Simple synthesis of nitrogen doped graphene/ordered mesoporous metal oxides hybrid architecture as high-performance electrocatalysts for biosensing study

Abstract

In the present work, a nitrogen doped graphene/ordered mesoporous metal oxides hybrid architecture (OMM-NGR) was prepared by a convenient procedure for the first time. During the preparation process, nitrogen doping, graphene reduction and ordered mesoporous metal oxides formation were successfully realized in one-step procedure. Transmission electron microscopy (TEM), energy-dispersive X-ray (EDX), elemental mapping, X-ray diffraction (XRD) and X-ray photoelectron spectroscopy (XPS) were employed to characterize the as-prepared nanocomposites. The hybrid architecture enables a good combination of electrochemically active metal oxide nanoparticles and graphene sheets, leading to the electrocatalytic advantages of both nitrogen doped graphene and ordered mesoporous metal oxides. The electrocatalytic performance of the nanocomposites was studied by using glucose, L-cysteine, uric acid (UA) and H₂O₂ as redox probes. The performance of OMM (M: Co, Fe)-NGR is found to be greatly improved compared to NGR. Furthermore, electrocatalytic quantitative and qualitative detection performance of OMM (M: Co, Fe)-NGR have been studied in detail. OMM (M: Co, Fe)-NGR exhibits outstanding electrocatalytic activity towards the probes with low detection limits, high reproducibility, good selectivity and long stability. The present OMM (M: Co, Fe)-NGR is a promising nanomaterial for electrochemical biosensing application.

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1. Introduction

Nanocomposites have attracted wide attention because of their potential to combine desirable properties of different nanoscale building blocks to improve mechanical, optical, electronic, or magnetic properties.^1–6 The disordered structure of the nanocomposites is due to chemical synthesis. To solve this problem, some researchers investigated other methods to prepare layered nanocomposites with ceramics, clays and graphite oxide nanoplatelets,^7–9 but these methods are time-consuming and difficult for nanomaterial synthesis. There are increasing numbers of studies in incorporating functional components, such as employing functional groups, polymers and nanoparticles on carbon nanomaterial. Metallic oxide nanoparticles, noble metal nanoparticles or non-metal loaded nanomaterial such as carbon nanotubes, ordered mesoporous carbon, g-C₃N₄ or graphene have been reported.^10–17 Combining the good electrical conductivity of carbon nanomaterial and high electrocatalytic performance of dopant, these nanocomposites show outstanding performance in electrochemical sensing, lithium rechargeable batteries, electrochemical biosensors area and so on.

Nevertheless, success has been limited to the following problems. First of all, the preparation methods are often time-consuming and difficult. It includes carbon nanomaterial preparation, dopant preparation and their combination. Moreover, the loading ability of carbon nanomaterial needs to be improved in order to increase the active site. Last but not least, the order of dopant is badly in need of improvement.

In order to overcome these problems and improve the electrocatalytic performance of nanocomposites, in the present work, for the first time, we intended to develop a simple one-step method to prepare novel nitrogen doped graphene/ordered mesoporous metal oxides nanocomposites.

Nitrogen doping can effectively ameliorate the electrocatalytic properties of graphene in the following aspects: (i) the doped nitrogen atoms can affect the spin density and charge distribution of adjacent carbon atoms in a great extent, producing active area on the graphene surface.¹⁸ (ii) Nitrogen doping can enhance the Fermi energy level of the conduction band, so as to enhance the conductivity of graphene.¹⁹ (iii) The nitrogen in graphene can interact with metal ions, preventing the aggregation of metal atoms and enhancing their dispersion.¹⁸ (iv) Nitrogen doping can produce more defect sites¹⁸ on the surface of graphene and increase the number of active catalytic sites. Researchers usually use two kinds of methods to prepare nitrogen doped graphene, one is the direct synthesis method and the other is the processing method. Direct synthesis method includes chemical vapor phase deposition method,²⁰ solvothermal method and arc discharge method. The processing method includes heat treatment, plasma treatment and hydrazine treatment method, etc. However, some complicated experimental steps are inevitable in these methods, moreover, the nitrogen doping degree and uniformity need to be further improved.

A variety of synthetic pathways have been proposed for the development of metal oxides nanostructures because of their potential applications.²¹ The use of soft templates^22–24 (chelating agents, surfactants, DNA, etc.) and hard templates^25–32 (anionic alumina, carbon nanotubes and mesoporous materials) has sparked wonderful contributions. Though various metal and semiconductor nanostructures have been successfully exploited,^22–24 uniform mesostructured crystallized metal oxide patterns are rarely reported.^30,31,33 This is probably a result of the difficulty of choosing the proper synthesis precursor, auxiliary reagents³⁴ and template.

In order to improve all the mentioned problems, including simplified synthetic method, realization of uniform nitrogen doping and high reduction degree of graphene, and formation of ordered metal oxides, in the present work, we have prepared nitrogen doped graphene/ordered mesoporous metal oxides nanocomposites by a simple and green method. Moreover, the preparation and combination of nitrogen doped graphene and ordered mesoporous metal oxides were finished in one step process. This work is of great significance in nanomaterial synthesis and electrocatalysis.

2. Experimental

2.1. Reagents and apparatus

D-(+)-Glucose, dopamine (DA), uric acid (UA), ascorbic acid (AA), L-cysteine, H₂O₂, TRIS, tetraethyl orthosilicate, Pluronic P123 (non-ionic triblock copolymer, EO₂₀PO₇₀EO₂₀), DMF, cobalt(II) nitrate hexahydrate and iron nitrate were purchased from Aladdin Industrial Corporation (shanghai, China). The serum samples of human were obtained from the local hospital (First central hospital of Baoding). All other reagents were of analytical grade and used as received without further purification. The 0.1 M NaOH and 0.1 M phosphate buffer saline (PBS, pH = 7.4) were employed as supporting electrolyte.

2.2. Instrumentation

Electrochemical measurements were carried out in a three electrode cell using a CHI 760E electrochemical workstation (CH Instruments, China). The modified glassy carbon electrode (GCE, 3 mm diameter) served as a working electrode, Pt wire and Ag/AgCl were used as the counter and reference electrodes, respectively. X-ray diffraction (XRD) patterns were recorded with a X-ray D/max-2200vpc (Rigaku Corporation, Japan) instrument operated at 40 kV and 20 mA and using Cu Kα radiation (κ = 0.15406 nm). Transmission electron microscopy (TEM) images of the samples were taken with a JEM-2100F transmission electron microscope (JEOL, Japan) operating at 200 kV. X-ray Photoelectron Spectrometer (XPS) was measured using Thermo ESCA LAB spectrometer (USA).

2.3. Material preparation

2.3.1. Preparation of GO/PDA. Graphene oxide (GO) was prepared by the modified Hummers method. For the synthesis of GO/PDA, 900 mg GO was dispersed in 900 mL deionized water and subjected to sonication. Then 1092 mg Tris was dispersed in above solution followed by sonication for 10 min. After that, 1800 mg of dopamine was added to the mixture which was continuously stirred at room temperature for 24 h. Subsequently, the product was centrifuged, washed for several times using deionized water and dried at 60 °C for 24 h, and the product is named GO/PDA.

2.3.2. Preparation of OMM (M: Co, Fe)-NGR. In a typical synthesis of OMM (M: Co, Fe)-NGR, 0.5823 g cobalt(II) nitrate hexahydrate (0.8081 g iron nitrate) was dissolved in 5 g ethanol under vigorous stirring, and then 0.15 g SBA-15 sample was added to the solution. After stirring for several hours, 0.06 g GO/PDA ethanol solution was added and the mixture was stirred for another 1 h. The solution was transferred to the beaker in order to evaporate the solvent at room temperature. After, when the sample was dried, it was calcined at 650 °C for 6 h. During this process, three problems including GO reduction, nitrogen doping and OMM (M: Co, Fe) formation were solved in one step. Finally, the product was refluxed gently with stirring at 85 °C for 2 h to remove SBA-15 template. Centrifugation was preferred for separation of the final product – OMM (M: Co, Fe)-NGR. The preparation process of OMM (M: Co, Fe)-NGR is shown in Scheme 1.

Scheme 1 Preparation process of nitrogen doped graphene/ordered mesoporous metal oxides hybrid architecture.

2.4. Electrode preparation

Prior to the modification, the bare GCE was polished with 1.0, 0.3 and 0.05 μm alumina slurry and sonicated successively in double-distilled water. Then, 2 mg of the as-prepared NGR and OMM (M: Co, Fe)-NGR samples were dispersed into 1 mL DMF to give a homogeneous suspension upon bath sonication. The NGR/GCE and OMM (M: Co, Fe)-NGR/GCE were prepared by casting 5 μL of the suspension on the surface of the well-polished GCE and the solvent was allowed to dry under an infrared lamp.

3. Results and discussion

3.1. Characterization of OMM (M: Co, Fe)-NGR nanocomposites

The morphology of the as-prepared OMCo-NGR and OMFe-NGR was characterized by TEM (Fig. 1). As can be seen, mesoporous OMCo and OMFe loaded on NGR have an ordered structure, an exact negative replica of SBA-15 with hexagonal arrangement of cylindrical tubes.³⁵ It is reasonable to believe that the ordered mesostructure has been formed in the OMCo-NGR and OMFe-NGR. Furthermore, energy-dispersive X-ray (EDX) spectra (inset of Fig. 1A and C) are displayed to determine the elements in OMCo-NGR and OMFe-NGR. Peaks corresponding to C, N, O and Co in OMCo-NGR and C, N, O and Fe in OMFe-NGR are shown in the EDX spectra, which confirm the successful doping of Co and Fe into OMCo-NGR and OMFe-NGR. As shown in Fig. 2, the elemental mapping images indicate the elemental distributions in OMCo-NGR and OMFe-NGR. The results show that nitrogen element is uniformly distributed in graphene. Moreover, Co and Fe elements can be clearly seen, which indicates the successful loading of OMCo and OMFe in NGR.

Fig. 1 (A) TEM image of OMCo-NGR nanocomposites. Inset: EDX spectra of OMCo-NGR nanocomposites. (B) TEM image of OMCo-NGR nanocomposites. Inset: megascopic TEM image. (C) TEM image of OMFe-NGR nanocomposites. Inset: EDX spectra of OMFe-NGR nanocomposites. (D) TEM image of OMFe-NGR nanocomposites. Inset: megascopic TEM image.

Fig. 2 (A) Elemental mapping image of OMCo-NGR nanocomposites. (B) Elemental mapping image of OMFe-NGR nanocomposites.

Fig. 3 shows typical XRD profiles for the as-prepared OMCo-NGR and OMFe-NGR. As exhibited in Fig. 3A, OMCo-NGR displays wide-angle patterns with the diffraction peaks at 2θ values of 21.06°, 31.20°, 36.62°, 44.78°, 55.88°, 59.10° and 65.32°. These values correspond to (111), (220), (311), (400), (422), (511) and (440) crystal plants of Co₃O₄ (JCPDS 42-1467),³⁶ indicating that Co₃O₄ has been formed successfully. The wide-angle XRD of OMFe-NGR shown in Fig. 3B indicates five characteristic peaks at 29.92°, 35.37°, 42.87°, 57.05° and 62.70°, which belong to (220), (311), (400), (511) and (440) crystallographic plane of Fe₃O₄.³⁵

Fig. 3 (A) XRD patterns of OMCo-NGR nanocomposites. (B) XRD patterns of OMFe-NGR nanocomposites.

The elemental composition of the synthesized products was further investigated by XPS. Compared with previous reports,^37,38 the peaks of C 1s, O 1s, N 1s, Co 2p and Fe 2p can be clearly seen in Fig. 4A and D. The C 1s region located at the binding energy of 280.885 and 279.043 eV is commonly attributed to carbon linked only to hydrogen or carbon (C–H or C–C).³⁹ The high resolution N 1s XPS spectra (Fig. 4B and E) show three nitrogen species: pyridinic N (N1, 398.0 eV), pyrrolic N (N2, 400.0 eV) and graphitic N (N3, 401.3 eV), which are typically observed in the case of N-doped carbons. Among the three nitrogen species (100%), the percentage of pyridinic N, pyrrolic N and graphitic N is 28.55%, 40.02%, and 31.42% in OMCo-NGR and 29.88%, 43.36% and 26.76% in OMFe-NGR samples, respectively. As can be seen, pyrrolic N is predominant in both OMCo-NGR and OMFe-NGR samples. Nevertheless, the pyridinic N portion is crucial for the promotion of the electrocatalytic reaction. It is because pyridinic N atoms with strong electron-accepting ability can create a net positive charge on the adjacent carbon atoms, favorable for the adsorption of small molecules, thus facilitating the electrochemical reaction and dominates the electrocatalytic activity.^40,41 Fig. 4C shows XPS spectrum of Co 2p in OMCo-NGR, which includes two peaks centered at 779.79 eV (Co 2p_3/2) and 794.92 eV (Co 2p_1/2). The peaks at 711.23 and 724.73 eV (Fig. 4F) are the characteristic peaks of Fe 2p_3/2 and Fe 2p_1/2 core-level spectra of iron oxide.⁴² These results testify the conclusion that OMCo-NGR and OMFe-NGR have been successfully synthesized.

Fig. 4 (A) XPS spectra of OMCo-NGR sample. (B) High resolution N 1s XPS spectra of OMCo-NGR sample. (C) XPS spectra of Co 2p_3/2 and Co 2p_1/2 for OMCo-NGR sample. (D) XPS spectra of OMFe-NGR sample. (E) High resolution N 1s XPS spectra of OMFe-NGR sample. (F) XPS spectra of Fe 2p_3/2 and Fe 2p_1/2 for OMFe-NGR sample.

3.2. Electrooxidation of glucose at the OMCo-NGR/GCE

Fig. 5A shows cyclic voltammograms (CVs) of the various electrodes (GCE and OMCo-NGR/GCE) in 0.1 M NaOH. There is almost no electrochemical response at bare GCE while OMCo-NGR/GCE shows four obvious redox peaks in 0.1 M NaOH. Two peaks located at 275 mV (peak I) and 567 mV (peak III) ascribe to Co(II)/Co(III) and Co(III)/Co(IV) redox transition, respectively, connected with diverse cobalt oxide species on the electrode surface.⁴³ Two other peaks located at 262 mV (peak II) and 513 mV (peak IV) are the considerable cathodic peaks associated with the reduction of cobalt nanoparticles which are formed in the anodic cycles.⁴⁴ This can be attributed to the production of Co(II) species.⁴⁴ These characteristic peaks of Co indicate that Co has been loaded on NGR successfully. To investigate the electrocatalytic performance of OMCo-NGR/GCE towards glucose, some other electrodes have been used for comparison. It can be clearly seen in Fig. 5B that GCE and NGR/GCE show weak electrocatalytic oxidation current towards glucose. However, OMCo-NGR/GCE exhibits a distinct peak at 588 mV, indicating that OMCo-NGR/GCE has stronger electrocatalytic activity towards the oxidation of glucose. Moreover, the electrocatalytic ability of OMCo-NGR/GCE was also investigated in 0.1 M PBS. Compared with 0.1 M NaOH, no obvious redox peak was observed in 0.1 M PBS (pH = 5, 6, 7, 8, 9) in the presence of 0.5 mM glucose. So 0.1 M NaOH was employed as supporting electrolyte. As shown in Fig. 5C, the response current increases with increasing concentration of glucose and thereby confirming the usefulness of OMCo-NGR/GCE for the quantitative detection of glucose.

Fig. 5 (A) CVs of GCE (a) and OMCo-NGR/GCE (b) in 0.1 M NaOH. (B) CVs of GCE (a and b), NGR/GCE (c and d) and OMCo-NGR/GCE (e and f) in 0.1 M NaOH in the absence (a, c and e) and presence of 0.5 mM glucose (b, d and f). Inset: CVs of OMCo-NGR/GCE in 0.1 M PBS (g to k: pH 5, 6, 7, 8, 9) in the presence of 0.5 mM glucose. (C) CVs of OMCo-NGR/GCE in 0.1 M NaOH (a) in the presence of 0.25 mM (b), 0.5 mM (c) and 0.75 mM (d) glucose, respectively. Scan rate: 50 mV s⁻¹.

The glucose oxidation mechanism was also researched. The pair of redox peaks I/II and peaks III/IV can be interpreted as below eqn (1) and (2).⁴⁵

Co₃O₄ + OH⁻+ H₂O ↔ 3CoOOH + e⁻ (1)

CoOOH + OH⁻ ↔ CoO₂ + H₂O + e⁻ (2)

The increase of current at peak III (CoOOH → CoO₂) with the addition of glucose, indicates that glucose electrooxidation is mainly mediated by CoOOH/CoO₂.

2CoO₂ + C₆H₁₂O₆ (glucose) → 2CoOOH + C₆H₁₀O₆ (gluconolactone) (3)

With the consumption of CoO₂ and the production of CoOOH, the extent of reaction (2) would greatly favor the forward reaction (CoOOH → CoO₂), thus resulting in an enhanced oxidation peak III upon the addition of glucose (Fig. 5C).³⁶

The effect of the scan rate on the CVs of the OMCo-NGR/GCE electrode is also investigated. Fig. 6A exhibits the CVs of OMCo-NGR/GCE in 0.1 M NaOH at different scan rates of 10, 20, 30, 40, 50, 70, 90, 120, 150, 180 and 210 mV s⁻¹. With the increase in the scan rate, the current response of four peaks increases. The redox peak currents increase linearly with the scan rates from 10 to 210 mV s⁻¹ (Fig. 6B), indicating a surface-controlled electrochemical process.³⁵

Fig. 6 (A) CVs of OMCo-NGR/GCE in 0.1 M NaOH at different scan rates of 10, 20, 30, 40, 50, 70, 90, 120, 150, 180 and 210 mV s⁻¹. (B) Plot of peak currents vs. scan rate.

To further explore the electrocatalytic performance of OMCo-NGR/GCE for the quantitative detection of glucose, amperometry has been employed. Fig. 7A shows amperometry of OMCo-NGR/GCE with successive additions of different concentrations of glucose into 0.1 M NaOH at an applied potential of +0.54 V. As directly perceived through current staircases, the response of OMCo-NGR/GCE is dramatically increased after each addition of glucose. The corresponding calibration plot for the oxidation of glucose at OMCo-NGR/GCE is shown in Fig. 7B. The linear response of glucose measurement is up to 152.139 μM on OMCo-NGR/GCE (Fig. 7B). The detection limit, based on a signal-to-noise ratio of 3, is estimated to be 0.26 μM on OMCo-NGR/GCE. From the slope of the calibration curve, much higher sensitivity of 1.98 μA μM⁻¹ cm⁻² is obtained.

Fig. 7 (A) Amperometry of OMCo-NGR/GCE with the additions of different concentration glucose to 0.1 M NaOH at an applied potential of +0.54 V. Inset: the magnification of (A). (B) The corresponding calibration curve of (A).

The selectivity of OMCo-NGR/GCE towards glucose detection has been also researched. Fig. 8A shows the differential pulse voltammetry (DPV) responses of three potential interfering substances in 0.15 mM glucose. It can be found that little interference is present when uric acid (UA), dopamine hydrochloride (DA) and ascorbic acid (AA) were added in 0.15 mM glucose. As shown in Fig. 8B, compared to the current response of glucose, the additions of 2 mM AA, UA and DA result in the minor increase of the current (2%, 4% and 3%, respectively), which indicates that the OMCo-NGR/GCE has a higher selectivity towards glucose determination.³⁶

Fig. 8 (A) DPVs of OMCo-NGR/GCE with (a) 0.15 mM glucose, (b) 0.15 mM glucose and 2 mM AA, (c) 0.15 mM glucose and 2 mM UA and (d) 0.15 mM glucose and 2 mM DA in 0.1 M NaOH. Pulse amplitude: 0.05 V. Pulse width: 0.05 V. Pulse period: 0.5 s. (B) The bar graph of (A).

3.3. Electrooxidation of Cys at the OMCo-NGR/GCE

Fig. 9A shows the CVs of GCE, NGR/GCE and OMCo-NGR/GCE in the presence and absence of 1 mM Cys in 0.1 M NaOH. As can be seen, GCE shows a pretty small current response while NGR/GCE exhibits larger background current, indicating the large electrical activity area of NGR. Interestingly, in the presence of 1 mM Cys, the current response on NGR/GCE is larger than that on GCE. As expected, OMCo-NGR/GCE displays the maximum response current, indicating the best electrocatalytic performance towards Cys detection. It is likely that the active Co(IV) species can accelerate the oxidation of ammo acids.⁴⁴ Fig. 9B shows that the current increases with the addition of Cys. This fact suggests that OMCo-NGR/GCE may be used for the quantitative detection of Cys.

Fig. 9 (A) CVs of GCE (a and b), NGR/GCE (c and d) and OMCo-NGR/GCE (e and f) in 0.1 M NaOH in the absence (a, c and e) and presence of 1 mM Cys (b, d and f). (B) CVs of OMCo-NGR/GCE in 0.1 M NaOH (a) in the presence of 1 mM (b) Cys, respectively. Scan rate: 50 mV s⁻¹.

The oxidation mechanism of Cys was researched.⁴⁶ Cys is oxidized by active Co(IV) moiety through a cyclic mediation redox process. The redox transition of cobalt species is:

Cys is oxidized via the following reaction:

Intermediate is further oxidized to the product:

Amperometry signal of OMCo-NGR/GCE for successive addition of the same concentration cysteine (0.3 μM) into 0.1 M NaOH is depicted in Fig. 10A. As directly perceived through current staircases, the response of OMCo-NGR/GCE is dramatically increased after each addition of Cys. Fig. 10B displays the amperometry at OMCo-NGR/GCE with successive additions of different concentration of Cys at +0.45 V. The current increases stepwise with each successive addition of Cys at the OMCo-NGR/GCE. A linear relationship of the OMCo-NGR/GCE is established in the range of 0.1 μM to 95.3 μM with the sensitivity of 0.06 μA μM⁻¹ and a detection limit of 0.05 μM (signal-to-noise ratio = 3).

Fig. 10 (A) Amperometry of OMCo-NGR/GCE with the additions of same concentration of Cys (0.3 μM) to 0.1 M NaOH at an applied potential of +0.45 V. (B) Amperometry of OMCo-NGR/GCE with the additions of different concentrations of Cys to 0.1 M NaOH at an applied potential of +0.45 V. Inset: the magnification of (B). (C) The corresponding calibration curve.

3.4. Electrooxidation of uric acid

Fig. 11A shows the CVs of GCE and OMFe-NGR/GCE in the presence and absence of UA in 0.1 M PBS. The high electrocatalytic performance of OMFe-NGR/GCE towards UA oxidation is indicated by the increased oxidation peak current compared to that of GCE. Fig. 11B shows the CVs for the various concentrations of UA at OMFe-NGR/GCE. Any increase in the concentration of UA causes a proportional and nearly linear enhancement of the peak current, indicating the possibility of quantitative detection of UA.

Fig. 11 (A) CVs of GCE (a and b) and OMFe-NGR/GCE (c and d) in 0.1 M PBS in the absence (a and c) and presence of 5 mM UA (b and d). (B) CVs of the OMFe-NGR/GCE in 0.1 M PBS (a) in the presence of 0.5 mM (b), 0.75 mM (c) and 1 mM UA (d), respectively. Scan rate: 50 mV s⁻¹.

The electrochemical oxidation mechanism of UA on OMFe-NGR/GCE can be explained in the following way: first, UA transports the mass to the electrode surface by diffusion. The next is the adsorption of UA on to the OMFe-NGR/GCE surface active sites. Then, OMFe(II)-NGR/GCE is electrochemically oxidized to OMFe(III)-NGR/GCE. Finally, the OMFe(III)-NGR/GCE reacted with adsorbed UA which causes an electrochemical oxidation of UA.⁴⁷

In addition, DPV response of OMFe-NGR/GCE towards UA is considered. Clearly, a series of the DPV curves were obtained from different concentrations of UA solutions in the PBS (Fig. 12A). It is clearly seen the oxidation peak current of UA increases linearly with the increase concentration of UA. In Fig. 12B, OMFe-NGR/GCE displays a linear range of 10–535 μM with a sensitivity of 0.042 μA μM⁻¹ cm⁻². The detection limit was calculated to be 5.38 μM with the signal to noise ratio of three (S/N = 3).

Fig. 12 (A) DPVs of different concentrations of UA in 0.1 M PBS. Pulse amplitude: 0.05 V. Pulse width: 0.05 V. Pulse period: 0.5 s. UA concentrations (a to p): 10, 15, 25, 40, 60, 85, 115, 150, 190, 235, 285, 335, 385, 435, 485 and 535 μM. (B) The linear relationship between peak currents and concentrations.

3.5. Electrocatalysis of H₂O₂

In order to further investigate the electrocatalytic activity of OMFe-NGR/GCE electrode, H₂O₂ was employed as the redox probe. Fig. 13A shows the CVs of H₂O₂ reduction on GCE, NGR/GCE and OMFe-NGR/GCE in 0.1 M PBS. It can be clearly seen, compared with other two electrodes, OMFe-NGR/GCE shows superior electrocatalytic response to H₂O₂ with lower over-potential and higher peak current. The quantitative detection of H₂O₂ was performed by amperometry.

Fig. 13 (A) CVs of GCE (a and b), NGR/GCE (c and d) and OMFe-NGR/GCE (e and f) in 0.1 M PBS in the absence (a, c and e) and presence of 3 mM H₂O₂ (b, d and f). (B) Amperometry of OMFe-NGR/GCE with the additions of different concentration (μM) H₂O₂ to 0.1 M PBS at an applied potential of −0.4 V. Inset: the magnification of (B). (C) The corresponding calibration curve of (B). Scan rate: 50 mV s⁻¹.

The electrocatalytic mechanism could be that H₂O₂ was instantly reduced by reacting with OMFe(II)-NGR as soon as H₂O₂ arrived at the OMFe(III)-NGR on the electrode surface (eqn (7)), and the resulting OMFe(III)-NGR was then reduced to OMFe(II)-NGR (eqn (8)) to facilitate the mediation effect for the reduction of H₂O₂.⁴⁸

OMFe(II)-NGR + H₂O₂ → OMFe(III)-NGR + H₂O (7)

OMFe(III)-NGR + e⁻ → OMFe(II)-NGR (8)

Amperometry was further employed to investigate the quantitative detection of H₂O₂ on OMFe-NGR/GCE by varying the H₂O₂ concentration. Fig. 13B displays the amperometric responses of OMFe-NGR/GCE for successive addition of different concentrations of H₂O₂ in 0.1 M PBS (pH = 7.4). The inset curve shows the current response of H₂O₂ at low concentrations on OMFe-NGR/GCE. As it is depicted, the current response is dramatically increased after each addition of H₂O₂ into PBS and rapidly reaches a platform. From Fig. 13C, a good linear relationship in the range of 13.6 μM to 2961.6 μM and a low detection limit of 2.29 μM (S/N = 3) are obtained for the detection of H₂O₂.

By taking the above experimental results into consideration, we may easily deduce that the high electrocatalytic performance of OMM-NGR could be attributed to the unique properties of OMM and NGR, especially the unusual host–guest synergy of OMM-NGR. First, nitrogen doping in NGR may provide more favorable active sites for electron transfer. Moreover it can increase the surface area and loading capacity of NGR. Additionally, the ordered mesoporous channel of OMM may greatly enhance the electrocatalytic performance by offering a favorite microenvironment for transferring species in solution. This would be beneficial for accelerating heterogeneous electron transfer between the electrode and species in solution. So, combing the advantages of OMM and NGR, OMM-NGR is a promising nanomaterial in electrocatalytic biosensing field.

3.6. Accuracy, reproducibility and stability of the OMM (M: Co, Fe)-NGR/GCE

Three glucose samples were employed for accuracy test. The relative errors for three samples are −2.0%, −4.0%, and 3.3%, respectively. The obtained results demonstrate the high accuracy of OMCo-NGR/GCE for the detection of glucose.

According to the reproducibility test, the relative standard deviation (RSD) is 1.12% at the OMCo-NGR/GCE by five successive measurements of 1.0 mM glucose. The RSD at the OMFe-NGR/GCE is 1.24% by five successive measurements of 150 μM UA. The experimental results show the OMM (M: Co, Fe)-NGR/GCE has acceptable reproducibility.

The stability of the prepared OMM (M: Co, Fe)-NGR/GCE was also investigated. The electrodes were stored at 4 °C for 4 days. After 4 days, the redox peak currents of glucose retain 91% of their initial response values on OMCo-NGR/GCE. Under the same experimental conditions, the peak currents of UA retain 90% of their initial response values on OMFe-NGR/GCE. These data suggest the long-term stability of the OMM (M: Co, Fe)-NGR/GCE.

3.7. Real sample analysis

In order to study the analytical performance of the prepared electrode for real sample, serum sample was used for H₂O₂ and glucose detection (ESI, Tables S1 and S2†). The obtained good recovery suggests that OMFe-NGR/GCE and OMCo-NGR/GCE can realize the actual sample detection.

4. Conclusions

We have developed the new design and simple synthesis of nitrogen doped graphene/ordered mesoporous metal oxides hybrid architecture (OMM-NGR). The resulting hybrid architecture enables a good combination of electrochemically active ordered metal oxide and graphene sheet, leading to remarkable electrochemical biosensing performance. The OMM (M: Co, Fe)-NGR sample exhibits improved electrocatalytic performance with low detection limits, high sensitivity and good selectivity towards glucose, UA, L-cysteine and H₂O₂ compared to bare GCE. This work is of great importance because it not only opens a new route for the synthesis of novel hybrid architecture material, but also broadens the application of nanocomposites material.

Acknowledgements

The authors gratefully acknowledge the support from the National Natural Science Foundation of China (No. 21505031), the Natural Science Foundation of Hebei Province (No. B2016201018) and the colleges and universities science technology research project of Hebei Province (No. Z2015096).

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c6ra19496f

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