Ion transport through a porphyrin-terminated hybrid bilayer membrane

Electrochimica Acta 56 (2011) 1076–1081

Contents lists available at ScienceDirect

Electrochimica Acta journal homepage: www.elsevier.com/locate/electacta

Ion transport through a porphyrin-terminated hybrid bilayer membrane Yongxin Li a,∗ , Yuanli Chen a , Lun Wang a , Yueping Fang b,∗∗ a b

College of Chemistry and Materials Science, Anhui Normal University, 1# Beijing East Rd, Wuhu 241000, Anhui, China Institute of Biomaterial, College of Science, South China Agricultural University, Guangzhou 510642, China

a r t i c l e

i n f o

Article history: Received 21 July 2010 Received in revised form 8 October 2010 Accepted 15 October 2010 Available online 21 October 2010 Keywords: Ion transport Hybrid bilayer membrane Glassy carbon electrode Electrochemistry

a b s t r a c t A hybrid bilayer membrane (HBM) has been prepared on a glassy carbon electrode (GCE). The HBM consists of an inner layer of n-hexadecylamine (HDA) covalently attached at the GCE and an out-layer of tetra-(N-hexadecylpyridiniurnyl)porphyrin (TC16 PyP). The HDA was covalently bounded on the carbon surfaces through its primary amine under cyclic voltammetric potential scans forming a self-assembled monolayer (SAM). The TC16 PyP forms a second layer on top of the HDA layer due to hydrophobic interaction. The TC16 PyP/HDA/GCE assembly has been utilized to study ion transport through hybrid bilayer membranes. The ion transport through this porphyrin-terminated HBM has been found to strongly dependent on pH. It allows negatively charged redox ions, Fe(CN)6 3− , to penetrate at pH <6 while repelling positively charged Ru(NH3 )6 3+ . However, no faradaic current has been detected at pH >6 with either ions. We believe that electrostatic interaction between the redox ions and the charged membrane is the main reason for this pH dependent ion transport. © 2010 Elsevier Ltd. All rights reserved.

1. Introduction Lipid films, liposomes, and black lipid membranes (BLM) have been used to study the structure and transport properties of biological membranes for many years [1–5]. More recently, there has been growing interest in solid-supported bilayer lipid membranes (ssBLMs). Two main ssBLMs have been widely utilized. The first ssBLM contains only phospholipids, which can be easily formed on metals, glassy carbon or hydrogel [2,6–8] The second kind of ssBLM is the HBM. A traditional HBM system contains of a phospholipids layer and a synthetic layer (e.g., alkanethiol) components, which can be easily formed by self-assembly on a metal surface [9–13]. There are a number of advantages of using HBMs. First, it increases the stability of membrane: HBM can be kept intact and studied for months, and they are predicted to have significantly more mechanical stability than suspended BLMs [12]. Second, by forming a HBM on an electrode, it allows for direct measurement of the molecular flux at the electrode [14–16] For example, the use of a gold electrode permits the application of electrochemical method for examining the insulating character of the lipid layers [6,8,17,18], and for assessing the activity of membrane protein pores [19], redox enzymes [20], proton translocators [21], and ionophores [22,23]. Moreover, because HBM is formed on a surface, many other sur-

face analysis techniques that had not been generally applied to biological membranes are now accessible [14–16]. Compared with metal electrodes, such as Pt and Au electrodes, the GCE is has many unique advantages in electrochemical experiments because of its inertion and wide potential window in aqueous solution. Modification of carbon materials is of interest to material science and electrochemistry [24–26]. Deinhammer et al. [27] reported a method for the covalent modification of GCE with primary amines. The attached molecules are shown to form a compact monolayer of amino groups [27–29]. Herein, we report the formation of a new HBM, as shown in Scheme 1, by first modifying the GCE with n-hexadecylamine followed by a second modification with tetra-(N-hexadecylpyridiniurnyl)porphyrin. This new HBM, which is extremely sensitive to the pH change, has allowed for direct measurement of the transport properties of porphyrin terminated lipid membranes. In acidic solutions, the HBM allows for fast transport of negatively charged ions, whereas it effectively blocks negatively charged ions in neutral and basic solutions. In addition, the pH dependent change in transport has been found to be reversible. This HBM-modified GCE can be used to assess the properties of artificial bio-membranes. 2. Experimental 2.1. Chemicals

∗ Corresponding author. Tel.: +1 86 553 3869302; fax: +1 86 553 3869303. ∗∗ Corresponding author. E-mail addresses: [email protected] (Y. Li), [email protected] (Y. Fang). 0013-4686/$ – see front matter © 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.electacta.2010.10.040

The tetra-(N-hexadecylpyridiniurnyl)porphyrin (TC16 PyP) was synthesized as reported [30], the molecular structure of which is schematically shown in Fig. 1. The compound n-hexadecylamine

Y. Li et al. / Electrochimica Acta 56 (2011) 1076–1081

1077

Scheme 1. A schematic representation of the HBM on a carbon electrode. Out-layer is TC16PyP and Inner layer is HDA monolayer. Substrate is a glassy carbon electrode.

(HDA) was purchased from Sigma and used without further purification. Analytical grade K3 Fe(CN)6 , disodium hydrogen phosphate, sodium dihydrogen phosphate, and lithium perchlorate were obtained from Beijing Chemical Reagent Factory (Beijing, China). Analytical grade Ru(NH3 )6 Cl3 was purchased from Aldrich. All other reagents were of analytical grade and used as supplied. All aqueous solutions were made using double distilled water. 2.2. Apparatus Electrochemical experiments were performed using a model CHI660A electrochemical workstation (CHI, Shanghai, China). A three-electrode system was used in all electrochemical measurements. A saturated calomel electrode (SCE) was used as the reference electrode, and a platinum coil was used as the counter electrode. The SCE reference electrode was separated from the bulk of the solution by a fritted glass disk. A 4.0-mm-diamter glassy carbon disk electrode (formal area = 0.126 cm2 ) was used to form HBMs. The effective surface area of this electrode was calibrated as 0.151 cm2 using cyclic voltammetry (CV) of K3 Fe(CN)6 solution. All experiments were performed at room temperature. The electrochemical solutions were thoroughly deoxygenated by N2 before sampling and an N2 atmosphere was maintained throughout the experiments.

Fig. 2. Multicircle cyclic voltammograms of 5 mM HDA in ethanol containing 0.1 M LiClO4 at a GCE. Scan rate: 20 mV/s.

Electrochemical impedance spectra (EIS) were measured in the frequency range of 0.1–100 kHz at CHI 660A. The measurements were conducted at the formal potential of Fe(CN)6 4−/3− redox couple in 0.1 M Fe(CN)6 4− + Fe(CN)6 3− (1:1) + 0.1 M KCl solution. X-ray photoelectron spectroscopy (XPS) measurements were performed with an ESCA lab MK2 (VG, UK) with a Mg K␣ radiation source at 50 and 0.05 eV per step. The elemental nitrogen-to-carbon ration (N/C) was used as a parameter for assessing the extent of modifier coverage. Values for N/C were calculated by dividing the total number of counts under the N (1s) band by that under the C (1s) band and multiplying the results by 100 after accounting for differences in sensitivity factors [31]. 2.3. Preparation of the HBM on a GCE A 2 mg/ml TC16 PyP/chloroform solution was prepared as solution A. A bare GCE was pretreated by polishing carefully to a mirror-like finish with sand papers of different grades, ultrasonicated for 5 min in ethanol and water, successively. Then, the electrode was immersed in a ethanol solution containing 5 mM HDA and 0.1 M LiClO4 (supporting electrolyte) for potential scanning in the range of 0.0 ∼1.6 V for several scans until the oxidation wave at ∼1.45 V almost disappeared, from which a monolayer of HDA was obtained covalently attached at the GCE, denoted as the HDA/GCE. After rinsing with ethanol, sonicating in pH 7.0 phosphate buffer solution (PBS), drying under highly purified nitrogen, the HDA/GCE was immersed in solution A to allow an adsorption of TC16 PyP for 30 s, then immediately transferred into pH 7.04 PBS for aging 20 min. The final electrode was denoted as the TC16 PyP/HDA/GCE. 3. Results and discussion 3.1. Covalent modification of HDA on GCE

Fig. 1. The molecular structure of tetra-(N-hexadecylpyridiniurnyl)porphyrin.

Fig. 2 shows typical voltammetric responses at a GCE in an ethanol solution containing 5 mM HDA and 0.1 M LiClO4 . In the first scan, HDA presents a single broad oxidation wave at 1.45 V (curve a); no reduction wave can be seen on the reverse scan even with scan rates as high as 2.0 V/s. This phenomenon indicates that the HDA species in the solution undergoes an electron transfer process followed by fast chemical reactions: HDA undergoes a one-electron oxidation step of the amino group and produces cation radicals, which will lose a proton by a fast chemical reaction process, and form a carbon–nitrogen bond in an ethanol solution [27–29,32–34]. In our case, the electrogenerated cation radicals react with aromatic

1078

Y. Li et al. / Electrochimica Acta 56 (2011) 1076–1081

Fig. 3. XPS of N(s) region for n-hexadecylamine monolayer-modified GCE (HDA/GCE).

moieties of GCE surface and form carbon–nitrogen bond, similar to the electrochemical oxidation of some other amine-containing compounds [27]. Furthermore, it can be seen from Fig. 2 that the oxidation wave current decreased (b–h) quickly in the successive scans and finally reached to a steady response for scan number larger than 8, which may be due to the passivation effect of GCE when HDA is immobilized on its surface [29]. The immobilization of HDA is verified by XPS, as shown in Fig. 3. A N(1s) peak was observed at 399.4 eV, which is consistent with a formation of carbon–nitrogen bond [25]. This wave remained for the HDA/GCE after sonicating the electrode in PBS for 20 min, demonstrating the strong bonding between HDA and GCE. For comparison, the XPS of bare GCE is also given (Supporting Information), and no peak appeared in N(1s) region. 3.2. The formation of TC16 PyP/HDA/GCE The TC16 PyP/HDA/GCE was fabricated by immersing the HDA/GCE in a TC16 PyP solution. The TC16 PyP layer was assembled by hydrophobic interactions between the HDA modified layer and the hydrophobic n-hexadecyl chains of the porphine, which can be shown in Scheme 1. Fig. 4 shows cyclic voltammograms of the modified GCE in a 5 mM K3 Fe(CN)6 solution. At a bare GCE, a couple of CV peaks appeared at midpoint potential (Em ) of 0.232 V with peakto peak separation (Ep ) of ∼65 mV (curve a). After modification with HDA, a sharp decrease in the peak current was observed in the voltammetric response. An increase in the peak-to-peak separation

Fig. 4. Cyclic voltammograms of 5 mM K3 Fe(CN)6 in 0.1 M KCl at GCE (a), HDA/GCE (b) and TC16 PyP/HDA/GCE (c). Scan rate: 50 mV/s.

Fig. 5. AC impedance spectroscopies of (a) HDA modified GCE, (b) TC16 PyP/HDA modified GCE in 1 mM Fe(CN)6 3−/4− , supporting electrolyte, 0.1 M KCl. Inset: AC impedance spectroscopy of bare GCE. pH, 7.0.

was also found (curve b), which indicates the formation of the HDA monolayer on the GCE. Further modification with TC16 PyP caused another decrease of the peak current (curve c). Electrochemical impedance spectroscopy is an effective method to probe the features of surface-modified electrodes. A lot of structural and functional information can be derived from impedance analysis [28,35]. Fig. 5 illustrates the results of impedance spectroscopy on bare GCE (inset), HDA/GCE (a) and TC16 PyP/HDA/GCE (b) in the presence of equivalent 1 mM Fe(CN)6 4−/3− + 0.1 M KCl, which are measured at the formal potential of Fe(CN)6 4−/3− . It can be seen at the bare GCE, a semicircle of about 280  diameter with an almost straight tail line are present, implying very low electron transfer resistance to the redox-probe dissolved in the electrolyte solution. The diameter of the high frequency semicircle was significantly enlarged by the surface deposition of the HDA layer (a), an Rct value of ∼6350  be estimated, which indicates an increased resistance to the anion redox reaction at the HDA/GCE. Furthermore, the diameter of the high frequency semicircle was obviously increased at the surface of TC16 PyP/HDA/GCE (b), a charge transfer resistance value of more than 30 k can be estimated, which indicates a huge increased resistance to the anion redox reaction at the TC16 PyP/HDA/GCE. The impedance change of the modification process indicated that the formation of hydride bilayer on the surface of GCE. To verify the presence of the TC16 PyP layer on the surface of HDA/GCE, the TC16 PyP/HDA/GCE was scanned in an acetonitrile solution containing 0.1 M NBu4 BF4 as supporting electrolyte after the electrode was rinsed and sonicated in water. As seen in Fig. 6, in the oxidative scan, an irreversible oxidation wave appeared at 1.37 V (vs SCE). This peak may be attributed to the formation of porphyrin cation radicals [36]. Another redox wave was shown at about −0.75 V in the reductive scan, which may be attributed to the formation of porphyrin anion radicals [36,37]. For comparison, a bare GC electrode was scanned from −1.6 V to 1.6 V in a 10 ␮g/ml TC16 PyP acetonitrile solution containing 0.1 M tetrabutylammonium tetrafluoroborate (NBu4 BF4 ), and the same oxidation and reduction waves were obtained (Supporting Information). It must be pointed out that the peak potential and the shape of TC16 PyP in this paper are different with the results reported previously, which may be due to the N-hexadecylpyridiniurnyl group on TC16 PyP though more details need investigate. It can be concluded that TC16 PyP has been formed on the surface of HDA/GCE forming a HBM. The surface concentration of TC16 PyP can be estimated using

Y. Li et al. / Electrochimica Acta 56 (2011) 1076–1081

Fig. 6. Cyclic voltammograms of TC16 PyP/HDA/GCE in acetonitrile/0.1 M NBu4 BF4. Scan rate: 50 mV/s.

1079

Fig. 8. Reproducibility of the peak current values of 5 mM K3 Fe(CN)6 in 0.1 M KCl measured using TC16 PyP/HDA/GCE.

the following equation [38], Q = nFA ∗

(1)

where F = 96,485, Q represents the charge under the peak at ∼0.75 V in Fig. 6,  * is the coverage of TC16 PyP, n and A stand for the electron number and the area of the GC electrode, respectively. The surface concentration of TC16 PyP was estimated to be 4.87 × 10−10 mol/cm2 , which is in agreement of monolayer adsorption [29]. 3.3. Ion transport though HBM Fig. 7 displays the voltammetric responses of the HBM coated electrode in a 5 mM K3 Fe(CN)6 solution containing 10 mM KCl. Curve a shows a reversible voltammetric response of K3 Fe(CN)6 at a bare GCE (pH 7.0). The redox reactions were significantly inhibited by the presence of the hybrid membrane at the TC16 PyP/HDA/GCE (curve b), indicating a strong blocking from the HBM at neutral pH. Curve c shows the voltammetric response of the same modified electrode at a pH of 2.0, the distinct redox waves were gained at potentials almost the same as those observed at the bare GCE. It can be seen that the ion transport of Fe(CN)6 3− through this HBM has been greatly promoted by decreasing the solution pH. For comparison, the transport behavior of HDA/GCE was also checked in the same solutions, the results were listed in the inset part of Fig. 10, it can be seen that Fe(CN)6 3− could not permeate through HDA/GCE in the absence of TC16 PyP layer in the selected pH ranges because

Fig. 7. Cyclic voltammograms of 5 mM K3 Fe(CN)6 in 0.1 M KCl at GCE (a) and at TC16 PyP/HDA/GCE. pH: b, 7.0; c, 2.0; d, 8.0; e, 4.0. Scan rate: 50 mV/s.

HDA layer can block the electron transfer between bulk solution and electrode surface. However, when TC16PyP layer modified on the surface of HAD/GCE, probably the long chain group in TC16PyP molecule, N-hexadecylpyridiniurnyl, can interact with HDA layer due to strongly hydrophobic interaction, and form nanopores or nanogaps for Fe(CN)6 3− to permeate. When the solution pH is higher than 6.0, the ion-transport of Fe(CN)6 3− is significantly inhibited possibly due to a dense packing of the hybrid membrane. The overall dissociation constant of tetra(4-N-methylpyridyl)porphyrin has been reported to be 4.97 [39]. Therefore, the out-layer membrane is mainly neutral at pH ≥6 creating a dense membrane. At lower pH, a significant amount of the porphyrin molecules are protonated creating relatively stronger electrostatic attraction between the HBM and the redox species, Fe(CN)6 3− [40–43]. So the anionic ions, Fe(CN)6 3− can permeate through the positively charged HBM to the electrode surface due to electrostatic attraction. Due to the large size of Fe(CN)6 3− [44–46], the redox ions most likely reach the electrode from spaces in between porphyrins instead of permeating through the porphyrin rings. The enhanced ion transport thus may be attributed to the electrostatic repulsion between the positively charged porphyrin rings, forming nanopores or nanogaps for Fe(CN)6 3− to permeate. It was demonstrated that the redox current of K3 Fe(CN)6 was inhibited again when the solution pH was adjusted to 8.0, indicating a reversible change in the HBM (shown in Fig. 7, curve d). When the solution pH was adjusted to 4.0, the voltammetric response was regenerated (Fig. 7, curve e). The behavior of the ion transport through the HBM can be reversibly controlled many times by adjusting the pH without obvious loss, as shown in Fig. 8. The following experiments were performed to confirm that electrostatic interaction is the main reason for the controlled ion transport. First, we selected Ru(NH3 )6 3+ as the oppositely charged marker ions to test the transport behavior of HBM. The results were given in Fig. 9. It can be seen that for the unmodified GCE welldefined reversible behavior was observed for both redox couples as expected (curve a). For the TC16 PyP/HDA/GCE, the electrochemistry of both redox couples is blocked by the HBM when the pH of the solution is 7.0 (curve b). In addition, no apparent redox current has been observed in a Ru(NH3 )6 3+ solution at pH of 2.0 (curve c). Comparing with the negatively charged Fe(CN)6 3− probe, we can find that the HBM shows permselectivity at low pH, only allowing the negatively charged Fe(CN)6 3− to reach the underlying electrode (Fig. 7) while excluding positively charged Ru(NH3 )6 3+ . This phenomenon supports our statement that the enhanced ion trans-

1080

Y. Li et al. / Electrochimica Acta 56 (2011) 1076–1081

bility. Moreover, the modified GCE has fast response (∼10 s) to the solution pH, which is advantageous over many traditional potentiometric pH sensors [47,48]. Furthermore, after being stored at a temperature of about −4 ◦ C for two weeks, the HBM-modified GCE can still be used to determine pH with almost the same detecting ability of freshly prepared sensor. This property shows that HBM-modified GCE can be used as a novel pH sensor. 4. Conclusions

Fig. 9. Cyclic voltammograms of 5 mM Ru(NH3 )6 3+ in 0.1 M KCl at the bare GCE (a) and at TC16 PyP/HDA/GCE. pH: b, 7.0; c, 2.0. Scan rate: 50 mV/s.

A new hybrid bilayer membrane, TC16 PyP/HDA, has been prepared on a glassy carbon electrode. The ion-transport behavior of the membrane was studied in redox solutions. The HBM has been found to allow for reversible transport of negatively charged redox species, Fe(CN)6 3− , by controlling the solution pH. The HBMmodified glassy carbon electrode offers potentials as novel pH sensors in pH range of 1–6, and can be used to study transport properties of artificial bio-membranes. Acknowledgements The authors gratefully acknowledge financial support from Natural Science Foundation of China (No. 20975002) and Anhui Normal University. YL thanks Prof. Bo Zhang (University of Washington) and Prof. Takashi Ito (Kansas State University) for their useful suggestion and discussion. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.electacta.2010.10.040. References

Fig. 10. Plot of anodic peak current (ipa ) of 5 mM K3 Fe(CN)6 vs pH at TC16 PyP/HDA/GCE in 0.1 M KCl. Inset: the plot of anodic peak current of 5 mM K3 Fe(CN)6 vs pH at HDA/GCE in 0.1 M KCl. Scan rate: 50 mV/s.

port of HBM at low pH was mainly due to electrostatic attraction between the charged membrane and the redox molecules. The effect of ionic strength on the transport behavior of HBM was also investigated by changing KCl concentration in a 5 mM K3 Fe(CN)6 solution at pH 2.0 (data not shown). The results showed that the oxidation peak current of K3 Fe(CN)6 at the HBM-modified GCE decreased gradually to about 26% with increasing KCl concentration from 0.05 to 2.0 M. At high ionic strength, the electrostatic interactions are significantly reduced between the charged membrane surfaces and the redox ions due to a smaller electrical double layer thickness. Therefore, less negatively charged redox ions can be brought to the electrode giving a smaller faradaic current. 3.4. Potential application of HBM supported on GCE for a pH sensor The voltammetric responses of HBM at GCE have been obtained in a Fe(CN)6 3− solution at a wide range of pH (1–8). Fig. 10 shows the plot of the oxidation peak current as a function of the solution pH. It can be seen that the peak current decreased with the increase of pH, reaching a plateau at about pH ∼6.3, the oxidation current of HBM-modified GCE displayed a linear response to the solution pH with a slope of about −3.3 ␮A/pH in the range of 1.0–6.0. The relative standard deviation (R.S.D.) of the HBM-modified GCE was 2.4% for seven parallel measurements in pH 3.0, indicating high repeata-

[1] F. Mueller, O.R. Donald, H.T. Tien, C.W. William, Reconstitution of cell membrane structure in vitro and its transformation into an excitable system, Nature 194 (1962) 979. [2] E. Sackman, Supported membranes: scientific and practical applications, Science 271 (1996) 43. [3] A. Ottova, V. Tvarozek, J. Racek, J. Sabo, W. Ziegler, T. Hianik, H.T. Tien, Selfassembled BLMs: biomembrane models and biosensor applications, Supramol. Sci. 4 (1997) 101. [4] K.J. Swartz, Sensing voltage across lipid membranes, Nature 456 (2008) 891. [5] E. Sparr, C. Aberg, P. Nilsson, H. Wennerstrom, Diffusional transport in responding lipid membranes, Soft Matter 5 (2009) 3225. [6] H.T. Tien, A.L. Ottova, From self-assembled bilayer lipid membranes (BLMs) to supported BLMs on metal and gel substrates to practical applications, Colloids Surf. A 149 (1999) 217. [7] H. Yuan, A.L. Ottova, H.T. Tien, An agarose-stabilized BLM: a new method for forming bilayer lipid membranes, Mater. Sci. Eng. C 4 (1996) 35. [8] D.P. Cherney, G.A. Myers, R.A. Horton, J.M. Harris, Optically trapping confocal Raman microscopy of individual lipid vesicles: kinetics of phospholipase A2 catalyzed hydrolysis of phospholipids in the membrane bilayer, Anal. Chem. 78 (2006) 6928. [9] A.L. Plant, Supported hybrid bilayer membranes as rugged cell membrane mimics, Langmuir 15 (1999) 5128. [10] H. Lang, C. Duschl, M. Gratzel, H. Vogel, Self-assembly of thiolipid molecular layers on gold surfaces: optical and electrochemical characterization, Thin Solid Films 210–211 (1992) 818. [11] A.L. Plant, Self-assembled phospholipid/alkanethiol biomimetic bilayers on gold, Langmuir 9 (1993) 2764. [12] E.L. Florin, H.E. Gaub, Painted supported lipid membranes, Biophys. J. 64 (1993) 375. [13] C.L. Wee, M.S.P. Sansom, S. Reich, E. Akhmatskaya, Improved sampling for simulations of interfacial membrane proteins: application of generalized shadow hybrid Monte Carlo to a peptide toxin/bilayer system, J. Phys. Chem. B 112 (2008) 5710. [14] M. Li, M. Chen, E. Sheepwash, C.L. Brosseau, H. Li, B. Pettinger, H. Gruler, J. Lipkowski, AFM studies of solid-supported lipid bilayers formed at a Au(111) electrode surface using vesicle fusion and a combination of Langmuir–Blodgett and Langmuir–Schaefer techniques, Langmuir 24 (2008) 10313. [15] I. Zawisza, X. Bin, J. Lipkowski, Potential-driven structural changes in Langmuir–Blodgett DMPC bilayers determined by in situ spectroelectrochemical PM IRRAS, Langmuir 23 (2007) 5180.

Y. Li et al. / Electrochimica Acta 56 (2011) 1076–1081 [16] D.A. Doshi, A.M. Dattelbaum, E.B. Watkins, C.J. Brinker, B.I. Swanson, A.P. Shreve, A.N. Parikh, J. Majewski, Neutron reflectivity study of lipid membranes assembled on ordered nanocomposite and nanoporous silica thin films, Langmuir 21 (2005) 2865. [17] M. Stelzle, G. Weissmüller, E. Sackmann, On the application of supported bilayers as receptive layers for biosensors with electrical detection, J. Phys. Chem. 97 (1993) 2974. [18] S. Lingler, I. Rubinstein, W. Knoll, A. Offenhäusser, Fusion of small unilamellar lipid vesicles to alkanethiol and thiolipid self-assembled monolayers on gold, Langmiur 13 (1997) 7085. [19] A.L. Plant, M. Guegeutchkeri, W. Yap, Supported phospholipid/alkanethiol biomimetic membranes: insulating properties, Biophys. J. 67 (1994) 1126. [20] J.D. Burgess, M.C. Rhoten, F.M. Hawkridge, Cytochrome c oxidase immobilized in stable supported lipid bilayer membranes, Langmiur 14 (1998) 2467. [21] K. Seifert, K. Fendler, E. Bamberg, Charge transport by ion translocating membrane proteins on solid supported membranes, Biophys. J. 64 (1993) 384. [22] B.A. Cornell, V.L.B. Braach-Maksvytis, L.G. King, P.D.J. Osman, B. Raguse, L. Wieczorek, R.J. Pace, A biosensor that uses ion-channel switches, Nature 387 (1997) 580. [23] A.T.A. Jenkins, R.J. Bushby, N. Boden, S.D. Evans, P.F. Knowles, Q. Liu, R.E. Miles, S.D. Ogier, Ion-selective lipid bilayers tethered to microcontact printed selfassembled monolayers containing cholesterol derivatives, Langmuir 14 (1998) 4675. [24] Y. Li, P. Wang, F. Li, X. Huang, L. Wang, X. Lin, Covalent immobilization of single-walled carbon nanotubes and single-stranded deoxyribonucleic acid nanocomposites on glassy carbon electrode: preparation, characterization, and applications, Talanta 77 (2008) 833. [25] P. Wang, F. Li, X. Huang, Y. Li, L. Wang, In situ electrodeposition of Pt nanoclusters on glassy carbon surface modified by monolayer choline film and their electrochemical applications, Electrochem. Commun. 10 (2008) 195. [26] Y. Li, X. Lin, Simultaneous electroanalysis of dopamine, ascorbic acid and uric acid by poly (vinyl alcohol) covalently modified glassy carbon electrode, Sens. Actuators B 115 (2006) 134. [27] R.S. Deinhammer, M. Ho, J.W. Anderegg, M.D. Porter, Electrochemical oxidation of amine-containing compounds: a route to the surface modification of glassy carbon electrodes, Langmuir 10 (1994) 1306. [28] X. Han, L. Wang, B. Qi, X. Yang, E. Wang, A strategy for constructing a hybrid bilayer membrane based on a carbon substrate, Anal. Chem. 75 (2003) 6566. [29] L. Zhang, X. Lin, Covalent modification of glassy carbon electrode with glutamic acid for simultaneous determination of uric acid and ascorbic acid, Analyst 126 (2001) 367. [30] Y. Okuno, W.E. Ford, M. Calvin, An improved synthesis of surfactant porphyrins, Synthesis 7 (1980) 537. [31] J.H. Scofield, Hartree–Slater subshell photoionization cross-sections at 1254 and 1487 eV, J. Electron. Spectrosc. Relat. Phenom. 8 (1976) 129.

1081

[32] C.K. Mann, Cyclic stationary electrode voltammetry of some aliphatic amines, Anal. Chem. 36 (1964) 2424. [33] B. Barbier, J. Pinson, G. Desarmot, M. Sanchez, Electrochemical bonding of amines to carbon fiber surfaces toward improved carbon-epoxy composites, J. Electrochem. Soc. 137 (1990) 1757. [34] A.J. Downard, A.B. Mohamed, Suppression of protein adsorption at glassy carbon electrodes covalently modified with tetraethylene glycol diamine, Electroanalysis 11 (1999) 418. [35] C. Saby, B. Ortiz, G.Y. Champagne, D. Belanger, Electrochemical modification of glassy carbon electrode using aromatic diazonium salts. 1. Blocking effect of 4-nitrophenyl and 4-carboxyphenyl groups, Langmuir 13 (1997) 6805. [36] K.M. Kadish, R.K. Rhodes, Reactions of metalloporphyrin. pi. radicals. 2. Thinlayer spectroelectrochemistry of zinc tetraphenylporphyrin cation radicals and dications in the presence of nitrogenous bases, Inorg. Chem. 20 (1981) 2961. [37] C.L. Lin, M.Y. Fang, S.H. Cheng, Substituent and axial ligand effects on the electrochemistry of zinc porphyrins, J. Electroanal. Chem. 531 (2002) 155. [38] A.J. Bard, L.R. Faulkner, Electrochemical Methods Fundamentals Applications, Wiley, New York, 1980. [39] B.P. Neri, G.S. Wilson, Electrochemical studies of mesotetra(4-Nmethylpyridyl)porphine in acid solution, Anal. Chem. 44 (1972) 1002. [40] G.L. Wang, A.K. Bohaty, I. Zharov, H.S. White, Photon gated transport at the glass nanopore electrode, J. Am. Chem. Soc. 128 (2006) 13553. [41] R.J. White, E.N. Ervin, T. Yang, X. Chen, S. Daniel, P.S. Cremer, H.S. White, Single ion-channel recordings using glass nanopore membranes, J. Am. Chem. Soc. 129 (2007) 11766. [42] T. Ito, Ion-channel-mimetic sensor for trivalent cations based on self-assembled monolayers of thiol-derivatized 4-acyl-5-pyrazolones on gold, J. Electroanal. Chem. 495 (2001) 87. [43] Q. Cheng, A. Brajter-Toth, Permselectivity and high sensitivity at ultrathin monolayers. Effect of film hydrophobicity, Anal. Chem. 67 (1995) 2767. [44] D.B. Brown, D.F. Shriver, Structures and solid-state reactions of Prussian blue analogs containing chromium, manganese, iron, and cobalt, Inorg. Chem. 8 (1969) 37. [45] A. Zahl, R.V. Eldik, T.W. Swaddle, Cation-independent electron transfer between ferricyanide and ferrocyanide ions in aqueous solution, Inorg. Chem. 41 (2002) 757. [46] H. Tsue, S. Nakashima, Y. Goto, H. Tatemitsu, S. Misumi, R.J. Abraham, T. Asahi, Y. Tanaka, T. Okada, N. Mataga, Y. Sakata, Synthesis of rigid porphyrin–quinone compounds for studying mutual orientation effects on electron transfer and their photophysical properties, Bull. Chem. Soc. Jpn. 67 (1994) 3067. [47] W.R. Heineman, H.J. Wieck, A.M. Yacynych, Polymer film chemically modified electrode as a potentiometric sensor, Anal. Chem. 52 (1980) 345. [48] N. Oyama, T. Hirokawa, S. Yamaguchi, N. Ushizawa, T. Shimomura, Hydrogen ion selective microelectrode prepared by modifying an electrode with polymers, Anal. Chem. 59 (1987) 258.