Sensors and Actuators B 236 (2016) 304–310

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Sensors and Actuators B: Chemical

journal homepage: www.elsevier.com/locate/snb

Nickel oxide-deposited cellulose/CNT composite electrode for

non-enzymatic urea detection
Nhi Sa Nguyen, Hyon Hee Yoon ∗
Department of Chemical and Bio Engineering, Gachon University, Gyeonggi-do 461-701, Republic of Korea

a r t i c l e i n f o a b s t r a c t

Article history: A highly sensitive non-enzymatic urea sensor was fabricated using nickel oxide deposited on
Received 12 April 2016 a cellulose/carbon nanotube (CNT) composite. The structure and morphological properties of
Received in revised form 28 May 2016 the NiO/cellulose/CNT composite were investigated using X-ray diffraction and scanning electron
Accepted 31 May 2016
microscopy. A thin film of the NiO/cellulose/CNT composite was coated on an indium tin oxide glass to
Available online 2 June 2016
fabricate an amperometric urea sensor. The composition of the modified electrode and detection condi-
tions (i.e., pH and temperature) were optimized. The prepared electrode exhibited an excellent sensitivity
Keywords:
of 371 ␮A mM−1 cm−2 with a fast response time of 4 s. The electrode also showed high stability with only
Urea sensor
Non-enzymatic
a 3.6% decrease in its sensitivity after storage for two months under ambient conditions. In addition, the
Nickel oxide feasibility for urea analysis in urine samples was demonstrated.
Cellulose © 2016 Elsevier B.V. All rights reserved.
Carbon nanotube

1. Introduction observed to exhibit good catalytic activity for the electrochemical

Monitoring the level of biologically important molecules, such
as urea, glucose, creatinine, and dopamine, in the human body is
a vital issue in biochemical and medical diagnostics [1–3]. Vari-
ous types of biosensors, including enzymatic biosensors, have been
used intensively in the detection of such molecules owing to its
selectivity, simplicity, and reliability [4–7]. However, the critical
issue encountered with the enzymatic biosensors is denaturation
of the enzyme, which results in poor stability. Moreover, enzy-
matic biosensors often fail to generate a strong electrical signal
and require mediators to transport electrons. In contrast, non-
enzymatic biosensors comprising metals and metal oxides are
considered to be potential substitute for enzymatic biosensors
because of their excellent redox flexibility and stability [8,9].
A variety of metals, alloys, and metal oxides, such as Pt, Rh,
Pt-Rh, Cu/Cu oxide, Zn/Zn oxides, Ni/Ni oxides, and Co/Co oxides,
have been widely used for the detection of different molecules.
Among them, Ni and Ni oxides have been extensively documented
for the direct electro-catalytic oxidation of biochemical molecules
such as urea [10,11] and glucose [12,13]. Nickel-based catalysts
are relatively cheaper and demonstrate superior catalytic proper-
ties for urea oxidation in alkaline medium as compared with other
metals/metal oxides. In our earlier study, a Ni-based catalyst was
∗ Corresponding author.
E-mail address: hhyoon@gachon.ac.kr (H.H. Yoon).
oxidation of urea [14]. The mechanism of electrochemical oxidation
of urea on Ni-based electrodes in alkaline media can be represented
with the following reactions [15]:
NiO + OH−  NiOOH + e− (1)
− −
Ni(OH)2 + OH  NiOOH + e (2)
− NiOOH
CO(NH2 )2 + 6OH → N2 + 5H2 O + CO2 + 6e− (3)
A common problem with Ni-based catalyts is the degrada-
tion and expansion of the catalyst structure during the oxidation
reaction, which adversely affects their catalytic performance.
Furthermore, such structures have a tendency to self-assemble,
forming agglomerations, which reduces their specific surface area
and decreases their electrochemical activity [16]. To overcome
such problems, supporting materials, such as carbon nanotubes,
graphene, and cellulose nanofibers, have been employed. Cellulose
and its derivatives, by virtue of their large functionality (hydroxyl,
ether, esters groups, etc.), have been found to be good supporting
materials for the anchorage of different metal/metal oxide struc-
tures, providing a good platform for the construction of bio-sensing
devices [17]. Mahadeva and Kim [18] deposited tin oxide onto a
cellulose scaffold to fabricate a urea sensor, which exhibited good
chemical stability and sensitivity.
Carbon nanotubes (CNTs) have also garnered much attention as
a supporting material for sensor fabrication due to their outstand-
ing properties such as high electron conductivity, high chemical
and thermal stability, and high surface area [19]. Amalgamations of

http://dx.doi.org/10.1016/j.snb.2016.05.165
0925-4005/© 2016 Elsevier B.V. All rights reserved.
 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310 305

cellulose with CNTs benefit from the advantages of the properties

of both the individual materials and thus can provide an improved
support material. Qi et al. [20] synthesized quasi-one-dimensional
CNT/cellulose composite fibers with good electrical conductivity.
The CNT networks formed with support of the functionalized cellu-
lose structure exhibited significant improvement in their electrical
and mechanical properties [21,22].
In the present work, a novel electrode employing nickel oxide
deposited on a cellulose/CNT composite was prepared for the
non-enzymatic sensing of urea. The structural, morphological, and
electrochemical properties of the electrode were investigated. The
prepared electrode exhibited excellent sensitivity for urea detec-
tion.

2. Experimental

2.1. Materials Fig. 1. XRD patterns of cellulose and a NiO/cellulose/CNT composite.

CNTs (multi-walled, diameter 20 nm, length 5 ␮m) weres pur-

chased from Carbon Nano Material Technology Co., Ltd. Sigmacell
cellulose (type 101, highly purified fibers), nickel nitrate hex-
ahydrate (98.5%) were purchased from Sigma-Aldrich. All other
chemicals used in this work were reagent grade products. Indium
tin oxide (ITO) glass (<20 X/square) was used as a base electrode
and was cut into a rectangular shape of 5 mm in width and 20 mm
in length. The working area of the ITO glass for the electrochemical
tests was controlled at 0.25 cm2 using masking tape.
2.2. Preparation of cellulose fibrils from orange peel
Natural cellulose was prepared from orange peel waste by
removing the oil, color, and pectin following a previous method
[23]. First, dried orange peel was immersed into a solution of
water/ethanol (15:85) at 80 ◦ C for 20 min to remove the colored
materials and oils, after which the solid portion was separated
by centrifugation. This extraction step was repeated until the
sample was free from color. The sample was then washed with
de-ionized (DI) water. Similarly, metal constituents were removed
using ammonium oxalate solution (50 mM in water) at 20 ◦ C for
1 h, and pectin was removed using 0.18 wt% HCl at 80 ◦ C for 1 h and
0.2 wt% NaOH at 5 ◦ C for 1 h. The sample was then washed with DI
water and dried in an oven overnight. The resulting cellulose was
denoted Cel-1. Commercial cellulose (Sigmacell cellulose), denoted
Cel-2, was used without further purification.
2.3. Preparation of NiO/cellulose/CNT composites
The NiO/cellulose/CNT composites were fabricated via a hydrox-
ide thermal decomposition method. First, a CNT-dispersed solution
in N,N-dimethylformamide (DMF, 1 mg mL−1 ) was prepared using a
sodium dodecyl sulfate surfactant. Similarly, a cellulose-dispersed
solution in DMF (1 mg/mL) was also prepared using a 2% cation-
ized starch. The CNT and cellulose solutions were mixed at
a CNT/cellulose ratio of 1:9, unless specified otherwise, and
homogenized for 2 h at room temperature. To this dispersion, a
pre-calculated amount of nickel nitrate hexahydrate was added,
and the dispersion was stirred for 3 h. Sodium borohydride (as a
reducing agent) was then slowly added with a Ni/Na ratio of 1:2.5,
and stirred for 1 h. The precipitated Ni(OH)2 /cellulose/CNT com-
posite material was filtered and washed with DI water and acetone
repeatedly. Finally, the precipitate was heated at 280 ◦ C for 4 h in
order to convert Ni(OH)2 on the cellulose/CNT network into NiO
and thus to obtain a NiO/cellulose/CNT composite.
2.4. Electrode fabrication
The electrodes were fabricated by a vacuum-assisted deposi-
tion method. First, the NiO/cellulose/CNT composite was dispersed
into DMF at a concentration of 1 mg mL−1 . An aliquot of the
NiO/cellulose/CNT dispersion was filtered through an Anodisc
membrane that was attached to a suction filter apparatus. After
filtration, the membrane was air-dried. The membrane was then
placed on the ITO glass, and dipped into a 0.1 M NaOH aqueous solu-
tion for 10 min during which the Anodisc membrane was etched
away leaving a uniformly NiO/cellulose/CNT coated ITO glass. It was
then washed repeatedly with DI water and dried in a vacuum oven
at 60 ◦ C overnight. The NiO/cellulose/CNT/ITO electrode was then
covered by an encapsulating membrane, a polyion complex (PIC) of
poly-l-lysine and polystyrene sulfonate, as described in a previous
report [14].
2.5. Analysis
The morphology and compositional analysis of the
NiO/cellulose-CNT composites were characterized using a scanning
electron microscope (SEM, Hitachi S-4700; Hitachi Ltd., Tokyo,
Japan), and energy dispersive X-ray spectroscopy (EDX). Powder
X-ray diffraction (XRD) analysis of the samples was performed
using an X-ray diffractometer (Rigaku D/MAX-2200, Japan) with
Cu-K␣ radiation at a wavelength of 1.5406 Å.
Cyclic voltammetry and chronoamperometry measurements
were carried out using a potentiostat-galvanostat (VSP, Biologic-
Science Instruments, 38640 CLAIX, France). All the electrochemical
measurements were conducted at room temperature using a
conventional three—electrode system (wherein Ag/AgCl was the
reference electrode, a platinum wire was the counter electrode,
and the sample was the working electrode).
3. Results and discussion
3.1. Characterization of NiO/cellulose/CNT composite
The XRD patterns of cellulose and the NiO/cellulose/CNT com-
posite are shown in Fig. 1. The cellulose sample (Cel-1) exhibits two
prominent peaks at 2␪ = 16.2◦ and 21.9◦ , attributed to the (110) and
(002) reflections of cellulose, respectively [24].
The NiO/cellulose/CNT composite showed peaks for cellulose,
(002) and (101) reflection peaks at 2␪ = 21.9◦ and 42.8◦ , respec-
tively; peaks for CNT; and characteristic peaks at 2␪ = 36.7◦ , 44.1◦ ,
306 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310

Fig. 2. SEM images of (a) NiO/Cel-1, (b) NiO/Cel-2, (c) NiO/Cel-1/CNT (×3 K), (d) NiO/Cel-1/CNT (×30 K), and (e) a cross-section of NiO/Cel-1/CNT on ITO glass. An EDX
elemental map of NiO/Cel-1/CNT composites for Ni (f).

62.5◦ , and 75.0◦ , corresponding to the (111), (200), (220), and (311)
reflections, respectively, for fcc phase NiO (JCPDS-47-1049).
Fig. 2a and b show the SEM images of the NiO/cellulose com-
posites. The NiO/Cel-1 (Fig. 2a) appeared to be more porous with
a larger surface area than the NiO/Cel-2 (Sigmacell, Fig. 2b). Fig. 2c
and d represent the SEM images of the NiO/Cel-1/CNT compos-
ite with a magnification of 3.0 K and 30.0 K, respectively. A porous
structure and a uniform distribution of NiO particles with an aver-
age diameter of ∼70 nm in the NiO/Cel-1/CNT composite were
observed. A cross-sectional SEM image of the NiO/Cel-1/CNT com-
posite layer on ITO glass exhibited a fibril structure with a thickness
of approximately 5 ␮m, as shown in Fig. 2e. The distribution of NiO
was further confirmed by EDX mapping for Ni (Fig. 2f).

3.2. Electrochemical characterization of NiO/cellulose-CNT/ITO

electrode Fig. 3. The chronoamperometric response of different electrodes to successive addi-
tions of urea solution at an applied potential of 0.4 V.

The effect of CNTs and cellulose on the electro-catalytic oxida-

tion of urea by a NiO-based electrode was investigated. Fig. 3 shows
the chronoamperometric response to the successive addition of
urea at an operating potential of 0.4 V. All the electrodes exhibited
a current increase as urea concentration increased, indicating that
the NiO/cellulose composite material had electro-catalytic activity
for urea oxidation. At 0.5 mM of urea, the current density val-
ues of NiO/Cel-1, NiO/Cel-2, and NiO/Cel-1/CNT were 75, 68, and
125 ␮A cm−2 , respectively. No detectable current was observed
in the absence of urea. Compared to the NiO/Cel-2 (Sigmacell
cellulose, a highly purified commercial cellulose), the NiO/Cel-
 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310
Fig. 4. (a) Oxidation peak currents and peak potentials at different loadings of
NiO/Cel-1/CNT composite on an ITO electrode, and (b) CVs of the NiO/Cel-1/CNT
composite electrode with different cellulose/CNT ratio at 20 mM urea in Tris–HCl
buffer (pH 8.0).
1 (cellulose from orange peel) showed a higher current density.
This result was mainly caused by the higher porosity of the Cel-
1 because the urea oxidation occurring on the electrode was a
diffusion controlled process, as discussed later. A further increase
of current density at a given urea concentration was obtained by
the addition of CNTs, due to better electrical conduction in the
electrode matrix. The enhanced conductivity and permittivity of
a CNT/cellulose composite has been previously reported [22].
The thickness of the structure of the NiO/Cel-1/CNT electrode
was optimized. An increase in the thickness causes an increase in
the loading of catalyst material, resulting in a fast reaction rate.
However, the diffusional resistance will increase with the increase
in the thickness. Fig. 4a shows the oxidation peak potentials and
the respective current of the NiO/Cel-1/CNT electrodes with vary-
ing thicknesses (the corresponding cyclic voltammograms (CVs)
are provided in Suppl. Fig. S1). The thickness of the NiO/Cel-1/CNT
composite material on the base electrode (ITO) was assumed to be
proportional to the loading amount of the composite material. The
oxidation peak current increased as the loading of the composite
material (i.e., the thickness) increased up to 30 ␮g (on 0.25 cm2 ITO
glass), as expected. After which, it decreased as the thickness fur-
ther increased, probably due to the increased diffusional resistance.
Therefore, the optimum thickness of the NiO/Cel-2/CNT composite
layer on the base ITO electrode was determined to be 30 ␮g per
0.25 cm2 , this loading was used for the subsequent studies. The
thickness at this loading was approximately 5 ␮m, as observed in
the SEM image (Fig. 2e). As shown in the inset of Fig. 4a, the oxida-
tion peak potential was as low as ∼0.4 V at the optimum thickness,
and thus oxygen-evolution reaction (OER) could be avoided [10,11].
307
Fig. 5. The oxidation peak currents at different pHs (a) and at different temperatures
(b) in the presence of 20 mM urea.
The effect of the cellulose and CNT ratio on the electrochem-
ical behavior of the NiO/cellulose/CNT composite electrode was
examined. Fig. 4b shows the CVs of the NiO/Cel-1/CNT compos-
ite electrode with different Cellulose/CNT ratio at 20 mM urea in
Tris–HCl buffer (pH 8.0). The NiO/Cel-1 (no CNT) exhibited the high-
est anodic peak current of 710 ␮A cm−2 (at a potential of 0.62 V),
whereas the NiO/CNT (no cellulose) showed the lowest anodic peak
current peak of 400 ␮A cm−2 (at a potential of 0.38 V). These results
suggested that the NiO catalyst particles on cellulose matrix were
more active for the urea oxidation than those on CNT, probably
due to the fact that cellulose possesses free hydroxyl groups that
can facilitate the deposition of metal ions on its surface [25]. How-
ever, the oxidation peak potential peak of NiO/Cel-1 was found
to be relatively high (0.62 V), which could be favored for OER. On
the other hand, the oxidation peak potential of NiO/CNT was rela-
tively low (0.38 V). As shown in Fig. 5, the NiO/Cel-1/CNT composite
with cellulose/CNT ratio of 9/1 exhibited a relatively high anodic
peak current at a relatively low peak potential, revealing that CNT
enhanced the electron transfer via providing conduction paths in
the cellulose matrices [21]. This composition was used through the
study.
The effects of the measurement conditions of the electrode, such
as pH and temperature, on the current signal were examined. As
shown in Fig. 5a, the pH of the medium significantly affected the
catalytic activity. As expected, the peak current intensity increased
as the pH increased up to ∼8, because the urea oxidation reaction
is favorable under alkaline conditions. However, a further increase
in pH above 8.0 adversely affected the current output due to the
308 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310

Table 1
Comparison of the analytical performance of amperometric urea biosensors.

Electrode material Sensitivity (␮A mM−1 cm−2 ) Response time (s) Detection limit(␮M urea) Linear range (mM urea) Stability (days) References
a
PAPP/urease – 25–50 – 6.3 × 10−3 –0.407 14 at b RT [6]
Rh/urease 1.85 15 50 1.75 27 at 4 ◦ C [30]
c
CPEs/Ur/GLDH 5.0 0.02–0.2 15 at 4 ◦ C [31]
d
PPy/Pt 1.11 × 103 <1 40 0.08–1.44 – [9]
3Dgraphene/NiCo2 O4 166 1 5 0.06–0.30 120 at RT [14]
e
H40-Au 7.48 × 10−3 3 11 1–3 120 at RT [32]
f
NiO NPs/Ur 21.3 5 830 0.83–16.65 140 at 4 ◦ C [29]
Ur/g PANI-Nf/Au 4.2 – 1 × 103 1–10 – [33]
Ur/h GLDH/MLG 10 1.3 1.6–16 40 at 4 ◦ C [34]
NiO/cellulose/CNT 371 4 7 0.01–1.4 80 at RT Current work
a
PAPP = poly(N-3-aminopropyl pyrrole-co-pyrrole).
b
RT = room temperature.
c
CPEs = carbon paste electrodes.
d
PPy = polypyrrole.
e
H40 = hyperbranched polyester-boltron.
f
NiO NPs = NiO nanoparticles.
g
PANI = polyaniline.
h
GLDH/MLG = glutamate dehydrogenase/multilayered graphene; and Ur = urease.

excess of OH− ions at high pH, which compete with urea and block
the access of urea molecules to the active sites [11]. It should be
noted that the electrode exhibited a significant current output at
physiological pH 7 (pH 7). Fig. 5b shows the effect of temperature
on the current output. In the temperature range of 20–50 ◦ C, the
oxidation peak current increased with temperature. Higher tem-
perature should be beneficial to the urea oxidation kinetics on the
electrode. However, as the temperature exceeded 50 ◦ C, deterio-
ration of the encapsulating membrane occurred and consequently
current loss was observed.

3.3. Electrochemical performance

Fig. 6a presents the CVs of the NiO/Cel-1/CNT electrode in the

presence of 20 mM urea in a Tris–HCl buffer solution (pH 8.0)
recorded at potential scan rates from 10 to 100 mVs−1 . A peak-
to-peak potential separation (Ep ) at a scan rate of 20 mVs−1 was
determined to be as low as 0.15 V. In addition, the redox peak poten-
tial did not vary with the scan rate. These results are the indication
of fast electron transfer kinetics in the electrode material [26]. The
redox peak currents varied linearly with the square root of the
scan rate, as shown in the inset of Fig. 6a, suggesting that the urea
oxidation kinetics on the NiO/Cel-1/CNT electrode surface was a
diffusion-controlled process. Using the slope (anodic peak current
(Ipa ) vs. scan rate (v)) and the Brown-Anson model [27], the sur-
face concentration of the absorbed electroactive species (C) on the
electrode can be estimated:
n2 F 2 CAv
Ipa = (4)
4RT
n is the number of electrons transferred (here, n = 1), A is the
electrode surface area (0.25 cm2 ), F is the Faraday constant
(96485C mol−1 ), R is the gas constant (8.314 J mol−1 K−1 ), and v
is the scan rate (20 mVs−1 ). The surface concentration was calcu-
lated to be 1.29 × 10−5 mol cm−2 , which is higher than the values Fig. 6. (a) CVs of the NiO/Cel-1/CNT electrode in the presence of 20 mM urea at
various scan rates, Inset: plot of anodic and cathodic peak currents vs. the square
reported previously [28], indicating a high surface coverage by root of the scan rate and (b) CVs of the NiO/Cel-1/CNT electrode at different urea
the large amount of electroactive species on the NiO/cellulose/CNT concentrations with a scan rate of 20 mVs−1 .
composites.
In order to determine the rate constants of electron transfer (ks ),
the Laviron equation,
The NiO/Cel-1/CNT electrode was further examined at differ-
Ep nF v
ks = (5) ent urea concentrations (Fig. 6b). The anodic oxidation peak results
RT from the oxidation process of Ni2+ to Ni3+ , whereas the cathodic
was used with the parameters as used above. The value of ks was peak maps to the reverse process [11]. The oxidation current
calculated to be 116 s−1 , which is greater than previously reported increased with urea concentration up to 100 mM. At 100 mM urea,
values [29]. the peak current density (at 0.4 V) was as high as 1.0 mA cm−2 , sug-
 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310
Fig. 7. Chronoamperometric response of the NiO/Cel-1/CNT electrode to successive
additions of urea solution at an applied potential of 0.4 V. Inset: a calibration curve
of current density vs. urea concentration.
gesting that the NiO/Cel-1/CNT electrode can be used as a highly
sensitive urea sensor and also as an anode for direct urea fuel cells.
To evaluate the NiO/cellulose/CNT electrode for its performance
in urea detection, chronoamperometric tests were carried out at
0.4 V, which was the urea oxidation peak potential on the electrode,
were carried out. As shown in Fig. 7, a steady-state signal can be
observed within only ∼4 s after the addition of urea solution, indi-
cating a stable and fast amperometric response. The calibration plot
of current signal vs. urea concentration was linear in the range of
10 ␮M to 1.4 mM with a correlation coefficient of 0.997 (inset Fig. 7).
The sensitivity determined within the linear range was found to be
371 ␮A mM−1 cm−2 , as calculated from the slope of the calibration
curve. The electrode showed a detectable response to urea con-
centration of ∼7 ␮M with a signal to noise ratio of 3. The overall
performance of the NiO/Cel-1/CNT electrode is competitive or far
better than those of other urea sensors (Table 1). In particular, the
sensitivity of the prepared electrode was remarkably higher than
any other reported values.
3.4. Interference, stability, and urine sample test
For the interference test of the NiO/Cel-1/CNT electrode in a real
scenario, the amperometric response to the consecutive additions
of different organic and ionic species at concentrations relevant to
those in urine and blood samples was investigated. Fig. 8 shows the
amperometric response to the addition of urea (370 ␮M) diluted
(1:500) in Tris–HCl buffer and other likely interfering species, such
as Na+ (60 ␮M), K+ (25 ␮M), Cl− (60 ␮M), thiourea (0.5 × 10−3 ␮M),
creatinine (0.01 ␮M), ascorbic acid (2 ␮M), glycine (30 × 10−3 ␮M),
uric acid (2 ␮M), and glucose (1 or 60 ␮M). The current response
appeared to vary mainly with the addition of urea, although small
responses were observed for ascorbic acid and uric acid. The current
Table 2
Analysis of urea in diluted urine samples.
Urine sample (mM) Urea diluted urine Urea spiked Determined by
sample (␮M) sample (␮M) standard methoda
(␮M)
186 372 372 372
392 399
412 403
432 444
452 469
472 477
a
Enzymatic colorimetric method [35].
309
Fig. 8. The current responses to the addition of urea and different interfering
species: 25 ␮M K+ , 60 ␮M Na+ , 60 ␮M Cl− , 0.5 × 10−3 ␮M thiourea (TU), 2 ␮M ascor-
bic acid (AA), 2 ␮M uric acid (UA), 0.01 ␮M creatinine (C), 1 ␮M glucose (Glu), and
30 × 10−3 ␮M glycine (Gly), and (2) with 60 ␮M glucose.
responses to the addition of ascorbic acid and uric acid addition
were less than 5% of the response to the addition urea, suggesting
a good selectivity of the NiO/Cel-1/CNT electrode to urea in the
presence of interferences in urine. However, a significantly high
interfering current was observed at a high concentration of glucose
(>60 ␮M), implying a possible interference when measuring blood
samples as the electrode is also responsive to glucose at appropriate
concentrations.
The shelf life of the NiO/Cel-1/CNT electrode was evaluated after
storage for two month under ambient conditions. The electrode
exhibited only a 3.6% reduction in its performance.
The feasibility of the use of the NiO/Cel-1/CNT electrode in the
real urine sample analysis was investigated. A human urine sample
was filtered to remove any protein aggregates prior to analysis. The
concentration of urea was 186 mM, as measured by an enzymatic
colorimetric method (Suppl. Fig. S2). The sample was diluted 500
times in Tris–HCl buffer and spiked with known concentrations of
urea solution, as summarized in Table 2. The spiked urine samples
were analyzed using the NiO/Cel-1/CNT electrode and also tested
using the standard colorimetric method for comparison [35]. The
results, presented in Table 2, showed good recoveries (95–101%)
across the different concentrations. The relative standard deviation
(RSD) of the five measurements was less than 2.52%, depending on
the concentration level. These results indicate a high reproducibil-
ity of the electrode in real sample measurements and a feasibility
for practical applications.
Determined by RSD (%) Recovery
sensor (␮M) (n = 5) (%)
361 2.52 101.88
380 1.34 98.47
409 1.28 99.27
411 0.98 95.14
422 2.12 97.35
452 1.98 97.88
310 N.S. Nguyen, H.H. Yoon / Sensors and Actuators B 236 (2016) 304–310
4. Conclusions
An electrode based on a composite of CNTs and cellulose impreg-
nated with NiO demonstrated a high electro-catalytic activity for
urea oxidation. The NiO/Cel-1/CNT electrode showed a high sensi-
tivity of 371 ␮A mM−1 cm−2 with a linear range of 10 ␮M to 1.4 mM,
and a fast response time of 4 s. The electrode was stable and exhib-
ited no significant loss in activity after 2 months of storage at room
temperature. The NiO/Cel-1/CNT electrode exhibited no significant
interference in urine samples. However, a significantly high inter-
fering current was observed at a high concentration of glucose
(>60 ␮M), implying a possible interference when measuring blood
samples. When applied to a real urine sample, the sensor exhibited
good recovery and repeatability. These results indicate that this-
NiO/cellulose/CNT composite electrode is highly promising for the
non-enzymatic analysis of urea with very high sensitivity.
Acknowledgment
This work was supported by the Gachon University research
fund of 2014 (GCU-2014-0154) and the Gyeonggi Regional
Research Center.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.snb.2016.05.165.
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Biographies
Nhi Sa Nguyen received the B.S degree in Chemical Engineering from Ho Chi Minh
City University of Technology, Vietnam in 2010, M.S. degrees in Chemical Engineer-
ing from Gachon University, South Korea in 2015. She is curently a Ph.D. candidate
in Chemical Engineering, Monash University, Australia. She has research interests
in nanomaterial based biosensors and ion exchange membranes.
Hyon Hee Yoon is a professor of Department of Chemical and Bio Engineering at
Gachon University, S. Korea. He is currently the Head of Advance Materials & Applied
Technology Research Center at Gachon University. He received his Ph.D degree from
Auburn University, USA. His research interests include biosensors, biofuel cells and
biogas-fueled SOFCs.