Selective sensing of cysteine on manganese dioxide nanowires and chitosan modified glassy carbon electrodes

Biosensors and Bioelectronics 24 (2009) 2985–2990

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Selective sensing of cysteine on manganese dioxide nanowires and chitosan modified glassy carbon electrodes Yu-Hui Bai, Jing-Juan Xu ∗ , Hong-Yuan Chen Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China

a r t i c l e

i n f o

Article history: Received 15 January 2009 Received in revised form 2 March 2009 Accepted 4 March 2009 Available online 17 March 2009 Keywords: Manganese dioxide Cysteine Electrochemical sensor Selectivity Detection

a b s t r a c t We report a simple approach for the selectively electrochemical sensing of cysteine (CySH) using ␤-MnO2 nanowires modified glassy carbon (GC) electrode via a direct and facile electrochemical co-deposition process with chitosan hydrogel. The electrode showed excellent electrocatalytic effect toward cysteine oxidation. A possible reaction mechanism related to the formation of surface complexes was proposed: the –SH group is the reacting group in the reaction between cysteine and MnO2 , while the –NH2 group acts as an anchor to stabilize the surface complexes. The electrocatalytic behavior is further developed as a sensitive detection scheme for cysteine by amperometric measurement, which shows a large determination range of 0.5–630 ␮M and a low detection limit of 70 nM. Other nineteen amino acids in a 20-fold concentration do not interfere in the detection of cysteine. Furthermore, other biological species containing –SH group or S S bonds, such as glutathione, cystine, and bovine serum albumin (BSA), show no interferences because of steric hindrance and inactive S S bonds. © 2009 Elsevier B.V. All rights reserved.

1. Introduction Cysteine, as a sulfur-containing amino acid, plays a significant role in biological systems. It provides a modality for the intramolecular crosslinking of proteins through disulfide bonds to support their secondary structures and functions, and it is also a disease associated physiological regulator (Stryer, 1995). A variety of detection methods for cysteine have emerged. Spectroscopic methods usually utilize gold nanomaterials based on the interactions between gold and thiols, for example, the thiol group can selectively functionalized onto the edges of Au nanorods (Sudeep et al., 2005), the cysteine can modulate the energy transfer between fluorophore and gold (Shang et al., 2007). Liquid chromatography methods together with other technologies have been used in the simultaneous detections of multiple components (Pelletier and Lucy, 2004; Hou and Wang, 1991). Electrochemical methods have also been developed extensively for the detection of thiols. Many carbonbased materials, such as boron-doped diamond, carbon nanotubes, and mesoporous carbon have been explored for the electrochemical oxidation and detection of cysteine (Spataru et al., 2001; Gong et al., 2005; Zhao et al., 2003; Zhou et al., 2007). Several metallic combined materials have also been applied on the detection of cysteine (Zen et al., 2001; Prasad et al., 2008). Although the electrochemical responses have been continuously improved, the development of an electrochemical method for sensitive and selective determination

∗ Corresponding author. Tel.: +86 25 83597294; fax: +86 25 83597294. E-mail address: [email protected] (J.-J. Xu). 0956-5663/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.bios.2009.03.008

of thiols still remains very challenging. Some electrodes contain certain disadvantages, such as leaching of modified materials, irreversible adsorption of substrates onto the electrode surface, narrow detection range or not enough sensitivity for real-sample analysis (Chen et al., 1990; Fawcett et al., 1994; Nekrassova et al., 2003). Thus, the choice of material is essential and important to construct sensors with excellent performances. Manganese dioxide is a kind of attractive inorganic material, and material scientists have made great efforts on the synthesis of MnO2 (Wang and Li, 2003, 2002a,b; Wang et al., 2006; De Guzman et al., 1994; Xiong et al., 2003). Recently, several kinds of MnO2 nanomaterials were utilized to construct chemical sensors or biosensors based on the electrocatalytic ability of MnO2 to H2 O2 (Lin et al., 2005; Yao et al., 2006; Turkusica et al., 2005; Luo et al., 2004b; Bai et al., 2007). Besides, the redox property of colloidal MnO2 has been used for the degradation of bioorganic species (Kabir-ud-Din et al., 2005; Khan et al., 2004; Andrabi and Khan, 2005). Afonso et al. studied the kinetics of reduction of soluble polymeric MnO2 by cysteine and glutathione (Herszage et al., 2003). Dos Santos Afonso and Khan found that the reactivity of –SH group with colloidal MnO2 is higher than –NH2 and –COOH groups (Andrabi and Khan, 2007). These previous studies were focused on the reaction between the colloidal MnO2 solution and cysteine or glutathione, however, the electrochemical detection of cysteine utilizing MnO2 nanomaterials modified electrode has not been investigated before, as far as we know. Our recent study has shown that the crystal MnO2 , especially the ␤-MnO2 nanowires, are suitable materials for constructing biosensors due to the high catalytic abilities and fast response to substrate (Bai et al., 2008). The one-step co-deposition

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is a very convenient and repetitive technique for the fabrication of sensors. The ␤-MnO2 nanowires together with the easy modification method can bring many advantages to the sensor, such as simplicity, low cost, fast response, high sensitivity and stability. Thus, the ␤-MnO2 nanowires are promising candidate for the electrochemical detections of organic molecules. In this paper, we report the detail and systematic investigation for the catalytic mechanism of cysteine oxidation at an easyprepared chitosan and ␤-MnO2 nanowires modified glassy carbon (GC) electrode. The structure property and electrocatalytic activity were characterized with transmission electron microscopy (TEM), X-ray diffraction (XRD), infrared spectrum (IR), nitrogen adsorption and cyclic voltammetry. The catalytic mechanism was applied in the determination of cysteine at trace level by the amperometric method. The present study enlarges the application range of MnO2 nanomaterials into detections of organic molecules using an electrochemical method. 2. Experimental 2.1. Chemicals and reagents All amino acids were purchased from Shanghai Bio Life Science & Technology Co., Ltd. Chitosan from crab shells (85% deacetylated), Bovine serum albumin (BSA), mercaptopropionic acid (MPA) were obtained from Sigma–Aldrich. All other chemicals were of analytical reagent grade. A borate saline buffer solution was prepared by adjusting 0.2 M boric acid with 0.05 M borax solution. An aqueous solution containing 5 mM cysteine was freshly prepared every day. All aqueous solutions were prepared in doubly distilled water from a Milli-Q water purifying system. Chitosan solutions were prepared as previously reported (Luo et al., 2004a). 2.2. Instruments Electrochemical experiments were performed on a CHI 660A electrochemical workstation (Shanghai Chenhua Apparatus, China). A three-electrode configuration was employed with a GC electrode (3 mm diameter) as a working electrode, a saturated calomel electrode (SCE) and a platinum wire as the reference and counter electrodes, respectively. TEM image analysis was carried out on a Joel JEM 2010 microscope. X-ray diffraction (XRD, VG-108R, Philips) was used for characterizing the structure of MnO2 nanomaterials. Infrared spectrum was obtained on American Avatar 360 apparatus.

solution for 3 min. The deposition solution was 2.0 mL of 0.4% chitosan solution with or without 1.0 mg/mL of MnO2 nanowires. At the negative potential, H+ was reduced to H2 at the cathode and released. This reaction leads to a high localized pH in the vicinity of the cathode surface. When the pH was higher than 6.3, the positively charged chitosan became insoluble, and chitosan hydrogel incorporated MnO2 were electrodeposited at the cathode surface as a result (see Fig. S1 in supplementary content). The modified electrodes were denoted as Chit/GC (without ␤-MnO2 nanowires) or ␤/Chit/GC (with ␤-MnO2 nanowires), respectively. The modified electrodes were dried in air at room temperature for about 5 h and then stored in a refrigerator (4 ◦ C) for use. 3. Results and discussion 3.1. Characterization of MnO2 nanomaterials The powder XRD pattern of MnO2 nanomaterials is displayed in Fig. 1. The major diffraction peaks can be indexed to JCPDS 24-0735. The sharp peaks indicate that the products are perfectly crystallized and no other impurity peaks were observed. Inset shows the typical transmission electron micrographs of ␤-MnO2 nanomaterials. A one-dimensional nanostructure was observed for ␤-MnO2 which displays the nanowires with diameters of 30 nm and lengths of ca. 1 ␮m. Fig. S2 shows the infrared spectroscopy of ␤-MnO2 nanomaterials. The broad peak at 3200–3600 cm−1 could be assigned to the stretching vibration of H2 O molecule and OH−1 in the lattice. The peaks of 1620 cm−1 were assigned to the bending vibration of H2 O molecule. This implied the hydroxyl group existed in MnO2 nanowires. The two peaks at about 520 and 720 cm−1 arose from the stretching vibration of the Mn O and Mn O Mn bonds. 3.2. CV behavior of cysteine at different electrodes Fig. 2 shows the CV responses of 0.1 mM cysteine on the bare GC electrode, the Chit/GC electrode, and the ␤/Chit/GC electrode in pH 7.8 borate saline buffer at a scan rate of 100 mV s−1 . At the bare GC electrode, cysteine starts to be electro-oxidized at more than +550 mV, and no oxidative peak displayed. At the Chit/GC electrode, the response of cysteine is reduced compared with that at the bare GC electrode, because the chitosan membrane prevents the cysteine molecule to arrive at the electrode surface. At the ␤/Chit/GC electrode, it displays a pair of small oxidative and reductive peaks

2.3. Preparation of MnO2 nanowires MnO2 nanowires were prepared according to the literature with a little modification (Wang et al., 2006). Briefly, hydrogen peroxide (1 mL, 30%) was added to an aqueous solution of manganese nitrate (12 mL, 0.3 mol/L) while stirring to form a homogeneous solution, and then ammonia (5 mL, 0.5 mol/L) was added to the mixture dropwise. A brown suspension was obtained and transferred into a 20 mL Teflon-lined autoclave up to 80% of the total volume. The sealed autoclave was heated to 200 ◦ C for 12 h. The resulting black solid product was collected, washed several times with distilled water and absolute ethanol, centrifuged, and dried at 105 ◦ C for 3 h. 2.4. Fabrication of biosensors The fabrication procedure is according to the previous reports (Luo et al., 2004a, 2005; Fernandes et al., 2003; Yi et al., 2005). A pair of polished and cleaned GC electrodes were connected respectively to a current power supply (−3.0 V) and dipped into a deposition

Fig. 1. XRD patterns of ␤-MnO2 nanomaterial. Inset: Typical TEM images of ␤-MnO2 nanowires.

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Fig. 2. Cyclic voltammograms with (solid lines) and without (dotted lines) 0.1 mM cysteine at different electrodes in pH 7.8 borate saline buffer. Scan rate, 100 mV s−1 .

between +250 and +520 mV in the absence of cysteine which are induced from the electro-redox of Mn species. With the addition of cysteine, the peak currents increase obviously. It demonstrates the effective catalytic behavior of ␤-MnO2 nanowires toward cysteine. Cyclic voltammograms of 0.3 mM cysteine at ␤/Chit/GC electrode in pH 7.8 borate saline buffer with different scan rates were tested (data not shown). The anodic peak currents are proportional to the scan rates which indicates a surface-controlled process and the anodic peak potentials (Ep,a ) are proportional to log  in the range less than 500 mV s−1 (see A and B in Fig. S3). On the basis of the model of Laviron (Laviron, 1979), the charge-transfer coefficient ˛ and electron number n were calculated to be 0.45 and 0.91, respectively, which indicates a one-electron process on the electrode surface. 3.3. Effect of pH Cyclic voltammograms of cysteine at different pH were tested at the ␤/Chit/GC electrode (data not shown). In Fig. 3, it shows the effect of pH on the anodic peak currents and potentials of cysteine at ␤/Chit/GC electrode. With the pH changing from 7.0 to 9.0, the anodic peak currents increase and reach a maximum around pH 7.8, and then decrease obviously above pH 8.6. The change trend of current is different from that at the lead ruthenate pyrochlore

Fig. 3. Plots of relative responses (square symbol) and oxidative peak potentials (triangle symbol) versus pH for 0.1 mM cysteine on the ␤/Chit/GC electrode. Scan rate, 100 mV s−1 .

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modified electrode reported before (Zen et al., 2001). Zen proposed that positively charged CySH2 + or the neutral CySH is responsible for the disulfide formation because relatively higher anodic current was noticed when pH < 5 and a decreasing trend was observed between pH 5.0 and 9.0. The present result is also different from the reports that the protonated forms of the thiols reacted more quickly with the anionic polymeric MnO2 (Herszage et al., 2003). With regard to the electroactive substance, Fujishima et al. noticed that the electroactive substance is CyS− in 0.5 M KHCO3 solution. Guo et al. observed that the changing trend of distribution fraction (ı) of CySH2 + and CyS− with pH was well consistent with the changing tendency of peak currents in the corresponding pH values, so the electroactive substance is CySH2 + or CyS− below or above pH 5.0, respectively (Zhou et al., 2007). In the present system, the electroactive substance should be CyS− which would react with MnO2 on the electrode. Both the existing forms of cysteine and MnO2 are closely related with pH, thus the redox reaction on the electrode surface has much relation with pH values. As a result, the oxidative currents are influenced by the pH values. The anodic peak potential shifted negatively along with the increasing pH. This is a consequence of the deprotonation involved in the oxidation process, which is facilitated at higher pH values. Plot of the formal potential versus pH (from 7.0 to 9.0) produced in a line with slope of −105.6 mV pH−1 (R = 0.997), which was close to two times the value of −59.0 mV pH−1 , indicating a two-proton and one-electron reaction process on the electrode surface. 3.4. Oxidation mechanism of cysteine at the ˇ/Chit/GC electrode To study the role of –NH2 , –COOH, and –SH groups present in the carbon chains in the oxidation of cysteine, parallel experiments were carried out using MPA and alanine under the same conditions. Cysteine, MPA and alanine all have three carbon atoms but with different functional groups (cysteine has –NH2 , –COOH, and –SH groups; MPA has –COOH and –SH groups; alanine has –NH2 and –COOH groups). According to the cyclic voltammetric and amperometric measurement, no electrochemical responses were noticed with the adding of alanine while a slight increase of oxidative current was observed with the adding of MPA on the ␤/Chit/GC electrode. Compared with the same concentration of MPA, cysteine can make fifty times increase of current on the ␤/Chit/GC electrode. These results indicate that the –SH group is responsible for the redox reaction between cysteine and MnO2 , while the –NH2 group may play certain special but necessary roles in the redox process. This result is different from the previous report that they thought only the –SH group alone got involved in the electrocatalytic oxidation process (Zen et al., 2001). It was reported that transition metal would form complexes with amino acids and peptides (Severin et al., 1998). Dos Santos Afonso studied the oxidation of cysteine by soluble MnO2 . They postulated that three surface complexes with different structures might form between cysteine and MnO2 , including one with the nitrogen coordinated to the metal center, and two with the oxygen coordinated to the metal center, although they did not make sure the most probably one (Herszage et al., 2003). Thus, it is reasonable to suggest that some surface complexes also exist in the present system: the –SH group of cysteine binds to the Mn atom, while the nitrogen of –NH2 group may be coordinated with Mn center, which acts as an effective assistance to stabilize the surface complexes. Other parallel experiments were also carried out using cystine, glutathione (reduced) and BSA by cyclic voltammetric and amperometric method. Cystine is the disulfide of cystine with one S S bond, glutathione is a tripeptide formed by glutamic acid, glycine and cysteine, and BSA is a protein with seventeen –S S bonds and one –SH in its structure. No electrochemical responses were noticed with the adding of BSA; while, a slight increase of oxidative cur-

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rent was observed with the addition of cystine or glutathione on the ␤/Chit/GC electrode. The fact that the response to glutathione is one thirtieth of that to cysteine can be attributed to steric hindrance. The complex structure of glutathione molecules makes it harder to approach and be adsorbed onto MnO2 . As regards the BSA, –SH is embedded in the extremely large molecule and cannot be exposed to MnO2 surfaces. The response to cystine is one percent of that to cysteine because of inactive S S bond and steric hindrance. These observations show that the –SH group has much higher activity with MnO2 than S S bond and the size of molecules has great influence on the activity. This is actually an advantage considering that larger molecules and proteins with –SH group or S S bond would not interfere in the detection of cysteine. Thus, selective detection of cysteine from glutathione can be achieved which shows the superiority of electrochemical method to spectral measurement based on the interaction between Au and –SH group. Considering the effects on the oxidation current that mentioned above and the disulfides as the main products (Herszage et al., 2003; Andrabi and Khan, 2007), the possible reaction mechanism should go through the following steps (see Scheme 1). With the addition of cysteine, chemical redox reaction takes place between MnO2 and cysteine on the electrode surface (Eq. (1)). Described in detail, first, H+ is adsorbed on the surface of MnO2 , forming more –OH groups on the modified electrode than the material itself (Eq. (2)). Then a precursor complex is formed on the electrode surface according to a ligand change. Similarly formation of such complexes has been proposed in the reaction of solid manganese (IV) oxides with sulfide (Yao and Millero, 1993). Accompanying with the deprotonation of the amino group, the N site of –NH2 group is coordinated to Mn atom which makes the intermediate more stable. Then the electron transfers from sulfur to Mn and the sulfhydryl radicals (RS• ) are formed as a result (Eq. (3)). The formation of RS• radicals was proposed by Wallace in his research on thiol oxidation with solid-phase MnO2 (Wallace, 1966), and they were also proposed as intermediates in reaction mechanisms to explain the oxidation of cysteine in solutions or at different modified electrodes (Spataru et al., 2001; Zhou et al., 2007; Zen et al., 2001; Herszage et al., 2003; Andrabi and Khan, 2007).

Scheme 1. Oxidation mechanism of cysteine at the ␤/Chit/GC electrode.

The formed RS• radicals dimerize to form the corresponding disulfide (Eq. (4)). Two ways are possible: one is that the radicals might dimerize after they are released to the solution, the other one is that the radicals might dimerize while they are still attached to Mn species. The length for a S S bond is approximately 2 Å in most molecules which contain two or more sulfur atoms, and the distances between vicinal Mn atoms in most manganese (IV) oxides is about 4 Å (Cotton and Wilkinson, 1967; Burns and Burns, 1979). The coordination group of –NH2 may work as an anchor allowing the dimerization of radicals in solution while the molecules are still attached to the electrode surface. These chemical reactions between MnO2 and cysteine are followed by electrode reactions. The product MnO(OH) is easily protonated because of the electronegativity of O atom. In the forward scan (from 0 to +1.0 V), the protonated MnO(OH) is electro-oxidized back to Mn species of quadrivalence (MnO2 ) at the electrode surface (Eq. (5)). This is a two-proton and one-electron reaction process on the electrode surface. Accordingly, in the reverse sweep (from +1.0 to 0 V), MnO2 is electro-reduced to Mn species of trivalence. The adsorption of cysteine onto MnO2 is described in the Eq. (3), which indicates that both the concentrations of protonated MnO2 and negatively charged CyS− have much effect on the formation of the surface complexes. It is noticed that low pH value is beneficial to the protonation of MnO2 , on the contrary, the distribution fraction of CyS− increases with the increase of pH values between pH 7.0 and 9.0. Thus, pH 7.8 at which the maximum anodic current is obtained (see Fig. 3) seems a good tradeoff between the two influence factors. The more surface complexes formed, the more cysteine chemically oxidized and more Mn species of lower states produced which would then be electro-oxidized back to higher states, and consequently, larger oxidation currents are displayed. 3.5. Analytical performance characteristics The electrochemical responses of electroactive substances, such as arginine, methionine, phenylalanine, tryptophan and tyrosine were tested at the bare GC electrode and the ␤/Chit/GC electrode to compare with that of cysteine. As Fig. S4 shows, all the six kinds of amino acids can be oxidized on the bare GC electrode, but the oxidative currents are extremely slight and with similar value so that the current induced by cysteine cannot distinguish itself. On the ␤/Chit/GC electrode, the responses to arginine and tyrosine become much lower than those on the bare GC electrode because the chitosan film confines the penetration of substrates to arrive at the electrode surface, the responses to phenylalanine and tryptophan change little compared with those on the bare GC electrode, and the response to methionine increases to two times that on the bare GC electrode which may be induced by the weak interaction of S S bond with MnO2 . Compared with the currents on the bare GC electrode, the currents of these five amino acids on the ␤/Chit/GC electrode only change within several nanoampere but the current of cysteine increases by nearly 10 times (note the break along the Y axis). It indicates that on one hand the chitosan film can confine the penetration of electroactive substrates to arrive at the electrode surface, and on the other hand MnO2 nanowires play a key role in amplifying the anodic current of cysteine based on the redox reaction between MnO2 and –SH group. One of the significant features of the present system is its ability to detect cysteine in the presence of various other amino acids. Fig. 4A shows the electrochemical responses upon the addition of cysteine and other nineteen amino acids in a 20-fold concentration of cysteine. The response to cysteine was strikingly larger than the other amino acids, indicating that the thiol functionality in cysteine is essential for the increased electrochemical signals. The anodic currents increase in the presence of 19 other amino acids with the concentration 20-fold are all less than 6% relative to that of cysteine.

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Fig. 4. (A) The electrochemical responses to cysteine and other amino acids (with the concentration 20-fold) on the ␤/Chit/GC electrode. Applied potential: +0.5 V. Taking the response of cysteine as 100%. (B) Successive amperometric response of the ␤/Chit/GC electrode to different concentrations of cysteine. Inset: The calibration curve for the detection of cysteine. Error bars represent S.D., n = 3. Table 1 Amperometric determination of cysteine in the presence of human urine. Sample

Amount added (␮M)

Amount found (␮M)

Recovery (%)

R.S.D.a (%)

1 2 3

3.13 6.25 12.50

3.09 6.52 12.39

98.7 104.3 99.1

2.4 3.2 3.8

a

R.S.D. (%) calculated from three separate experiments.

The successive detections of cysteine were carried out at pH 7.8 at room temperature. Fig. 4B presents the current–time plots with the ␤/Chit/GC electrode upon the addition of varying amounts of cysteine. It takes less than 3 s for the ␤/Chit/GC electrode to reach 95% of the maximum current. The catalytic currents showed a linear relationship with the concentration of cysteine in the range of 0.5–630 ␮M (r = 0.998, n = 13) (see inset of Fig. 4B). The detection limit is about 70 nM (three times the signal-to-noise ratio), which suggests that the present approach has great potential for diagnostic purposes because the healthy plasma total cysteine concentrations are about 240–360 ␮M (Refsum et al., 1989; Seshadri et al., 2002). The reproducibility of the biosensor was measured in 0.2 M borate buffer (pH 7.8). The relative standard deviations (R.S.D.) were found to be 4.2% to 0.05 mM cysteine for seven successive measurements on ␤/Chit/GC electrode. The ␤/Chit/GC electrode was stored dry at room temperature and measured at intervals of five days. They remained about 80% of their original responses after 30 days. As a practical use, the ␤/Chit/GC electrode was also used to detect cysteine in the presence of human urine samples. Urine samples were collected from a healthy female. The standard addition was carried out by spiking a certain amount of the cysteine standard solution to urine samples. The electrochemical signal increases apparently after urine samples were spiked by standard solutions containing different concentrations of cysteine. The recoveries of cysteine are determined by standard addition and the corresponding results are given in Table 1. These results suggest that the proposed method is promising for the detection of cysteine in biological fluids. 4. Conclusions In this work, we investigated the electrochemical mechanisms for cysteine oxidation at the ␤-MnO2 wires and chitosan modified GC electrode. The ␤/Chit/GC electrode showed excellent electrocatalytic effect toward cysteine oxidation. Experimental results indicate that –SH group is the reacting group in the reaction of cysteine with MnO2 , while the –NH2 group acts as an effective

assistance to stabilize the surface complexes according to the coordination of N atom with Mn atom. The electroactive substance should be negatively charged CyS− between pH 7.0 and 9.0. A possible reaction mechanism was proposed involving the formation of surface complexes. The results obtained in the analysis of human urine samples indicate that the method is promising for the detection of cysteine in biological fluids. The present study enlarges the application range of MnO2 nanomaterials into detections of organic molecules. Acknowledgments The financial support from the National Natural Science Foundation (Nos. 20890020, 20775033), the National Natural Science Funds for Creative Research Groups (20821063), and the 973 Program (2007CB936404), the 863 program (2007AA022001), and the program for New Century Excellent Talents in University of China is gratefully acknowledged. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.bios.2009.03.008. References Andrabi, S.M.Z., Khan, Z., 2005. Colloid Polym. Sci. 284, 36. Andrabi, S.M.Z., Khan, Z., 2007. Colloid Polym. Sci. 285, 389–396. Bai, Y.H., Du, Y., Xu, J.J., Chen, H.Y., 2007. Electrochem. Commun. 9, 2611–2616. Bai, Y.H., Zhang, H., Xu, J.J., Chen, H.Y., 2008. J. Phys. Chem. C 112, 18984–18990. Burns, R.G., Burns, M.B., 1979. Marine Minerals. In: Burns, R.G. (Ed.), Reviews in Mineralogy 6. Mineralogical Society of America, Washington, DC, pp. 1–46. Chen, X., Xia, B., He, P.J., 1990. J. Electroanal. Chem. 281, 185–198. Cotton, F.A., Wilkinson, G., 1967. Advanced Inorganic Chemistry, 2nd ed. John Wiley & Sons-Interscience, New York, pp. 530–531. De Guzman, R.N., Shen, Y.F., Neth, E.J., Suib, S.L., O’Young, C.L., Levine, S., Newsam, J.M., 1994. Chem. Mater. 6, 815–821. Fawcett, W.R., Fedurco, M., Kovacova, Z., Borkowska, Z., 1994. J. Electroanal. Chem. 368, 265–274. Fernandes, R., Wu, L.Q., Chen, T., Yi, H., Rubloff, G.W., Ghodssi, R., Bentley, W.E., Payne, G.F., 2003. Langmuir 19, 4058. Gong, K., Zhu, X., Zhao, R., Xiong, S., Mao, L., Chen, C., 2005. Anal. Chem. 77, 8158–8165. Herszage, J., Dos Santos Afonso, M., Luther, G.W., 2003. Environ. Sci. Technol. 37, 3332. Hou, W.Y., Wang, E.K., 1991. Talanta 38 (5), 557–560. Kabir-ud-Din, Iqubal, S.M.S., Khan, Z., 2005. Colloid Polym. Sci. 283, 504. Khan, Z., Kumar, P., Kabir, U.D., 2004. Colloids Surf. A Physicochem. Eng. Asp. 248, 25. Laviron, E., 1979. J. Electroanal. Chem. 101, 19–28. Lin, Y.H., Cui, X.L., Li, L.Y., 2005. Electrochem. Commun. 7, 166–172. Luo, X.L., Xu, J.J., Du, Y., Chen, H.Y., 2004a. Anal. Biochem. 334, 285. Luo, X.L., Xu, J.J., Zhao, W., Chen, H.Y., 2004b. Biosens. Bioelectron. 19, 1295–1297. Luo, X.L., Xu, J.J., Wang, J.L., Chen, H.Y., 2005. Chem. Commun. 16, 2169–2171.

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