Introduction

Glucose detection is highly important in the clinical diagnosis of the diabetes mellitus [1, 2]. To develop effective methods for rapid, sensitive and reliable detection of glucose in blood is thus of key significance for diabetes patients to receive timely treatments. Among the many approaches [1,2,3,4,5,6] established for that purpose, the electrochemical method [1, 2, 7] has received particular attention due to its characteristics of high sensitivity, fast response, outstanding accuracy, low cost and easy operation. It has been reported [7] that there are two types of electrochemical sensors for glucose detection, i.e., the enzymatic and the non-enzymatic glucose sensors. However, the enzymatic glucose sensor suffers from inevitable disadvantages such as requirement for expensive enzyme, tedious enzyme immobilization process and unsatisfactory chemical and thermal stabilities [8], whereas the non-enzymatic glucose sensor can overcome those drawbacks to some extent through direct electrocatalytic oxidation of glucose.

During the past few decades, a lot of metals or alloys and metal oxides at nanometer scale have been explored for the non-enzymatic glucose detection, and the unique features and architecture of the nanomaterials were carefully designed and utilized [7]. In comparison with the noble metals and alloys, copper oxide (CuO)- and cuprous oxide (Cu2O)-based nanomaterials have superiorities of inexpensiveness together with high activity of non-enzymatic direct electrooxidation of glucose [7, 9, 10]. The Cu-based oxides were fabricated into different kinds of nanoscaled architectures in order to make utilization of their exceptional optical and electrical properties [9, 10]. During glucose detection, Cu2O generates Cu(III)/Cu(II) redox couple, which can then transform glucose into glucolactone through catalytic reaction to realize glucose detection [7]. Lately, the well-known two-dimensional material of graphene oxide (GO) has been widely employed to prepare Cu-based composite sensors [11]. The large surface area and excellent conductivity of GO [12] will greatly improve the electrochemical performances of the prepared composite or hybrid materials [6, 7, 11, 13,14,15,16,17]. In order to take advantage of the characteristics of GO, metal oxides are anticipated to grow on the matrix of GO to expose more active sites of the obtained sensor to improve its electrochemical performances [11]. However, GO can hardly form framework of a porous material [12] to provide firm matrix for the growth of the metal oxides.

It has been reported [18] that there is strong π–π stacking interaction between the aromatic groups of polystyrene (PS) and GO sheets. In this work, a water-in-oil emulsion was prepared by dispersing GO aqueous solution in PS/CH solution, and self-assembly of the PS chains and the GO sheets was expected to take place at the interface of the emulsion. Meanwhile, the PS chains would be precipitated at the interface during the cooling step of the preparation process, which facilitated formation of a firm framework. After complete solvent evaporation of the emulsion, a porous composite of PS/GO was thus prepared, where water droplets in the emulsion served as sacrificial templates. Copper compounds were then loaded on the PS/GO composite. After carbonization to remove the polymer matrix, a free-standing and porous Cu2O/rGO nanocomposite film was thus obtained. The preparation and the microstructures of the obtained materials were carefully analyzed and discussed. The electrochemical performances of the Cu2O/rGO composite were characterized, and its amperometric responses to glucose were evaluated. Moreover, the stability of the porous Cu2O/rGO composite and its selective amperometric responses to glucose were measured to indicate its potential application in non-enzymatic glucose detection.

Experimental section

Materials

Polystyrene (PS, weight-average molecular weight M w = 240 kg/mol, polydispersity index d = 1.80) was obtained from the former Hüls AG. Analytical grade reagents of copper sulfate, sodium hydroxide, cyclohexane (CH), sodium dodecyl benzene sulfonate (SDS), glucose, uric acid (UA), ascorbic acid (AA), sodium chloride (NaCl) and dopamine (DA) were purchased from Sinopharm Chemical Reagent Co. Ltd (Shanghai, China) and were used directly without further purification. Deionized water was used throughout. Graphene oxide (GO) was synthesized from graphite powder using an improved Hummers method [19]. The obtained GO is about 1.2 nm in thickness according to our previous work [20].

Preparation of PS/GO composite

PS was firstly dissolved in CH under 50 °C with a concentration of 20 mg/mL. GO and SDS were ultrasonic oscillated in water for 15 min under 50 °C to obtain a mixture having GO content of 2 mg/mL and SDS content of 50 mg/mL. Then, the mixture was dropped into the PS/CH solution having twice volume of the former under vigorously stirring. A water-in-oil emulsion was obtained after stirring for 4 h, during which the slurry was naturally cooled down to 25 °C. The emulsion was thereafter cast onto a clean glass substrate to evaporate CH at room temperature and then completely dried in a ventilation oven at 50 °C. The obtained sponge-like composite was washed thoroughly with water and ambient-dried.

Preparation of porous Cu2O/rGO composite

The PS/GO composite was soaked in a 50 mM copper sulfate aqueous solution for 12 h and then dipped into a 100 mM sodium hydroxide aqueous solution. After drying at ambient conditions, it was washed with water until neutral and heat-treated at 500 °C in a tube furnace (GSL-1500X, Kejing, China) under argon atmosphere for 3 h. During the thermal treatment, the polymer template of PS was removed, the GO was reduced into reduced graphene oxide (rGO) [19], and Cu2O was formed at the same time. In this manner, the Cu2O/rGO composite was fabricated.

Characterizations

The obtained samples were sputter-coated with a thin layer of gold (about 2 nm) and then examined with a scanning electron microscope (SEM, SU-70, Hitachi, Japan). X-ray diffraction (XRD) patterns of the obtained materials were recorded using a Bruker D8 ADVANCE X-ray diffractometer (Bruker, Germany) with Cu-Kα radiation. The samples were continuously scanned from 10° to 80° (2θ) at a speed of 0.0167° s−1.

Electrochemical performances and glucose detection

All electrochemical experiments were performed on an Autolab PGSTAT302N electrochemical work station (Metrohm, Switzerland). The electrochemical properties of the obtained composite were evaluated using a three-electrode system at room temperature, where the counter and reference electrodes were Pt foil and saturated calomel electrode (SCE), respectively. The working electrode was glass carbon electrode (GCE) modified with the Cu2O/rGO composite. The GCE (3 mm in diameter) was in turn polished by 1, 0.3 and 0.05 μm alumina powder and then washed repeatedly with ethanol and water. Four mg of the as-prepared Cu2O/rGO composite was dispersed in 1 mL 0.5% Nafion (Aldrich, USA) solution and then sonicated for 10 min to obtain a homogeneous black suspension. Four μL of the suspension was dropped onto the aforementioned GCE. After air-dried, the GCE modified with the Cu2O/rGO composite was served as the working electrode.

Cyclic voltammetry (CV) experiments for the detection of glucose were carried out in the potential window of −0.8 to 0.8 V with a series of scan rates (10, 20, 50, 80, 100, 150 and 200 mV/s) using 0.1 M NaOH solution as the electrolyte. Chronoamperometry experiments were implemented at a constant potential of 0.6 V in 0.1 M NaOH electrolyte.

Results and discussion

Preparation of the porous Cu2O/rGO composite

For the purpose of the fabrication of the final Cu2O/rGO porous composite material, a water-in-oil emulsion slurry was firstly prepared. Based on the strong π–π stacking interaction between the aromatic groups of PS and the GO sheets [18], the PS chains and the GO sheets assemble at the interface of the emulsion. The PS chains are in a relatively extended conformation in CH at 50 °C, which is much higher than the θ temperature (about 34.5 °C) of the PS/CH binary system [21]. With the temperature of the emulsion gradually decreasing during stirring, the PS chain coils start to shrink. As the temperature of the emulsion decreases below the θ temperature, the PS chains are precipitated and enriched at the interfaces [21]. After complete evaporation of the solvents (CH and water) of the casting slurry, a sponge-like film containing PS and GO was obtained. Figure 1 shows the morphology of the PS/GO composite film and indicates an irregular microparticle randomly packed microstructure.

Figure 1
figure 1

SEM image of the surface of the PS/GO film

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According to the above analysis, the obtained PS/GO composite has a hollow microcapsular microstructure, where the PS and the GO sheets form the walls of the microcapsules. This is confirmed by the SEM images illustrated in Fig. 2, which shows the microstructure of the PS/GO composite after thermal treatment at 500 °C in Argon for 3 h. Figure 2a shows a porous microstructure of the obtained material, and Fig. 2b reveals that the microcapsules constitute the matrix of the material. The microcapsules have a scale at hundreds of nanometers to several micrometers. It is interesting to note that most of the microcapsules are to some extent collapsed, which is attributed to evaporation of the water-droplet templates during drying of the PS/GO composite and to the removal of PS during the following thermal treatment. Therefore, the GO remained to form the framework of the obtained material after carbonization of the PS/GO composite film. It is worth mentioning that the obtained material is a free-standing film.

Figure 2
figure 2

SEM images of the PS/GO composite after thermally treated at 500 °C in Argon for 3 h

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Through immersing the as-prepared PS/GO composite film in an aqueous solution of copper sulfate and subsequent drying under ambient conditions, Cu2+ was loaded on the framework in order to take advantage of the specific microstructure. Then, it was soaked in sodium hydroxide aqueous solution to transfer the Cu2+ into Cu(OH)2. After washing until neutral, the Cu(OH)2-loaded film was thermally treated at 500 °C in argon atmosphere for 3 h to remove the polymer template. During the thermal treatment, the GO was reduced to reduced graphene oxide (rGO) [18] and Cu2O was formed [22] simultaneously, so that the Cu2O/rGO composite was prepared.

Figure 3 shows the SEM images of the obtained Cu2O/rGO composite. As illustrated in Fig. 3a, the composite retains its microporous morphology. More detailed observations (Fig. 3b) suggest that numerous nanoparticles at the scale of 100 nm decorate the matrix of the composite. Figure 4 displays the XRD pattern of the obtained composite. The peaks at 29.5°, 36.3°, 42.4°, 61.5° and 73.8° can be indexed to the (110), (111), (200), (220) and (311) crystal faces of Cu2O [8, 13], respectively. The sharp and strong peaks indicate that the Cu2O is highly crystalline, which may have positive influence on the electrochemical activity of the composite [23]. The weak peaks at 43.4°, 50.6° and 74.2° are corresponding to the crystalline planes of (111), (200) and (220) for Cu metal. However, there is no peak for crystalline CuO in the pattern. The results demonstrate that the loaded copper compounds have mainly been transferred into Cu2O during the thermal treatment, which is known as the carbothermal reaction procedure [24,25,26]. The wide band centered around 23.8° is attributed to the layered rGO, which was reduced from GO during the thermal treatment [19]. Therefore, the Cu2O/rGO composite was successfully prepared to combine the advantages of the microporous structure, the conductivity of rGO, and the electrochemical performance of nano-Cu2O, implying excellent potential application in electrochemical sensing.

Figure 3
figure 3

SEM images of the Cu2O/rGO composite under different magnification

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Figure 4
figure 4

XRD pattern of the Cu2O/rGO composite

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Glucose detection

The porous Cu2O/rGO composite was used to modify the GCE to serve as the working electrode. Figure 5 displays the CV curves of the electrode in 0.1 M NaOH aqueous solution in the absence and in the presence of 5 mM glucose at a scan rate of 50 mV/s. The CV curve in the absence of extra glucose has a sharp peak loop at the potential over 0.6 V, which could be attributed to the oxidation of copper ions [13, 27, 28]. After addition of 5 mM glucose in the electrolyte, the CV curve exhibits an obvious cathodic stage in the range of 0.3–0.7 V, which is considered to be due to the oxidation of glucose. It has been widely reported that the oxidation of glucose could be catalyzed by Cu2O, and the most acceptable mechanism is that Cu3+ ions produced from the oxidation of Cu2+ may act as electron transmitter to oxidize glucose into glucolactone [7, 11, 27, 28]. As illustrated in the CV curve, the present composite material has an obvious amperometric response to glucose.

Figure 5
figure 5

CV curves of the Cu2O/rGO composite in 0.1 M NaOH aqueous solution in the absence and in the presence of 5 mM glucose at scan rate of 50 mV/s

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Figure 6a shows the CV curves of the Cu2O/rGO composite material in 0.1 M NaOH aqueous solution in the presence of 5 mM glucose under different scan rates (10–200 mV/s). It is obvious that the profiles of the CV curves are similar under the investigated scan rates. Moreover, it is found that the glucose oxidation current increases with increasing the scan rate, so the current at 0.6 V has been correlated with the square root of the corresponding scan rate. As shown in Fig. 6b, the experimental data points can be linearly fitted with a correlation coefficient (R) of 0.9991. The linear dependence suggests that the oxidation of glucose is determined by its diffusion to the electrode [8]. This means that the composite has electrochemical response to glucose in a wide range of scan rates.

Figure 6
figure 6

CV curves of the Cu2O/rGO composite material in 0.1 M NaOH aqueous solution with 5 mM glucose under different scan rates (a), and dependence of the glucose oxidation current at 0.6 V on square root of scan rate (b). The line in b is the linear fitting of the experimental data with a correlation coefficient (R) of 0.9991

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The sensitivity of the composite to glucose has been evaluated. Figure 7a shows the amperometric responses of the Cu2O/rGO composite to successive addition of glucose at various concentrations under a constant working potential of 0.6 V. In order to eliminate the diffusion effect, the electrolyte was magnetically stirred to realize homogeneous distribution of glucose in the electrolyte. A well-defined and stable amperometric response has been observed during the successive addition of glucose. Moreover, it took within 10 s to achieve constant current upon addition of glucose, suggesting fast detection of glucose by the Cu2O/rGO material. The high electrocatalytic activity of Cu2O/rGO toward glucose could be attributed to the large surface area and high conductivity as well as fast electron transfer provided by graphene.

Figure 7
figure 7

Amperometric responses of the Cu2O/rGO composite to the successive addition of glucose with various concentrations in 0.1 M NaOH at 0.6 V (a), and dependence of the current on glucose concentration (b). The data points in b are linearly fitted in glucose concentration below and over 200 μM, respectively

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The dependence of the current on the glucose concentration is illustrated in Fig. 7b. It is interesting to see that the data points below and above 200 μM of glucose can be linearly fitted, respectively. The linear fitting of the data points in the glucose concentration window between 20 and 200 μM gives the dependence of I (μA) = 0.1381 + 37.93C (mM) with a correlation coefficient (R) of 0.9977. The sensitivity is thus obtained to be 525.1 μA/mM cm2, and the detection limit is calculated to be 4.74 μM (S/N = 3) of glucose. For the glucose concentration over 200 μM, the dependence reads I (μA) = 4.8583 + 13.96C (mM) (R = 0.9992) with a sensitivity of 198.2 μA/mM cm2. It is noted that the sensitivity in the higher glucose concentration region is much lower. This could be ascribed to the intermediates absorbed onto the surface of the electrode due to high concentration of glucose, leading to less direct contact between glucose and the sensor [17]. The observation of multiple linear response of the present Cu2O/rGO composite to glucose is similar as those reported in literatures [7, 13, 17]. The glucose detection performances of the present material is compared with those reported in some literature works, as collected in Table 1. The comparison indicates that the present sensor has relatively large linear range and low detection limit and provides satisfactory sensitivity.

Table 1 Comparison of the performances of the Cu2O-based non-enzymatic glucose sensors
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In addition, the specific response of the Cu2O/rGO composite to glucose is important for the practical condition. For that purpose, the electrochemical signal of the electrode to glucose was measured by successive addition of additional interferents such as AA, DA, UA and NaCl, which are the main electrolytes in human blood. Figure 8 shows the amperometric responses of the Cu2O/rGO composite to successively addition of 0.1 mM glucose, 0.05 mM DA, 0.05 mM AA, 0.05 mM UA and 0.05 mM NaCl under the working potential of 0.6 V in 0.1 M NaOH. It can be seen that the signals for DA, AA, UA and NaCl are all far less than that for glucose, indicating satisfactory selectivity toward glucose. Furthermore, 0.1 mM glucose was added after the interferents, and the response intensity was comparable with that of the first addition of glucose, suggesting excellent interference resistance. The stability of the Cu2O/rGO composite was tested by storing it in 0.1 M NaOH solution and to check its response to 0.1 mM glucose after specific intervals. It has been found that the amperometric responsive current retained 92% of its initial value after two weeks, indicating high stability. Therefore, such a porous Cu2O–rGO composite would be feasible for the fabrication of non-enzymatic glucose sensor with high stability, sensitivity and specificity.

Figure 8
figure 8

Amperometric responses of the Cu2O/rGO composite upon successive addition of 0.1 mM glucose, 0.05 mM DA, 0.05 mM AA, 0.05 mM UA, 0.05 mM NaCl and 0.1 mM glucose again under the working potential of 0.6 V in 0.1 M NaOH

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Conclusions

In this work, a double-sacrificial-template method was employed to successfully prepare a free-standing Cu2O/rGO porous nanocomposite. The results indicate that the rGO sheets constitute the walls of the matrix of the composite material and highly crystalline Cu2O nanoparticle at the scale of 100 nm decorates the rGO walls. The porous Cu2O/rGO composite was applied to modify the GCE to serve as the working electrode, which shows fast amperometric response to glucose. The Cu2O/rGO composite exhibits linear amperometric responses to glucose, which reads I (μA) = 0.1381 + 37.93C (mM) in the glucose concentration range of 20–200 μM, and I (μA) = 4.8583 + 13.96C (mM) when the glucose concentration is over 200 μM. The sensitivities are 525.1 and 198.2 μA/mM cm2 in the two glucose concentration regions, respectively. The satisfactory electrochemical response of the Cu2O/rGO porous nanocomposite to glucose can be attributed to the porous microstructure, the conductivity of rGO, and the electrochemical performance of crystalline nano Cu2O. Moreover, the Cu2O/rGO composite displays selective amperometric response to glucose and excellent stability in electrolyte solution, suggesting its feasibility for fabrication of non-enzyme electrochemical sensors for glucose detection.