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Highly sensitive electrogenerated chemiluminescence produced at Ru(bpy)32+ in Eastman-AQ55D-carbon nanotube composite film electrode
Journal of
Electroanalytical Chemistry Journal of Electroanalytical Chemistry 592 (2006) 63–67 www.elsevier.com/locate/jelechem
Highly sensitive electrogenerated chemiluminescence produced at RuðbpyÞ2þ 3 in Eastman-AQ55D-carbon nanotube composite film electrode Lihua Zhang, Zhihui Guo, Zhiai Xu, Shaojun Dong
*
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Graduate School of the Chinese Academy of Science, Changchun 130022, China Received 13 August 2005; received in revised form 7 March 2006; accepted 7 April 2006 Available online 24 May 2006
Abstract The electrochemistry and electrogenerated chemiluminescence (ECL) of tris(2,2 0 -bipyridyl)ruthenium(II) ion-exchanged in EastmanAQ-carbon nanotube (CNT) composite films were investigated at a glassy carbon (GC) electrode. Eastman-AQ55D is a poly (ester sulfonic acid) cation exchanger available in a commercial dissolved form. It is much more hydrophilic than Nafion due to its unique structure, so RuðbpyÞ2þ 3 does not diffuse into the hydrophobic region where it may lose its electroactivity as that in Nafion. The interfused CNT could act as electronic wires that connect the electrode with RuðbpyÞ2þ 3 , which made the composite film much more electronically 2þ conductive. Besides, the negatively charged CNT could also absorb some RuðbpyÞ2þ 3 which finally led to the increasing of RuðbpyÞ3 . 2þ Moreover, the strong electrostatic interaction between AQ and RuðbpyÞ3 made the composite films much more stable. The combination of AQ and CNT brings excellent sensitivity with the detection limit as low as 3 · 1011 M for TPA. 2006 Elsevier B.V. All rights reserved. Keywords: Electrogenerated chemiluminescence; Tris(2,2 0 -bipyridyl)ruthenium(II); Eastman-AQ-carbon nanotube composite film
1. Introduction Electrogenerated chemiluminescence (ECL) is the emission from an excited molecule generated by an electrochemical redox reaction [1]. Many chemiluminescent regents were applied in ECL reactions, such as luminol, polyaromatic hydrocarbons (PAH) and RuðbpyÞ2þ [2]. 3 Among them, RuðbpyÞ2þ -based ECL attracts more atten3 tion due to its sensitive and selective detection method in analytic science. Besides, the oxidation–reduction reaction 2þ mechanism for RuðbpyÞ3 ECL system postulated by 2þ Rubinstein and Bard reveals that RuðbpyÞ3 could be regenerated on the electrode surface during the ECL reac2þ tion process [3]. So the immobilization of RuðbpyÞ3 on the surface of the electrode exhibits a lot of advantages, such as reducing the consumption of reagents and avoiding the use *
Corresponding author. Tel.: +86 431 5262101; fax: +86 431 5689711. E-mail address: [email protected] (S. Dong).
0022-0728/$ - see front matter 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.jelechem.2006.04.003
of an extra pump to deliver the reagents to the electrochemical cell in the case of detection under flowing condition. Recently many methods have been developed to immo2þ bilize RuðbpyÞ3 on electrode surface to fabricate a regenerable ECL sensor. For example, the derivatives of 2þ RuðbpyÞ3 have been immobilized on an electrode surface as Langmiur–Blodgett films and self-assembled monolayers [4,5]. However, because of their instability in positively biased potential and desorption in organic solvent, these methods can not be applied widely [6]. Another method 2þ is the immobilization of RuðbpyÞ3 in sol–gel [7], for exam2þ ple RuðbpyÞ3 was immobilized in titania sol–gel membranes [8], silica thin films [9], or PSS-silica-Triton X-100 2þ composite films [10]. But RuðbpyÞ3 is likely to leak out of the film due to the porous structure of the sol–gel. Nafion, a cation exchange polymer, was also used as a proper 2þ support for immobilizing RuðbpyÞ3 [11]. But the diffusion of electroactive cations into Nafion is relatively slow and the compact Nafion film is unfavorable for analyte to
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diffuse in. Therefore, alternative matrix of composite film consisting Nafion and other substances were reported to improve the sensitivity and long-term stability of ECLbased sensor. For example, both Nafion–silica [12] and TiO2–Nafion composite films [6] are more porous than pure Nafion, which led to the faster diffusion of analyte into the film and quickened the charge transportation. Moreover, the incorporation of CNT into the Nafion made the composite film more electronically conductive [13]. These ECL sensors showed faster response, higher sensitivity and better stability. However, the structure of Nafion 2þ made the RuðbpyÞ3 likely partition into more hydrophobic region, thus resulting in charge transport difficult within the film [14,15]. Although the properties of the reported ECL sensors based on composite film have been improved, new materials and immobilization methods are still needed in order to promote both sensitivity and long-term stability of ECL-based sensor. What we report here is an alternative matrix of composite film consisting Eastman-AQ polymers (AQ) and CNT 2þ for immobilization of RuðbpyÞ3 on a glassy carbon (GC) electrode surface. Eastman-AQ55D is a poly (ester sulfonic acid) cation exchanger available in a commercial dissolved form. Films of this polymer exhibit attractive permselectivity, ion-exchange, and antifouling properties. Although the complete structure of AQ55D is unclear right now, the proposed backbone is as follows [16]: O O C
O C
H2 O C
H2 C
2. Experimental 2.1. Reagents Ru(bpy)3Cl2 Æ 6H2O, tripropylamine (TPA, 99%) and Eastman-AQ55D (28% dispersion; AQ55D) were purchased from Aldrich and used as received. The multi-wall carbon nanotubes with 80% purity were purchased from Shenzhen Nanotech. Port. Co. Ltd. China and they were around 1.4 nm in diameter and 1–10 lm in length. It was purified and functionalized by the published procedure [23]. Under such circumstance, the CNT was negatively charged. All of other reagents were of analytical grade. All aqueous solutions were prepared with distilled water. 2.2. Apparatus Cyclic voltammetric experiments were performed with a CH Instruments 600 voltammetric analyzer. All experiments were carried out with a conventional three-electrode system. The working electrode was glassy carbon coated with CNT–AQ composite film. A platinum wire was the counter electrode, and an Ag/AgCl (saturated KCl) worked as a reference electrode. All the potential were measured and reported according to this reference electrode. The ECL signal produced in the electrolytic cell was detected and recorded by a flow injection chemiluminesH2 C
O
x
O O C
H2 C
O C z
y SO 3 Na + AQ55D
AQ has relatively high molecular weight amorphous polyesters with sulfonic groups on aromatic dicarboxylic acid unit, it is much more hydrophilic than Nafion [17]. 2þ Thus RuðbpyÞ3 immobilized in AQ polymer is prevented from partitioning into hydrophobic region, which could improve the stability of ECL sensor. With their low cost, strong adherence to the surface and rapid response [18], AQ polymers appear to be attractive material for sensor development. Moreover, CNT represents a considerably significant group of nanomaterial with unique geometrical, mechanical, electronic and chemical properties [19,20]. High sensitivity and stability could be obtained when CNT was used to modify the electrode. The improved sensitivity was attributed to the electrocatalytic activity of CNT and the interfacial accumulation onto its surface, while the better stability was attributed to the minimization of surface fouling at CNT surface [21]. Therefore, the interfusion of negatively charged CNT into the AQ increases not only the amount of the RuðbpyÞ2þ 3 but also the electronic conductivity of the composite film [22]. In general, the AQ–CNT composite-modified electrode exhibits greatly improved stability and sensitivity.
cence analyzer (IFFD, xi’an Remax electronic Science Tech. Co. Ltd. China), which was operated by a personal computer. ECL intensities were measured through the bottom of the cell with PMT window and all of them were enclosed in a light-tight box. The photomultiplier tube (PMT) was operated in current mode. Unless noted, otherwise, the PMT was biased at 600 V. 2.3. Preparation of ECL sensor Certain amount of CNT was dispersed in acetone solution. Then mixed it with AQ55D-acetone solution. The mixture was ultrasonicated for about 20 min until a homogeneous solution of CNT–AQ–acetone complex was obtained and the concentration of the CNT in this solution is about 0.5 mg/ml. The GC electrode was polished with 0.3 and 0.05 lm aluminum slurry, respectively, rinsed thoroughly with redistilled water, and ultrasonicated in redistilled water for 1 min, allowed to dry in room temperature. About 1.5 ll composite was hand-cast on the surface of a GC electrode and the film was dried for 1 h at room temperature. The modified GC electrodes were
L. Zhang et al. / Journal of Electroanalytical Chemistry 592 (2006) 63–67
then placed in 1 mM RuðbpyÞ2þ aqueous solution for 3 2þ about 30 min to incorporate RuðbpyÞ3 . With chronocoul2þ ometry method, the surface coverage of RuðbpyÞ3 ðC Þ of the AQ–CNT composite film was estimated to be 8.5 · 108 mol/cm2. 3. Results and discussion 3.1. Electrochemistry and ECL of immobilized RuðbpyÞ2þ 3 2þ
The electrochemical behavior of RuðbpyÞ3 immobilized in AQ–CNT composite film electrode has been investigated using cyclic voltammetry to provide information about reagent electroactivity and stability. AQ is an effective ion exchanger, so RuðbpyÞ2þ 3 could be easily incorporate into the composite film through ion-exchange process and electrostatic adsorption. Fig. 1 shows the cyclic voltammo2þ grams (CVs) of 0.5 mM RuðbpyÞ3 in phosphate buffer (pH 7.6) at a pure AQ film electrode and AQ–CNT composite film electrode with the scan rate of 100 mV/s. The CVs were recorded after reaching steady state. In general, the two CVs are similar in shape, but they are much different in current intensity. At the pure AQ film electrode, only a weak peak due to the oxidation of RuðbpyÞ2þ 3 in solution can be observed. Whereas a distinguishable response occurs at the AQ–CNT composite film electrode. Its oxidation peak current is much higher than that at AQ film electrode. This could be attributed to the structure of the composite film. AQ is a good medium for preconcentration 2þ of RuðbpyÞ3 . Negatively charged CNT could also absorb 2þ some RuðbpyÞ2þ 3 . Moreover, the oxidation of RuðbpyÞ3 was shifted negatively about 33 mV at AQ–CNT composite electrode compared with the pure AQ electrode. That is because the existence of CNT increases the electronic conductivity of the composite film, which quickens the charge transportation and decreases iR effects.
Fig. 1. Cyclic voltammograms (CVs) of 0.5 mM RuðbpyÞ2þ 3 in phosphate buffer (pH 7.6) with the scan rate of 100 mV/s, at a pure AQ modified electrode (dashed line) and AQ–CNT modified electrode (solid line).
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incorporated in the The ECL behavior of RuðbpyÞ2þ 3 AQ–CNT composite film electrode has been studied with tripropylamine (TPA) as a representative analyte, since 2þ the RuðbpyÞ3 –TPA system has been fully investigated and given stronger ECL signal than other reductants [24]. 2þ Fig. 2 exhibits CVs of RuðbpyÞ3 immobilized in AQ– CNT composite film electrode in the absence and presence of 3 · 109 M TPA at the scan rate of 100 mV/s in phosphate buffer solution (pH 7.6). In the presence of the 2þ TPA, the oxidation current of RuðbpyÞ3 increases clearly while the reduction current decreases. Meanwhile, the ECL signal increases considerately in the presence of TPA. The 2þ ECL behavior of immobilized RuðbpyÞ3 on AQ–CNT composite film electrode is illustrated in Fig. 3. The onset of luminescence occurs near 1 V, and then the ECL intensity rises steeply until it reaches a maximum near 1.15 V, which is consistent with the oxidation potential of 2þ RuðbpyÞ3 . This means that the oxidation of immobilized 2þ RuðbpyÞ3 plays a key role in the process of ECL. ECL intensity could be as high as about 1330 a.u. when the concentration of TPA is only 3 · 109 M. This could be attributed to the electronic conductivity of CNT and the ionexchange ability of Eastman-AQ55D. All of these results indicate that an energetic electron-transfer reaction occurs between electrogenerated RuðbpyÞ3þ 3 and the strong reducing intermediate, the deprotonated form of TPA (TPA). The ECL reaction of the system can be expressed as the follow equations [24]: 3þ RuðbpyÞ2þ 3 ! RuðbpyÞ3 þ e 3þ RuðbpyÞ3
þ TPA ! TPA ! TPAþ þ e
2þ RuðbpyÞ3
ð1Þ þ TPA
þ
ð2aÞ ð2bÞ
Fig. 2. Cyclic voltammograms of RuðbpyÞ2þ immobilized in AQ–CNT 3 composite-modified electrode in the absence (dashed line) and presence (solid line) of 3 · 109 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
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Fig. 3. Corresponding ECL–potential curves for AQ–CNT composite film in phosphate buffer solution (pH 7.6) containing 3 · 109 M TPA at the scan rate of 10 mV/s.
TPAþ ! TPA þ Hþ
ð3Þ
3þ 2þ RuðbpyÞ3 þ TPA ! RuðbpyÞ3 2þ 2þ RuðbpyÞ3 ! RuðbpyÞ3 þ hm
ð4Þ ð5Þ
We investigated the effect of the amount of CNT interfused into the AQ film. Fig. 4. showed the ECL response varied with the amount of CNT in the presence of 107 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s. The ECL response increased as the amount of CNT in AQ film increased up to 0.5 mg/ ml, which may due to the increased adsorption of 2þ RuðbpyÞ3 onto negatively charged CNT surface. However, as the amount of CNT increased further, the ECL response decreased gradually. The CNT could not dispersed evenly in the composite films if the amount of
Fig. 4. Effect of the CNT amount in the AQ film on ECL intensity in the presence of 107 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
CNT was more than 0.5 mg/ml, which finally led to poor film fabrication. Therefore, an optimum amount of CNT at 0.5 mg/ml was used in the subsequent experiments. A pH study has been carried out to examine the pH 2þ effect on the RuðbpyÞ3 –TPA system. The ECL signals for TPA obtained at the AQ–CNT composite-modified electrode increases from 5.5 to 7.6 considerably and then decreases rapidly at higher pH. This shows the similar result as that of the Nafion–CNT composite films [13] and AQ–silica composite film [25]. The ECL signal increases from 5.5 to 7.6, implying that deprotonation of TPA is needed as demonstrated in Eq. (3). While as the pH increasing continuously, some decomposition of species would be expected, leading to a diminished ECL reagent available for ECL reaction. Therefore, the ECL intensity decreases [26]. The scan rate effect on the ECL intensity is also investigated. The ECL intensity decreases with the increasing of scan rate in the range from 30 to 200 mV s1. The ECL signal changes with the scan rate depended on two factors: the chemical kinetics of the system and the rate of TPA diffusion in the film. Because the ECL signal increases as the scan rate decreasing, the chemical kinetics of the system plays a major role in this process. In order to evaluate the sensitivity of the composite film modified electrode, TPA is used as the probes. Calibration curve for TPA is demonstrated in Fig. 5. The result shows that the ECL signal varies linearly with the concentration of TPA in two ranges with different slopes. The first one is from 1010 to 107 M with the slope of 0.01 and the correlation coefficient of 0.9986. The second one is from 107 to 105 M with the slope of 0.031 and the correlation coefficient of 0.9982. To our knowledge, among many methods to immobilize RuðbpyÞ2þ onto the electrode surface, the 3 lowest detection limit has been 109 M for TPA occurred at Nafion-CNT composite film electrode so far. While the
Fig. 5. Calibration of TPA at the RuðbpyÞ2þ immobilized AQ–CNT 3 composite film electrode in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
L. Zhang et al. / Journal of Electroanalytical Chemistry 592 (2006) 63–67
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into the solution. Furthermore, according to the structure 2þ of AQ, it is more hydrophilic than Nafion, so RuðbpyÞ3 is strongly concentrated in the whole AQ film by ionexchange and keeps its electroactivity. 4. Conclusion
Fig. 6. ECL emission of immobilized RuðbpyÞ2þ 3 in AQ–CNT composite film electrode in phosphate buffer solution (pH 7.6) containing 3 · 109 M TPA at the scan rate of 100 mV/s.
AQ–CNT composite films have proven to be an effective matrix for the immobilization of RuðbpyÞ2þ at electrode 3 surface to prepare an ECL sensor. Because of the special character of the AQ and CNT, this sensor shows excellent sensitivity with detection limit of 3 · 1011 M for TPA. Moreover, since the AQ is a proper support for some enzymes, the AQ–CNT composite film can be used as a good matrix for coimmobilization of RuðbpyÞ2þ 3 and some enzymes(such as alcohol dehydrogenase) to fabricate a highly sensitive RuðbpyÞ2þ 3 ECL based biosensor. Acknowledgement
detection limit from the present experiments is 3 · 1011 M for TPA, which is two orders of magnitude lower than that of Nafion–CNT film. The excellent sensitivity could be explained as follows: firstly, the presence of CNT makes the composite film much more electronically conductive, which is advantageous for electron transfer. Besides, the negatively charged CNT could also absorb some 2þ RuðbpyÞ3 through electrostatical interaction. Secondly, AQ is much more hydrophilic than Nafion, so RuðbpyÞ2þ 3 does not diffuse into the hydrophobic region where it may lose its electroactivity as that in Nafion. The long-term stability of the AQ–CNT composite film 2þ electrode immobilized with RuðbpyÞ3 was investigated over two weeks. The composite film electrode was stored in dry state at room temperature and monitored by cyclic voltammetry measurements in a phosphate buffer (pH 7.6) periodically. The peak potential was essentially unchanged during this period, and the oxidation peak current decreased less than 5% compared with the initial steady state value after two weeks of storage. Moreover, when the CV measurements were run continuously for 100 cycles in phosphate buffer (pH 7.6) at the scan rate of 100 mV/s, the ECL signal could decrease about 10.8%. 2þ Fig. 6 shows ECL emission of immobilized RuðbpyÞ3 in AQ–CNT composite film electrode in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s containing 3 · 109 M TPA for 10 continuous cycles. Peak intensity displays good reproducibility with the relative standard 2þ deviation less than 1%. The good stability of RuðbpyÞ3 in the composite film may arise from that the 2þ RuðbpyÞ3 is a quite stable CL regent, and the ion2þ exchange selectivity of AQ for RuðbpyÞ3 is so high that 2þ it could prevent immobilized RuðbpyÞ3 from leaching
This work was supported by National Natural Science Foundation of China (20275036, 20275037). References [1] L.R. Faulkner, A.J. Bard, in: A.J. Bard (Ed.), Electroanalytical chemistry, vol. 10, Marcel Dekker, NewYork, 1997, p. 1. [2] K.A. Fa¨hnrich, M. Pravda, G.G. Guilbault, Talanta 54 (2001) 531. [3] I. Rubinstein, A.J. Bard, J. Am. Chem. Soc 103 (1981) 512. [4] C.J. Miller, P. Mccord, A.J. Bard, Langmuir 7 (1991) 2781. [5] Y.S. Obeng, A.J. Bard, Langmuir 7 (1991) 195. [6] H.N. Choi, S.H. Cho, W.Y. Lee, Anal. Chem. 75 (2003) 4250. [7] M.M. Collinson, B. Novak, S.A. Martin, J.S. Taussig, Anal. Chem. 72 (2000) 2914. [8] Y.F. Zhuang, H.X. Ju, Electroanalysis 16 (2004) 1401. [9] L. Armelao, R. Bertoncello, S. Gross, D. Badocco, P. Pastore, Electroanalysis 15 (2003) 803. [10] H.Y. Wang, G.B. Xu, S.J. Dong, Analyst 126 (2001) 1095. [11] I. Rubinstein, A.J. Bard, J. Am. Chem. Soc 102 (1980) 6641. [12] A.N. Khramov, M.M. Collinson, Anal. Chem. 72 (2000) 2943. [13] Z.H. Guo, S.J. Dong, Anal. Chem. 76 (2004) 2683. [14] T.M. Downey, T.A. Nieman, Anal. Chem. 64 (1992) 261. [15] W.Y. Lee, T.A. Nieman, Anal. Chem. 67 (1995) 1789. [16] S.J. Dong, G.L. Che, Y.W. Xie (Eds.), Chemically Modified Electrode, Science Press, Beijing, China, 2003. [17] A.F. Martin, T.A. Nieman, Biosens. Bioelectrotron. 12 (1997) 479. [18] J. Wang, T. Golden, Anal. Chem. 61 (1989) 1397. [19] C.N. Rao, B.C. Satishkumar, A. Govindaraj, M. Nath, ChemPhyChem 2 (2001) 79. [20] R.H. Baughman, A. Zakhidov, W.A. de Heer, Science 297 (2002) 787. [21] J. Wang, G. Chen, M.P. Chatrathi, M. Musameh, Anal. Chem. 76 (2004) 298. [22] P.M. Ajayan, Chem. Rev. 99 (1999) 1787. [23] Z. Shi, Y. Lian, X. Zhou, Z. Gu, Y. Zhang, S. Lijima, H. Li, K.T. Yue, S.L. Zhang, J. Phy. Chem. B. 103 (1999) 8698. [24] J.B. Noffsinger, N.D. Danielson, Anal. Chem. 59 (1987) 865. [25] H.Y. Wang, G.B. Xu, S.J. Dong, Anal. Chim. Acta 480 (2003) 285. [26] J.K. Leland, M.J. Powell, J. Electrochem. Soc. 137 (1990) 3127.
Electroanalytical Chemistry Journal of Electroanalytical Chemistry 592 (2006) 63–67 www.elsevier.com/locate/jelechem
Highly sensitive electrogenerated chemiluminescence produced at RuðbpyÞ2þ 3 in Eastman-AQ55D-carbon nanotube composite film electrode Lihua Zhang, Zhihui Guo, Zhiai Xu, Shaojun Dong
*
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Graduate School of the Chinese Academy of Science, Changchun 130022, China Received 13 August 2005; received in revised form 7 March 2006; accepted 7 April 2006 Available online 24 May 2006
Abstract The electrochemistry and electrogenerated chemiluminescence (ECL) of tris(2,2 0 -bipyridyl)ruthenium(II) ion-exchanged in EastmanAQ-carbon nanotube (CNT) composite films were investigated at a glassy carbon (GC) electrode. Eastman-AQ55D is a poly (ester sulfonic acid) cation exchanger available in a commercial dissolved form. It is much more hydrophilic than Nafion due to its unique structure, so RuðbpyÞ2þ 3 does not diffuse into the hydrophobic region where it may lose its electroactivity as that in Nafion. The interfused CNT could act as electronic wires that connect the electrode with RuðbpyÞ2þ 3 , which made the composite film much more electronically 2þ conductive. Besides, the negatively charged CNT could also absorb some RuðbpyÞ2þ 3 which finally led to the increasing of RuðbpyÞ3 . 2þ Moreover, the strong electrostatic interaction between AQ and RuðbpyÞ3 made the composite films much more stable. The combination of AQ and CNT brings excellent sensitivity with the detection limit as low as 3 · 1011 M for TPA. 2006 Elsevier B.V. All rights reserved. Keywords: Electrogenerated chemiluminescence; Tris(2,2 0 -bipyridyl)ruthenium(II); Eastman-AQ-carbon nanotube composite film
1. Introduction Electrogenerated chemiluminescence (ECL) is the emission from an excited molecule generated by an electrochemical redox reaction [1]. Many chemiluminescent regents were applied in ECL reactions, such as luminol, polyaromatic hydrocarbons (PAH) and RuðbpyÞ2þ [2]. 3 Among them, RuðbpyÞ2þ -based ECL attracts more atten3 tion due to its sensitive and selective detection method in analytic science. Besides, the oxidation–reduction reaction 2þ mechanism for RuðbpyÞ3 ECL system postulated by 2þ Rubinstein and Bard reveals that RuðbpyÞ3 could be regenerated on the electrode surface during the ECL reac2þ tion process [3]. So the immobilization of RuðbpyÞ3 on the surface of the electrode exhibits a lot of advantages, such as reducing the consumption of reagents and avoiding the use *
Corresponding author. Tel.: +86 431 5262101; fax: +86 431 5689711. E-mail address: [email protected] (S. Dong).
0022-0728/$ - see front matter 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.jelechem.2006.04.003
of an extra pump to deliver the reagents to the electrochemical cell in the case of detection under flowing condition. Recently many methods have been developed to immo2þ bilize RuðbpyÞ3 on electrode surface to fabricate a regenerable ECL sensor. For example, the derivatives of 2þ RuðbpyÞ3 have been immobilized on an electrode surface as Langmiur–Blodgett films and self-assembled monolayers [4,5]. However, because of their instability in positively biased potential and desorption in organic solvent, these methods can not be applied widely [6]. Another method 2þ is the immobilization of RuðbpyÞ3 in sol–gel [7], for exam2þ ple RuðbpyÞ3 was immobilized in titania sol–gel membranes [8], silica thin films [9], or PSS-silica-Triton X-100 2þ composite films [10]. But RuðbpyÞ3 is likely to leak out of the film due to the porous structure of the sol–gel. Nafion, a cation exchange polymer, was also used as a proper 2þ support for immobilizing RuðbpyÞ3 [11]. But the diffusion of electroactive cations into Nafion is relatively slow and the compact Nafion film is unfavorable for analyte to
64
L. Zhang et al. / Journal of Electroanalytical Chemistry 592 (2006) 63–67
diffuse in. Therefore, alternative matrix of composite film consisting Nafion and other substances were reported to improve the sensitivity and long-term stability of ECLbased sensor. For example, both Nafion–silica [12] and TiO2–Nafion composite films [6] are more porous than pure Nafion, which led to the faster diffusion of analyte into the film and quickened the charge transportation. Moreover, the incorporation of CNT into the Nafion made the composite film more electronically conductive [13]. These ECL sensors showed faster response, higher sensitivity and better stability. However, the structure of Nafion 2þ made the RuðbpyÞ3 likely partition into more hydrophobic region, thus resulting in charge transport difficult within the film [14,15]. Although the properties of the reported ECL sensors based on composite film have been improved, new materials and immobilization methods are still needed in order to promote both sensitivity and long-term stability of ECL-based sensor. What we report here is an alternative matrix of composite film consisting Eastman-AQ polymers (AQ) and CNT 2þ for immobilization of RuðbpyÞ3 on a glassy carbon (GC) electrode surface. Eastman-AQ55D is a poly (ester sulfonic acid) cation exchanger available in a commercial dissolved form. Films of this polymer exhibit attractive permselectivity, ion-exchange, and antifouling properties. Although the complete structure of AQ55D is unclear right now, the proposed backbone is as follows [16]: O O C
O C
H2 O C
H2 C
2. Experimental 2.1. Reagents Ru(bpy)3Cl2 Æ 6H2O, tripropylamine (TPA, 99%) and Eastman-AQ55D (28% dispersion; AQ55D) were purchased from Aldrich and used as received. The multi-wall carbon nanotubes with 80% purity were purchased from Shenzhen Nanotech. Port. Co. Ltd. China and they were around 1.4 nm in diameter and 1–10 lm in length. It was purified and functionalized by the published procedure [23]. Under such circumstance, the CNT was negatively charged. All of other reagents were of analytical grade. All aqueous solutions were prepared with distilled water. 2.2. Apparatus Cyclic voltammetric experiments were performed with a CH Instruments 600 voltammetric analyzer. All experiments were carried out with a conventional three-electrode system. The working electrode was glassy carbon coated with CNT–AQ composite film. A platinum wire was the counter electrode, and an Ag/AgCl (saturated KCl) worked as a reference electrode. All the potential were measured and reported according to this reference electrode. The ECL signal produced in the electrolytic cell was detected and recorded by a flow injection chemiluminesH2 C
O
x
O O C
H2 C
O C z
y SO 3 Na + AQ55D
AQ has relatively high molecular weight amorphous polyesters with sulfonic groups on aromatic dicarboxylic acid unit, it is much more hydrophilic than Nafion [17]. 2þ Thus RuðbpyÞ3 immobilized in AQ polymer is prevented from partitioning into hydrophobic region, which could improve the stability of ECL sensor. With their low cost, strong adherence to the surface and rapid response [18], AQ polymers appear to be attractive material for sensor development. Moreover, CNT represents a considerably significant group of nanomaterial with unique geometrical, mechanical, electronic and chemical properties [19,20]. High sensitivity and stability could be obtained when CNT was used to modify the electrode. The improved sensitivity was attributed to the electrocatalytic activity of CNT and the interfacial accumulation onto its surface, while the better stability was attributed to the minimization of surface fouling at CNT surface [21]. Therefore, the interfusion of negatively charged CNT into the AQ increases not only the amount of the RuðbpyÞ2þ 3 but also the electronic conductivity of the composite film [22]. In general, the AQ–CNT composite-modified electrode exhibits greatly improved stability and sensitivity.
cence analyzer (IFFD, xi’an Remax electronic Science Tech. Co. Ltd. China), which was operated by a personal computer. ECL intensities were measured through the bottom of the cell with PMT window and all of them were enclosed in a light-tight box. The photomultiplier tube (PMT) was operated in current mode. Unless noted, otherwise, the PMT was biased at 600 V. 2.3. Preparation of ECL sensor Certain amount of CNT was dispersed in acetone solution. Then mixed it with AQ55D-acetone solution. The mixture was ultrasonicated for about 20 min until a homogeneous solution of CNT–AQ–acetone complex was obtained and the concentration of the CNT in this solution is about 0.5 mg/ml. The GC electrode was polished with 0.3 and 0.05 lm aluminum slurry, respectively, rinsed thoroughly with redistilled water, and ultrasonicated in redistilled water for 1 min, allowed to dry in room temperature. About 1.5 ll composite was hand-cast on the surface of a GC electrode and the film was dried for 1 h at room temperature. The modified GC electrodes were
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then placed in 1 mM RuðbpyÞ2þ aqueous solution for 3 2þ about 30 min to incorporate RuðbpyÞ3 . With chronocoul2þ ometry method, the surface coverage of RuðbpyÞ3 ðC Þ of the AQ–CNT composite film was estimated to be 8.5 · 108 mol/cm2. 3. Results and discussion 3.1. Electrochemistry and ECL of immobilized RuðbpyÞ2þ 3 2þ
The electrochemical behavior of RuðbpyÞ3 immobilized in AQ–CNT composite film electrode has been investigated using cyclic voltammetry to provide information about reagent electroactivity and stability. AQ is an effective ion exchanger, so RuðbpyÞ2þ 3 could be easily incorporate into the composite film through ion-exchange process and electrostatic adsorption. Fig. 1 shows the cyclic voltammo2þ grams (CVs) of 0.5 mM RuðbpyÞ3 in phosphate buffer (pH 7.6) at a pure AQ film electrode and AQ–CNT composite film electrode with the scan rate of 100 mV/s. The CVs were recorded after reaching steady state. In general, the two CVs are similar in shape, but they are much different in current intensity. At the pure AQ film electrode, only a weak peak due to the oxidation of RuðbpyÞ2þ 3 in solution can be observed. Whereas a distinguishable response occurs at the AQ–CNT composite film electrode. Its oxidation peak current is much higher than that at AQ film electrode. This could be attributed to the structure of the composite film. AQ is a good medium for preconcentration 2þ of RuðbpyÞ3 . Negatively charged CNT could also absorb 2þ some RuðbpyÞ2þ 3 . Moreover, the oxidation of RuðbpyÞ3 was shifted negatively about 33 mV at AQ–CNT composite electrode compared with the pure AQ electrode. That is because the existence of CNT increases the electronic conductivity of the composite film, which quickens the charge transportation and decreases iR effects.
Fig. 1. Cyclic voltammograms (CVs) of 0.5 mM RuðbpyÞ2þ 3 in phosphate buffer (pH 7.6) with the scan rate of 100 mV/s, at a pure AQ modified electrode (dashed line) and AQ–CNT modified electrode (solid line).
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incorporated in the The ECL behavior of RuðbpyÞ2þ 3 AQ–CNT composite film electrode has been studied with tripropylamine (TPA) as a representative analyte, since 2þ the RuðbpyÞ3 –TPA system has been fully investigated and given stronger ECL signal than other reductants [24]. 2þ Fig. 2 exhibits CVs of RuðbpyÞ3 immobilized in AQ– CNT composite film electrode in the absence and presence of 3 · 109 M TPA at the scan rate of 100 mV/s in phosphate buffer solution (pH 7.6). In the presence of the 2þ TPA, the oxidation current of RuðbpyÞ3 increases clearly while the reduction current decreases. Meanwhile, the ECL signal increases considerately in the presence of TPA. The 2þ ECL behavior of immobilized RuðbpyÞ3 on AQ–CNT composite film electrode is illustrated in Fig. 3. The onset of luminescence occurs near 1 V, and then the ECL intensity rises steeply until it reaches a maximum near 1.15 V, which is consistent with the oxidation potential of 2þ RuðbpyÞ3 . This means that the oxidation of immobilized 2þ RuðbpyÞ3 plays a key role in the process of ECL. ECL intensity could be as high as about 1330 a.u. when the concentration of TPA is only 3 · 109 M. This could be attributed to the electronic conductivity of CNT and the ionexchange ability of Eastman-AQ55D. All of these results indicate that an energetic electron-transfer reaction occurs between electrogenerated RuðbpyÞ3þ 3 and the strong reducing intermediate, the deprotonated form of TPA (TPA). The ECL reaction of the system can be expressed as the follow equations [24]: 3þ RuðbpyÞ2þ 3 ! RuðbpyÞ3 þ e 3þ RuðbpyÞ3
þ TPA ! TPA ! TPAþ þ e
2þ RuðbpyÞ3
ð1Þ þ TPA
þ
ð2aÞ ð2bÞ
Fig. 2. Cyclic voltammograms of RuðbpyÞ2þ immobilized in AQ–CNT 3 composite-modified electrode in the absence (dashed line) and presence (solid line) of 3 · 109 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
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Fig. 3. Corresponding ECL–potential curves for AQ–CNT composite film in phosphate buffer solution (pH 7.6) containing 3 · 109 M TPA at the scan rate of 10 mV/s.
TPAþ ! TPA þ Hþ
ð3Þ
3þ 2þ RuðbpyÞ3 þ TPA ! RuðbpyÞ3 2þ 2þ RuðbpyÞ3 ! RuðbpyÞ3 þ hm
ð4Þ ð5Þ
We investigated the effect of the amount of CNT interfused into the AQ film. Fig. 4. showed the ECL response varied with the amount of CNT in the presence of 107 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s. The ECL response increased as the amount of CNT in AQ film increased up to 0.5 mg/ ml, which may due to the increased adsorption of 2þ RuðbpyÞ3 onto negatively charged CNT surface. However, as the amount of CNT increased further, the ECL response decreased gradually. The CNT could not dispersed evenly in the composite films if the amount of
Fig. 4. Effect of the CNT amount in the AQ film on ECL intensity in the presence of 107 M TPA in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
CNT was more than 0.5 mg/ml, which finally led to poor film fabrication. Therefore, an optimum amount of CNT at 0.5 mg/ml was used in the subsequent experiments. A pH study has been carried out to examine the pH 2þ effect on the RuðbpyÞ3 –TPA system. The ECL signals for TPA obtained at the AQ–CNT composite-modified electrode increases from 5.5 to 7.6 considerably and then decreases rapidly at higher pH. This shows the similar result as that of the Nafion–CNT composite films [13] and AQ–silica composite film [25]. The ECL signal increases from 5.5 to 7.6, implying that deprotonation of TPA is needed as demonstrated in Eq. (3). While as the pH increasing continuously, some decomposition of species would be expected, leading to a diminished ECL reagent available for ECL reaction. Therefore, the ECL intensity decreases [26]. The scan rate effect on the ECL intensity is also investigated. The ECL intensity decreases with the increasing of scan rate in the range from 30 to 200 mV s1. The ECL signal changes with the scan rate depended on two factors: the chemical kinetics of the system and the rate of TPA diffusion in the film. Because the ECL signal increases as the scan rate decreasing, the chemical kinetics of the system plays a major role in this process. In order to evaluate the sensitivity of the composite film modified electrode, TPA is used as the probes. Calibration curve for TPA is demonstrated in Fig. 5. The result shows that the ECL signal varies linearly with the concentration of TPA in two ranges with different slopes. The first one is from 1010 to 107 M with the slope of 0.01 and the correlation coefficient of 0.9986. The second one is from 107 to 105 M with the slope of 0.031 and the correlation coefficient of 0.9982. To our knowledge, among many methods to immobilize RuðbpyÞ2þ onto the electrode surface, the 3 lowest detection limit has been 109 M for TPA occurred at Nafion-CNT composite film electrode so far. While the
Fig. 5. Calibration of TPA at the RuðbpyÞ2þ immobilized AQ–CNT 3 composite film electrode in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s.
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into the solution. Furthermore, according to the structure 2þ of AQ, it is more hydrophilic than Nafion, so RuðbpyÞ3 is strongly concentrated in the whole AQ film by ionexchange and keeps its electroactivity. 4. Conclusion
Fig. 6. ECL emission of immobilized RuðbpyÞ2þ 3 in AQ–CNT composite film electrode in phosphate buffer solution (pH 7.6) containing 3 · 109 M TPA at the scan rate of 100 mV/s.
AQ–CNT composite films have proven to be an effective matrix for the immobilization of RuðbpyÞ2þ at electrode 3 surface to prepare an ECL sensor. Because of the special character of the AQ and CNT, this sensor shows excellent sensitivity with detection limit of 3 · 1011 M for TPA. Moreover, since the AQ is a proper support for some enzymes, the AQ–CNT composite film can be used as a good matrix for coimmobilization of RuðbpyÞ2þ 3 and some enzymes(such as alcohol dehydrogenase) to fabricate a highly sensitive RuðbpyÞ2þ 3 ECL based biosensor. Acknowledgement
detection limit from the present experiments is 3 · 1011 M for TPA, which is two orders of magnitude lower than that of Nafion–CNT film. The excellent sensitivity could be explained as follows: firstly, the presence of CNT makes the composite film much more electronically conductive, which is advantageous for electron transfer. Besides, the negatively charged CNT could also absorb some 2þ RuðbpyÞ3 through electrostatical interaction. Secondly, AQ is much more hydrophilic than Nafion, so RuðbpyÞ2þ 3 does not diffuse into the hydrophobic region where it may lose its electroactivity as that in Nafion. The long-term stability of the AQ–CNT composite film 2þ electrode immobilized with RuðbpyÞ3 was investigated over two weeks. The composite film electrode was stored in dry state at room temperature and monitored by cyclic voltammetry measurements in a phosphate buffer (pH 7.6) periodically. The peak potential was essentially unchanged during this period, and the oxidation peak current decreased less than 5% compared with the initial steady state value after two weeks of storage. Moreover, when the CV measurements were run continuously for 100 cycles in phosphate buffer (pH 7.6) at the scan rate of 100 mV/s, the ECL signal could decrease about 10.8%. 2þ Fig. 6 shows ECL emission of immobilized RuðbpyÞ3 in AQ–CNT composite film electrode in phosphate buffer solution (pH 7.6) at the scan rate of 100 mV/s containing 3 · 109 M TPA for 10 continuous cycles. Peak intensity displays good reproducibility with the relative standard 2þ deviation less than 1%. The good stability of RuðbpyÞ3 in the composite film may arise from that the 2þ RuðbpyÞ3 is a quite stable CL regent, and the ion2þ exchange selectivity of AQ for RuðbpyÞ3 is so high that 2þ it could prevent immobilized RuðbpyÞ3 from leaching
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