Anal. Chem. 2003, 75, 6341-6345
Carbon Fiber Nanoelectrodes Modified by Single-Walled Carbon Nanotubes Rong-Sheng Chen, Wei-Hua Huang, Hua Tong, Zong-Li Wang, and Jie-Ke Cheng*
Department of Chemistry, Wuhan University, Wuhan 430072, China
Microelectrode voltammetry has been considered to be a powerful technique for single biological cell analysis and brain research. In this paper, we have developed a simple method to get highly sensitive carbon fiber nanoelectrodes (CFNE) modified by single-walled carbon nanotubes (SWNTs) on the basis of our previous work. The electrochemical behavior of SWNTs/CFNE was characterized by potassium ferricyanide, dopamine (DA), epinephrine (E), and norepinephrine (NE) using cyclic voltammetry (CV). Compared with CFNE, SWNTs/CFNE has a much larger available internal surface area per external geometric area, which is supported by SEM images. The modified electrodes show very high sensitivity and favorable electrochemical behavior toward these neurotransmitters. The peak current increases linearly with the concentration of DA, E, and NE in the range of 1.0 × 10-7-1.0 × 10-4, 3.0 × 10-7-1.0 × 10-4, and 5.0 × 10-7-1.0 × 10-4 M, respectively. The CV detection limit (S/N ) 3) of DA, E, and NE is 7.7 × 10-9, 3.8 × 10-8, and 4.2 × 10-8 M, respectively. The modified electrode exhibited almost the same electrochemical behavior after 15 days, indicating that SWNTs/CFNE is pretty stable and has good reproducibility. Microelectrodes have evoked increasing interest and have been studied comprehensively and intensively due to their intrinsic characteristics. Many chemical messengers that transport information in biological systems are electrochemically active molecules. Microelectrode voltammetry has been demonstrated as a powerful tool for in vivo and in vitro measurements, especially for single-cell analysis and brain research.1-7 The electroactive molecules include catecholamines,1-3,8-9 glucose,7,10-11 phenolic * To whom correspondence should be addressed. E-mail:
[email protected]. Tel: +086-27-87218954. Fax: +086-27-87647617. (1) Troyer, K. P.; Wightman, R. M. Anal. Chem. 2002, 74, 5370-5375. (2) Wightman, R. M.; Finnegan, J. M.; Pihel, K. Trends Anal. Chem. 1995, 14, 154-158. (3) Huang, L.; Kennedy, R. T. Trends Anal. Chem. 1995, 14, 158-164. (4) Malinski, K.; Taha, Z. Nature 1992, 358, 676-678. (5) Wightman, R. M.; May, L. J.; Michael, A. C. Anal. Chem. 1988, 60, 169A179A. (6) Shram, N. F.; Netchiporouk, L. I.; Martelet, C.; Jaffrezic-Renault, N.; Bonnet, C.; Cespuglio, R. Anal. Chem. 1998, 70, 2618-2622. (7) Netchiporouk, L. I.; Shram, N. F.; Jaffrezic-Renault, N.; Martelet, C.; Cespuglio, R. Anal. Chem. 1996, 68, 4358-4364. (8) Downard, A. J.; Roddick, A. D.; Bond, A. M. Anal. Chim. Acta 1995, 317, 303-310. (9) Cahill, P. S.; Walker, Q. D.; Finnegan, J. M.; Mickelson, G. E.; Travis, E. R.; Wightman, R. M. Anal. Chem. 1996, 68, 3180-3186. 10.1021/ac0340556 CCC: $25.00 Published on Web 10/11/2003
© 2003 American Chemical Society
antioxidants,12 heavy metals,13 hydrogen peroxide,14,15 lactate,6 amino acids,16 nitric oxide,4,17 and ascorbic acid.8,18 Microelectrodes have exhibited their impressive advantages in capillary electrophoresis (CE), especially in microchip electrophoresis.19-22 However, the sensitivity of electrochemical detection is not desirable enough in contrast with that of laser-induced fluorescence. Since the discovery of carbon nanotubes,23-25 they have attracted much attention because of their unique mechanical and electrical properties.26-28 There are two main types of carbon nanotubes that have high structural perfection. Single-walled carbon nanotubes (SWNTs) consist of a single graphite sheet seamlessly wrapped into a cylindrical tube. Multiwalled carbon nanotubes comprise an array of such nanotubes that are concentrically nested like rings of a tree trunk. Applications of the carbon nanotubes as an electrode material have been demonstrated,29,30 (10) McRipley, M. A.; Linsenmeier, R. A. J. Electroanal. Chem. 1996, 414, 235246. (11) Dixon, B. M.; Lowry, J. P.; O’Neill, R. D. J. Neurosci. Methods 2002, 119, 135-142. (12) Agui, M. L.; Reviejo, A. J.; Ya´nˇez-Sedenˇo, P.; Pingarro´n, J. M. Anal. Chem. 1995, 67, 2195-2200. (13) Sanna, G.; Pilo, M. I.; Piu, P. C.; Tapparo, A.; Seeber, R. Anal. Chim. Acta 2000, 415, 165-173. (14) Cso ¨regl, E.; Gorton, L.; Marko-Varga, G. Anal. Chem. 1994, 66, 36043610. (15) Dressman, S. F.; Garguilo, M. G.; Sullenberger, E. F.; Michael, A. C. J. Am. Chem. Soc. 1993, 115, 7541-7542. (16) Akhtar, P.; Too, C. O.; Wallace, G. G. Anal. Chim. Acta 1997, 339, 211223. (17) Kitamura, Y.; Uzawa, T.; Oka, K.; Komai, Y.; Ogawa, H.; Takizawa, N.; Kobayashi, H.; Tanishita, K. Anal. Chem. 2000, 72, 2957-2962. (18) Janda, P.; Weber, J.; Dunsch, L.; Lever, A. B. P. Anal. Chem. 1996, 68, 960-965. (19) Zeng, Y.; Chen, H.; Pang, D. W.; Wang, Z. L.; Cheng, J. K. Anal. Chem. 2002, 74, 2441-2445. (20) Martin, R. S.; Ratzlaff, K. L.; Huynh, B. H.; Lunte, S. M. Anal. Chem. 2002, 74, 1136-1143. (21) Baldwin, R. P.; Roussel, T. J., Jr.; Crain, M. M.; Bathlagunda, V.; Jackson, D. J.; Gullapalli, J.; Conklin, J. A.; Pai, R.; Naber, J. F.; Walsh, K. M.; Keynton, R. S. Anal. Chem. 2002, 74, 3690-3697. (22) Backofen, U.; Matysik, F. M.; Lunte, C. E. Anal. Chem. 2002, 74, 40544059. (23) Iijima, S. Nature 1991, 354, 56-58. (24) Ijjima, S.; Ichihashi, T. Nature 1993, 363, 603-605. (25) Bethune, D. S.; Kiang, C. H.; de Vries M. S.; Gorman, G.; Savoy, R. J.; Vazquez, J.; Beyers, R. Nature 1993, 363, 605-607. (26) Ajayan, P. M. Chem. Rev. 1999, 99, 1787-1799. (27) Odom, T. W.; Huang, J. L.; Kim, P.; Lieber, C. M. J. Phys. Chem. B 2000, 104, 2794-2809. (28) Baughrman, R. H.; Zakhidov, A. A.; Heer, W. A. Science 2002, 297, 787792. (29) Davis, J. J.; Coles, R. J.; Hill, H. A. O. J. Electroanal. Chem. 1997, 440, 279-282. (30) Joseph, K.; Compbell, J. J.; Sun, L.; Crooks, R. M. J. Am. Chem. Soc. 1999, 121, 3779-3780.
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Figure 2. Cyclic voltammograms at a CFNE (a) and on its SWNTmodified electrode (b-f) in a 25 mM Tris-HCl buffer solution (pH 7.4) at a scan rate of 100 mV/s between -0.2 and 0.5 V. (b) The first, (c) the second, (d) the third, and (e) the thousandth cycle, (f) after 15 days. CFNE tip diameter, 200 nm; CFNE length, 560 µm.
Figure 1. SEM images of (a) single-walled nanotubes, (b) the general view of a CFNE, (c) cross section of a CFNE, (d) cross section of a SWNTs/CFNE, and (e) tip of a SWNTs/CFNE (tip diameter, 200 nm).
but the voltammograms have not been satisfctory. Moreover, carbon nanotubes, generally with diameters in the range of 1-50 nm, are difficult to manipulate one by one. Electrodes modified by carbon nanotubes have shown well-resolved voltammograms and favorable electrocatalytic behavior toward the oxidation of some biomolecules.31-34 Because of their conventional dimensions, these modified electrodes are not suitable for measurements in ultrasmall sample volumes or as detectors in a CE system. The carbon fiber microelectrode is most widely used. In this paper, we have developed a simple method to prepare highly sensitive carbon fiber microelectrodes and nanoelectrodes (CFNE) modified by SWNTs on the basis of our previous work.35 Without (31) Luo, H. X.; Shi, Z. J.; Li, N. Q.; Gu, Z. N.; Zhuang, Q. K. Anal. Chem. 2001, 73, 915-920. (32) Wang, J. X.; Li, M. X.; Shi, Z. J.; Li, N. Q.; Gu, Z. N. Anal. Chem. 2002, 74, 1993-1997. (33) Musameh, M.; Wang, J.; Merkoci, A.; Lin, Y. Electrochem. Commun. 2002, 4, 743-746. (34) Wang, G.; Xu, J. J.; Chen, H. Y. Electrochem. Commun. 2002, 4, 506-509. (35) Huang, W. H.; Pang, D. W.; Tong, H.; Wang, Z. L.; Cheng, J. K. Anal. Chem. 2001, 73, 1048-1052.
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epoxy involved, the carbon fiber was flame fuse-sealed in the tip of the capillary glass. The tip of the carbon fiber is easily etched to form CFNE with tip dimensions ranging from 100 to 300 nm in diameter. The CFNE is suitable for the measurements of ultrasmall sample volumes, such as monitoring cell release with high spatial and temporal resolution, analysis of the insides of cells, direct measurements of the single exocytotic events, and probing into the synaptic cleft. Highly purified SWNTs were dispersed in aqueous sodium dodecyl sulfate (SDS) surfactant solution to obtain a stable suspension. The CFNE was immersed in the SWNT suspension and then dried to get a modified electrode. The SWNTs/CFNE was characterized by potassium ferricyanide, dopamine (DA), epinephrine (E), and norepinephrine (NE) using cyclic voltammetry. SWNTs/CFNE have showed very high sensitivity and favorable electrochemical behaviors toward these neurotransmitters. In comparison with CFNE, SWNTs/ CFNE has much larger available internal surface per external geometric area, which is supported by scanning electron microscopy (SEM) images. EXPERIMENTAL SECTION Instruments and Chemicals. Cyclic voltammograms were obtained using a CHI660A potentiostat (CH Instruments, Shanghai, China) in conjunction with a computer. A two-electrode system was employed, and a Ag/AgCl reference electrode was used throughout the experiment. All the potentials given in this paper are the potential versus Ag/AgCl. A scanning electron microscope (SEM X-650, Hitachi, Tokyo, Japan) was used for observation of the carbon nanotubes and electrodes. SWNTs were obtained by a direct current carbon arc discharge method36 and purified to get high-purity samples.37-41 DA, NE, (36) Ebbesen, T. W.; Ajayan, P. M. Nature 1992, 358, 220-222. (37) Tsang, S. C. Nature 1994, 372, 159-162. (38) Wang, Q.; Jiang, N.; Li, N. Q. Microchem. J. 2001, 68, 77-85. (39) Kang, T. F.; Shen, J. L.; Yu, R. Q. Anal. Chim. Acta 1997, 356, 245-251. (40) Zhao, H.; Zhang, Y. Z.; Yuan, Z. B. Analyst 2001, 126, 358-360. (41) Fiaccabrino, G. C.; Tang, X. M.; Skinner, N.; de Rooij, N. F.; Koudelka-Hep, M. Sens. Actuators, B 1996, 35, 247-254.
Figure 3. Cyclic voltammograms of (A) 1.5 mM potassium ferricyanide, (B) 0.1 mM DA, (C) 0.1 mM E, and (D) 0.1 mM NE at a CFNE (a) and its SWNT-modified electrode (b) in 25 mM Tris-HCl buffer solution (pH 7.4) at a scan rate of 10 mV/s. CFNE tip diameter, 150 nm; CFNE length, 450 µm.
and E were obtained from Sigma. All other reagents were of analytical grade. Stock solutions of the neurotransmitters (1 mM) were prepared by 25 mM Tris-HCl buffer (pH 7.4) and stored in a refrigerator. Stock solutions were diluted with 25 mM Tris-HCl buffer (pH 7.4) before use. All buffer solutions were prepared in doubly distilled, deionized water. Fabrication of CFNE and SWNTs/CFNE. The method to fabricate CFNE was previously described.35 A 0.9-mm-inner diameter glass capillary was pulled on the flame of the gas lamp to form a tip with ∼20-µm inner diameter. A cleaned carbon fiber (7 µm in diameter, Goodfellow Co., Oxford, U.K.) was connected to a copper wire with silver print conductive paint (GC Thorsen, 1801 Morgan St., Rockford, IL). The carbon fiber-copper wire was inserted from the other end of the glass capillary into the tip. A 1-cm length of the carbon fiber was exposed from the tip. The capillary tip was fused on the flame to seal the carbon fiber. The protruding carbon fiber was etched slowly on the bottom of the flame to get a desired length (usually 100-1000 µm) and a nanometer-scale (about 100-300 nm in diameter) tip. A total of 3 mg of purified SWNTs was dispersed with the aid of ultrasonic agitation in 10 mL of aqueous SDS surfactant solution (1 mg/mL) to obtain a black suspension. After the cyclic voltammograms at a CFNE were done, the CFNE was washed carefully by sonication for 5 min in acetone, alcohol, and doubly distilled water, respectively. The CFNE was then dried under an infrared lamp. The SWNT suspension was dropped on the tip of the CFNE, which was set on a clean planar glass, and then the protruding carbon fiber was immersed in SWNT suspension. The SWNTs/CFNE was prepared by evaporating the solution under an infrared lamp. The modified electrode was washed carefully
in doubly distilled water before use. All the electrodes are washed carefully with doubly distilled water between use in different solutions. RESULTS AND DISCUSSION SEM Characterization. The SEM images of SWNTs, CFNE, and SWNTs/CFNE are shown in Figure 1. Many SWNT bundles with diameters of ∼20 nm can be observed in Figure 1a. Some bundles twist together. The lengths of SWNTs are unmeasurable. The image shows the high purity of a SWNT sample (>95%). A smooth needle-shaped carbon fiber is shown in Figure 1b. The tip of CFNE was carefully cut to expose the cross section of the electrode (Figure 1c). The cross section of SWNTs/CFNE is shown in Figure 1d. In comparing the surface of a CFNE with that of a SWNTs/CFNE, it can be seen that the carbon fiber is wrapped with SWNT sheets (Figure 1c) while the surface of the CFNE is quite smooth (Figure 1d). It suggests that the modified electrode has much larger internal surface per external geometric area. The tip diameter of the SWNTs/CFNE is ∼200 nm (Figure 1e). Voltammetric Characteristics of SWNTs/CFNE. Figure 2 shows cyclic voltammograms of a CFNE and its SWNT-modified electrode in a 25 mM Tris-HCl buffer solution (pH 7.4) at a scan rate of 100 mV/s. The background current of the modified electrode is much larger than that of a bare carbon fiber electrode. This might be due to the large accessible surface of the modified electrode. After the electrochemical experiment, the modified electrode was removed from the solution, rinsed in water, and exposed to the air for 15 days, and then the same experiment was performed again. The first, the second, the third, and the Analytical Chemistry, Vol. 75, No. 22, November 15, 2003
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Figure 4. Cyclic voltammograms at a CFNE (B) and its SWNT-modified electrode (A) of 0.1 mM DA in 25 mM Tris-HCl buffer (pH 7.4) at scan rates of 0.01, 0.02, 0.05, and 0.1 V/s between -0.2 and +0.5 V. CFNE tip diameter, 150 nm; CFNE length, 450 µm.
Figure 5. Cyclic voltammograms of DA at a SWNTs/CFNE in 25 mM Tris-HCl buffer (pH 7.4) at a scan rate of 100 mV/s. DA concentration (nM): a, 0; b, 50; c, 500; d, 1000. CFNE tip diameter, 300 nm; CFNE length, 420 µm.
thousandth cyclic voltammograms and the cyclic voltammogram after 15 days are shown in Figure 2. It indicates that SWNTs/ CFNE is quite stable and has good reproducibility. Figure 3 shows the cyclic voltammograms of potassium ferricyanide, DA, E, and NE at a CFNE and its SWNT-modified electrode in 25 mM Tris-HCl buffer solution (pH 7.4) at a scan rate of 10 mV/s. Both electrodes exhibit quasi-steady-state behaviors. The cyclic volatmmograms take on a sigmoidal shape, and the currents are diffusion limited. The limiting currents at the SWNTs/CFNE are almost doubled in comparison with that of the CFNE, because of larger geometry and surface area of the SWNTs/CFNE. The increasing geometry and surface area should be due to SWNT sheets coated on the carbon fiber. Figure 4 shows the cyclic voltammograms of 0.1 mM dopamine at a CFNE and its SWNTs/CFNE at different scan rates. At such scan rates, the modified electrode exhibits different CV behaviors in comparison with that of CFNE, at which the currents rely little on the scan rates. It is known that the shape of cyclic voltammograms at microelectrodes relate to scan rates. The cyclic voltammograms at SWNTs/CFNE become peaked as the scan rates over 20 mV/s, while cyclic voltammograms at CFNE are a sigmoidal shape at these scan rates. CFNE is too small to deplete electroactive molecules in the surrounding solution at such scan rates, in contrast to SWNTs/CFNE. The voltammetric peak separation is ∼50 mV at SWNTs/CFNE, which is apparently affected by coupled surface interactions of DA. In epinephrine and 6344
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Figure 6. Relationship of ipa and c of DA (A), E (B), and NE(C) in 25 mM Tris-HCl buffer (pH 7.4) at a scan rate of 100 mV/s between -0.2 and 0.5 V on a CFNE modified by SWNTs. CFNE tip diameter, 300 nm; CFNE length, 420 µm.
norepinephrine solutions, similar electrochemical behaviors are exhibited at SWNTs/CFNE. Detection Limits of the Neurotransmitters at SWNTs/ CFNE. The modified electrode shows very high sensitivity toward neurotransmitters such as DA, E, and NE. Cyclic voltammograms of 1 µM, 0.5 µM, and 50 nM DA at a scan rate of 100 mV/s at
Table 1. Comparison of CV Detection Limits for DA electrodes fibera
carbon carbon fiberb Au diskc glassy carbon electroded glassy carbon electrodee microelectrode arraysf
detection limits (M) 10-8
7.6 × 7.7 × 10-9 3.0 × 10-6 9.0 × 10-8 3.0 × 10-8 5.0 × 10-8
ref 35 this work 38 39 40 41
a Carbon fiber nanoelectrode. b SWNT-modified carbon fiber nanoelectrode. c The thiolactic acid self-assembled monolayer-modified gold electrode. dElectropolymerized tetraaminophthalocyanatonickle(II) (pNiTAPc) film-coated electrode. e Poly(2-picolinic acid) chemically modified electrode. f Interdigitated microelectrode arrays based on sputtered carbon thin films.
SWNTs/CFNE are shown in Figure 5. The anodic currents increase linearly with the concentration of DA, E, and NE in the range of 1.0 × 10-7-1.0 × 10-4, 3.0 × 10-7-1.0 × 10-4, and 5.0 × 10-7-1.0 × 10-4 M, respectively. The relationship of ipa on the SWNT-modified CFNE and concentration of DA, E, and NE is shown in Figure 6. The linear regression equations of DA, E, and NE are expressed as Ip/nA ) 0.3923 + 0.7184c/µM (correlation coefficient r ) 0.9985), Ip/nA ) 0.8051 + 0.8461c/µM (r ) 0.9989), and Ip/nA ) -0.1216 + 1.1483c/µM (r ) 0.9996), respectively. The CV detection limits (S/N ) 3) of DA, E, and NE are 7.7 × 10-9, 3.8 × 10-8, and 4.2 × 10-8 M, respectively. As compared to CV detection limits of the previous works for DA, SWNTs/CFNE exhibits a higher sensitivity (Table 1). This should be attributed
to the large available surface area of the modified electrode and excellent electrical properties of SWNTs. CONCLUSION In this work, we have developed a simple method to fabricate SWNTs/CFNE. This nanometer-scale electrode displays excellent electrochemical characters and shows very high sensitivity toward neurotransmitters such as dopamine, epinephrine, and norepinephrine. The modified electrode is quite stable and has good reproducibility. This method offers a simple, economical, and convenient way to obtain high-quality detectors for bioelectroanalysis, capillary electrophoresis, miniaturized total analysis systems, and other fields. Owing to the nanometer scale, CFNE is suitable for direct monitoring the secretion from a single cell with high spatial and temporal resolution. However, some components in a single cell are too difficult to be detected. SWNTs/CFNE would be likely to solve the problem. The study is in progress in our laboratory. ACKNOWLEDGMENT This work was supported by the National Natural Science Foundation of China (20299034).
Received for review January 20, 2003. Accepted August 6, 2003. AC0340556
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