Structure and Electrochemical Performance of Nitrogen-Doped

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Structure and Electrochemical Performance of Nitrogen-Doped Carbon Film Formed by Electron Cyclotron Resonance Sputtering Tomoyuki Kamata,† Dai Kato,† Shigeru Hirono,‡ and Osamu Niwa*,† †

National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan MES-Afty Corporation, 2-35-2 Hyoe, Hachioji, Tokyo 192-0918, Japan.

‡

S Supporting Information *

ABSTRACT: A nitrogen-doped nanocarbon film electrode with mixed sp2 and sp3 bonds formed using the electron cyclotron resonance (ECR) sputtering method was studied with respect to the relationship between nitrogen concentration and electrochemical performance. The film (N-ECR) has a nanocrystalline structure, and the sp3 content increases with increasing nitrogen concentration unlike the recently reported nitrogen-containing tetrahedral amorphous carbon film.1 The film has a very smooth surface with an average roughness of 0.1 to 0.2 nm, which is almost independent of nitrogen concentration. In contrast, the ratio of nitrogen-containing graphite-like bonding is high at low nitrogen concentrations, and then pyridine-like bonding increases as the nitrogen concentration increases. These variations in the chemical structures and the sp2 and sp3 content greatly change the electrochemical performance. The N-ECR electrode shows a wider potential window (∼3.8 V) than a pure nanocarbon electrode (∼3.1 V) due to its higher sp3 content. The N-ECR electrode (N = 9.0 at. %) shows improved electrochemical activity because the lowest peak separation of Fe(CN)63−/4− was observed at this nitrogen concentration. The oxygen and hydrogen peroxide (H2O2) reduction potentials at the N-ECR electrode shifted about 0.3 and 0.15 V, respectively, and the peak height of H2O2 is greatly increased. As a result, a linear relationship was obtained from 0.2 to 17 mM for the reductive current detection of H2O2. The N-ECR electrode also shows better activity for oxidizing certain biomolecules. The oxidation potentials of guanosine and adenosine decreased about 0.1 V, suggesting that the N-ECR electrode is suitable for use as a biosensing platform.

C

at nitrogen-containing carbon materials.4,5 More recently, nanostructured carbon materials with high electrocatalytic activity for ORR have been reported, including nitrogencontaining carbon nanotubes (N-CNTs) prepared by pyrolyzing iron(II) phthalocyanine at 800−1100 °C on a quartz glass plate or nitrogen-doped graphene by chemical vapor deposition in the presence of NH3 (at 1000 °C).6,7 Since these carbon materials have a large surface area because they are typically powder, flake, or fiber-like structures, they are very suitable as electrodes for fuel cells and batteries. However, such flake or fiber-like electrode materials are not always suitable for the electroanalysis of biomolecules because their large surface area generally increases the background noise level, which is inconvenient as regards improving the detection limit (or improving the S/N ratio). These materials are not freestanding, and so they are usable after they have been modified on solid electrodes such as GC or metal. In addition, their high temperature fabrication process limits the usable solid electrode materials.

arbon materials such as glassy carbon (GC) and borondoped diamond (BDD) are widely used for the electrochemical measurement of biomolecules because of their advantageous properties for electroanalytical measurements.2 For example, they exhibit a wider potential window and lower noise than metal electrodes such as Au and Pt. Since the electrochemical activity of carbon electrodes depends on their structure, it is very important to fabricate carbon electrodes by controlling their structures with higher electrochemical activity. For example, it is well-known that an edge plane is much more active than a basal plane.3 Therefore, to make carbon materials containing a crystalline structure and a large quantity of surface edge planes is a common approach.3 The other approach is to dope the carbon materials with other elements (Fe, Co, B, and N) to realize high activity of the oxygen reduction reaction (ORR) as used for the cathodes of fuel cells. In particular, a nitrogen-containing carbon alloy usually obtained by the carbonization of nitrogen-containing organic precursors such as phthalocyanine and polyimides at high temperature (600− 1000 °C) can be expected to provide an alternative to a conventional Pt-based cathode (Pt) as a metal-free electrocatalyst for fuel cells. This is because various researchers have reported that the ORR potential shifted in a positive direction © 2013 American Chemical Society

Received: July 30, 2013 Accepted: September 20, 2013 Published: September 20, 2013 9845

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On the other hand, thin film carbon electrodes have more advantages for electroanalytical applications since the film can be fabricated with excellent reproducibility using low temperature processes such as the sputtering method. The film form is also advantageous for fabricating any shape or size of electrode by conventional micro or nanofabrication processes. Tetrahedral amorphous carbon thin film (ta-C) thin film is a potent carbon thin film material that can be deposited at relatively low temperatures (25−100 °C).8 Since ta-C thin film typically has an amorphous structure and consists of 40−60% sp3-bonded carbon, it exhibits low electrode activity. To improve aspects of the electrochemical performance, some groups have studied the effect of nitrogen doping on ta-C (ta-C:N) thin films.9,10 For example, Yoo et al. demonstrated that ta-C:N films exhibit a wide potential window (∼3 V), good electrode activity for Ru(NH3)6+3/+2 .11 More recently, Swain et al. reported the relationship between the carbon structure and electrochemical properties of ta-C:N thin films deposited by pulsed laser-arc deposition (PLD).1 They demonstrated that the conductivity of ta-C:N films and their sp2 bonds increases with increasing nitrogen concentration. They also reported a wide potential window (∼3 V) and improved activity for Fe(CN)63−/4− with increased nitrogen concentration. In addition, the surface roughness increased with increasing nitrogen concentration. In contrast, we have developed a carbon film electrode formed by using electron cyclotron resonance (ECR) sputtering. This method offers high-density ion irradiation at the growth surface of the film that allows us to form an ultraflat nanocrystalline carbon film at a relatively low temperature.12 This film electrode consists of a nanocrystalline sp2 and sp3 mixed bond structure and exhibits excellent electrochemical performance.13,14 Swain et al. also commented in their report that ta-C and ta-C:N films exhibit certain similarities with our ECR nanocarbon film.1 With the above as background, we recently obtained a nitrogen-doped ECR (N-ECR) nanocarbon film and investigated the relationship between nitrogen concentration and the structure/mechanical properties of NECR nanocarbon films.15 Our previous results clearly demonstrated that the sp3-bonded carbon content in N-ECR nanocarbon film increases with increasing nitrogen concentration. This result is quite unlike that recently reported by Swain’s group for ta-C:N thin films where the sp3-content decreased with increases in doped nitrogen. In addition, the surface flatness of N-ECR nanocarbon films is almost unchanged by increasing nitrogen concentration, which is advantageous for comparing the electrokinetic parameters of NECR nanocarbon films with different nitrogen concentrations without taking account of surface roughness. In our previous report, we also found that nitrogen atoms are incorporated into the graphene structure in the low nitrogen concentration region,15 which is expected to show high electrode activity. In addition, we reported that the pyridine-like structure, which is considered to have low electrode activity, increases with further increases in the nitrogen concentration. The above structural information makes it possible to study the relationship between the structure and electrochemical performance of N-ECR electrodes and thus design carbon thin film electrodes with an optimized structure and superior electrochemical activity. Here, we studied the structure and electrochemical properties of N-ECR electrodes with different content of sp3 bonds and nitrogen concentrations using some redox species. We also investigated the relationship between the structure of N-ECR electrodes and the electrocatalytic performance, including the

ORR and H2O2 reduction. Finally, we demonstrated the advantages of N-ECR electrodes for detecting certain biologically important analytes, namely nucleic acids, by comparison with a pure-ECR electrode.

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EXPERIMENTAL SECTION Electrode Preparation. A pure-ECR nanocarbon film electrode and an N-ECR nanocarbon film electrode were deposited on a boron-doped silicon (100) substrate by ECR sputtering at room temperature. A sintered carbon target was used. An argon (Ar) and nitrogen (N2) mixture gas was used with the N2 gas content controlled in the 0 to 40% range and the total pressure maintained at 5.0 × 10−2 Pa. The film thickness was 40 nm. No special catalyst metal was used during the deposition. Film Characterization. X-ray photoelectron spectroscopy (XPS) was performed using a Shimadzu/Kratos model AXIS Ultra (Al Kα 1486.6 eV) spectrometer to determine the elemental composition and the quantity of chemical bonds in the carbon film electrode surface. The nitrogen concentration of the N-ECR electrode was estimated from the peak area ratio of the N 1s corrected using instrumental sensitivity factors. The C 1s and N 1s spectra were fitted by Gaussian−Lorentzian sum components after the background had been subtracted according to Shirley’s method.16 Atomic force microscopy (AFM) measurements were performed with an SPI4000 (SII NanoTechonolgy, Inc.) using a silicon cantilever in air at room temperature. The images were recorded in the dynamic force AFM mode at scan rates of 0.28 Hz with 256 × 256 pixels. Electrochemical Measurements. Cyclic voltammograms (CVs) and square-wave voltammograms (SWVs) were obtained using a three-electrode configuration with an electrochemical analyzer Model ALS/CHI 730C (CH Instruments, Inc.). In all the experiments, platinum wire and an Ag/ AgCl electrode were used as auxiliary and reference electrodes, respectively. The electrolyte solutions were purged with pure Ar gas for 20 min prior to the measurement and blanketed with Ar during the measurement in the deoxygenated experiments. For an oxygen (O2) reduction reaction (ORR), the solutions were purged with pure O2 for 5 min prior to the measurement. The H2O2 reduction was measured from CVs obtained after adding different amounts of H2O2 to the sample solutions. All the SWV measurements were performed with an amplitude of 25 mV and a ΔE of 5 mV at 10 Hz. Chemicals. All the chemicals were analytical grade and were used as received. Hexaammineruthenium(III) chloride was purchased from Sigma-Aldrich. Potassium ferricyanide was obtained from Wako Pure Chemical Industries, Ltd. (Japan). KH2PO4, Na2HPO4, and sulfuric acid were purchased from Kanto Chemical. We prepared 100 mM phosphate buffer,17 which contained 100 mM KH2PO4 and Na2HPO4. We prepared a fresh concentration of H2O2 with 0.2 to 17 mM phosphate buffer (pH 7.0). A 50 mM acetate buffer (pH 5.0) was used as the electrolyte solution for electrochemical nucleoside detection. Acetate buffer is the most appropriate for obtaining well-defined peaks with high sensitivity.18 Adenosine and guanosine (0.1 mM) were purchased from Sigma-Aldrich. Ultrapure water was used in all the experiments.

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RESULTS AND DISCUSSION Chemical and Surface Structures. Recently, Swain’s group reported the structure of ta-C:N film electrodes when 9846

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they changed the N2 flow rate, which is proportional to the nitrogen concentration as shown in the Supporting Information of their paper.1 The sp2 bonds account for about 44% when the N2 flow rate is between 0 to 10 sccm, and then it increases to 58% when the flow rate is increased to 50 sccm. The ID/IG value obtained with Raman spectroscopy increases from 0.64 to 1.12 with increasing nitrogen concentration, indicating a disorder structure in the nitrogen-rich film. Particle-like structures on the film surface increase with increasing nitrogen concentration and finally make the surface much rougher than that of the ta-C (pure carbon film). We prepared N-ECR nanocarbon film electrodes with different nitrogen concentrations by changing the N2 gas content. Table 1 shows the variation in the amounts of each

We obtained transmission electron microscopy (TEM) images with and without doping the film with nitrogen atoms, as shown in Figure 1. Without nitrogen atoms, a nanometer order layered structure could be frequently observed in the film, indicating that the film contained a large quantity of nanocrystalline graphite planes, which is consistent with the film containing a large quantity of sp2 bonds, as in our previous reports.13 On the other hand, the layered structure decreased and the circled and closed structures increased with increasing nitrogen concentration. Since a similar structure with a lower sp2 bond content of 55.2% can be observed for a pure-ECR electrode by increasing the ion acceleration voltage between the target and substrate, the decreases in the layered structure in the TEM image is related to the reduction in sp2 content. This difference between our N-ECR electrode and the previously reported ta-C:N electrode could be due to the difference in the atomic diffusion on the substrate. With ECR sputtering, highdensity ion irradiation on the substrate increases the energy of the carbon atoms, which could cause a rearrangement of the carbon film structures such as an increase in graphite nanocrystalline structures. The relatively small Ra of ECR carbon films could also be due to the rearrangement of carbon atoms on the deposited film surface. In contrast, PLD forms a relatively amorphous structure similar to diamond-like carbon films, which contain a relatively large quantity of sp3 bonds. The quantity of nanocrystalline graphite structures in the pure-ECR electrode decreases with increases in the nitrogen concentration due to the defects formation by the incorporation of nitrogen atoms in the graphite plane. In fact, nitrogen-containing graphite-like bonding and pyridine-like bonding structures greatly increases with increasing nitrogen concentration. Since the content of the former structure is very similar to that of sp2 bonds, the latter structure (pyridine-like) will help to reduce the sp2 bond content. It has been reported that nitrogencontaining carbon structures influence the electrocatalytic performance of carbon electrodes, and therefore, control of the nitrogen concentration could be very important in terms of obtaining better electrochemical performance. Potential Window and Basic Electrochemical Properties. We have studied the electrochemical properties of pureECR electrodes by changing the ion acceleration voltage. With a pure-ECR electrode, the potential window became wider with increasing sp3 bond content when we increased the acceleration voltage during sputtering. 12 Figure 2 compares cyclic voltammograms (CVs) of pure-ECR and N-ECR electrodes

Table 1. Surface Properties of Pure-ECR and N-ECR with Different Nitrogen Concentrationsa N concentration (at. %) C 1s (%) sp2 bonding sp3 bonding N 1s (%) graphite-like bonding pyridine-like bonding O/C Ra (nm)

0%

2.5 %

10 %

40 %

0 81.5 18.5 0 0 0.04 0.1

8.89 64.3 35.7 63.2 36.8 0.05 0.2

20.19 55.2 44.8 56.9 43.1 0.07 0.2

30.39 46.2 53.8 44.5 55.5 0.13 0.2

a

Chemical components of C, N, and O were obtained and analyzed from XPS analysis. Ra values were obtained from AFM measurements.

chemical bond and the average roughness (Ra) for our pureECR and N-ECR electrodes. Each chemical bond was estimated from high-resolution C 1s and N 1s XPS spectra as shown in Figure S1 because XPS is a more suitable approach for our films than Raman measurement.15 Unlike the results of Swain’s group, the sp2 content was 81.5% in the pure carbon film and decreased to 55.2% when the nitrogen concentration increased to 20.2 at. %. Figure S2 presents AFM images (3 μm × 3 μm) for the pure-ECR and N-ECR nanocarbon films. No clear surface differences could be observed. Indeed, the Ra value remained almost unchanged (between 0.1 to 0.2 nm) when the nitrogen concentration was increased to 30.4 at. %. Note that the roughness of our N-ECR electrodes did not change with the increasing nitrogen concentration, which is also different from the previous report.1

Figure 1. Plan views of pure-ECR and N-ECR (N = 9.0 at. %) observed by TEM. TEM images of films deposited at N2 gas content 0 and 2.5%. Scale bar = 5 nm. 9847

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Figure 2. Cyclic voltammograms of pure-ECR and N-ECR electrodes in 0.05 M H2SO4 deoxygenated with Ar.

with different nitrogen concentrations (N = 0, 9.0, 20.2, 30.4 at. %) obtained in a 0.05 M H2SO4 solution. The potential window is defined as the potential range between current limits that do not exceed 500 μA/cm2, as previously reported by Swain et al.19 The potential windows of the pure-ECR and N-ECR (N = 9.0 at. %) electrodes are 3.3 and 3.8 V, respectively. This increase in the potential window is due to the increase in the sp3 bond content caused by nitrogen doping. A similar potential window increase was observed with a pure-ECR electrode when we increased the sp3 bond concentration by changing the sputtering conditions.13 However, the potential windows of the N-ECR electrodes decreased from 3.8 to 3.3 V when the nitrogen concentration exceeded 9.0 at. %. We estimated the electrochemical properties of N-ECR electrodes by measuring typical redox species. Figure 3a shows CVs of Ru(NH3)62+/3+ obtained at the carbon electrode with different nitrogen concentrations. The peak separation (ΔEp) is almost constant at about 60 mV at nitrogen concentrations below 20.2 at. %, but it increases to 86 mV at a nitrogen concentration of 30.4 at. %. Since Ru(NH3)62+/3+ is classified as an outer-sphere redox couple and is relatively insensitive to the surface microstructure such as surface oxide and the monolayers adsorbed on the sp2 carbon electrode, the increased ΔEp at higher nitrogen concentrations is considered to indicate increased film resistance.20,21 This is because the sp2 bond ratio is only 46.2% in a film with a 30.4 at. % nitrogen concentration. It is reported that Fe(CN)63−/4− is an inner-sphere redox couple and that the k0 value is strongly influenced by several parameters including (1) the density of the electronic state near E0, (2) surface cleanliness, and (3) surface chemistry. Swain’s group reported that ΔEp decreased from 119 to 84 mV with increasing nitrogen concentration in the ta-C:N film electrodes. (Their film was formed at a 50 sccm nitrogen flow rate and contained 13% nitrogen.) With our N-ECR electrode, the ΔEp value was 167 mV for a pure-ECR electrode and decreased to 101 mV for a film with a 9.0 at. % nitrogen concentration, which is a similar tendency to that reported by Swain’s group except for the relatively higher ΔEp values. There are two possible reasons for this, as shown below. One is the greatly superior flatness of our electrode surface despite the nitrogen concentration being different from that of their films. The other reason could relate to the surface oxygen concentration. The O/C atomic ratio of the ta-C:N electrode is about 0.13, which is independent of the nitrogen concentration. In contrast, the O/C ratio of a pure-ECR and N-ECR electrode is a very low ratio (∼0.07) when the nitrogen concentration is below 20.2 at. %. Since an increase in the surface oxygen-containing groups often increases the electron transfer rates for some redox

Figure 3. (a) Cyclic voltammograms of pure-ECR and N-ECR electrodes in 1.0 mM Ru(NH3)62+/3+ in 1.0 M KCl. (b) Cyclic voltammograms of pure-ECR and N-ECR electrodes in 1.0 mM Fe(CN)63−/4− in 1.0 M KCl.

species, the lower surface oxygen concentration of the N-ECR carbon film electrode is suitable for studying the effect of nitrogen atoms in the film without interfering with other effects. When the nitrogen concentration was increased to 20.2 and 30.4 at. %, the ΔEp values greatly increased to 143 and 541 mV. The better electrode activity with a relatively low nitrogen concentration can be considered as follows in terms of the variation in the chemical structure of the electrodes. At an NECR electrode with a 9.0 at. % nitrogen concentration, the sp2 ratio and nitrogen-containing graphite-like bonding, which are considered to improve electrochemical activity, are both above 63%, but they then decrease significantly when the nitrogen concentration is increased, as shown in Table 1. Oxygen and H2O2 Reduction. Recently, several groups have reported that nitrogen-doped carbon materials exhibit an 9848

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overpotential decrease in the ORR.22 The well-known carbon alloys prepared by pyrolyzing nitrogen-containing aromatic polymers exhibit a large positive shift in the ORR potential compared with that of pure carbon materials.23 More recently, it was also reported that N-CNTs show excellent ORR performance.6 A positive ORR shift was also observed by using a chemical reaction to introduce a nitrogen-containing group onto the carbon material surfaces.24 Most of the previously reported nitrogen-containing carbons are bulk, flake, or fiber-like structures and are mainly designed as cathodic materials for fuel cells. In contrast, thin film carbon films have greater potential for chemical or biochemical sensor applications. This is because a flat surface with a decreased overpotential realizes a shaper response with a lower background noise level for electroactive molecules. Therefore, we primarily studied the ORR characteristics for N-ECR electrodes to clarify the effect of nitrogen at our carbon film electrode compared with that at a pure-ECR electrode. Figure 4 shows

which to study the mechanism of the ORR performance. For this, we must evaluate our developed N-ECR electrode in terms of the catalytic reduction of the oxygen process including the number of electrons by using a rotating disk electrode method. The electrochemical reduction of H2O2 is also very important for biosensor applications because various kinds of oxidase enzymes have been used to develop electrochemical biosensors. Figure 5a shows voltammograms of 0.2 mM H2O2

Figure 5. (a) Cyclic voltammograms of 0.2 mM H2O2 at pure-ECR and N-ECR (N = 9.0 at. %) electrodes measured in PB solution. Dotted lines are background scans. (b) Calibration lines of H2O2 at both electrodes. Concentration range is from 0.2 to 17 mM.

Figure 4. Cyclic voltammograms of pure-ECR and N-ECR (N = 9.0 at. %) electrodes for oxygen reduction reaction in O2 saturated 0.5 M H2SO4. Dotted lines are background scans.

obtained with pure-ECR and N-ECR (N = 9.0 at. %) electrodes. A very clear reduction peak was observed at −0.65 V for the N-ECR electrode. In contrast, only a broad peak was observed at −0.80 V for a pure-ECR electrode, indicating that the overpotential and sensitivity of the H2O2 reduction was also improved by doping nitrogen atoms without losing the low background noise level. Figure 5b compares the calibration curves of H2O2 at the two electrodes. Both electrodes show good linearity between 0.2 and 17 mM, and the current of the N-ECR electrode is 7 times larger than that of the pure-ECR electrode. The oxidation current of H2O2 has often been measured by using metal electrodes such as platinum, but the signal interfered with those of other biochemicals including L-ascorbic acid and uric acid. In contrast, the reduction of H2O2 will be useful without the effect of such interfering molecules, particularly for biological fluid samples. In addition, the obtained N-ECR electrode exhibited a low background current noise level similar to that of the pure-ECR electrode because the surface roughness was almost unchanged, as described above. Therefore, the effect of increasing the background current was negligible in our case, which is unlike other previous reports.26,27 Indeed, nitrogen doping also causes an increase in surface roughness that in turn increased the background current.26,27 Our N-ECR electrode is highly advantageous in terms of measuring lower concentrations of H2O2. Effect on Electrochemical Oxidation for Biomolecules. The interaction between analyte molecules and electrode

voltammograms of oxygen reduction at pure-ECR and N-ECR (N = 9.0 at. %) electrodes in O2 saturated 0.5 M H2SO4. The pure-ECR and N-ECR electrodes showed peak reduction currents at −0.8 and −0.5 V (versus Ag/AgCl), respectively. This indicates that the ORR overpotential was improved significantly by doping nitrogen atoms while maintaining a lower background noise level. However, further increases in the nitrogen concentration caused a negative shift in the ORR peak (0.2 V). Okamoto reported that an increase of sp2 carbons next to nitrogen atoms in the graphite structure improves oxygen adsorption on the carbon surface by first-principles molecular dynamics simulation.25 In our results, nitrogen-containing graphite-like bonding is at its maximum (63.2%) when the nitrogen concentration is 9.0 at. %, but it decreases to 56.9% at a nitrogen concentration of 20.2 at. %. These results agreed well with the simulated results reported by Okamoto. Although the positive shift in the ORR is insufficient compared with those of previous reports,4,5 we did not study the electrochemical properties of an N-ECR electrode with a nitrogen concentration of less than 9.0 at. %. Further improvement in the ORR performance could be expected in this nitrogen concentration region because we can expect a more ordered structure (or a higher graphite structure content) in carbon films with a lower nitrogen concentration. The structures of the carbon film electrodes are easier to analyze by controlling the parameters including the nitrogen concentration and sp3/sp2 ratio, and so carbon film electrodes will constitute a suitable material with 9849

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CONCLUSION In this paper, we studied the structure and electrochemical performance of an N-ECR nanocarbon film electrode and compared it with a ta-C:N electrode. The sp2 ratio decreased with increasing nitrogen concentration, whereas the results for ta-C:N were the opposite. The film surface remained very smooth and almost unchanged when the nitrogen concentration was increased. These differences with respect to the NECR electrode formed by ECR sputtering and PLD techniques could be due to the high ion irradiation of the ECR sputtering, which increases the atomic mobility on the substrate and rearranges the surface structure such as increasing the graphite crystalline content. The relatively flat surface despite the nitrogen concentration is also due to the effect of ion irradiation during sputtering. In spite of such difference in the structures, the electrochemical response of redox species such as Ru(NH3)62+/3+ and Fe(CN)63‑/4‑ at the N-ECR electrode exhibit a similar tendency to those of ta-C:N, suggesting that nitrogen doping improves the electron transfer at the electrode surface. We also studied the reduction of oxygen (ORR) and H2O2 and observed a significant positive shift in the reduction peaks could be observed at the N-ECR electrode compared with the pure-ECR electrode. These results are similar to the properties of carbon alloys and N-CNTs, although our results were not optimized. We also used the NECR electrode to measure DNA bases and observed a reduced overpotential and improved peak height without losing the low background noise level. This suggests a wide variety applications for detecting biomolecules.

surface could be different between pure-ECR and N-ECR electrodes. This is because surface functional groups on the nitrogen-containing carbon should be different from those of pure carbon, and the biomolecules also have polar groups such as amino and carboxyl groups. Therefore, we expected improved electrochemical responses even for biomolecules. We compare electrochemical responses of DNA bases at both carbon films since we previously studied the electrochemistry of such molecules and understood the kinetic parameters on ECR nanocarbon films.28,29 Figure 6 shows background-subtracted

Figure 6. Background-subtracted SWVs of 100 μM guanosine (dotted) and adenosine (solid) at pure-ECR and N-ECR (N = 9.0 at. %) electrodes measured in 50 mM acetate buffer (pH 5.0).

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SWVs of 0.1 mM of guanosine and adenosine obtained with pure-ECR and N-ECR (N = 9.0 at. %) electrodes. Sharp oxidation peaks of 1.08 and 1.46 V for guanosine and adenosine, respectively, were observed with the N-ECR electrode. In contrast, the shapes of both the guanosine and adenosine peaks were broader and lower with significant peak shifts about 0.1 V at the pure-ECR electrode. The most important point is that the N-ECR electrode exhibited larger currents at lower oxidation potential while maintaining the background current noise level (Figure S3), which is unlike other previous reports.26,27 The N-ECR electrode used for this measurement had a larger sp3 ratio (35.7%) than the pure-ECR electrode (18.5%), which is not advantageous with respect to increasing the electron transfer because the carbon electrode with a higher sp2 ratio has more edge planes. In fact, we previously reported that electrode activity in relation to DNA bases was dependent on the π−π interaction between aromatic DNA bases and the carbon surface.28,30 Moreover, we also reported that increasing the surface oxygen on the ECR electrode improved the electrochemical responses of the DNA bases.29 Given that the sp2 content was decreased and the surface oxygen was almost unchanged by nitrogen doping, which was unlike the case with the ECR electrode described above, this improved electrochemical response at the N-ECR electrode was presumably because nitrogen doping (including the change in surface wettability) allowed us to obtain a catalytic effect with respect to DNA base oxidation. Although we reported only two examples of the improved response of biomolecules using the N-ECR electrode, the application of our film electrodes to other analytes will be promising for achieving the highly sensitive measurement of some biologically important molecules.

ASSOCIATED CONTENT

S Supporting Information *

Additional information as noted in text. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Fax: +81-29-861-6177. Tel: +81-29861-6158. Author Contributions

All authors contributed to this manuscript and approved the final version. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was supported by a Grant-in-Aid for Challenging Exploratory Research (24655071). REFERENCES

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dx.doi.org/10.1021/ac402385q | Anal. Chem. 2013, 85, 9845−9851