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Talanta 71 (2007) 1096–1102
Abstract
An electrochemical sensing platform was developed based on the magnetic loading of carbon nanotube (CNT)/nano-Fe3 O4 composite on
electrodes. To demonstrate the concept, nano-Fe3 O4 was deposited by the chemical coprecipitation of Fe2+ and Fe3+ in the presence of CNTs in an
alkaline solution. The resulting magnetic nanocomposite brings new capabilities for electrochemical devices by combining the advantages of CNT
and nano-Fe3 O4 and provides an alternative way for loading CNT on electrodes. The fabrication and the performances of the magnetic nanocomposite
modified electrodes have been described. Cyclic voltammetry (CV) and constant potential measurement indicated that the incorporated CNT
exhibited higher electrocatalytic activity toward the redox processes of hydrogen peroxide. In addition, chitosan (CTS) has also been introduced
into the bulk of the CNT/nano-Fe3 O4 composite by coprecipitation to immobilize glucose oxidase (GOx) for sensing glucose. The marked
electrocatalytic activity toward hydrogen peroxide permits effective low-potential amperometric biosensing of glucose, in connection with the
incorporation of GOx into CNT/Fe3 O4 /CTS composite. The accelerated electron transfer is coupled with surface renewability. TEM images and
XRDs offer insights into the nature of the magnetic composites. The concept of the magnetic loading of CNT nanocomposites indicates great
promise for creating CNT-based biosensing devices and expands the scope of CNT-based electrochemical devices.
© 2006 Elsevier B.V. All rights reserved.
Keywords: Carbon nanotube; Magnetic loading; Electrochemical sensing; Composite; Magnetite; Hydrogen peroxide; Glucose
0039-9140/$ – see front matter © 2006 Elsevier B.V. All rights reserved.
doi:10.1016/j.talanta.2006.06.003
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S. Qu et al. / Talanta 71 (2007) 1096–1102 1097
approach to lowering the oxidation potential of hydrogen per- acetic acid while GOx solution was made by dissolving 50 mg
oxide has relied on the use of redox mediators with low formal GOx in 10 mL of 50 mM phosphate buffer (pH 7.4). 5 mg/mL of
potentials [24]. glutaraldehyde solution was prepared by the appropriate dilution
Magnetic loading provides an alternative approach to load of 25 wt.% solution of glutaraldehyde using 50 mM phosphate
various materials on electrodes. Willner et al. have reported the buffer (pH 7.4).
magnetic control of enzymatic reactions using magnetic parti-
cles functionalized with redox relay units [25,26]. Ferrocene- 2.2. Apparatus
functionalized magnetic particles were thus used for electri-
cally contacting glucose oxidase (GOx) and the electrode. The Cyclic voltammetry (CV) and constant potential measure-
use of magnetic fields to align CNT [27] and to self-assemble ments were performed with a CHI 830B electrochemical ana-
CNT devices [28] has been reported. Recently, Musameh and lyzer (Shanghai Chen-Hua Instruments Co., Shanghai, China)
Wang have demonstrated that the magnetic and catalytic prop- in a three-electrode system consisted of a 2.2 mm diameter
erties of CNT can be exploited for the magnetoswitchable con- graphite-epoxy composite electrode with small magnets accom-
trol of electron-transfer reactions [29]. Herein, we report on modated inside, a platinum auxiliary electrode, and a saturated
a novel electrochemical-sensing platform for the detection of calomel electrode (SCE) as the reference electrode. XRD mea-
hydrogen peroxide based on the magnetic loading of carbon surements were carried out using a Rigaku D/max-rB diffrac-
nanotube/nano-Fe3 O4 (CNT/Fe3 O4 ) composite on electrodes. tometer (Rigaku, Tokyo, Japan) with Cu K␣1 radiation (40 kV,
The magnetic nanocomposite dispersed in water could be loaded 60 mA). The size and morphology of magnetic nanoparticles
onto the surface of magnetic electrode with the aid of the Fe3 O4 were observed by TEM using a JEOL model JEM-2011(HR)
nanoparticles. Although CNT/Fe3 O4 nanocomposites have been at 200 kV. The sample for TEM analysis was obtained by dip-
synthesized to adsorb heavy metal ions [30] and to minimize the ping a holy grid to nanoparticles-dispersed ethanol solution and
sedimentation of Fe3 O4 particles [31], there is no earlier report evaporated in air at room temperature.
on the magnetic loading of CNT/Fe3 O4 composite on electrodes
for electrochemical sensing. 2.3. Preparation of CNT/Nano-Fe3 O4 composites
In the present work, it has been demonstrated that mag-
netic CNT/nano-Fe3 O4 composite prepared by coprecipita- Further purification of the multiwalled carbon nanotubes was
tion exhibited excellent electrocatalytic ability in the reduction accomplished by stirring the CNT in concentrated nitric acid at
and oxidation of hydrogen peroxide. In addition, CNT/nano- 60 ◦ C for 12 h. Subsequently, the obtained CNT was dried at
Fe3 O4 /chitosan composite has also been prepared by copre- 110 ◦ C for 4 h. The CNT/Fe3 O4 nanoparticle composite was
cipitation to immobilize glucose oxidase covalently for sensing prepared by the in situ chemical coprecipitation of Fe2+ and
glucose. The attractive analytical performances of the magnetic Fe3+ in alkaline solution in the presence of CNTs. The molar
nanocomposite modified electrodes, along with the character- ratio of Fe2+ :Fe3+ was 1:2. The magnetic composite was pre-
ization of the nanocomposites, are illustrated in the following pared by suspending 1.0 g purified CNT in 200 mL of solution
sections. containing 1.7 g (4.33 mmol) (NH4 )2 Fe(SO4 )·6H2 O and 2.51 g
(8.66 mmol) NH4 Fe(SO4 )2 ·12H2 O at 50 ◦ C under N2 atmo-
2. Experimental sphere. After the solution was sonicated (30 W, 40 kHz) for
10 min, 10 mL of 8 M NH4 OH aqueous solution was added
2.1. Reagent and solutions dropwise to precipitate the iron oxides while the mixture solu-
tion was sonicated. The pH of the final mixture should be in
-d-Glucose and glucose oxidase (EC 1.1.3.4, from the range of 11–12. To promote the complete growth of the
Aspergillus niger, 158 units/mg) were obtained from Sigma (St. nanoparticle crystals, the reaction was carried out at 50 ◦ C for
Louis, MO, USA). Glutaraldehyde (25 wt.% solution in water) 30 min under constant mechanical stirring. The precipitate was
was the product of Merck (Germany). Chitosan (CTS, 75–85% isolated in the magnetic field, and the supernatant was separated
deacetaylation) was purchased from Aldrich (Milwaukee, WI, from the precipitate by decantation. Impurities (such as sulfate
USA). Multiwall carbon nanotubes (MWCNT, 30–60 nm diam- and ammonia) in the CNT/Fe3 O4 samples were removed by
eter, 5–15 m long), with a purity of 95%, were provided by washing with copious amounts of doubly distilled water and the
Shenzhen Nanoport Company (Shenzhen, China). Ammonium precipitate was isolated by a permanent magnet. The obtained
ferrous sulfate and ammonium ferric sulfate were both pur- CNT/Fe3 O4 nanocomposite was then washed with 150 mL of
chased from SinoPharm (Shanghai, China). Other chemicals absolute alcohol for three times. Subsequently, the composite
were all analytical grade. was dried under vacuum. The content of CNT in the compos-
The supporting electrolyte for the electrochemical investi- ite was approximately 50 wt.%. If no CNT was added to the
gations was 50 mM phosphate buffer (pH 7.4). Stock solutions coprecipitation solution of Fe2+ :Fe3+ , pure Fe3 O4 nanoparticles
of 1 M glucose were allowed to mutarotate for 24 h before use could be obtained for the control experiments. The chemical
and subsequently kept in a 4 ◦ C refrigerator, while solutions of reaction of the Fe3 O4 precipitation occurring in the container is
hydrogen peroxide (1 M) were prepared by diluting a 30 wt.% as follows:
a stocking solution with doubly distilled water. CTS solution
(0.2 wt.%) was prepared by dissolving 0.4 g of CTS in 10 mg/mL 2Fe3+ + Fe2+ + 8NH3 ·H2 O = Fe3 O4 + 4H2 O + 8NH4 +
中国科技论文在线 http://www.paper.edu.cn
1098 S. Qu et al. / Talanta 71 (2007) 1096–1102
2.4. Magnetic electrode fabrication The cyclic voltammetry measurements were carried out in the
desired potential range at a scan rate of 100 mV/s and no solu-
Fig. 1 illustrated schematic diagrams of the magnetic elec- tion agitation was necessary during the run. Measurements of the
trode. Epoxy resin and hardener (Zhejian Cixi Tiandong Adhe- amperometric response to the sample solution were performed
sive Co. Ltd., Ningbo, China) were mixed thoroughly at a weight at a desired potential while the solution was being magnetically
ratio of 2:1. Graphite powder (Aldrich, Wilwaukee, WI, USA) stirred at 300 rpm continuously. Because the supporting elec-
and epoxy resin/hardener were hand-mixed at a ratio of 1:1 trolytes were magnetically stirred, the magnetic electrode should
(w/w). The graphite-epoxy composite was subsequently packed be inserted into the electrolyte through a hole inside the rub-
into a glass tube (Fig. 1(b), 2.2 mm i.d., 5 mm o.d., 60 mm long) ber plug on a cylinder electrochemical cell (20 mm i.d., 50 mm
by pressing the end into a sample of the composite to a depth of height) to prevent it from vibrating. The surface of the magnetic
∼3 mm. A stainless-steel rod (1.5 mm diameter) was employed electrode was downwards and should be immersed into the solu-
to press the graphite-epoxy composite (Fig. 1(d)) inside the tube tion for no more than 5 mm so that the influence of the magnetic
to acquire a flat inner surface. A piece of copper wire (Fig. 1(a), stirrer could be minimized.
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S. Qu et al. / Talanta 71 (2007) 1096–1102 1099
3. Results and discussion that the CNTs in the composite were coated with nano-Fe3 O4
that facilitated the magnetic loading of CNT on the surface of
In order to load the magnetic nanocomposite on the electrode, the electrodes.
a specially designed magnetic electrode has been prepared by Fig. 2C displays the X-ray diffraction (XRD) patterns of CNT
using NdFeB permanent magnets (Fig. 1). The magnets facili- and the CNT/nano-Fe3 O4 composite. Diffraction peaks assigned
tate the magnetic loading of the nanocomposite and can prevent to CNTs at 2θ = 26.5◦ [32] can be clearly seen for both pure
the composite from falling off in the solution. The magnetic CNT and CNT/nano-Fe3 O4 composite, indicating that the CNTs
electrode can carry a 0.5 g iron screw on its surface. When the structure was not destroyed after the chemical coprecipitation of
magnets were taken away or put back, the magnetic compos- Fe3 O4 . As shown in Fig. 2C, the other two weak peak of CNT
ite loaded on the electrode would fall off or could be reloaded, merged with that of Fe3 O4 . Six characteristic peaks for Fe3 O4
allowing a simple and fast method for renewing the modifica- (2θ = 30.2◦ , 35.6◦ , 43.3◦ , 53.7◦ , 57.3◦ and 62.8◦ ), marked by
tion layer on the electrodes. Transmission electron microscopy their indices ((2 2 0), (3 1 1), (4 0 0), (4 2 2), (5 1 1) and (4 4 0))
(TEM) was performed to observe the microstructure of the mag- [33], were observed for the CNT/nano-Fe3 O4 composite, indi-
netic composite powders. Fig. 2A illustrates the TEM image cating that the resultant Fe3 O4 particles in the composite were
of the CNT/nano-Fe3 O4 composite. The diameter of Fe3 O4 pure Fe3 O4 with a spinel structure.
nanoparticles in Fig. 2A is in the range of 10–30 nm. A higher The magnetic lodging of magnetic CNT composite permits
resolution TEM image of the CNT/nano-Fe3 O4 composite as a variety of manipulations, including the modification of elec-
shown in Fig. 2B illustrates the nature of the composite mate- trode surfaces and the fabrication of biosensors. The CNT/Fe3 O4
rial. Although the composites had been sonicated in ethanol association does not impair the electrocatalytic properties of
before TEM measurements, a great amount of Fe3 O4 nanopar- CNT. Such behavior has been demonstrated for the redox pro-
ticles were found on the surface of CNTs, indicating the strong cesses of hydrogen peroxide that are of considerable interest
interaction between CNT and nano-Fe3 O4 . It could be concluded for the operation of oxidase-based amperometric biosensors.
Fig. 2. (A and B) Typical transmission electron micrographs (TEMs) of the MWCNT/nano-Fe3 O4 composite and (C) XRD pattern of CNTs and the CNT/Fe3 O4
composites. (a) MWCNT and (b) Fe3 O4 nanoparticles.
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1100 S. Qu et al. / Talanta 71 (2007) 1096–1102
H2 O2 + 2H2 O − 2e → 2H3 O+ + O2
Fig. 3. Cyclic voltammograms of (a, b and e) 0, (f) 1, (g) 2, (c, d and h) 3, (i) 4,
(j) 5 mM hydrogen peroxide in 0.05 M phosphate buffer solution (pH 7.4) at the
bare (a and c), Fe3 O4 nanopartile modified (b and d) and CNT/Fe3 O4 composite H2 O2 + 2H3 O+ + 2e → 4H2 O
modified (e–j) electrodes in the potential range of 0 to 1.0 (A) and 0.3 to −0.5 V
(B). The amount of the loaded magnetic materials, 20 g; scan rate, 100 mV/s. The amount of CNT/Fe3 O4 plays a crucial role in the electrocat-
alytic reduction of hydrogen peroxide. Fig. 4A shows the effect
of the amount of CNT/nano-Fe3 O4 composite on the amper-
Fig. 4. (A) Effect of the amount of CNT/nano-Fe3 O4 composite on the current response of 5 mM hydrogen peroxiode at potential of −0.1 V (vs. SCE) and (B)
hydrodynamic voltammograms (HDVs) for 1 mM hydrogen peroxide at a bare (a) and a CNT/nano-Fe3 O4 composite (b) electrode. The amount of the loaded
composite, 20 g; electrolyte, 50 mM phosphate buffer (pH 7.4); stirring rate, 300 rpm.
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S. Qu et al. / Talanta 71 (2007) 1096–1102 1101
Fig. 5. Amperometric response of the bare (a) and the CNT/Fe3 O4 composite modified (b) electrodes to the successive addition of 1 mM hydrogen peroxide at the
potential of (A) −0.1 and (B) +0.4 V (vs. SCE). (C) Amperometric response of the bare (a) and the magnetic electrode loaded with Fe3 O4 /CNT/CTS–GOx composite
(b) to the successive addition of 1 mM glucose at the potential of −0.1 V (vs. SCE). Also shown (in the insets) are the plots of amperometric currents versus the
concentrations of hydrogen peroxide (A and B) and glucose (C) at the electrodes modified with magnetic CNT composite. Other conditions, as in Fig. 4.
ometric current response of 5 mM hydrogen peroxide at the concentration in the steps of 1 mM at the bare (a) and the
constant potential of −0.1 V (versus SCE). The current increases CNT/Fe3 O4 coated (b) electrodes at the detection potential of
to a value of 16.8 A, upon raising the nanocomposite amount +0.4 V. The greatly enhanced amperometric response and the
between 10 and 20 g, and decreases gradually to 15.9 A upon short response time (2–3 s) at the CNT-based device reflects
increasing the amount to 30 g. The dropping of the current the accelerated oxidation of hydrogen peroxide, in accordance
implies that the excess composite may disturb the mass trans- with the HDV data illustrated in Fig. 4B. Although the oxida-
formation on the surface of the electrode. Fig. 4B compares the tion potential has been significantly reduced to 0.3–0.5 V by
hydrodynamic voltammograms (HDVs) for 1 mM hydrogen per- the CNT/nano-Fe3 O4 composite, some coexistent substances
oxide at the bare electrode (a) and the CNT/Fe3 O4 composite (such as dopamine, ascorbic acid, uric acid, etc.) in the body
modified electrode (b). It is obvious that the current response fluids at physiological levels may interfere with the determi-
for the oxidation and the reduction of hydrogen peroxide at nation because they are electroactive at the potentials above
the bare electrode was poor. A small response was observed 0.2 V. So the glucose biosensor based on the magnetic loading of
at more positive or more negative potentials for the oxidation the CNT/Fe3 O4 /CTS/GOx composite on the base electrode was
or reduction of hydrogen peroxide, respectively. In contrast, the allowed to run at potential of −0.1 V in reduction mode so that
CNT/Fe3 O4 composite modified electrode exhibited enhanced the interferences of dopamine, ascorbic acid and uric acid were
current response with significant decrease in the overvoltages. minimized. Fig. 5C illustrates the amperometric response of the
Such electrocatalytic activities facilitate the low-potential magnetic electrode loaded with Fe3 O4 /CNT/CTS/GOx compos-
amperometric measurements of hydrogen peroxide. Fig. 5A ite to the successive addition of 1 mM glucose. Well-defined
compares the amperometric response (at −0.1 V) of the bare (a) current responses for glucose were obtained at the magnetic
and the CNT/Fe3 O4 modified (b) electrodes to the successive biosensor. The reaction occurring at the biosensor is very fast to
additions of 1 mM hydrogen peroxide, along with the resulting reach a dynamic equilibrium upon each addition of the sample
calibration plots (insets). As expected from the voltammetric solution, generating a steady-state current signal within 5–6 s.
data, the bare and the Fe3 O4 modified electrodes are not respon- The high sensitivity is accompanied by a low noise level (despite
sive to these concentration changes using this low detection the large voltammetric background current). In order to test the
potential. However, well-defined and fast amperometric signals selectivity of the magnetic biosensor, amperometric responses
are observed for these 1 mM-H2 O2 additions at the CNT/Fe3 O4 were obtained by a batch addition of interfering species (0.2 mM
modified electrode, generating steady-state signals within 2–3 s. ascorbic acid, 0.2 mM uric acid and 0.2 mM dopamine) after the
The CNT-based magnetic electrode offers substantially larger addition of 5 mM glucose. The interferences of dopamine, ascor-
signals, reflecting the electrocatalytic activity of CNT (b). In bic acid and uric acid, have not been found because they are not
addition, the hydrogen peroxide response was not affected by electroactive at the detection potential of −0.1 V. Because chi-
regenerating the surface of the electrode. Such renewability tosan dissolved in the acidic solution can be precipitated in alka-
was indicated from a series of seven successive measurements line solution, CTS was coprecipapted with Fe3+ and Fe2+ in the
each recorded on a freshly loaded surface, yielding satisfactory presence of CNT to form magnetic CNT/Fe3 O4 /CTS composite,
reproducible signals with relative standard derivation (R.S.D.) a versatile biosensing material. The CNTs inside the composite
of 5.3% (not shown). form a conductive network that can accommodate Fe3 O4 , CTS
The CNT/nano-Fe3 O4 coated electrodes exhibit highly elec- and other functional components. The primary amino groups in
trocatalytic activity toward not only the reduction but also the CTS facilitate the further immobilization of enzyme, DNA or
oxidation of hydrogen peroxide. Fig. 5B illustrates the current- proteins, while the magnetic moiety of the composite provides
time recordings obtained by increasing the hydrogen peroxide conveniences for magnetic loading and magnetic isolation.
中国科技论文在线 http://www.paper.edu.cn
1102 S. Qu et al. / Talanta 71 (2007) 1096–1102
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