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Synthesis and characterization of ferrocene modified Fe3O4@Au magnetic nanoparticles and its application
Biosensors and Bioelectronics 24 (2009) 2649–2653
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Synthesis and characterization of ferrocene modified Fe3 O4 @Au magnetic nanoparticles and its application Jian-Ding Qiu ∗ , Meng Xiong, Ru-Ping Liang, Hua-Ping Peng, Fen Liu Department of Chemistry, Nanchang University, Xue Fu Da Dao 999, Nanchang 330031, China
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Article history: Received 20 October 2008 Received in revised form 16 January 2009 Accepted 19 January 2009 Available online 29 January 2009 Keywords: Fe3 O4 @Au nanoparticles 6-Ferrocenylhexanethiol Dopamine Magnetic carbon paste electrode
a b s t r a c t A novel dopamine sensor was fabricated by forming the 6-ferrocenylhexanethiol (HS(CH2 )6 Fc) functionalized Fe3 O4 @Au nanoparticles (NPs) films on the surface of a carbon paste electrode with the aid of a permanent magnet. HS(CH2 )6 Fc, which acted as the redox mediator, was self-assembled to Fe3 O4 @Au NPs via Au–S bond. Transmission electron microscopy, UV–visible absorption spectroscopy, Fourier transform infrared spectra, and cyclic voltammetry were used to characterize the properties of the Fe3 O4 @Au NPs/HS(CH2 )6 Fc nanocomposite. It is shown that the prepared ferrocene-functionalized Fe3 O4 @Au NPs composite shuttled electrons between analyte and electrode, increased the mediator loading, and more importantly prevented the leakage of the mediator during measurements, which resulted in the substantially enhanced stability and reproducibility of the modified electrode. The electrooxidation of dopamine could be catalyzed by Fc/Fc+ couple as a mediator and had a higher electrochemical response due to the unique performance of Fe3 O4 @Au NPs. The nanocomposite modified electrode exhibited fast response (3 s) and the linear range was from 2.0 × 10−6 to 9.2 × 10−4 M with a detection limit of 0.64 M. This immobilization approach effectively improved the stability of the electron transfer mediator and is promising for construction of other sensors and bioelectronic devices. © 2009 Elsevier B.V. All rights reserved.
1. Introduction Magnetic nanoparticles (NPs) owe their popularity to their numerous attributes such as their magnetic properties that enable them to be directed by an external magnetic field, the possibility to separate them from a reaction mixture, in addition to their low toxicity and biocompatibility. Magnetic NPs as special biomolecule carriers via a suitable immobilization process offer promise as sensitive sensors, and have been used in immunoassays and in various reactions involving enzymes, proteins, and DNA for magnetically controlled transport and target delivery of anticancerous drugs (Niemeyer, 2001). Their reduced size and ability to be transported in biological systems and reacting medium is an advantage over conventional support systems. Unfortunately, iron nanoparticles with a large surface area are easily oxidized, i.e. they react vigorously with oxygen present in the air and also react between themselves forming aggregates. To overcome these limitations in the research of such nanoparticles, the main protocol is to modify the surface during the synthesis or coating process (Zou et al., 2008; Kim et al., 2001; Kouassi and Irudayaraj, 2006). Many techniques and applications for the surface modification such as silica-coated (Fang et al., 2007), silver-coated (Tang et al., 2006), and titania-
∗ Corresponding author. Tel.: +86 791 3969518. E-mail addresses: [email protected], [email protected] (J.-D. Qiu). 0956-5663/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.bios.2009.01.022
coated (Chen and Chen, 2005) magnetic shell/core NPs have been reported. Considerable interest over the past two decades has been directed toward the functionalization of gold NPs because of their excellent biocompatibility, stability, and established synthesis protocols. Furthermore, the use of thiol chemistry on a gold surface allows the attachment of molecules with a relative ease using a variety of thiol linkers (Minard-Basquin et al., 2005; Demers et al., 2000). Hence, if the magnetic particles are provided with a gold coating, then the combined benefits of the robust chemistry for gold surfaces and the uniqueness of magnetic NPs could be realized, in additional, the magnetic cores could be protected from oxidation and corrosion (Demers et al., 2000; Zhou et al., 2001; Lin et al., 2001; Kim et al., 2002; Mikhaylova et al., 2004). Recently, Fan et al. (2005), taking advantage of a magnetic separation/mixing process and the amplification feature of colloidal gold label, used gold-coated magnetic beads for immunoassay development. Pham et al. (2008) has reported that magnetic separation of biological molecules using Au-coated magnetic oxide composite NPs was examined after attachment of protein immunoglobulin (IgG) through electrostatic interactions. Min et al. (2007) has reported that the acetylcholinesterase (AChE) biosensor was fabricated by immobilizing of AChE on the surface of Au-doped magnetic Fe3 O4 NPs and used to determine organophosphorus pesticides. The immobilization of single-stranded biotinylated oligonucleotides onto gold-coated magnetic NPs as a DNA sensor was also conducted (Kouassi and Irudayaraj, 2006).
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Ferrocene and its derivatives are well-known substances as mediators (Petersson, 1986; Kulys and Dcosta, 1991; Escorcia and Dhirani, 2007), due to their various very desirable properties of relatively low molecular mass, good electrochemical reversibility, regeneration at low potential, and generation of stable redox forms (Pham et al., 2008; Qiu et al., 2007a,b; Fernandez and Carrero, 2005). However, for small electron transfer mediators, ferrocene and its derivatives modified electrodes are unstable and difficult to be controlled because of their weak adsorption on electrode surface, and leakage has been a main problem for the ferrocene and its derivatives modified electrodes. This problem can be resolved by covalent linking of mediator with polymer, high molecular weight compounds, or nanoparticles (Masuko et al., 2005; Okawa et al., 1999; Chen et al., 2006; Qiu et al., 2007a,b) before immobilization. Dopamine (DA) is one of the most important neurotransmitters and is ubiquitous in the mammalian central nervous system. The change in the level of DA has been proved to be a very effective route toward brain functions, and the loss of dopamine-containing neurons may result in serious disease such as Parkinson’s disease (Pihel et al., 1996; Cha et al., 1998). Hence, selective and sensitive determination of DA has become important, and electrochemical techniques have been proved to be one of the most advantageous ways in the determination of DA (Pihel et al., 1996; Selvaraju and Ramaraj, 2003; Zhang et al., 2005). It is well known that detection of DA directly with plain electrodes, such as carbon and metallic electrodes (e.g., Au, Pt), is ineffective. Various methods, mainly based on the chemical modification of traditional electrode materials, have been developed to resolve the problem (Wring et al., 1990; Lyons et al., 1991; Leal et al., 1993; Mao and Pickup, 1989). However, develop a simple and rapid electrochemical method to selectively detect DA is still a long-standing goal. Herein, we demonstrated the synthesis of bifunctional nanocomposites using Fe3 O4 @Au NPs covalently bound with HS(CH2 )6 Fc via Au–S bond, opening wide application in biocatalysis and biosensors. The prepared Fe3 O4 @Au NPs composite possessed high surface area, good mechanical stability, and good conductivity, which provided a compatible microenvironment for maintaining the activity of the immobilized mediator, increased the mediator loading, and more importantly prevented the leakage of HS(CH2 )6 Fc. Together with low cost and ease of preparation, the designed Fe3 O4 @Au NPs/HS(CH2 )6 Fc nanocomposite modified electrode was successfully applied for the sensitive determination of dopamine. 2. Materials and methods 2.1. Chemicals Dopamine, HAuCl4 ·4H2 O and Nafion solution (∼5% in a mixture of lower aliphatic alcohols and water) were purchased from Sigma and used as received. 6-Ferrocenylhexanethiol (HS(CH2 )6 Fc) was synthesized according to the literature procedure (Creager and Rowe, 1994). A permanent magnet (3 mm i.d. and 5 mm depth, 0.2 T
at the surface) was purchased from As One Ltd. (Osaka, Japan). 0.1 M phosphate buffer solution (PBS, pH 6.86, containing 0.1 M KClO4 ) was used as supporting electrolyte. All other chemicals were of analytical grade and doubly distilled water was used throughout. 2.2. Preparation of the ferrocene-functionalized Fe3 O4 @Au NPs The synthesis of Fe3 O4 magnetic NPs was achieved in a typical procedure (Kang et al., 1996). Core–shell Fe3 O4 @Au NPs (Fe3 O4 @Au NPs) were prepared by growing Au layers onto the surface of the Fe3 O4 as described by Williams (Lyon et al., 2004). The ferrocene-functionalized Fe3 O4 @Au magnetic NPs were prepared by mixing 1.5 mL Fe3 O4 @Au NPs (0.5 mg/mL) with 0.5 mL HS(CH2 )6 Fc (5.0 mM) with shaking for 48 h under nitrogen surrounding. The products were rinsed with copious amounts of doubly distilled water and hexane, finally suspended in 2.0 mL water. The schematic procedure of the preparation of the ferrocenefunctionalized Fe3 O4 @Au magnetic NPs is described in Scheme 1. 2.3. Preparation of modified electrode A magnetic carbon paste electrode (MCPE) was prepared using literature procedure (Tang et al., 2006). A copper wire was put into the glass tube (3 mm in diameter and 50 mm in depth) initially, and a nummular magnet (3 mm in diameter and 2 mm in thickness, 0.2 T at the surface) was embedded with a depth of 5 mm from the surface of electrode. Paraffin oil (1.2 mL) was mixed with graphite powder (10 mg) thoroughly to get homogeneous paste, and then a portion of the resulting paste was stuffed into the glass tube to immobilize the magnet. The obtained MCPEs were dried and stored for a day at 4 ◦ C. Before modification, the MCPE was polished with ultrafine emery paper until a smooth surface was obtained and cleaned with doubly distilled water. Then 5 L ferrocene-functionalized Fe3 O4 @Au NPs were spread evenly onto the MCPE surface with syringe and allowed to dry to form a ferrocene-functionalized Fe3 O4 @Au NPs film. Prior to use, the modified electrode was washed with PBS, the ferrocene-functionalized Fe3 O4 @Au NPs were tightly attached on the surface of the MCPE in the presence of magnetic force. 2.4. Characterization Transmission electron micrographs (TEM) of magnetic NPs (0.05 mg/mL) were obtained with a Hitach-600 transmission electron microscope (Hitachi). Fourier transform infrared spectra (FTIR) were recorded on a Nicolet 5700 FTIR spectrometer (Nicolet). A UV2450 spectrophotometer (Shimadzu) was used to collect UV–vis data. All the electrochemical experiments were performed on an Autolab PGSTAT30 electrochemical workstation (Eco Chemie). A three-electrode system was used including an Ag/AgCl reference electrode, a platinum wire auxiliary electrode, and the modified MCPE as the working electrode. The electrolyte solutions were
Scheme 1. Schematic illustration of the preparing procedures of the ferrocene-functionalized Fe3 O4 @Au magnetic NPs.
J.-D. Qiu et al. / Biosensors and Bioelectronics 24 (2009) 2649–2653
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purged with N2 for at least 10 min to remove O2 and kept under N2 atmosphere during the measurements. 3. Results and discussion 3.1. Characterization of the magnetic nanoparticles The Fe3 O4 and Fe3 O4 @Au magnetic nanoparticle sizes were characterized by TEM. The TEM photograph illustrates that the average diameter of Fe3 O4 was about 8 ± 2 nm (Fig. 1a). After modification of the Fe3 O4 with Au shell, the resulted core–shell Fe3 O4 @Au composite retained the spherical structure with the average diameter of 30 ± 5 nm (Fig. 1b). The thickness of the Au shell was estimated to be 11 nm. As reported, the thickness of the shell layer could be changed by varying the experimental parameters (Brown et al., 2000). However, proper shell thickness is essential to obtain high enough magnetic-field intensity and stability of the magnetic NPs. The coating of Au on the magnetic NPs facilitated the
Fig. 2. UV–vis absorption spectra of Fe3 O4 (a) and Fe3 O4 @Au (b) nanoparticles.
dispersion of Fe3 O4 NPs. Also the presence of Au shell surface was helpful to complete the further modification of these magnetic NPs (Niemeyer, 2001). When an external magnetic field was applied, the Fe3 O4 @Au NPs can be separated from the solution, which implied that this prepared magnetic NPs can be stably immobilized on the surface of CPE by the magnetic force. Thus, the further ferrocenefunctionalized Fe3 O4 @Au NPs could be firmly immobilized on the surface of CPE in the presence of magnetic field, and the leakage of ferrocene would be efficiently avoided. The UV–vis absorption spectra of the dispersion of pure Fe3 O4 and core–shell Fe3 O4 @Au NPs are shown in Fig. 2. The Fe3 O4 absorption spectra were increased with the wavelength decrease in the range 800–300 nm (Fig. 2a), which was in agreement with the literature (Tang et al., 2006). However, after the Fe3 O4 magnetic NPs were coated with Au shell, 550 nm of the absorption peak was remarkably observed (Fig. 2b), indicating that the core–shell Fe3 O4 @Au NPs were formed by the deposition precipitation method, which was in good agreement with the result in Fig. 1. The formation of ferrocene-functionalized Fe3 O4 @Au NPs was also confirmed by FTIR measurements (not shown). In contrast to the bands in the spectra of Fe3 O4 @Au NPs, the characteristic absorptions of ferrocene appeared at 2926 cm−1 , 1406 cm−1 , 1106 cm−1 , 1043 cm−1 and 810 cm−1 in the spectrum of the ferrocene-functionalized Fe3 O4 @Au NPs (Guan et al., 2005). These results confirmed that the Fe3 O4 @Au NPs were successfully modified with 6-ferrocenylhexanethiol to form ferrocene-functionalized Fe3 O4 @Au conjugate. 3.2. Electrochemical behavior of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE
Fig. 1. TEM images of pure Fe3 O4 (a) and core–shell Fe3 O4 @Au (b) nanoparticles.
Due to the presence of external magnetic field, the ferrocenefunctionalized Fe3 O4 @Au NPs clusters were attracted to the MCPE surface to form an even layer for robust electrochemical determination. Fig. 3, inset A displays the cyclic voltammograms (CVs) of the modified MCPE in PBS. No redox peak at the Fe3 O4 @Au NPs modified MCPE was observed (curve a), whereas the ferrocenefunctionalized Fe3 O4 @Au NPs modified electrode exhibited a pair of well-defined peaks at +353 mV and +388 mV (curve b), which is attributed to the redox of ferrocene group in the ferrocenefunctionalized Fe3 O4 @Au NPs conjugate. The voltammetric stability of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE was checked by the repetitive potential sweep at a scan rate of 50 mV s−1 in PBS. The results showed that the peak currents remained about 95% of the initial one after 100 repeated scans, indicating the ferrocene-functionalized Fe3 O4 @Au NPs modified
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Fig. 4. CVs of bare MCPE (a and b) and ferrocene-functionalized Fe3 O4 @Au NPs modified electrodes (c and d) in the absence (a and c) and presence (b and d) of 1.0 mM DA at a scan rate of 10 mV s−1 . Fig. 3. CVs of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE in PBS at scan rates of 5–1000 mV s−1 (from internal to external). Inset: (A) CVs of Fe3 O4 @Au (a) and ferrocene-functionalized Fe3 O4 @Au NPs (b) modified MCPE in PBS at 50 mV s−1 . (B) Plot of anodic peak current (a) and cathodic peak current (b) vs. v1/2 .
MCPE was fairly stable. The reproducibility of the ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE was also checked carefully by CVs. Results showed that the relative standard deviation of peak currents on the CVs were less than 6.0% for a ferrocene-functionalized Fe3 O4 @Au NPs modified MCPEs after successive six times of surface-renew with coating the same amount of ferrocene-functionalized Fe3 O4 @Au NPs composite, indicating good reproducibility of such kind of electrodes operated by the present method. With an increasing scan rate, the CV peak currents of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE increased and the values of Ep were increased slightly in the scan rates range of 5–1000 mV s−1 (Fig. 3). Inset B shows that the cathodic and anodic peak currents increased linearly with the increase of the square root of scan rates, suggesting that the electrochemical reaction of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE is a nonsurface controlled electrode process, which might be attributed to a slow electron hoping across the matrix of the composite membrane. The result was consistent with other previous studies (Tripathi et al., 2006; Kandimalla et al., 2006).
functionalized Fe3 O4 @Au NPs modified MCPE exhibits excellent electrocatalytic activity toward the oxidation of DA. The electrocatalytic reactions can be explained by the possible reaction scheme (Jiao et al., 2007): Fc → Fc+ + e− (on electrode)
(1)
2Fc+ + DA → 2Fc + DOQ + 2H+
(2)
(DOQ = dehydrodopamine) 3.4. Amperometric response Based on the voltammetric results described above, amperometric current–time response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE was recorded with successive addition of DA in a stirred PBS (Fig. 5). The time required to reach 95% of the maximum steady-state current was less than 3 s, indicating a fast response, which was mainly due to the enhanced electron transfer of DA due to the existence of HS(CH2 )6 Fc and the well conductive properties of the Fe3 O4 loaded Au NPs nanocomposite. With the increasing DA concentration, the amperometric response increased linearly in the range of 2.0 × 10−6 to 9.2 × 10−4 M with a correlation coefficient of 0.999. The detection limit was 0.64 M at a signal to noise of 3, which was much lower than those reported
3.3. Electrochemical response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE to dopamine Since the modified electrode exhibits stable and reversible electrochemical response, it can be used in electrocatalysis as electron transfer mediator to shuttle electrons between analytes and substrate electrodes. DA was selected to evaluate the electrocatalytical ability of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE. Fig. 4 shows the CVs of bare MCPE and ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE in the absence and presence of 1.0 mM DA at a scan rate of 10 mV s−1 . As can be seen, at the bare MCPE, no peak was observed in the absence of DA (Fig. 4a) and a broad and irreversible oxidation peak was obtained at about 0.58 V in the presence of DA (Fig. 4b). When the ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE was placed into PBS, a pair of redox peaks attributing to oxidation and reduction of ferrocene group occurred (Fig. 4c). After DA was added, the anodic peak current increased noticeably and the cathodic peak current decreased (Fig. 4d). The behavior is typical of that expected for mediated oxidation of an irreversible process. The anodic peak potential was 0.38 V, which was about 200 mV lower than that of the bare MCPE. These results indicated that the ferrocene-
Fig. 5. Typical amperometric response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE at 0.38 V after successive addition of DA in 0.1 M PBS. Inset: (A) the magnified curve from 400 s to 1040 s and (B) calibration curve.
J.-D. Qiu et al. / Biosensors and Bioelectronics 24 (2009) 2649–2653 Table 1 Observed recovery of DA in urine samples. Number
[DA] Added (M)
[DA] Found (M)
Recovery (%)
1 2 3 4
– 20.0 50.0 100.0
– 20.0 48.9 103
– 100 97.8 103
previously (47.8 M (Moccelini et al., 2008), 25 M (Tembe et al., 2008), 10 M (Wang et al., 2006), 50 M (Pandey et al., 2001)). 3.5. Interferences In order to assess the possibility of interference from some other bio-compounds, the effects of some common interfering substances such as ascorbic acid (AA) and uric acid (UA) in the determination of DA were evaluated. It was found that 250 M UA did not cause any observable interference, whilst, 80 M AA caused a few interference to the modified electrode response to DA. However, it is well known that Nafion film can effectively eliminate the interferences of anions (Wang and Li, 1989; Dong and Kuwana, 1991; Yu et al., 2003). Thus, Nafion film covered modified electrode was prepared by dropping 5 L 0.5 wt% Nafion solution (5% Nafion was diluted with alcohols) on the surface of the electrode, the response current and response time of DA are the same as the ferrocene-functionalized Fe3 O4 @Au NPs modified electrode, and the interference of UA and AA are omissible. We also studied interference of other compounds. To 1.0 × 10−5 M DA, the following compounds were investigated: cysteine (50), glucose (200), NaCl (400), KCl (400), and CaCl2 (200), where the numbers in parentheses represent the concentration ratios to 1.0 × 10−5 M DA. No shift in the oxidation peak potential of DA was observed in the presence of all the interferents, and the current response of DA was nearly not affected. These results indicating that the modified electrode prepared by this method can be used to selective determination of DA in real samples by controlling the applied potential (0.38 V). 3.6. Determination of DA in biological fluid It is essential to monitor the applicability of the proposed method in biological samples. For this purpose, urine samples were 100-fold diluted with PBS, in order to fit into the linear range and reduce the matrix effect of real samples. Samples were spiked with different quantities of DA as found in the samples. The results obtained are tabulated in Table 1. It was noted that DA was not detectable in urine samples. The recovery values of DA have been found to be in good agreement and very close to their spiked values. This indicates that the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPEs have high selectivity for DA after coated a layer of Nafion, and can be used in a biological matrix where the large amount of AA and UA are present. 4. Conclusions In summary, we have shown a novel bifunctional gold-coated magnetic NPs covered with a large number of ferrocenylalkanethiol molecules, which could be effectively employed for the fabrication of dopamine biosensors by forming the ferrocenefunctionalized Fe3 O4 @Au NPs films on MCPEs. The use of Fe3 O4 @Au NPs as a ferrocene supporter resulted in the substantially enhanced stability and reproducibility and without electroactive mediator leaching during the electrochemical procedure. This composite not
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only provides a suitable environment to accelerate the electronic communication between dopamine and electrode, but also offers the potential for the construction of biosensors and bioelectronic devices. Acknowledgements This work was supported by grants from the National Natural Science Foundation of China (20605010; 20865003; 20805023), the Jiangxi Province Natural Science Foundation (0620039; 2007JZH2644) and the Opening Foundation of State Key Laboratory of Chem/Biosensing and Chemometrics of Hunan University (2006022; 2007012). References Brown, K.R., Walter, D.G., Natan, M.J., 2000. Chem. Mater. 12, 306–313. Cha, C.S., Chen, J., Liu, P.F., 1998. Biosens. Bioelectron. 13, 87–94. Chen, C.T., Chen, Y.C., 2005. Anal. Chem. 77, 5912–5919. Chen, J., Tang, J.H., Yan, F., Ju, H.X., 2006. Biomaterials 27, 2313–2321. Creager, S.E., Rowe, G.K., 1994. J. Electroanal. Chem. 370, 203–211. Demers, L.M., Mirkin, C.A., Mucic, R.C., Reynolds, R.A., Leitsinger, R.L., Viswanadham, G.A., 2000. Anal. Chem. 72, 5535–5541. Dong, S.J., Kuwana, T., 1991. Electroanalysis 3, 485–491. Escorcia, A., Dhirani, A.A., 2007. J. Electroanal. Chem. 601, 260–268. Fan, A., Lau, C., Lu, J., 2005. Anal. Chem. 77, 3238–3242. Fang, H., Ma, C.Y., Wan, T.L., Zhang, M., Shi, W.H., 2007. J. Phys. Chem. C 111, 1065–1070. Fernandez, L., Carrero, H., 2005. Electrochim. Acta 50, 1233–1240. Guan, L.H., Shi, Z.J., Li, M.X., Gu, Z.N., 2005. Carbon 43, 2780–2785. Jiao, S.F., Li, M.G., Wang, C., Chen, D.L., Fang, B., 2007. Electrochim. Acta 52, 5939–5944. Kandimalla, V.B., Tripathi, V.S., Ju, H., 2006. Biomaterials 27, 1167–1174. Kang, Y.S., Risbud, S., Rabolt, J.F., Stroeve, P., 1996. Chem. Mater. 8, 2209–2211. Kim, D.K., Zhang, Y., Kehr, J., Klason, T., Bjelke, B., Muhammed, M., 2001. J. Magn. Magn. Mater. 225, 256–261. Kim, D.K., Mikhaylova, M., Toprak, M., Zhang, Y., Bjelke, B., Kehr, J., Muhammed, M., 2002. Mater. Res. Soc. Symp. Proc. 704, W6.28.1. Kouassi, G.K., Irudayaraj, J., 2006. Anal. Chem. 78, 3234–3241. Kulys, J., Dcosta, E.J., 1991. Anal. Chim. Acta 243, 173–178. Leal, J.M., Domingo, P.L., Garcia, B., Ibeas, S., 1993. J. Chem. Soc., Faraday Trans. 89, 3571–3577. Lin, J., Zhou, W., Kumbhar, A., Weimann, J., Fang, J., Carpenter, E.E., O’Connor, C.J., 2001. J. Solid State Chem. 159, 26–31. Lyon, J.L., Fleming, D.A., Stone, M.B., Schiffer, P., Williams, M.E., 2004. Nano Lett. 4, 719–723. Lyons, M.E.G., Breen, W., Cassidy, J., 1991. J. Chem. Soc., Faraday Trans. 87, 115–123. Mao, H., Pickup, P.G., 1989. J. Electroanal. Chem. 265, 127–142. Masuko, T., Minami, A., Iwasaki, N., Majima, T., Nishimura, S., Lee, Y.C., 2005. Biomacromolecules 6, 880–884. Mikhaylova, M., Kim, D.K., Berry, C.C., Zogorodni, A., Toprak, M., Curtis, A.S.G., Muhammed, M., 2004. Chem. Mater. 16, 2344–2354. Min, H., Qu, Y.H., Li, X.H., Xie, Z.H., Wei, Y.Y., Jin, L.T., 2007. Acta Chim. Sin. 65, 2303–2308. Minard-Basquin, C., Kügler, R., Matsuzawa, N.N., Yasuda, A., 2005. IEEE ProcNanobiotechnol. 152, 97–103. Moccelini, S.K., Fernandes, S.C., Vieira, I.C., 2008. Sens. Actuators B 133, 364–369. Niemeyer, C.M., 2001. Angew. Chem. Int. Ed. 40, 4128–4158. Okawa, Y., Nagano, M., Hirota, S., Kobayashi, H., Ohno, T., Watanabe, M., 1999. Biosens. Bioelectron. 14, 229–235. Pandey, P.C., Upadhyay, S., Tiwari, I., Singh, G., Tripathi, V.S., 2001. Sens. Actuators B 75, 48–55. Petersson, M., 1986. Anal. Chim. Acta 187, 333–338. Pham, T.T.H., Cao, C., Sim, S.J., 2008. J. Magn. Magn. Mater. 320, 2049–2055. Pihel, K., Walker, Q.D., Wightman, R.M., 1996. Anal. Chem. 68, 2084–2089. Qiu, J.D., Guo, J., Liang, R.P., Xiong, M., 2007a. Electroanalysis 19, 2335–2341. Qiu, J.D., Peng, H.P., Liang, R.P., 2007b. Electrochem. Commun. 9, 2734–2738. Selvaraju, T., Ramaraj, R., 2003. Electrochem. Commun. 5, 667–672. Tang, D.P., Yuan, R., Chai, Y.Q., 2006. J. Phys. Chem. B 110, 11640–11646. Tembe, S., Kubal, B.S., Karve, M., Souza, S.F.D., 2008. Anal. Chim. Acta 612, 212–217. Tripathi, V.S., Kandimalla, V.B., Ju, H., 2006. Biosens. Bioelectron. 21, 1529–1535. Wang, H., Wang, L.J., Shi, Z.F., Guo, Y., Cao, X.P., Zhang, H.L., 2006. Electrochem. Commun. 8, 1779–1783. Wang, J., Li, R., 1989. Talanta 36, 279–284. Wring, S.A., Hart, J.P., Birch, B.J., 1990. Anal. Chim. Acta 229, 63–70. Yu, J.H., Liu, S.Q., Ju, H.X., 2003. Biosens. Bioelectron. 19, 401–409. Zhang, M., Gong, K., Zhang, H., Mao, L., 2005. Biosens. Bioelectron. 20, 1270– 1276. Zhou, W.L., Carpenter, E.E., Kumbhar, J.A.L., Sims, J., O’Connor, C.J., 2001. Eur. Phys. J. D 16, 289–292. Zou, Z.Q., Ibisate, M., Zhou, Y., Aebersold, R., Xia, Y.N., Zhang, H., 2008. Anal. Chem. 80, 1228–1234.
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Synthesis and characterization of ferrocene modified Fe3 O4 @Au magnetic nanoparticles and its application Jian-Ding Qiu ∗ , Meng Xiong, Ru-Ping Liang, Hua-Ping Peng, Fen Liu Department of Chemistry, Nanchang University, Xue Fu Da Dao 999, Nanchang 330031, China
a r t i c l e
i n f o
Article history: Received 20 October 2008 Received in revised form 16 January 2009 Accepted 19 January 2009 Available online 29 January 2009 Keywords: Fe3 O4 @Au nanoparticles 6-Ferrocenylhexanethiol Dopamine Magnetic carbon paste electrode
a b s t r a c t A novel dopamine sensor was fabricated by forming the 6-ferrocenylhexanethiol (HS(CH2 )6 Fc) functionalized Fe3 O4 @Au nanoparticles (NPs) films on the surface of a carbon paste electrode with the aid of a permanent magnet. HS(CH2 )6 Fc, which acted as the redox mediator, was self-assembled to Fe3 O4 @Au NPs via Au–S bond. Transmission electron microscopy, UV–visible absorption spectroscopy, Fourier transform infrared spectra, and cyclic voltammetry were used to characterize the properties of the Fe3 O4 @Au NPs/HS(CH2 )6 Fc nanocomposite. It is shown that the prepared ferrocene-functionalized Fe3 O4 @Au NPs composite shuttled electrons between analyte and electrode, increased the mediator loading, and more importantly prevented the leakage of the mediator during measurements, which resulted in the substantially enhanced stability and reproducibility of the modified electrode. The electrooxidation of dopamine could be catalyzed by Fc/Fc+ couple as a mediator and had a higher electrochemical response due to the unique performance of Fe3 O4 @Au NPs. The nanocomposite modified electrode exhibited fast response (3 s) and the linear range was from 2.0 × 10−6 to 9.2 × 10−4 M with a detection limit of 0.64 M. This immobilization approach effectively improved the stability of the electron transfer mediator and is promising for construction of other sensors and bioelectronic devices. © 2009 Elsevier B.V. All rights reserved.
1. Introduction Magnetic nanoparticles (NPs) owe their popularity to their numerous attributes such as their magnetic properties that enable them to be directed by an external magnetic field, the possibility to separate them from a reaction mixture, in addition to their low toxicity and biocompatibility. Magnetic NPs as special biomolecule carriers via a suitable immobilization process offer promise as sensitive sensors, and have been used in immunoassays and in various reactions involving enzymes, proteins, and DNA for magnetically controlled transport and target delivery of anticancerous drugs (Niemeyer, 2001). Their reduced size and ability to be transported in biological systems and reacting medium is an advantage over conventional support systems. Unfortunately, iron nanoparticles with a large surface area are easily oxidized, i.e. they react vigorously with oxygen present in the air and also react between themselves forming aggregates. To overcome these limitations in the research of such nanoparticles, the main protocol is to modify the surface during the synthesis or coating process (Zou et al., 2008; Kim et al., 2001; Kouassi and Irudayaraj, 2006). Many techniques and applications for the surface modification such as silica-coated (Fang et al., 2007), silver-coated (Tang et al., 2006), and titania-
∗ Corresponding author. Tel.: +86 791 3969518. E-mail addresses: [email protected], [email protected] (J.-D. Qiu). 0956-5663/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.bios.2009.01.022
coated (Chen and Chen, 2005) magnetic shell/core NPs have been reported. Considerable interest over the past two decades has been directed toward the functionalization of gold NPs because of their excellent biocompatibility, stability, and established synthesis protocols. Furthermore, the use of thiol chemistry on a gold surface allows the attachment of molecules with a relative ease using a variety of thiol linkers (Minard-Basquin et al., 2005; Demers et al., 2000). Hence, if the magnetic particles are provided with a gold coating, then the combined benefits of the robust chemistry for gold surfaces and the uniqueness of magnetic NPs could be realized, in additional, the magnetic cores could be protected from oxidation and corrosion (Demers et al., 2000; Zhou et al., 2001; Lin et al., 2001; Kim et al., 2002; Mikhaylova et al., 2004). Recently, Fan et al. (2005), taking advantage of a magnetic separation/mixing process and the amplification feature of colloidal gold label, used gold-coated magnetic beads for immunoassay development. Pham et al. (2008) has reported that magnetic separation of biological molecules using Au-coated magnetic oxide composite NPs was examined after attachment of protein immunoglobulin (IgG) through electrostatic interactions. Min et al. (2007) has reported that the acetylcholinesterase (AChE) biosensor was fabricated by immobilizing of AChE on the surface of Au-doped magnetic Fe3 O4 NPs and used to determine organophosphorus pesticides. The immobilization of single-stranded biotinylated oligonucleotides onto gold-coated magnetic NPs as a DNA sensor was also conducted (Kouassi and Irudayaraj, 2006).
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Ferrocene and its derivatives are well-known substances as mediators (Petersson, 1986; Kulys and Dcosta, 1991; Escorcia and Dhirani, 2007), due to their various very desirable properties of relatively low molecular mass, good electrochemical reversibility, regeneration at low potential, and generation of stable redox forms (Pham et al., 2008; Qiu et al., 2007a,b; Fernandez and Carrero, 2005). However, for small electron transfer mediators, ferrocene and its derivatives modified electrodes are unstable and difficult to be controlled because of their weak adsorption on electrode surface, and leakage has been a main problem for the ferrocene and its derivatives modified electrodes. This problem can be resolved by covalent linking of mediator with polymer, high molecular weight compounds, or nanoparticles (Masuko et al., 2005; Okawa et al., 1999; Chen et al., 2006; Qiu et al., 2007a,b) before immobilization. Dopamine (DA) is one of the most important neurotransmitters and is ubiquitous in the mammalian central nervous system. The change in the level of DA has been proved to be a very effective route toward brain functions, and the loss of dopamine-containing neurons may result in serious disease such as Parkinson’s disease (Pihel et al., 1996; Cha et al., 1998). Hence, selective and sensitive determination of DA has become important, and electrochemical techniques have been proved to be one of the most advantageous ways in the determination of DA (Pihel et al., 1996; Selvaraju and Ramaraj, 2003; Zhang et al., 2005). It is well known that detection of DA directly with plain electrodes, such as carbon and metallic electrodes (e.g., Au, Pt), is ineffective. Various methods, mainly based on the chemical modification of traditional electrode materials, have been developed to resolve the problem (Wring et al., 1990; Lyons et al., 1991; Leal et al., 1993; Mao and Pickup, 1989). However, develop a simple and rapid electrochemical method to selectively detect DA is still a long-standing goal. Herein, we demonstrated the synthesis of bifunctional nanocomposites using Fe3 O4 @Au NPs covalently bound with HS(CH2 )6 Fc via Au–S bond, opening wide application in biocatalysis and biosensors. The prepared Fe3 O4 @Au NPs composite possessed high surface area, good mechanical stability, and good conductivity, which provided a compatible microenvironment for maintaining the activity of the immobilized mediator, increased the mediator loading, and more importantly prevented the leakage of HS(CH2 )6 Fc. Together with low cost and ease of preparation, the designed Fe3 O4 @Au NPs/HS(CH2 )6 Fc nanocomposite modified electrode was successfully applied for the sensitive determination of dopamine. 2. Materials and methods 2.1. Chemicals Dopamine, HAuCl4 ·4H2 O and Nafion solution (∼5% in a mixture of lower aliphatic alcohols and water) were purchased from Sigma and used as received. 6-Ferrocenylhexanethiol (HS(CH2 )6 Fc) was synthesized according to the literature procedure (Creager and Rowe, 1994). A permanent magnet (3 mm i.d. and 5 mm depth, 0.2 T
at the surface) was purchased from As One Ltd. (Osaka, Japan). 0.1 M phosphate buffer solution (PBS, pH 6.86, containing 0.1 M KClO4 ) was used as supporting electrolyte. All other chemicals were of analytical grade and doubly distilled water was used throughout. 2.2. Preparation of the ferrocene-functionalized Fe3 O4 @Au NPs The synthesis of Fe3 O4 magnetic NPs was achieved in a typical procedure (Kang et al., 1996). Core–shell Fe3 O4 @Au NPs (Fe3 O4 @Au NPs) were prepared by growing Au layers onto the surface of the Fe3 O4 as described by Williams (Lyon et al., 2004). The ferrocene-functionalized Fe3 O4 @Au magnetic NPs were prepared by mixing 1.5 mL Fe3 O4 @Au NPs (0.5 mg/mL) with 0.5 mL HS(CH2 )6 Fc (5.0 mM) with shaking for 48 h under nitrogen surrounding. The products were rinsed with copious amounts of doubly distilled water and hexane, finally suspended in 2.0 mL water. The schematic procedure of the preparation of the ferrocenefunctionalized Fe3 O4 @Au magnetic NPs is described in Scheme 1. 2.3. Preparation of modified electrode A magnetic carbon paste electrode (MCPE) was prepared using literature procedure (Tang et al., 2006). A copper wire was put into the glass tube (3 mm in diameter and 50 mm in depth) initially, and a nummular magnet (3 mm in diameter and 2 mm in thickness, 0.2 T at the surface) was embedded with a depth of 5 mm from the surface of electrode. Paraffin oil (1.2 mL) was mixed with graphite powder (10 mg) thoroughly to get homogeneous paste, and then a portion of the resulting paste was stuffed into the glass tube to immobilize the magnet. The obtained MCPEs were dried and stored for a day at 4 ◦ C. Before modification, the MCPE was polished with ultrafine emery paper until a smooth surface was obtained and cleaned with doubly distilled water. Then 5 L ferrocene-functionalized Fe3 O4 @Au NPs were spread evenly onto the MCPE surface with syringe and allowed to dry to form a ferrocene-functionalized Fe3 O4 @Au NPs film. Prior to use, the modified electrode was washed with PBS, the ferrocene-functionalized Fe3 O4 @Au NPs were tightly attached on the surface of the MCPE in the presence of magnetic force. 2.4. Characterization Transmission electron micrographs (TEM) of magnetic NPs (0.05 mg/mL) were obtained with a Hitach-600 transmission electron microscope (Hitachi). Fourier transform infrared spectra (FTIR) were recorded on a Nicolet 5700 FTIR spectrometer (Nicolet). A UV2450 spectrophotometer (Shimadzu) was used to collect UV–vis data. All the electrochemical experiments were performed on an Autolab PGSTAT30 electrochemical workstation (Eco Chemie). A three-electrode system was used including an Ag/AgCl reference electrode, a platinum wire auxiliary electrode, and the modified MCPE as the working electrode. The electrolyte solutions were
Scheme 1. Schematic illustration of the preparing procedures of the ferrocene-functionalized Fe3 O4 @Au magnetic NPs.
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purged with N2 for at least 10 min to remove O2 and kept under N2 atmosphere during the measurements. 3. Results and discussion 3.1. Characterization of the magnetic nanoparticles The Fe3 O4 and Fe3 O4 @Au magnetic nanoparticle sizes were characterized by TEM. The TEM photograph illustrates that the average diameter of Fe3 O4 was about 8 ± 2 nm (Fig. 1a). After modification of the Fe3 O4 with Au shell, the resulted core–shell Fe3 O4 @Au composite retained the spherical structure with the average diameter of 30 ± 5 nm (Fig. 1b). The thickness of the Au shell was estimated to be 11 nm. As reported, the thickness of the shell layer could be changed by varying the experimental parameters (Brown et al., 2000). However, proper shell thickness is essential to obtain high enough magnetic-field intensity and stability of the magnetic NPs. The coating of Au on the magnetic NPs facilitated the
Fig. 2. UV–vis absorption spectra of Fe3 O4 (a) and Fe3 O4 @Au (b) nanoparticles.
dispersion of Fe3 O4 NPs. Also the presence of Au shell surface was helpful to complete the further modification of these magnetic NPs (Niemeyer, 2001). When an external magnetic field was applied, the Fe3 O4 @Au NPs can be separated from the solution, which implied that this prepared magnetic NPs can be stably immobilized on the surface of CPE by the magnetic force. Thus, the further ferrocenefunctionalized Fe3 O4 @Au NPs could be firmly immobilized on the surface of CPE in the presence of magnetic field, and the leakage of ferrocene would be efficiently avoided. The UV–vis absorption spectra of the dispersion of pure Fe3 O4 and core–shell Fe3 O4 @Au NPs are shown in Fig. 2. The Fe3 O4 absorption spectra were increased with the wavelength decrease in the range 800–300 nm (Fig. 2a), which was in agreement with the literature (Tang et al., 2006). However, after the Fe3 O4 magnetic NPs were coated with Au shell, 550 nm of the absorption peak was remarkably observed (Fig. 2b), indicating that the core–shell Fe3 O4 @Au NPs were formed by the deposition precipitation method, which was in good agreement with the result in Fig. 1. The formation of ferrocene-functionalized Fe3 O4 @Au NPs was also confirmed by FTIR measurements (not shown). In contrast to the bands in the spectra of Fe3 O4 @Au NPs, the characteristic absorptions of ferrocene appeared at 2926 cm−1 , 1406 cm−1 , 1106 cm−1 , 1043 cm−1 and 810 cm−1 in the spectrum of the ferrocene-functionalized Fe3 O4 @Au NPs (Guan et al., 2005). These results confirmed that the Fe3 O4 @Au NPs were successfully modified with 6-ferrocenylhexanethiol to form ferrocene-functionalized Fe3 O4 @Au conjugate. 3.2. Electrochemical behavior of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE
Fig. 1. TEM images of pure Fe3 O4 (a) and core–shell Fe3 O4 @Au (b) nanoparticles.
Due to the presence of external magnetic field, the ferrocenefunctionalized Fe3 O4 @Au NPs clusters were attracted to the MCPE surface to form an even layer for robust electrochemical determination. Fig. 3, inset A displays the cyclic voltammograms (CVs) of the modified MCPE in PBS. No redox peak at the Fe3 O4 @Au NPs modified MCPE was observed (curve a), whereas the ferrocenefunctionalized Fe3 O4 @Au NPs modified electrode exhibited a pair of well-defined peaks at +353 mV and +388 mV (curve b), which is attributed to the redox of ferrocene group in the ferrocenefunctionalized Fe3 O4 @Au NPs conjugate. The voltammetric stability of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE was checked by the repetitive potential sweep at a scan rate of 50 mV s−1 in PBS. The results showed that the peak currents remained about 95% of the initial one after 100 repeated scans, indicating the ferrocene-functionalized Fe3 O4 @Au NPs modified
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Fig. 4. CVs of bare MCPE (a and b) and ferrocene-functionalized Fe3 O4 @Au NPs modified electrodes (c and d) in the absence (a and c) and presence (b and d) of 1.0 mM DA at a scan rate of 10 mV s−1 . Fig. 3. CVs of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE in PBS at scan rates of 5–1000 mV s−1 (from internal to external). Inset: (A) CVs of Fe3 O4 @Au (a) and ferrocene-functionalized Fe3 O4 @Au NPs (b) modified MCPE in PBS at 50 mV s−1 . (B) Plot of anodic peak current (a) and cathodic peak current (b) vs. v1/2 .
MCPE was fairly stable. The reproducibility of the ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE was also checked carefully by CVs. Results showed that the relative standard deviation of peak currents on the CVs were less than 6.0% for a ferrocene-functionalized Fe3 O4 @Au NPs modified MCPEs after successive six times of surface-renew with coating the same amount of ferrocene-functionalized Fe3 O4 @Au NPs composite, indicating good reproducibility of such kind of electrodes operated by the present method. With an increasing scan rate, the CV peak currents of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE increased and the values of Ep were increased slightly in the scan rates range of 5–1000 mV s−1 (Fig. 3). Inset B shows that the cathodic and anodic peak currents increased linearly with the increase of the square root of scan rates, suggesting that the electrochemical reaction of ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE is a nonsurface controlled electrode process, which might be attributed to a slow electron hoping across the matrix of the composite membrane. The result was consistent with other previous studies (Tripathi et al., 2006; Kandimalla et al., 2006).
functionalized Fe3 O4 @Au NPs modified MCPE exhibits excellent electrocatalytic activity toward the oxidation of DA. The electrocatalytic reactions can be explained by the possible reaction scheme (Jiao et al., 2007): Fc → Fc+ + e− (on electrode)
(1)
2Fc+ + DA → 2Fc + DOQ + 2H+
(2)
(DOQ = dehydrodopamine) 3.4. Amperometric response Based on the voltammetric results described above, amperometric current–time response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE was recorded with successive addition of DA in a stirred PBS (Fig. 5). The time required to reach 95% of the maximum steady-state current was less than 3 s, indicating a fast response, which was mainly due to the enhanced electron transfer of DA due to the existence of HS(CH2 )6 Fc and the well conductive properties of the Fe3 O4 loaded Au NPs nanocomposite. With the increasing DA concentration, the amperometric response increased linearly in the range of 2.0 × 10−6 to 9.2 × 10−4 M with a correlation coefficient of 0.999. The detection limit was 0.64 M at a signal to noise of 3, which was much lower than those reported
3.3. Electrochemical response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE to dopamine Since the modified electrode exhibits stable and reversible electrochemical response, it can be used in electrocatalysis as electron transfer mediator to shuttle electrons between analytes and substrate electrodes. DA was selected to evaluate the electrocatalytical ability of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE. Fig. 4 shows the CVs of bare MCPE and ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE in the absence and presence of 1.0 mM DA at a scan rate of 10 mV s−1 . As can be seen, at the bare MCPE, no peak was observed in the absence of DA (Fig. 4a) and a broad and irreversible oxidation peak was obtained at about 0.58 V in the presence of DA (Fig. 4b). When the ferrocenefunctionalized Fe3 O4 @Au NPs modified MCPE was placed into PBS, a pair of redox peaks attributing to oxidation and reduction of ferrocene group occurred (Fig. 4c). After DA was added, the anodic peak current increased noticeably and the cathodic peak current decreased (Fig. 4d). The behavior is typical of that expected for mediated oxidation of an irreversible process. The anodic peak potential was 0.38 V, which was about 200 mV lower than that of the bare MCPE. These results indicated that the ferrocene-
Fig. 5. Typical amperometric response of the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPE at 0.38 V after successive addition of DA in 0.1 M PBS. Inset: (A) the magnified curve from 400 s to 1040 s and (B) calibration curve.
J.-D. Qiu et al. / Biosensors and Bioelectronics 24 (2009) 2649–2653 Table 1 Observed recovery of DA in urine samples. Number
[DA] Added (M)
[DA] Found (M)
Recovery (%)
1 2 3 4
– 20.0 50.0 100.0
– 20.0 48.9 103
– 100 97.8 103
previously (47.8 M (Moccelini et al., 2008), 25 M (Tembe et al., 2008), 10 M (Wang et al., 2006), 50 M (Pandey et al., 2001)). 3.5. Interferences In order to assess the possibility of interference from some other bio-compounds, the effects of some common interfering substances such as ascorbic acid (AA) and uric acid (UA) in the determination of DA were evaluated. It was found that 250 M UA did not cause any observable interference, whilst, 80 M AA caused a few interference to the modified electrode response to DA. However, it is well known that Nafion film can effectively eliminate the interferences of anions (Wang and Li, 1989; Dong and Kuwana, 1991; Yu et al., 2003). Thus, Nafion film covered modified electrode was prepared by dropping 5 L 0.5 wt% Nafion solution (5% Nafion was diluted with alcohols) on the surface of the electrode, the response current and response time of DA are the same as the ferrocene-functionalized Fe3 O4 @Au NPs modified electrode, and the interference of UA and AA are omissible. We also studied interference of other compounds. To 1.0 × 10−5 M DA, the following compounds were investigated: cysteine (50), glucose (200), NaCl (400), KCl (400), and CaCl2 (200), where the numbers in parentheses represent the concentration ratios to 1.0 × 10−5 M DA. No shift in the oxidation peak potential of DA was observed in the presence of all the interferents, and the current response of DA was nearly not affected. These results indicating that the modified electrode prepared by this method can be used to selective determination of DA in real samples by controlling the applied potential (0.38 V). 3.6. Determination of DA in biological fluid It is essential to monitor the applicability of the proposed method in biological samples. For this purpose, urine samples were 100-fold diluted with PBS, in order to fit into the linear range and reduce the matrix effect of real samples. Samples were spiked with different quantities of DA as found in the samples. The results obtained are tabulated in Table 1. It was noted that DA was not detectable in urine samples. The recovery values of DA have been found to be in good agreement and very close to their spiked values. This indicates that the ferrocene-functionalized Fe3 O4 @Au NPs modified MCPEs have high selectivity for DA after coated a layer of Nafion, and can be used in a biological matrix where the large amount of AA and UA are present. 4. Conclusions In summary, we have shown a novel bifunctional gold-coated magnetic NPs covered with a large number of ferrocenylalkanethiol molecules, which could be effectively employed for the fabrication of dopamine biosensors by forming the ferrocenefunctionalized Fe3 O4 @Au NPs films on MCPEs. The use of Fe3 O4 @Au NPs as a ferrocene supporter resulted in the substantially enhanced stability and reproducibility and without electroactive mediator leaching during the electrochemical procedure. This composite not
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only provides a suitable environment to accelerate the electronic communication between dopamine and electrode, but also offers the potential for the construction of biosensors and bioelectronic devices. Acknowledgements This work was supported by grants from the National Natural Science Foundation of China (20605010; 20865003; 20805023), the Jiangxi Province Natural Science Foundation (0620039; 2007JZH2644) and the Opening Foundation of State Key Laboratory of Chem/Biosensing and Chemometrics of Hunan University (2006022; 2007012). References Brown, K.R., Walter, D.G., Natan, M.J., 2000. Chem. Mater. 12, 306–313. Cha, C.S., Chen, J., Liu, P.F., 1998. Biosens. Bioelectron. 13, 87–94. Chen, C.T., Chen, Y.C., 2005. Anal. Chem. 77, 5912–5919. Chen, J., Tang, J.H., Yan, F., Ju, H.X., 2006. Biomaterials 27, 2313–2321. Creager, S.E., Rowe, G.K., 1994. J. Electroanal. Chem. 370, 203–211. Demers, L.M., Mirkin, C.A., Mucic, R.C., Reynolds, R.A., Leitsinger, R.L., Viswanadham, G.A., 2000. Anal. Chem. 72, 5535–5541. Dong, S.J., Kuwana, T., 1991. Electroanalysis 3, 485–491. Escorcia, A., Dhirani, A.A., 2007. J. Electroanal. Chem. 601, 260–268. Fan, A., Lau, C., Lu, J., 2005. Anal. Chem. 77, 3238–3242. Fang, H., Ma, C.Y., Wan, T.L., Zhang, M., Shi, W.H., 2007. J. Phys. Chem. C 111, 1065–1070. Fernandez, L., Carrero, H., 2005. Electrochim. Acta 50, 1233–1240. Guan, L.H., Shi, Z.J., Li, M.X., Gu, Z.N., 2005. Carbon 43, 2780–2785. Jiao, S.F., Li, M.G., Wang, C., Chen, D.L., Fang, B., 2007. Electrochim. Acta 52, 5939–5944. Kandimalla, V.B., Tripathi, V.S., Ju, H., 2006. Biomaterials 27, 1167–1174. Kang, Y.S., Risbud, S., Rabolt, J.F., Stroeve, P., 1996. Chem. Mater. 8, 2209–2211. Kim, D.K., Zhang, Y., Kehr, J., Klason, T., Bjelke, B., Muhammed, M., 2001. J. Magn. Magn. Mater. 225, 256–261. Kim, D.K., Mikhaylova, M., Toprak, M., Zhang, Y., Bjelke, B., Kehr, J., Muhammed, M., 2002. Mater. Res. Soc. Symp. Proc. 704, W6.28.1. Kouassi, G.K., Irudayaraj, J., 2006. Anal. Chem. 78, 3234–3241. Kulys, J., Dcosta, E.J., 1991. Anal. Chim. Acta 243, 173–178. Leal, J.M., Domingo, P.L., Garcia, B., Ibeas, S., 1993. J. Chem. Soc., Faraday Trans. 89, 3571–3577. Lin, J., Zhou, W., Kumbhar, A., Weimann, J., Fang, J., Carpenter, E.E., O’Connor, C.J., 2001. J. Solid State Chem. 159, 26–31. Lyon, J.L., Fleming, D.A., Stone, M.B., Schiffer, P., Williams, M.E., 2004. Nano Lett. 4, 719–723. Lyons, M.E.G., Breen, W., Cassidy, J., 1991. J. Chem. Soc., Faraday Trans. 87, 115–123. Mao, H., Pickup, P.G., 1989. J. Electroanal. Chem. 265, 127–142. Masuko, T., Minami, A., Iwasaki, N., Majima, T., Nishimura, S., Lee, Y.C., 2005. Biomacromolecules 6, 880–884. Mikhaylova, M., Kim, D.K., Berry, C.C., Zogorodni, A., Toprak, M., Curtis, A.S.G., Muhammed, M., 2004. Chem. Mater. 16, 2344–2354. Min, H., Qu, Y.H., Li, X.H., Xie, Z.H., Wei, Y.Y., Jin, L.T., 2007. Acta Chim. Sin. 65, 2303–2308. Minard-Basquin, C., Kügler, R., Matsuzawa, N.N., Yasuda, A., 2005. IEEE ProcNanobiotechnol. 152, 97–103. Moccelini, S.K., Fernandes, S.C., Vieira, I.C., 2008. Sens. Actuators B 133, 364–369. Niemeyer, C.M., 2001. Angew. Chem. Int. Ed. 40, 4128–4158. Okawa, Y., Nagano, M., Hirota, S., Kobayashi, H., Ohno, T., Watanabe, M., 1999. Biosens. Bioelectron. 14, 229–235. Pandey, P.C., Upadhyay, S., Tiwari, I., Singh, G., Tripathi, V.S., 2001. Sens. Actuators B 75, 48–55. Petersson, M., 1986. Anal. Chim. Acta 187, 333–338. Pham, T.T.H., Cao, C., Sim, S.J., 2008. J. Magn. Magn. Mater. 320, 2049–2055. Pihel, K., Walker, Q.D., Wightman, R.M., 1996. Anal. Chem. 68, 2084–2089. Qiu, J.D., Guo, J., Liang, R.P., Xiong, M., 2007a. Electroanalysis 19, 2335–2341. Qiu, J.D., Peng, H.P., Liang, R.P., 2007b. Electrochem. Commun. 9, 2734–2738. Selvaraju, T., Ramaraj, R., 2003. Electrochem. Commun. 5, 667–672. Tang, D.P., Yuan, R., Chai, Y.Q., 2006. J. Phys. Chem. B 110, 11640–11646. Tembe, S., Kubal, B.S., Karve, M., Souza, S.F.D., 2008. Anal. Chim. Acta 612, 212–217. Tripathi, V.S., Kandimalla, V.B., Ju, H., 2006. Biosens. Bioelectron. 21, 1529–1535. Wang, H., Wang, L.J., Shi, Z.F., Guo, Y., Cao, X.P., Zhang, H.L., 2006. Electrochem. Commun. 8, 1779–1783. Wang, J., Li, R., 1989. Talanta 36, 279–284. Wring, S.A., Hart, J.P., Birch, B.J., 1990. Anal. Chim. Acta 229, 63–70. Yu, J.H., Liu, S.Q., Ju, H.X., 2003. Biosens. Bioelectron. 19, 401–409. Zhang, M., Gong, K., Zhang, H., Mao, L., 2005. Biosens. Bioelectron. 20, 1270– 1276. Zhou, W.L., Carpenter, E.E., Kumbhar, J.A.L., Sims, J., O’Connor, C.J., 2001. Eur. Phys. J. D 16, 289–292. Zou, Z.Q., Ibisate, M., Zhou, Y., Aebersold, R., Xia, Y.N., Zhang, H., 2008. Anal. Chem. 80, 1228–1234.