APPLIED PHYSICS LETTERS 101, 023703 (2012)
1,3,a)
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,2,a)
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Ved Varun Agrawal, and B. D. Malhotra
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Surendra K. Yadav,
Department of Science and Technology Centre on Biomolecular Electronics, Biomedical Instrumentation
Jay Singh,
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Section, National Physical Laboratory, Dr. K.S. Krishnan Marg, New Delhi 110012, India
Department of Physics, Motilal Nehru National Institute of Technology, Allahabad, Allahabad 211004, India
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Department of BIN Fusion Technology and Department of Polymer-Nano Science and Technology, Chonbuk
National University, Jeonju, Jeonbuk 561-756, Korea
Department of Biotechnology, Delhi Technological University, Shahbad Daulatpur, Delhi 110042, India
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(Received 15 May 2012; accepted 22 June 2012; published online 13 July 2012)
Xanthine oxidase (XO ) has been physisorbed onto nanostructured nickel oxide (n-NiO, ꢀ41 nm),
x
electrophoretically deposited onto indium-tin-oxide (ITO) glass substrate using poly-ethylene glycol
via co-precipitation. The electrochemical response studies of the XO /n-NiO/ITO bioelectrode show
x
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linear response in the xanthine concentration range as 10–200lM (R ¼ 0.9989), sensitivity as
2
0
.6214 lA/lM cm , detection limit as 2.989 lM, standard deviation as 0.6192 lA, response time
of 15 s with the stability of 100 days. The observed low value (0.0306 mM) Michaelis–Menten
x
The past decade has seen considerable rise in the
demand for fish as a nutritious and healthy commodity both
ethylene glycol (PEG) used as a surfactant. The observed
green precipitate washed with deionized water and ethanol
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in developing and the developed countries. There is thus an
ꢁ
for 4–5 times is dried at 70 C for about 12 h and is then cal-
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urgent need for the availability of a device that can be used
for evaluation of fish freshness. In this context, xanthine has
been found to be a major metabolite in degradation of adeno-
sine triphosphate (ATP) contained in the fish meat, whose
cined at 400 C for about 8 h. The n-NiO (10 mg/ml) dis-
persed in double distilled water is ultrasonicated for about
30 min and is then electrophoretically deposited (EPD) at
35 V for 60 s to form a uniform film onto an indium-tin-oxide
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concentration increases with storage. Xanthine can thus be
(ITO) (0.25 cm ) glass surface. XO (0.2 unit/ml), in phos-
x
used as an indicator for estimation of fish freshness. Various
analytical methods like high performance liquid chromatog-
raphy (HPLC) and high performance capillary electrophore-
sis (HPCE) are being used for detection and quantification of
phate buffer, 50 mM, pH 7.0) is immobilized onto n-NiO/
ITO electrode via physisorption and is kept overnight in a
humid chamber for drying. The higher activity of XOx/
n-NiO/ITO bioelectrode toward the substrate (xanthine) is
obtained at pH 7.0 (Fig. 2, inset (ii)).
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xanthine. However, these methods are time-consuming,
expensive, and require expertise. Enzyme biosensor based
on xanthine oxidase (XO ) has recently been suggested for
The n-NiO particles have been characterized using
X-ray diffraction (Rikagu) Fourier transform infrared spectra
(FTIR, PerkinElmer, Spectrum BX II), scanning electron mi-
croscopy (SEM, LEO-440), transmission electron micros-
copy (TEM, Hitachi Model H-800) with a tungsten filament
at an accelerating voltage of 200 kV the electrochemical
analysis has been conducted by an Autolab Potentiostat/Gal-
vanostat (Eco Chemie, The Netherlands) using a three-
electrode system with XOx/n-NiO/ITO bioelectrode as the
working electrode, a platinum (Pt) wire as the counter elec-
trode, and saturated Ag/AgCl electrode as a reference elec-
trode in phosphate buffer saline (PBS) (50 mM, pH 7.0,
x
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estimation of xanthine. In this context, immobilization of
XO onto a desired matrix including nanostructured metal
x
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oxide is considered very important. Among the various
nanostructured metal oxides, nano-structured nickel oxide
has aroused much interest for fabrication of a desired biosen-
sor. This has been attributed to its large surface area, high
conductivity, efficient catalytic activity, wide band gap, high
carrier mobility, good biocompatibility, chemical stability,
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and excellent electrochemical properties. Moreover, the
high isoelectric point (IEP) of NiO (ꢀ10.7) can be helpful to
immobilize a given biomolecules with low IEP via electro-
static interactions. We report results of the studies relating to
fabrication of a fish freshness biosensor based on nanostruc-
tured nickel oxide.
3ꢂ/4ꢂ
0.9% NaCl) containing 5 mM [Fe(CN)6]
. The contact
angle (Dataphysics, Model OCA15EC) measurements have
been done by Sessile drop method to delineate the hydro-
philic/hydrophobic nature of the electrode.
All chemicals of analytical grade have been purchased
from Sigma Aldrich. Nanostructured n-NiO nanoparticles
have been prepared by using Ni(NO ) . 6H O of (0.5 M) and
The XRD pattern of n-NiO nanoparticles (Fig. 1) per-
formed in the 2h range exhibits characteristic peaks indexed
at (111), (200), (220), (222), and (311) corresponding to
nickel oxide (JCPDS file 78-0429) having fcc crystal struc-
ture. The average crystallite size has been obtained by
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NaOH of (2 M) are separately dissolved in 100 ml distilled
water and n-NiO has been prepared using co-precipitation
method to adjust pH up to ꢀ12 by utilizing 2–3 ml of poly-
˚
Debye–Scherrer equation (k ¼ 1.5406 A) and it turns out to be
˚
41nm, lattice parameter (4.166 A). The strain in the lattice
ꢀ
of the n-NiO has been evaluated via Hall–Williamson rela-
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a)
S. K. Yadav and J. Singh contributed equally to this work.
b)
Author to whom correspondence should be addressed. Electronic mail:
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tion using Scherrer constant (k¼ 0.94), (k ¼ 1.5406A) and
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bansi.malhotra@gmail.com.
of linear fit of the data points gives (g ¼ ꢂ3.46 ꢃ 10 ). The
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Yadav, Surendra K.
