nanomaterials

Article
One-Step Electrochemical Fabrication of Reduced
Graphene Oxide/Gold Nanoparticles
Nanocomposite-Modified Electrode for Simultaneous
Detection of Dopamine, Ascorbic Acid, and Uric Acid
Chang-Seuk Lee † , Su Hwan Yu † and Tae Hyun Kim *
Department of Chemistry, Soonchunhyang University, Asan 31538, Korea; eriklee0329@sch.ac.kr (C.-S.L.);
shsh422@naver.com (S.H.Y.)
* Correspondence: thkim@sch.ac.kr; Tel.: +82-41-530-4722
† These authors contributed equally to this work.

Received: 23 November 2017; Accepted: 28 December 2017; Published: 30 December 2017

Abstract: Here, we introduce the preparation of the hybrid nanocomposite-modified electrode

consisting of reduced graphene oxide (RGO) and gold nanoparticles (AuNPs) using the one-step
electrochemical method, allowing for the simultaneous and individual detection of dopamine (DA),
ascorbic acid (AA), and uric acid (UA). RGO/AuNPs nanocomposite was formed on a glassy carbon
electrode by the co-reduction of GO and Au3+ using the potentiodynamic method. The RGO/AuNPs
nanocomposite-modified electrode was produced by subjecting a mixed solution of GO and Au3+ to
cyclic sweeping from −1.5 V to 0.8 V (vs. Ag/AgCl) at a scan rate 10 mV/s for 3 cycles. The modified
electrode was characterized by scanning electron microscopy, Raman spectroscopy, contact angle
measurement, electrochemical impedance spectroscopy, and cyclic voltammetry. Voltammetry results
confirm that the RGO/AuNPs nanocomposite-modified electrode has high catalytic activity and
good resolution for the detection of DA, AA, and UA. The RGO/AuNPs nanocomposite-modified
electrode exhibits stable amperometric responses for DA, AA, and UA, respectively, and its detection
limits were estimated to be 0.14, 9.5, and 25 µM. The modified electrode shows high selectivity
towards the determination of DA, AA, or UA in the presence of potentially active bioelements.
In addition, the resulting sensor exhibits many advantages such as fast amperometric response,
excellent operational stability, and appropriate practicality.

Keywords: electrochemically reduced graphene oxide (ERGO); gold nanoparticles (AuNPs); hybrid
nanocomposites; dopamine; ascorbic acid; uric acid

1. Introduction
Dopamine (DA) plays a key role in the central nervous, renal, and hormonal systems of human
bodies [1]. Abnormal levels of DA are diagnostic factors of several diseases such as schizophrenia [2],
Parkinson’s disease [3], Alzheimer’s disease [4], and Huntington’s disease [5]. Therefore, a highly
sensitive determination of DA levels is needed in the early diagnosis of neurological disorders.
Many researchers have proposed different methods for detection of DA, including fluorescence [6,7],
surface enhanced Raman scattering [8], chromatography [9], microdialysis [10,11], and electrochemical
methods [12–15]. Among them, electrochemical methods have a lot of advantages such as simplicity,
low cost, short time of operation, high sensitivity, and availability of in-situ monitoring. However,
electrochemical detection of DA can be disturbed by other biological molecules such as ascorbic
acid (AA), and uric acid(UA) [16]. The high levels of AA can overwhelm the electrochemical signal
of DA. Moreover, the voltammetric response of UA is similar to that of DA, making it difficult to
detect DA selectively. To overcome this limitation, several approaches have been developed using the

Nanomaterials 2018, 8, 17; doi:10.3390/nano8010017 www.mdpi.com/journal/nanomaterials

Nanomaterials 2017, 8, 17 2 of 14

DA, making it difficult to detect DA selectively. To overcome this limitation, several approaches
Nanomaterials 2018, 8, 17
have been developed using the modified electrodes based on catalytic nanomaterials. Combination 2 of 13
of various nanomaterials such as nanoparticles [17], bimetallic nanocomposites [18], ionic liquids
[19], polymers [20], MoS2 [21], and graphene [22–31] have been applied to modify the electrodes for
modified electrodes based on catalytic nanomaterials. Combination of various nanomaterials such
DA determination.
as nanoparticles [17], bimetallic nanocomposites [18], ionic liquids [19], polymers [20], MoS2 [21],
Among those nanomaterials, graphene has been extensively used as a material for modifying
and graphene [22–31] have been applied to modify the electrodes for DA determination.
electrodes, owing to its unique structural and electronic properties [22,23]. Considering the
Among those nanomaterials, graphene has been extensively used as a material for modifying
characteristic of the graphene and its inherent electrocatalytic property, modified electrodes with
electrodes, owing to its unique structural and electronic properties [22,23]. Considering the characteristic
graphene-related nanomaterials such as pristine graphene [24], chemically reduced graphene
of the graphene and its inherent electrocatalytic property, modified electrodes with graphene-related
oxide (CRGO) [25–27], and electrochemically reduced graphene oxide (ERGO) [28–30] have been
nanomaterials such as pristine graphene [24], chemically reduced graphene oxide (CRGO) [25–27],
reported to effectively detect DA, AA, and UA. On the other hand, the modified electrodes of metal
and electrochemically reduced graphene oxide (ERGO) [28–30] have been reported to effectively
nanoparticles (NPs) have received foremost interest in electroanalysis due to good biocompatibility,
detect DA, AA, and UA. On the other hand, the modified electrodes of metal nanoparticles (NPs)
large surface area, and excellent catalytic property [31]. Besides, graphene decorated with metal NPs
have received foremost interest in electroanalysis due to good biocompatibility, large surface area,
has become increasingly important, because the graphene-metal hybrid nanocomposites can show
and excellent catalytic property [31]. Besides, graphene decorated with metal NPs has become
the synergic effect of electrocatalytic behavior of both graphene and metal NPs. In particular, RGO
increasingly important, because the graphene-metal hybrid nanocomposites can show the synergic
can provide a versatile scaffold for NPs to form hybrid nanocomposite with improved properties,
effect of electrocatalytic behavior of both graphene and metal NPs. In particular, RGO can provide
owing to its defects and oxygen functional groups (–OH, C=O, –COOH). In recent times, many types
a versatile scaffold for NPs to form hybrid nanocomposite with improved properties, owing to
of graphene-metal hybrid nanocomposites [20,21] have been suggested for the modification
its defects and oxygen functional groups (–OH, C=O, –COOH). In recent times, many types of
materials of the electrochemical biosensor. The nanocomposite-modified electrodes are usually
graphene-metal hybrid nanocomposites [20,21] have been suggested for the modification materials of
prepared using RGO, which is, however, obtained mostly by chemical reduction with toxic reducing
the electrochemical biosensor. The nanocomposite-modified electrodes are usually prepared using
agents. This can cause problems for human health and the environment. Moreover, electrode
RGO, which is, however, obtained mostly by chemical reduction with toxic reducing agents. This can
modifications with RGO are commonly achieved using the drop-casting method [32,33], which can
cause problems for human health and the environment. Moreover, electrode modifications with
cause variations in film thickness or internal structure, due to differences in evaporation rates across
RGO are commonly achieved using the drop-casting method [32,33], which can cause variations
the substrate or concentration fluctuations. Furthermore, the preparation to composite RGO with
in film thickness or internal structure, due to differences in evaporation rates across the substrate
NPs may require multiple and time-consuming steps of preparation or more sensitive handling.
or concentration fluctuations. Furthermore, the preparation to composite RGO with NPs may
Therefore, the eco-friendly and simple preparation of graphene-based hybrid nanocomposites is
require multiple and time-consuming steps of preparation or more sensitive handling. Therefore,
very necessary.
the eco-friendly and simple preparation of graphene-based hybrid nanocomposites is very necessary.
In this study, we report a simple electrochemical fabrication of RGO/AuNPs nanocomposite
In this study, we report a simple electrochemical fabrication of RGO/AuNPs nanocomposite
modified electrode by one-step electrochemical co-reduction of RGO and Au3+ 3+. It is worth noting
modified electrode by one-step electrochemical co-reduction of RGO and Au . It is worth noting
that AuNPs were utilized here to provide a highly effective surface area and better mass transport of
that AuNPs were utilized here to provide a highly effective surface area and better mass transport of
target analytes to the electrocatalyst [34,35]. The as-prepared electrode was used for the
target analytes to the electrocatalyst [34,35]. The as-prepared electrode was used for the simultaneous
simultaneous and individual detection of DA, AA, and UA. Scheme 1 shows the overall process for
and individual detection of DA, AA, and UA. Scheme 1 shows the overall process for fabricating the
fabricating the RGO/AuNPs nanocomposite modified electrode; as can be seen, the fabrication was
RGO/AuNPs nanocomposite modified electrode; as can be seen, the fabrication was performed in a
performed in a mixed solution of GO and Au3+ using voltammetric cycling, which is a simple, fast,
mixed solution of GO and Au3+ using voltammetric cycling, which is a simple, fast, and eco-friendly
and eco-friendly process. Notably, this is one-step fabrication method via co-electrodeposition
process. Notably, this is one-step fabrication method via co-electrodeposition through simple
through simple voltammetric scanning exists without the need for any other processes, such as
voltammetric scanning exists without the need for any other processes, such as drop-casting and
drop-casting and drying. The resulting electrode exhibited good electrocatalytic behavior, allowing
drying. The resulting electrode exhibited good electrocatalytic behavior, allowing for good sensitivity
for good sensitivity and selectivity with respect to individual and simultaneous determination of
and selectivity with respect to individual and simultaneous determination of DA, AA, and UA without
DA, AA, and UA without requiring any additional treatments.
requiring any additional treatments.

Scheme 1. Fabrication of RGO/AuNPs nanocomposite modified electrode via one-step electrochemical

co-reduction for simultaneous detection of DA, AA, and UA.
Nanomaterials 2017, 8, 17 3 of 14

Scheme 1. Fabrication of RGO/AuNPs nanocomposite modified electrode via one-step

electrochemical co-reduction for simultaneous detection of DA, AA, and UA.
Nanomaterials 2018, 8, 17 3 of 13

2. Results and Discussion

2. Results and Discussion
2.1. Preparation and Structure Characterization of RGO/AuNPs Nanocomposite-Modified Electrode
2.1. Preparation and Structure
Figure 1 shows Characterization
the typical of RGO/AuNPs
cyclic voltammetry Nanocomposite-Modified
(CV) curve recorded during theElectrode
co-reduction of
GO and Au13+shows
Figure on glassy carboncyclic
the typical electrode (GCE) in
voltammetry (CV)10 curve
mM PBS buffer
recorded solution
during (pH 7) containing
the co-reduction of GO
0.3
andmg/mL3+
Au onofglassy
GO and 0.8 mM
carbon HAuCl
electrode 4. There
(GCE) in 10ismM
a large reduction
PBS buffer peak(pH
solution (a) at approximately
7) containing −1.4 V
0.3 mg/mL
due
of GOto and
the reduction
0.8 mM HAuClof the4oxygen
. There functional groups ofpeak
is a large reduction GO, (a)
which increases gradually
at approximately −1.4 Vafter
duecycles,
to the
differently from previous electrochemical reduction of GO [36]. This can be
reduction of the oxygen functional groups of GO, which increases gradually after cycles, differently explained by the
deposition
from previousof AuNPs with higher
electrochemical conductivity
reduction on GO
of GO [36]. This[37].
canThe reductionby
be explained peak
the (b) at approximately
deposition of AuNPs
0.52 V is related to the reduction process of Au 3+, which leads to the formation of AuNPs on
with higher conductivity on GO [37]. The reduction peak (b) at approximately 0.52 V is related to the
electrode
reduction surface.
process of Au3+results
These , whichindicate
leads toRGO/AuNPs
the formationnanocomposite was electrodeposited
of AuNPs on electrode surface. Theseonresults
GCE
after the RGO/AuNPs
indicate voltammetric nanocomposite
cycling. was electrodeposited on GCE after the voltammetric cycling.

100

-100
8
Current / μA

-200 4
Current / μA

-300 0

-4
-400
-8 (b)
-500 0.0 0.2 0.4 0.6 0.8 1.0
(a)
Potential / V vs. Ag/AgCl
-600
-1.5 -1.0 -0.5 0.0 0.5 1.0
Potential / V vs. Ag/AgCl
Figure 1. CV curve obtained on GCE in 10 mM PBS buffer solution
solution (pH
(pH 7)
7) containing
containing 0.3
0.3 mg/mL
mg/mL GO
and 0.8 mM HAuCl44 at
at aa scan
scan rate
rate of
of 10
10 mV/s.
mV/s.

The surfacemorphology
The surface morphology of theofas-prepared
the as-prepared nanocomposite-modified
nanocomposite-modified electrode waselectrode was
characterized
characterized
and comparedand compared
using scanningusing scanning
electron electron
microscopy microscopy
(SEM) with those(SEM)
of RGO with those
and AuNPsof RGO and
modified
AuNPs modified electrodes, prepared in same voltammetric condition
electrodes, prepared in same voltammetric condition with RGO/AuNPs nanocomposite. Figure 2 with RGO/AuNPs
nanocomposite.
shows SEM images Figure 2 shows
of (a) bare GCE,SEM(b) images
RGO of (a) bareGCE
modified GCE, (b) RGO modified
(RGO-GCE), (c) AuNPsGCE (RGO-GCE),
modified GCE
(c)
(AuNPs-GCE), and (d) RGO/AuNPs nanocomposite modified GCE (RGO/AuNPs-GCE). The GCE
AuNPs modified GCE (AuNPs-GCE), and (d) RGO/AuNPs nanocomposite modified SEM
(RGO/AuNPs-GCE).
image of GCE (a) displaysThe SEM image
relatively of GCE
smooth (a) displays
surface, while therelatively
surface ofsmooth
RGO-GCE surface, while the
(b) is gauze-like
surface
shape withof RGO-GCE
wrinkles,(b) is gauze-like
which providesshape
large with wrinkles,area
electroactive which
on provides large electroactive
the electrode. The AuNPs-GCE area
on the electrode. The AuNPs-GCE (c) shows spherical-shaped and homogeneously
(c) shows spherical-shaped and homogeneously distributed deposits with diameters of 38.7 ± 3.5 nm distributed
deposits with diameters
on the surface. The SEMofimage38.7 ± of
3.5RGO/AuNPs-GCE
nm on the surface. (d) Theshows
SEM image
that theof RGO/AuNPs-GCE
AuNPs were densely (d)
shows that the decorated
and uniformly AuNPs were alongdensely and uniformly
the surface of RGO. decorated along
These results the surface
clearly confirmofthe
RGO. These
formation
results clearly confirm the formation
of RGO/AuNPs nanocomposite on GCE. of RGO/AuNPs nanocomposite on GCE.
The structural property of RGO/AuNPs nanocomposite was investigated by Raman spectroscopy.
Figure 3 shows the Raman spectra of GO, RGO, and RGO/AuNPs nanocomposite, respectively.
The D band (~1354 cm−1 ) corresponds to the disorder in the sp2 carbon network, and the G band
(~1607 cm−1 ) is associated with the tangential vibrations of the sp2 carbon atoms in the hexagonal
planes [38]. The intensity ratio of the D and G bands (ID /IG ) was employed to calculate the structural
disorder; this ratio increased from 0.65 to 1.18 during the reduction of GO to RGO, suggesting a
decrease in the average size of sp2 domains owing to the removal of the oxygen functional groups.
Nanomaterials 2018, 8, 17 4 of 13

ID /IG (1.69) of the RGO/AuNPs is found to be higher, compared to that in GO, confirming the
deoxygenation
Nanomaterials 2017, during
8, 17 the co-reduction of GO and Au3+ . 4 of 14

Nanomaterials
Figure 2. 2017,
SEM 8, 17
images of (a) bare GCE, (b) RGO-GCE, (c) AuNPs-GCE, and (d) RGO/AuNPs-GCE. 5 of 14
Figure 2. SEM images of (a) bare GCE, (b) RGO-GCE, (c) AuNPs-GCE, and (d) RGO/AuNPs-GCE.

The structural property of RGO/AuNPs nanocomposite was investigated by Raman

spectroscopy. Figure 3 shows the Raman spectra of GO, RGO, and RGO/AuNPs nanocomposite,
respectively. The D band (~1354 cm−1) corresponds to the disorder in the sp2 carbon network, and the
G band (~1607 cm−1) is associated with the tangential vibrations of the sp2 carbon atoms in the
hexagonal planes [38]. The intensity ratio of the D and G bands (ID/IG) was employed to calculate the
structural disorder; this ratio increased from 0.65 to 1.18 during the reduction of GO to RGO,
suggesting a decrease in the average size of sp2 domains owing to the removal of the oxygen
functional groups. ID/IG (1.69) of the RGO/AuNPs is found to be higher, compared to that in GO,
confirming the deoxygenation during the co-reduction of GO and Au3+.
To analyze the surface property of RGO/AuNPs nanocomposite, the changes in the wettability
were determined through contact angle measurements. Figure 4 shows the wetting characteristics of
bare GCE, RGO-GCE, AuNPs-GCE, and RGO/AuNPs-GCE, and their average equilibrium static
contact angles are 76°, 80°, 69°, and 60°, respectively. GO is hydrophilic due to its oxygen containing
functionalities, and bare GCE is slightly hydrophobic. However, after electrochemical reductive
deposition of GO onto GCE, the contact angle of RGO-GCE shows more hydrophobic character than
the bare GCE, which is attributed to the de-oxygenation or de-hydroxylation of GO. On the other
Figure 3. Raman spectra of GO, RGO, and RGO/AuNPs nanocomposites.
hand, AuNPs-GCEFigureand RGO/AuNPs-GCE
3. Raman spectra of GO,show hydrophilic
RGO, characters.
and RGO/AuNPs This should be due to the
nanocomposites.
gold surface oxidation during consecutive electro-oxidation and reduction cycling for the deposition
To analyze the surface property of RGO/AuNPs nanocomposite, the changes in the wettability
of AuNPs [39]. The more hydrophilic property of RGO/AuNPs-GCE is attributed to the more
were determined through contact angle measurements. Figure 4 shows the wetting characteristics
immobilization of AuNPs on the wrinkled surface of RGO with large surface area.
of bare GCE, RGO-GCE, AuNPs-GCE, and RGO/AuNPs-GCE, and their average equilibrium static
contact angles are 76◦ , 80◦ , 69◦ , and 60◦ , respectively. GO is hydrophilic due to its oxygen containing
functionalities, and bare GCE is slightly hydrophobic. However, after electrochemical reductive
Nanomaterials 2018, 8, 17 5 of 13

deposition of GO onto GCE, the contact angle of RGO-GCE shows more hydrophobic character
than the bare GCE, which is attributed to the de-oxygenation or de-hydroxylation of GO. On the
other hand, AuNPs-GCE and RGO/AuNPs-GCE show hydrophilic characters. This should be due
to the gold surface oxidation during consecutive electro-oxidation and reduction cycling for the
deposition of AuNPs [39]. The more hydrophilic property of RGO/AuNPs-GCE is attributed to the
Figure 3. Raman spectra of GO, RGO, and RGO/AuNPs nanocomposites.
more immobilization of AuNPs on the wrinkled surface of RGO with large surface area.

Figure 4. Contact angle analysis of (a) bare GCE, (b) RGO-GCE, (c) AuNPs-GCE, and (d) RGO/AuNPs-GCE.
Figure 4. Contact angle analysis of (a) bare GCE, (b) RGO-GCE, (c) AuNPs-GCE, and
(d) RGO/AuNPs-GCE.
2.2. Electrochemical Properties of RGO/AuNPs Nanocomposite-Modified Electrode
2.2. Electrochemical Properties of
CV and electrochemical RGO/AuNPs
impedance Nanocomposite-Modified
spectroscopy (EIS) measurements Electrode
were carried out to analyze
the electrochemical property of RGO/AuNPs-GCE using K4 [Fe(CN)6 ] as the electrochemical probe.
CV and electrochemical impedance spectroscopy (EIS) measurements were carried out to
Figure 5a
analyze thedisplays CV curves
electrochemical for bare
property GCE, AuNPs-GCE,using
of RGO/AuNPs-GCE RGO-GCE, and6] RGO/AuNPs-GCE
K4[Fe(CN) as the electrochemical in
0.1 M KNO solution containing 2 mM [Fe(CN) ] 3− at a scan rate of 50 mV/s, exhibiting a characteristic
probe. Figure 3 5a displays CV curves for bare GCE, 6 AuNPs-GCE, RGO-GCE, and RGO/AuNPs-GCE
voltammetric response for [Fe(CN) ] 3−/4− . However, the electron transfer reaction of [Fe(CN) ]3−
6
in 0.1 M KNO3 solution containing 2 mM [Fe(CN)6]3− at a scan rate of 50 mV/s, exhibiting 6 a
after the immobilization of AuNPs, RGO, or both on GCE was more
characteristic voltammetric response for [Fe(CN)6]3−/4−. However, the electron transfer reaction of facilitated with smaller peak
separations
[Fe(CN) of 70 mV, 87 mV, and 71 mV, respectively, compared to that in the bare GCE with 135 mV.
6]3− after the immobilization of AuNPs, RGO, or both on GCE was more facilitated with
In addition,
smaller peakAuNPs-GCE
separations of and 70RGO/AuNPs-GCE
mV, 87 mV, and 71revealed significantly
mV, respectively, higher redox
compared peak
to that in currents,
the bare
GCE with 135 mV. In addition, AuNPs-GCE and RGO/AuNPs-GCE revealed significantly activity
indicating the better electrochemical performance. This should come from the electrocatalytic higher
along with the large effective surface area of the modified electrodes.
redox peak currents, indicating the better electrochemical performance. This should come from the The effective surface areas of
AuNPs-GCE, RGO-GCE, and RGO/AuNPs-GCE was 0.123 cm 2 , 0.083 cm2 , and 0.121 cm2 , respectively,
electrocatalytic activity along with the large effective surface area of the modified electrodes. The
as calculated
effective surfaceusing theof
areas RandlesSevcik
AuNPs-GCE,equation RGO-GCE, [40]and
from the CV curves obtained
RGO/AuNPs-GCE was 0.123 using
cm0.1 M KNO
2, 0.083 cm23,
containing 2 mM [Fe(CN) ] 4 − at different scan rates (Figure 5b). The capability of electron transfer
6
and 0.121 cm , respectively, as calculated using the Randles-Sevcik equation [40] from the CV
2
of the RGO/AuNPs-GCE
curves obtained using 0.1 M was KNO further examined by EIS, as shown in Figure 5c. With respect to
3 containing 2 mM [Fe(CN)6]4− at different scan rates (Figure 5b).
barecapability
The GCE, a decrease in the
of electron charge
transfer transfer
of the resistance (Rctwas
RGO/AuNPs-GCE ) by further
a factorexamined
of ~2 times was as
by EIS, observed
shown
in Figure 5c. With respect to bare GCE, a decrease in the charge transfer resistance (Rct) byelectrode
for the RGO/AuNPs-GCE, suggesting fast electron transfer occurred on the modified a factor
surface.
of ~2 times Thewaselectrocatalytic
observed for oxidations of DA, AA, and suggesting
the RGO/AuNPs-GCE, UA on the bare fast GCE, AuNPs-GCE,
electron RGO-GCE,
transfer occurred on
and RGO/AuNPs-GCE were primarily assessed by CV in 0.1 M PBS buffer
the modified electrode surface. The electrocatalytic oxidations of DA, AA, and UA on the bare GCE, solution (pH 7.4) containing
100 µM UA, 1RGO-GCE,
AuNPs-GCE, mM AA, and and 100 µM DA at a scan were
RGO/AuNPs-GCE rate ofprimarily
50 mV/s.assessed
As shown by in
CVFigure
in 0.15d, in contrast
M PBS buffer
with the bare GCE, AuNPs-GCE, and RGO-GCE, the RGO/AuNPs-GCE
solution (pH 7.4) containing 100 μM UA, 1 mM AA, and 100 μM DA at a scan rate of 50 mV/s. As exhibited three well-resolved
oxidation peaks corresponding to DA, AA, and UA. The ability of the RGO/AuNPs-GCE to promote
the voltammetric resolution of DA, AA, and UA could be ascribed to the synergistic effect between RGO
and AuNPs. The existence of oxide functional groups on the RGO would enable the nanocomposite to
selectively interact with DA, AA, and UA via hydrogen bonds with the proton-donating group such
as –NH and –OH [41,42]. In addition, π–π stacking interactions could induce facile electron transfer
between RGO and target analytes [42]. Meanwhile, well-distributed AuNPs on the surface of RGO
(Figure 2d) enhanced the catalytic activity of the nanocomposite by integrating AuNPs with RGO,
which lead to the increases of electrode’s surface area and active sites for the oxidations of DA, AA,
and UA [28]. These results evidenced that the RGO/AuNPs-GCE exhibits excellent electrocatalytic
 functional groups on the RGO would enable the nanocomposite to selectively interact with DA, AA,
and UA via hydrogen bonds with the proton-donating group such as –NH and –OH [41,42]. In
addition, π–π stacking interactions could induce facile electron transfer between RGO and target
analytes [42]. Meanwhile, well-distributed AuNPs on the surface of RGO (Figure 2d) enhanced the
catalytic activity of the nanocomposite by integrating AuNPs with RGO, which lead to the increases
Nanomaterials 2018, 8, 17 6 of 13
of electrode’s surface area and active sites for the oxidations of DA, AA, and UA [28]. These results
evidenced that the RGO/AuNPs-GCE exhibits excellent electrocatalytic activity towards the
oxidation
activity of DA,
towards the AA, and UA.
oxidation Thus,
of DA, AA,RGO/AuNPs-GCE could be used to could
and UA. Thus, RGO/AuNPs-GCE efficiently discriminate
be used to efficiently
between DA,
discriminate AA, and
between UA,AA,
DA, based
andonUA,
its voltammetric responses.
based on its voltammetric responses.

Figure5.5. (a)
Figure (a) CV
CV curves
curvesofofbare GCE,
bare GCE,AuNPs-GCE,
AuNPs-GCE, RGO-GCE, and RGO/AuNPs-GCE
RGO-GCE, and RGO/AuNPs-GCEin 0.1 M in
KNO0.13 M
containing 2 mM [Fe(CN) 6]3− with350 mV/s; (b) relationship between peak current and square
KNO3 containing 2 mM [Fe(CN)6 ] − with 50 mV/s; (b) relationship between peak current and square root of
scan rate for oxidation of bare GCE, AuNPs-GCE, RGO-GCE, and RGO/AuNPs-GCE in 0.1 M KNO3
root of scan rate for oxidation of bare GCE, AuNPs-GCE, RGO-GCE, and RGO/AuNPs-GCE in
containing 2 mM [Fe(CN)6]3−; (c) Nyquist plots of bare GCE, RGO/AuNPs-GCE in 0.1 M KCl
0.1 M KNO3 containing 2 mM [Fe(CN)6 ]3− ; (c) Nyquist plots of bare GCE, RGO/AuNPs-GCE in
containing 2 mM [Fe(CN)6]3−; (d) CV curves of bare GCE, AuNPs-GCE, RGO-GCE, and
0.1 M KCl containing 2 mM [Fe(CN)6 ]3− ; (d) CV curves of bare GCE, AuNPs-GCE, RGO-GCE,
RGO/AuNPs-GCE in 0.1 M PBS buffer solution (pH 7.4) containing 100 μM UA, 1 mM AA, and 100
and RGO/AuNPs-GCE in 0.1 M PBS buffer solution (pH 7.4) containing 100 µM UA, 1 mM AA,
μM DA with 50 mV/s scan rate. Ag/AgCl electrode and Pt wire were introduced as reference and
and 100 µM DA with 50 mV/s scan rate. Ag/AgCl electrode and Pt wire were introduced as reference
counter electrode, respectively.
and counter electrode, respectively.
Since the electroactivity of DA is pH dependent [43], the effect of pH on the oxidation process
of Since
DA was the investigated
electroactivitybyofrecording
DA is pHCV dependent
curves in[43], the effect
various of pH on (from
pH solutions the oxidation process
4.0 to 9.0) that of
DA was investigated
contained 10 mM ofby DA.recording
As shownCVincurves
Figurein6,various
anodic pH peaksolutions (from 4.0 with
current increased to 9.0)
anthat contained
increase in
pH up to 7.4, and then decreased at higher pH. Therefore, pH 7.4 was chosen
10 mM of DA. As shown in Figure 6, anodic peak current increased with an increase in pH up to for the rest of
experiments
7.4, to detect DA.
and then decreased In addition,
at higher the anodicpH
pH. Therefore, peak
7.4 potential
was chosen(Epafor
) shifted to negative
the rest values to
of experiments
with DA.
detect increasing pH. The
In addition, relationship
the anodic peak between the pH
potential (Epaand oxidation
) shifted potentialvalues
to negative was linear, and the
with increasing
related
pH. regression equation
The relationship betweenwas thefound
pH andto be Epa (V) =potential
oxidation 0.463−0.047
waspH (R2 =and
linear, 0.95)the
forrelated
DA. The slope
regression
was 47 mV·pH −1, which is close to the theoretical value 2of 59 mV·pH−1 [44]. This result suggests that −1
equation was found to be Epa (V) = 0.463−0.047 pH (R = 0.95) for DA. The slope was 47 mV·pH ,
the oxidation
which is close toofthe
DAtheoretical
at RGO/AuNPs-GCE
value of 59involves
mV·pH−an equalThis
1 [44]. number
resultofsuggests
protons andthat electrons [45], of
the oxidation
i.e., two protons and two electrons were involved in the oxidation process [46].
DA at RGO/AuNPs-GCE involves an equal number of protons and electrons [45], i.e., two protons
Nanomaterials 2017, 8, 17 7 of 14
and two electrons were involved in the oxidation process [46].

0.30 1.2
0.25 1.0

0.20 0.8
Ipa / μA
Epa / V

0.15 0.6

0.10 0.4
0.2
0.05
0.0
0.00
-0.2
4 5 6 7 8 9
pH

Figure 6. Effect
Figure of pH
6. Effect on on
of pH thethe
peak current
peak currentand
and oxidation potentialofofDA
oxidation potential DAat at RGO/AuNPs-GCE.
RGO/AuNPs-GCE.

2.3. Selective Determination of DA, AA, and UA

To further gain insight into detection and discrimination ability of RGO/AuNPs-GCE, differential
pulse voltammetry (DPV) was performed in a 0.1 M PBS (pH 7.4) solution containing different
concentration of DA, AA, and UA. As seen in Figure 7, the DPV curved towards DA, AA, or UA, when
 4 5 6 7 8 9
pH

Figure 6. Effect of pH on the peak current and oxidation potential of DA at RGO/AuNPs-GCE.

Nanomaterials 2018, 8, 17 7 of 13
2.3. Selective Determination of DA, AA, and UA
To
2.3.further
Selectivegain insight into
Determination detection
of DA, AA, andandUAdiscrimination ability of RGO/AuNPs-GCE, differential
pulse voltammetry (DPV) was performed in a 0.1 M PBS (pH 7.4) solution containing different
To further gain insight into detection and discrimination ability of RGO/AuNPs-GCE, differential
concentration of DA, AA,
pulse voltammetry and UA.
(DPV) was As seen in Figure
performed in a 0.17, M
thePBS
DPV (pHcurved towardscontaining
7.4) solution DA, AA, or UA, when
different
the increasing concentrations
concentration of DA, AA, of andtheUA.
target molecule
As seen were7,added
in Figure the DPV to the mixed
curved solution
towards DA,ofAA,
theorother
UA,two
analytes
when the increasing concentrations of the target molecule were added to the mixed solution of are
at constant concentrations. It can be seen that the oxidation waves of the three compounds
well separated
the other twoandanalytes
their current intensities
at constant rise with the
concentrations. increase
It can be seenofthat
species’ concentration.
the oxidation waves As shown
of the
in Figure
three7a,d, the oxidation
compounds are wellpeak currents
separated andoftheir
DAcurrent
linearlyintensities
increasesrise
with increasing
with concentrations
the increase of species’ of
concentration. As shown in Figure 7a,d, the oxidation peak currents of DA
DA in the range from 0.1 to 100 μM with limit of detection (LOD) of 0.69 μM (S/N = 3). Notably, linearly increases with the
increasing concentrations of DA in the range from 0.1 to 100 µM with limit
changes of DA concentration have negligible influence on the oxidation behaviors of AA and UA. of detection (LOD) of In a
0.69
similar (S/Noxidation
µM the
way, = 3). Notably,
peakthe changesofofAA
currents DA (Figure
concentration
7b,e) orhave
UA negligible
(Figure influence on the oxidation
7c,f) increased linearly with
behaviors of AA and UA. In a similar way, the oxidation peak currents of AA (Figure 7b,e) or UA
increasing concentration in the range from 0.01 to 1 mM and 0.1 to 100 μM with LODs of 5.7 μM and
(Figure 7c,f) increased linearly with increasing concentration in the range from 0.01 to 1 mM and 0.1 to
2.2 μM, respectively. These results confirm that the electrode exhibits simultaneous detection of DA,
100 µM with LODs of 5.7 µM and 2.2 µM, respectively. These results confirm that the electrode exhibits
AA, and UA, as the oxidation waves of these analytes are well resolved.
simultaneous detection of DA, AA, and UA, as the oxidation waves of these analytes are well resolved.

Figure 7. DPV
Figure curves
7. DPV of RGO/AuNPs-GCE
curves of RGO/AuNPs-GCEinin0.1 0.1MMPBS
PBS (pH 7.4)mixed
(pH 7.4) mixedsolution
solution containing
containing (a) 1(a)
mM 1 mM
AA, μM
AA, 100 100 µM
UA,UA,
andand
DADA in in differentconcentrations
different concentrations (0,
(0, 1,
1, 3,
3, 5,
5, 10,
10,30,
30,50,
50,100
100µM);
μM);(b)(b)
100100
µMμM DA,DA,
100 µM UA, and AA in different concentrations (0, 100, 300, 500, 1000 µM); and (c)
100 μM UA, and AA in different concentrations (0, 100, 300, 500, 1000 μM); and (c) 1 mM AA, 1001 mM AA, 100 µM μM
DA, and UA in different concentrations (0, 10, 30, 50, 100 µM). Calibration curves of (d) DA, (e) AA,
DA, and UA in different concentrations (0, 10, 30, 50, 100 μM). Calibration curves of (d) DA, (e) AA,
and (f) UA.
and (f) UA.

2.4. Amperometric Responses of DA, AA, and UA

To evaluate the analytical feasibility of RGO/AuNPs-GCE, chronoamperometric measurements were
performed in 0.1 M PBS (pH 7.4) at fixed potentials under constant stirring. A stock solution of the target
analyte was added into the stirred 10 mL of 0.1 M PBS (pH 7.4) at 50 s intervals, and the final concentration
and composition of the solution were adjusted to the desired value. The amperometric response of the
RGO/AuNPs-GCE was monitored before and after the addition of the target analyte at selected working
potential (0.2, −0.02, and 0.265 V for DA, AA, and UA, respectively). The amperometric current-time
plots of DA, AA, and UA at the RGO/AuNPs-GCE are displayed in Figure 8a–c. The current signal
rose rapidly with each addition of the target analyte, then reaching a steady-state current within 2 s,
indicating a fast oxidation response behavior. The RGO/AuNPs-GCE show linear responses towards
the oxidation of DA, AA, and UA. Their corresponding calibration curves are shown in Figure 8d–f;
the relationship between the concentration of analyte and its amperometric currents are linear. Sensing
performance analyses from amperometric and DPV methods are summarized in Table 1.
are displayed in Figure 8a–c. The current signal rose rapidly with each addition of the target analyte,
then reaching a steady-state current within 2 s, indicating a fast oxidation response behavior. The
RGO/AuNPs-GCE show linear responses towards the oxidation of DA, AA, and UA. Their
corresponding calibration curves are shown in Figure 8d–f; the relationship between the
concentration
Nanomaterials 2018,of analyte and its amperometric currents are linear. Sensing performance analyses
8, 17 8 of 13
from amperometric and DPV methods are summarized in Table 1.

Figure
Figure 8.8. Amperometric
Amperometric responses
responses of of RGO/AuNPs-GCE
RGO/AuNPs-GCE to to successive
successiveaddition
additionof of (a)
(a) DA,
DA, (b)
(b) AA,
AA,
and
and (c) UA in 0.1 M PBS (pH 7.4) under stirring. Working potentials were 0.2, −0.02, 0.265 V (vs.
(c) UA in 0.1 M PBS (pH 7.4) under stirring. Working potentials were 0.2, −0.02, 0.265 V (vs.
Ag/AgCl reference
Ag/AgCl reference electrode)
electrode) for
for DA,
DA, AA,
AA, and UA, respectively.
and UA, respectively. Calibration
Calibration plot
plot ofof steady-state
steady-state
currents obtained at
currents obtained at the
the RGO/AuNPs-GCE
RGO/AuNPs-GCE againstagainst concentrations
concentrations of
of (d)
(d) DA,
DA, (e)
(e) AA,
AA, and
and (f)
(f) UA.
UA.

Table 1.
Table 1. Comparison
Comparisonofoflimits
limitsofofdetection
detection(LOD) and
(LOD) andsensitivity values
sensitivity withwith
values respect to DA,
respect AA, AA,
to DA, and
UA as determined by amperometric and DPV analysis.
and UA as determined by amperometric and DPV analysis.
Targets DA AA UA
Targets
ip (μA) = 7.6 ×DA
10−8 + 0.43C (μM) AA UA
ip (µA) = 7.6 ×
R210=− 8 + 0.43C (µM)
0.954
R2 = 0.954 ip (μA) = 4.8 × 10−8 + ip (μA) = 2.1 × 10−8 +
Linear regression (0.14 –100 μM), ip (µA) = 4.8 × 10−8 + ip (µA) = 2.1 × 10−8 +
Linear regression (0.14–100 µM), 0.0081 C (μM) 0.0035 C (μM)
equation 0.0081 C (µM) 0.0035 C (µM)
ip (µA) = 1.4 × 10−8 ++ 0.023C
i p (μA) = 1.4 × 10 0.023C(µM)
(μM)
−8
equation
RR2 == 0.988
0.988 R2R= =0.979
0.979
2 2
Amperometry
Amperometry R
2 2 = 0.968
R = 0.968
(100–700µM)
(100–700 μM)
LOD
LOD (μM)
(µM) 0.14
0.14 9.5
9.5 2525
Sensitivity
Sensitivity 0.43
0.43 8.1 10−−33
8.1××10 3.53.5 −3−3
× 10
× 10
(µA·µM−
(μA·μM −11))

ipip(µA)
(μA)==1.2 1.2××1010−7 ++ ip i(µA)
p (μA) = 6.0
×× 1010
−7 ++
−7 −7
Linear −−7
7+ = 6.0
Linearregression
regression ipip(µA)
(μA)= =1.8
1.8××1010 + 0.27
0.27CC(µM)
(μM)
2 0.0031
0.0031 C C (µM)
(μM) 0.051CC(µM)
0.051 (μM)
equation
equation RR ==0.935
2 0.935
RR22 == 0.989
0.989 R2R=
2 =0.977
0.977
DPV
DPV
LOD
LOD (μM)
(µM) 0.69
0.69 5.7
5.7 2.2
2.2
Sensitivity
Sensitivity 0.27
0.27 3.1××10
3.1 10 −−3
3 0.051
0.051
(µA·µM−
(μA·μM −11)
)

It is worth comparing the proposed modified electrode with other similar electrodes for DA
determination [24–30]. Table 2 summarizes some characteristics of the proposed electrode and other
modified electrodes, including LOD, detectable range, advantages, and disadvantages. Although
some of the modified electrodes have some advantages, the proposed electrode has made some
improvements in terms of analytical performance such as having a good linear range and low LOD.
Most notably, our fabrication method is based on a one-pot simultaneous reduction of GO and gold
precursor without any other processes such as drop-casting and drying, and is more facile and less
time consuming than other methods.
Nanomaterials 2018, 8, 17 9 of 13

Table 2. Comparison of various graphene-based, composite-modified electrodes towards the detection

of dopamine.

Electrode Material LOD (µM) Sensitivity (µA/µM) Range (µM) Remarks Reference
Multi-step, ERGO,
RGO/Au nanoplate 1.4 1.0 6.8–41 [28]
Drop-casting
Multi-step, CRGO,
RGO-AuNPs-CSHMs 0.3 0.048 1–200 [25]
Drop-casting
Multi-step, CRGO,
AgNPs/rGO 5.4 0.39 10–800 [26]
Drop-casting
Two-step, ERGO,
ERGO 0.5 0.482 0.5–60 [29]
Drop-casting
Two-step, CRGO,
Pt/RGO 0.45 0.0391 10–170 [27]
Drop-casting
Multi-step,
MgO/Gr/Ta 0.15 1.191 0.1–7 [24]
CVD-Graphene
ERGO/CFE 0.77 0.1024 1.5–224.82 One-step, ERGO [30]
RGO/AuNPs 0.137 0.43 0.14–700 One-step, ERGO This work

2.5. Interference Study

In the development of electrochemical biosensor, the study of selectivity by interfering species is
very important factor. Various foreign species were tested to examine whether they would interfere
with the detection of DA, AA, and UA. The RGO/AuNPs-GCE was dipped in a stirring solution and
its amperometric response was monitored in the presence of 10 µM DA, 100 µM AA, and 10 µM UA
with different interferents such as citric acid, glucose, sodium nitrate, and sodium carbonate (Figure 9).
The applied potentials were −0.02, 0.2, and 0.265 V for DA, AA, and UA, respectively. Noticeably,
no significant change of the response current was found in the presence of interferent. This clearly
Nanomaterials 2017, 8, 17 10 of 14
demonstrates that RGO/AuNPs-GCE performs with an excellent selectivity and little interference.

Figure 9. Amperometric responses of RGO/AuNPs-GCE

RGO/AuNPs-GCE upon uponsuccessive
successiveaddition
additionofof (a)
(a) 10
10 μM
µM DA,
DA,
100 µM
(b) 100 μMAA,
AA,(c)(c)1010
µMμM UA,
UA, andand other
other chemicals
chemicals of 10ofµM
10toμM
0.1 to
M 0.1
PBSM PBS (pH
buffer buffer (pH
7.4). The7.4). The
applied
applied potentials
potentials for AA,
for AA, DA, andDA, are −
UA and UA are0.2,
0.02, −0.02,
and0.2, and
0.265 V, 0.265 V, respectively.
respectively.

3. Materials and Methods

3.1. Materials
Graphite powder, dopamine (DA) hydrochloride, L-(+)-ascorbic acid (AA), uric acid (UA), citric
Nanomaterials 2018, 8, 17 10 of 13

3. Materials and Methods

3.1. Materials
Graphite powder, dopamine (DA) hydrochloride, L-(+)-ascorbic acid (AA), uric acid (UA), citric
acid (CA), glucose, potassium hexacyanoferrate (K3 [Fe(CN)6 ]), phosphate buffer saline (PBS, pH 7.4,
0.1 M), and hydrogen tetrachloroaurate (III) (HAuCl4 ) were purchased from Sigma-Aldrich Co.
(St. Louis, MO, USA). All the other chemicals used such as sodium hydroxide (NaOH), sodium
nitrate (NaNO3 ), sodium carbonate (Na2 CO3 ), potassium permanganate (KMnO4 ), hydrogen peroxide
(H2 O2 ), and sulfuric acid (H2 SO4 ) were obtained from Duksan Pure Chemicals Co. (Ansan, Korea).
All the reagents were analytical grade and were used without further purification. Deionized (DI)
water was used throughout the work.

3.2. Instruments
The electrochemical measurements were performed with a CHI 660D electrochemical workstation
(CH Instruments, Inc., Austin, TX, USA). A conventional three-electrode cell was used at room
temperature, including glassy carbon electrode (GCE) as working electrode, Pt wire as counter
electrode, and Ag/AgCl electrode as reference electrode, respectively. Electrochemical impedance
spectroscopy (EIS) was performed in 2.0 mM K3 Fe(CN)6 with 0.1 M KCl as supporting electrolyte.
The morphology of the modified electrode was examined using scanning electron microscopy (SEM)
system (JEOL, JSM 5600 LV, Tokyo, Japan). Contact angle images were taken on a PHOENIX-MINI
(SEO Co. Ltd., Suwon, Korea) system. Raman spectroscopy was performed using a EnSpectr R532
Raman spectrometer (Enhanced Spectrometry, Inc., Torrance, CA, USA).

3.3. Preparation of RGO/AuNPs Nanocomposite Electrode

Graphene oxide (GO) was synthesized using graphite powders by the modified Hummers
method [47,48]. The synthesized GO was dispersed as 1.0 mg/mL in DI water, and the solution was
ultrasonicated for 1 h. Before the co-reduction of GO and Au3+ ion, GCE was polished with 1.0, 0.3,
and 0.05 µm alumina powder and then sonicated for 5 min in ethanol and DI water successively.
After that, the GCE was electrochemically polished in 0.25 mM H2 SO4 solution with CV; potential was
scanned from −1.0 V to 1.0 V with 50 mV/s scan rate for 20 cycles. The GCE was immersed into a
10 mM of PBS buffer solution (pH 7.4) containing 0.3 mg/mL of GO and HAuCl4 . The co-reduction of
GO and Au3+ ion was performed with CV from −1.5 V to 0.8 V (vs. Ag/AgCl) at a scan rate 10 mV/s
for 3 cycles.

3.4. Electrochemical Measurements

A 0.1 M PBS (pH 7.4) was used as the supporting electrolyte for electrochemical determination of
DA, AA and UA, respectively. Before the measurement, the solution was deoxygenated with pure N2
gas for 10 min.

4. Conclusions
We proposed one-step electrochemical preparation of RGO and AuNPs nanohybrid on a GCE by
CV in 10 mM PBS buffer (pH 7) containing 0.3 mg/mL of GO and 0.8 mM HAuCl4 . The analytical feasibility
of the RGO/AuNPs-GCE was investigated by using CV, DPV, and amperometric measurements.
The RGO/AuNPs-GCE exhibited good catalytic activity toward AA, DA, and UA oxidation, displaying
the well-resolved potential peak separation and enhanced peak currents for the oxidation of the
three analytes. The proposed electrode also showed excellent electrochemical sensing performance
such as low LOD, wide linear range, fast response time, as well as good selectivity and sensitivity,
which indicates the RGO/AuNPs can be a potential candidate for detection of AA, DA, and UA.
Nanomaterials 2018, 8, 17 11 of 13

The fabrication of the proposed electrode is simple and efficient that may contribute to develop a high
performance electrochemical sensor.

Acknowledgments: This work was supported by the Basic Science Research Program through the National
Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2016R1D1A1B03931362).
This work is also supported by the Soonchunhyang University Research Fund.
Author Contributions: T.H.K. and C.-S.L. conceived and designed the experiments; S.H.Y. and C.-S.L. performed
the experiments; T.H.K., S.H.Y. and C.-S.L. analyzed the data; T.H.K. and C.-S.L. wrote the paper.
Conflicts of Interest: The authors declare no conflict of interest.

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