disadvantages such as expensive
instrumentation requirement, highly trained personnel, time
consumption, and difficulties in adaptation for field analysis.
In recent years enzyme electrodes show remarkable
advances for the detection of toxic compounds based on
enzymatic reactions. Tyrosinase enzyme is one of the
appropriate enzymes to detect the pesticide residue depending
on the inhibition mechanism directly related to decrease in
the amount of enzymatically produced quinones while
substrate (catechol) is consumed [3] as shown in Figure 1.
Figure 2. The illustration of tyrosinase immobilized on Fe 3 O 4 -
chitosan nanocomposite
The morphology of the (a) Fe 3 O 4 nanoparticles, (b)
chitosan film, (c) Fe 3 O 4 nanoparticle-chitosan and (d)
tyrosinase-Fe 3 O 4 -chitosan nanobiocomposite films were
characterized with AFM. The binding mechanism of each
material was characterized by using FTIR spectra. TGA and
DSC were performed to reveal the formation of
nanobiocomposite material.
Corresponding author: 0Tfehimeozkan@iyte.edu.tr
Figure 1. Basic reaction mechanism of tyrosinase on catecol
The effective immobilization of tyrosinase on the electrode
surface is the key issues for the development of tyrosinase
biosensors. The immobilization methods can be classified as
chemical (covalent cross-linking and covalent binding) and
physical (physical entrapment, micro encapsulation, and
adsorption) methods [4]. In recent years magnetic
nanoparticles are becoming the focus of researchers due to
lots of properties. Especially Fe 3 O 4 has significant properties
such as strong superparamagnetism, low toxicity and large
surface area provides high enzyme loading [5]. However this
nanoparticle tends to aggregate and therefore stabilizers such
as surfactants, metal nanoparticles and polymeric compounds
have been used. As a natural polymer, chitosan has the
characteristic of biodegradable, biocompatible, bioactive,
nontoxic, film forming ability, physiological inertness and
high mechanical strength [6].
In this study, Fe3O 4 -chitosan nanocomposite was prepared
to immobilize the tyrosinase enzyme on it. This
nanobiomaterial was characterized for the development of
tyrosinase biosensor used in determination of pesticide
residue as the future work.
For this aim tyrosinase solution was prepared in phosphate
buffer solution (PBS) at pH 6.5. Acetic acid was used for the
preparation of chitosan solution. The Fe 3O 4 nanoparticle
suspensions was prepared by dispersing the particle in double
distilled water with ultrasonication. Then tyrosinase
solutions, chitosan solutions and Fe 3 O 4 nanoparticles were
mixed with a specific volume ratio in order to obtain
tyrosinase-Fe 3 O 4 -chitosan nanobiocomposite film (Figure 2).
[1] Ingrid Walz I., Schwack W., 2007. Cutinase inhibition by
means of insecticidal organophosphates and carbamates Part 1:
Basics in development of a new enzyme assay, Eur. Food. Res.
Technol., 225: 593-601.
[2] Poerschmann, J., Zhang, Z., Kopinke, F.D., Pawliszyn, J.,
1997. Solid Phase Microextraction for Determining the
Distribution of Chemicals in Aqueous Matrixes, Anal. Chem., 69:
597–600
[3] Munoz J.L.-Molina F., Varon R., Rodriguez-Lopez
-Canovas F., Tudela J., 2008 Calculating molar
absorptivities for quinones: Application to the measurement of
tyrosinase activity, J. Comput. Aided Mol. Des., 22: 299-309.
[4] Buerk D.G., 1993. Biosensors; Theory and Applications.
U.S.A.
[5] Gu, H. W.; Xu, K. M.; Xu, C. J.; Xu, B., 2006. Biofunctional
Magnetic Nanoparticles for Protein Separation and Pathogen
Detection, Chem. Commun., 941-949.
[6]Barbara K. , 2004. Application of chitin- and chitosan-based
materials for enzyme immobilizations: a review, Enzyme and
Microbial Technology, 35: 126–139
.
6th Nanoscience and Nanotechnology Conference, zmir, 2010 331