A Chitosan-Multiwall Carbon Nanotube Modified Electrode for Simultaneous Detection of Dopamine and Ascorbic Acid

2004 The Japan Society for Analytical Chemistry 1055 A Chitosan-Multiwall Carbon Nanotube Modified Electrode for Simultaneous Detection of Dopamine and Ascorbic Acid Lingyan JIANG,* Chuanyin LIU,* Liping JIANG,* Zhen PENG,** and Guanghan LU* *College of Chemistry, Central China Normal University, Wuhan 430079, People s Republic of China **Department of Chemistry, Caohu College, Anhui 238000, People s Republic of China A chemically modified electrode based on a chitosan-multiwall carbon nanotube (MWNT) coated glassy carbon electrode (GCE) is described, which exhibits an attractive ability to determine dopamine (DA) and ascorbic acid (AA) simultaneously. The modified electrode exhibited a high differential pulse voltammetry (DPV) current response to DA at 0.144 V and AA at 0.029 V (vs. SCE) in a 0.1 mol l 1 phosphate buffer solution (ph = 7.2). The properties and behaviors of the chitosanmultiwall carbon nanotube modified electrode (MC/GCE) were characterized using cyclic voltammetry (CV) and DPV methods. The mechanism for the discrimination of dopamine from ascorbic acid at MC/GCE is discussed. The linear calibration range for DA and AA were 5 10 7 mol l 1 to 1 10 4 mol l 1 (r = 0.997), and 5 10 6 mol l 1 to 1 10 3 mol l 1 (r = 0.996), respectively. The MC/GCE showed good sensitivity, selectivity and stability. (Received February 23, 2004; Accepted April 28, 2004) Dopamine (DA) is a very important neurotransmitter in the mammalian central nervous system. Because low levels of DA have been found in patients with Parkinson s disease, 1 its detection with high selectivity and sensitivity is of great significance in investigating its physiological functions and the diagnose of nervous diseases resulting from an abnormal metabolite. When solid electrodes are used to detect DA, the main and foremost difficulty is the interference of ascorbic acid (AA), which is oxidized at almost the same potential as DA. 2 It is thus very important to develop an electrochemical method of high selectivity and sensitivity for the determination of DA. Chitosan, which is derived from chitin, has excellent biologic compatibility and can be used as a modifier because it has many amino and hydroxyl groups. 3 8 Besides, using a natural polymer as a dispersant is a requirement of environment protection. Carbon nanotubes have attracted much attention because they have the ability to promote electron transfer reactions when used as electrode materials, 9,10 but carbon nanotubes are very stable and insoluble in most solvents, which restrained its application in electroanalysis. Thus, in this study, a multiwall carbon nanotube (MWNT)-chitosan modifier was obtained by putting MWNT into a chitosan solution (0.5%). The MWNTchitosan modified electrode (MC/GCE) has many advantages with regard to a low detection limit, a fast response, excellent reproducibility for the detection for DA and AA simultaneously. Experimental Apparatus All of the electrochemical measurements were carried out with a bioanalytical system CV-50W electrochemical analyzer. The three-electrode system consisted of a modified GCE, a saturated calomel reference electrode (SCE), and a platinum To whom correspondence should be addressed. E-mail: ghlu@ccnu.edu.cn auxiliary electrode. All potentials were referred to the SCE. The transmission electron microscopy image was done with a TEM-100CXII. Reagents Dopamine, ascorbic acid (Fluka Corp.). A chitosan solution (0.5%) was prepared by dissolving 0.05 g of chitosan in 10 ml acetic acid (2 mol l 1 ). 11 The MWNT used in this work (obtained from the Institute of Nanometer, Central China Normal University) was synthesized by catalytic pyrolysis and purified followed the method given in a reference. 12 Preparation for a MWNT-chitosan modified electrode Upon putting 1 mg of MWNT into 5 ml of a chitosan solution (0.5%), ultrasonic agitation for a few minutes gave 0.2 mg ml 1 of a black suspension. Before being modified, the GCE was polished with 0.3 µm and 0.05 µm aluminum slurry, rinsed thoroughly with redistilled water, then ultrasonically rinsed with alcohol, redistilled water for 1 min each, and dried under an infrared lamp. After the GCE was cooled, it was smeared evenly with 10 µl of a MWNT-chitosan (0.5%) solution by a micro-syringe, and then dried under an infrared lamp for 10 min. After cooling, the MC/GCE could be used. Experimental procedure A 0.1 mol l 1 phosphate buffer solution (ph = 7.2) was used as the supporting electrolyte for the determination of DA and AA. Before and after every measurement, the MC/GCE was activated by successive cyclic voltammetric sweeps between 0.20 V and 0.5 V at 100 mv s 1 in a blank phosphate buffer solution (ph = 7.2). Results and Discussion Redox behavior of dopamine at MC/GCE In a phosphate buffer solution (0.1 mol l 1, ph = 7.2), the

1056 ANALYTICAL SCIENCES JULY 2004, VOL. 20 Fig. 1 Cyclic voltammograms of 1 10 4 mol l 1 DA at bare and MC/GCE, in a 0.1 mol l 1 phosphate buffer solution (PBS) (ph 7.2). Scan rate, 100 mv s 1. Fig. 3 Effect of the solution ph at MC/GCE. Fig. 4 Effect of the modifier amount at MC/GCE in a 0.1 mol l 1 PBS (ph 7.2). rate, suggesting that the redox reaction of DA at the MC/GCE was an absorption-controlled behavior. 13 Fig. 2 Cyclic voltammograms of 1 10 4 mol l 1 DA in a 0.1 mol l 1 PBS (ph 7.2) at MC/GCE and the relationship of the peak currents and scan rates. Scan rates in : 1, 25; 2, 50; 3, 75; 4, 100; 5, 125; 6, 150; 7, 175; 8, 200 mv s 1. electrochemical behavior of DA was investigated. Figure 1 shows the cathodic peak and anodic peak of DA that emerge in the curves. Compared with the case at a bare electrode, the cathodic peak potential of DA (E pc) shifted positively from 0.082 V to 0.117 V, and the anodic peak potential (E pa) shifted negatively from 0.252 V to 0.183 V; the difference between E pa and E pc ( E p) decreased from 170 mv to 66 mv. All of these results showed that the reversibility of the MC/GCE became better than that of the bare electrode. Meanwhile, the cathodic peak current (i pc) and anodic peak currents (i pa) both increased, the figure of the curve became better. These results indicated that the MWNT-chitosan modified electrode exerted an obvious electrocatalysis effect on DA. The effect of the scan rate on the redox peak currents of DA was studied by cyclic voltammetric sweep (shown as in Fig. 2). It was also found that the redox peak currents of DA were directly proportional to the scanning Effect of the supporting electrolyte and solution ph It was discovered that the redox peak current of DA at MC/GCE in the phosphate buffer solution was higher than that in other supporting electrolytes, such as a HAc NaAc, B R buffer solution. Besides, its peak current also depended on the phosphate concentration; the highest current was obtained with a 0.1 mol l 1 phosphate buffer solution. The effect of the solution ph on the electrochemical behavior of DA was studied in the ph range of 3.2 8.2. It was also found that the E pa of DA shifted to a more negative value with increasing the ph; this is a consequence of a deportonation step involved in all oxidation processes that are facilitated at higher ph values. The relationship of E pa and the ph obeyed the following equation: E pa = 0.571 0.056pH (r = 0.998); the slope of equation was 56 mv/ph, all of which suggested that two protons took part in the electrode reaction of DA, which agreed with the reference. 14,15 As shown in Fig. 3, the highest oxidation peak current of DA was observed in a ph 7.2 phosphate buffer solution. Because in this condition, DA carries positive charge, while a small quantity of NH 2 in chitosan can be converted into NH 3+ for a low concentration of H +, the electrostatic repulsion between DA and chitosan is very faint, and it is easier for DA to have access to the electrode. Thus the following experiments were carried out in a ph 7.2 phosphate buffer solution. Effect of the amount of MWNT-chitosan modifier at MC/GCE The quantity of the modifier on the surface of the electrode affected the peak current of DA. It is shown in Fig. 4 that the i pa

1057 Fig. 5 Cyclic voltammograms of DA and DA + AA at MC/GCE in a 0.1 mol l 1 PBS (ph 7.2). a, 1 10 4 mol l 1 DA; b, 1 10 4 mol l 1 DA + 1 10 3 mol l 1 AA. Scan rate, 100 mv s 1. Fig. 7 DPV recordings of DA at MC/GCE in a 0.1 mol l 1 PBS (ph 7.2) in the presence of 10 4 mol l 1 AA. DA concentrations (10 5 mol l 1 ): 1, 1.5; 2, 3; 3, 4.5; 4, 6; 5, 7.5; 6, 9. Scan rate, 20 mv s 1 ; pulse amplitude, 50 mv; pulse width, 50 ms; pulse period, 20 s. Fig. 6 Differential pulse voltammograms for (1) 1 10 3, (2) 3 10 3, (3) 4 10 3, (4) 5 10 3 mol l 1 AA + 1 10 4 mol l 1 DA at MC/GCE. Scan rate, 20 mv s 1 ; pulse amplitude, 50 mv; pulse width, 50 ms; pulse period, 20 s. of DA was enhanced when the amount of MWNTs increased, which was probably because of the presence of a threedimensional reaction zone, which increased with the amount of MWNTs. 16 When the amount of MWNTS surpassed 45 µg/cm 2, the capacitive current enhanced, while the i pa of DA fell. This was possibly because the film on the electrode surface became thicker, which blocked the electron transfer between DA and electrode. In this study, the amount of MWNTS at MC/GCE was 38 µg/cm 2 because the figure of curve was the best when using this amount. Electrochemical behavior of coexisting DA and AA at MC/GCE A cyclic voltammetric curve of a mixture containing DA (10 4 mol l 1 ) and AA (10 3 mol l 1 ) was recorded by applying a cyclic voltammetric sweep from 0.20 V to 0.50 V. By reason that MC/GCE can also catalyze the oxidation of AA, 17,18 which lead to the anodic peak potential of AA shifted negatively by 250 mv. As a result, two totally divided anodic peak (p 1, p 2) were observed in the CV figure (E pda = 182 mv, E paa = 20 mv, as shown in Fig. 5). Because the charging current contribution to the background current is a limiting factor in the analytical determination of any electroactive species, experiments were carried out using differential pulse voltammetry (DPV) modes. 19 When the DPV was applied instead of CV in the experiments, an obvious improvement in the height of the DA and AA peaks was observed, which was quite important in the detection of low concentrations of DA and AA. The analytical experiments were carried out either by varying the AA concentration in the presence of 10 4 mol l 1 DA (Fig. 6) or by varying the DA concentration in the presence of 10 4 mol l 1 AA (Fig. 7) or varying the DA and AA concentrations simultaneously (Fig. 8) in the phosphate buffer solution (ph = 7.2). As shown in Figs. 6 and 7, DA or AA exhibited excellent DPV responses with the signal height of the other ingredients remaining unchanged, thus proving that the responses to DA and AA at the MC/GCE were relatively independent. It can also be observed in Fig. 6 that AA did not interfere with the detection for DA under condition that the AA concentration was 50-times that of DA. When the concentration of AA was 100-times that of DA, the i pa of DA decreased by 9.8%, while the E pa of DA did not shift. Accordingly, simultaneous measurements of low concentration levels of DA and AA at the MC/GCE were possible. The current-to-concentration relationship for DA was linear from 5 10 7 1 10 4 mol l 1, with a correlation coefficient of 0.997 and a detection limit of 2 10 7 mol l 1 (3σ). The linearity of the AA signal ranged from 5 10 6 1 10 3 mol l 1, with a correlation coefficient of 0.996 and a detection limit of 2 10 6 mol l 1 (3σ) (shown in Figs. 8, 8(c)). TEM images of H 2O-MWNT and chitosan-mwnt The transmission electron microscopy images of H 2O-MWNT and chitosan-mwnt are shown as in Fig. 9. Figure 9 shows that different lengths of MWNTS twist each other, and purification of MWNTS is very high. It can also be observed in Fig. 9 that the twisted MWNTS were dispersed equably in the chitosan solution, and their average diameters ranged from 30 nm to 40 nm. Mechanism of electrocatalysis towards DA and AA by the MC/GCE Chitosan is derived from chitin by deacetylation, and it has many active groups, such as amino groups and hydroxy groups, which have high reactivity to several functional groups. 20,21 When chitosan was used to modify GCE as the dispersant of MWNT, there was an intensive H-bond interaction between it and OH, which existed in the structures of DA and AA. Meanwhile, the subtle carbon nanotube provided many active sites enhancing the transfer of charge between the electrodes and the reagents, 22 all of which lead to great eletrocatalysis towards DA and AA. Furthermore, in a neutral solution, a small quantity of NH 2 in chitosan could be converted into NH 3+, and DA carried positive charge while AA carried negative charge under this condition. 23,24 Chitosan would electrostaticly attract AA, which promoted the access of AA to the electrode, and further catalyzed the oxidation of AA, resulting in that the E pa of

1058 ANALYTICAL SCIENCES JULY 2004, VOL. 20 (c) Fig. 9 Transmission electron microscopy image of H 2O-MWNT and chitosan-mwnt. Magnification: 19000. Fig. 8 DPV recordings of coexisting DA and AA and relationship of ip and concentration for DA and AA (c). Concentration of DA from 1 to 8, 5 10 7 1 10 4 mol l 1 ; concentration of AA from 1 to 8, 5 10 6 1 10 3 mol l 1. Scan rate, 20 mv s 1 ; pulse amplitude, 50 mv; pulse width, 50 ms; pulse period, 20 s. AA shifted negatively to 0.20 V. Although there was electrostatic repulsion between DA and chitosan, its blocking effect on the oxidation of DA was very faint because the NH 3+ concentration was low in a ph 7.2 solution. Also, the H-bond interaction between DA and chitosan was more dominant than the electrostatic repulsion effect, so DA was still catalyzed by the modifier. As a result, the separation between the two anodic peak potentials ( E pa) of DA and AA was 212 mv in CV and 185 mv in DPV, which were large enough to determine DA and AA simultaneously, and AA did not interfere with the determination of DA. The reproducibility of MC/GCE Chitosan-MWNT film was very homogeneous and stable on the surface of the electrode. The reproducibility of MC/GCE was studied. The RSD was 4.07% for ten successive determinations of 1 10 4 mol l 1 DA. For the same electrode, modified ten times, respectively, and determining 1 10 4 mol l 1 DA, the RSD was found to be 5.87%. After every measurement, the MC/GCE was refreshed by 40 successive cyclic voltammetric sweeps between 0.2 V and 0.5 V at 100 mv s 1 in a blank phosphate buffer solution (ph = 7.2). The MC/GCE could be used for at least seven days. Determination of samples Different amounts of DA injection and an AA standard solution were transferred into three volumetric flasks of 50 ml; Table 1 Sample No. then some redistilled water was added in to give three samples, in which the concentrations of DA were 2.021 10 5 mol l 1, 2.636 10 5 mol l 1, 3.515 10 5 mol l 1, and the AA concentrations were 5 10 5 mol l 1, 1.5 10 4 mol l 1, 2 10 4 mol l 1. The results of determination are summarized in Table 1. A natural polymer, chitosan, can be used as a dispesant of MWNT. Chitosan-MWNT film was very homogeneous and stable on the surface of the electrode. The MC/GCE can be used to determine DA and AA simultaneously and its sensitivity, reproducibility and stability are satisfactory. References Determination of samples DA found/ RSD, % Relative mol L 1 n = 6 error, % AA found/ mol L 1 RSD, % n = 6 Relative error, % 1 2.097 10 5 3.65 3.76 5.170 10 5 3.48 3.40 2 2.592 10 5 3.15 1.67 1.521 10 4 2.78 1.40 3 3.460 10 5 3.67 1.56 1.948 10 4 4.07 2.60 1. M. Wightman, L. J. May, and A. C. Michael, Anal. Chem., 1988, 60, 769A. 2. A. Ciszewaki and G. Milizanek, Anal. Chem., 1999, 71, 1055. 3. B. Li, Y. F. Lian, Z. J. Shi, and G. X. Shen, Chem. J. Chinese Universities [J], 2000, 21, 1633. 4. K. Sugawara, H. Kuramita, S. Hoshi, K. Akatsuka, and S. Tanaka, Anal. Sci., 2002, 18, 195. 5. A. Hiratsuka, H. Muguruma, S. Sasaki, K. Ikebukuro, and I. Karube, Electroanalysis, 1999, 11, 1098.

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