Surface-bound membranes

Evaluation of Surface-Bound Membranes with Electrochemical Impedance Spectroscopy... 

To assemble the surface-bound membrane structure, we first form a hydrophobic monolayer by using alkylsilanization to covalently attach long-chained hydrocarbon chains to hydroxyl groups in the oxide layer on the electrode surface. Such a hydrophobic surface can be thought of as one leaflet of a membrane bilayer. Alkylsilane-modified surfaces have been widely used as substrates for lipid monolayers deposited by Langmuir-Blodgett techniques. The lipids in these monolayers have mobilities like those of lipids... 

Model of Surface-Bound Membrane on Pt Electrode. From the foregoing discussion of experimental results we can develop a physical model of the surface-bound membrane that consists of two layers, as schematically depicted in Figure 7. The porous, hydrophobic OTS layer provides a structure to anchor the reconstituted membrane layer. Protein molecules with bound lipid may insert into the pores in the OTS layer. The... 

Figure 7. Model of a single surface-bound membrane formed by detergent dialysis on an alkylsilanated electrode surface.

An equivalent circuit can be derived for the surface-bound membrane formed in this work similar in a manner to the approach taken for porous anodic films and porous electrodes (41-46). An equivalent circuit network, proposed in Figure 8a, corresponds to the model in Figure 7. This network has three RC subnetworks that represent the oxide layer, the surface-bound membrane layer, and the double layer. Cox and Rox are the capacitance and resistance of oxide. and Rdl are the double-layer capacitance and the polarization resistance, known as the charge transfer resistance at the membrane-water interface. For the subnetwork of the surface-bound membrane layer, one branch represents a tightly packed alkylsilane and lipid bilayer in series, and the other branch represents the pores and defects through the bilayer. Calk, Clip and Ralk, Rhp are the capacitances and resistances of... 

Figure 8. (a) Proposed equivalent circuit for surface-bound membrane electrode interface, (b) Simplified equivalent circuit valid at higher frequency region. 

The simulation spectra of the PtO electrodes with and without the surface-bound membrane are shown in Figure 9 for comparison with the experimental data of Figure 6. The parameters used in the simulation are listed in Table I. The first column lists the values used for curve fitting the experimental spectra, and the second column gives the corresponding values normalized for unit area. 

This simulation reproduces the essential features of the experimental data. A calculated capacitance for the tightly packed part of the surface-bound membrane (Cbl) can be obtained by treating Calk and Clip in series. The resistance can be similarly calculated from Ralk and Ru. The calculated values of 0.52 fiF/cm2 and 1325 ft cm2 are in good agreement with literature values for natural membranes (47-48). The best curve fit for the coverage factor, 0, was 0.97, which indicates formation of a relatively complete membrane by the detergent dialysis approach. 

In this simplified form, the membrane capacitance, Cm, is in series with Cox. Thus, Cm = CoxCt/(Ct - Cox), where Ct is the total capacitance of the surface-bound membrane electrode and Cox is the measured capacitance of the electrode before membrane formation. The membrane capacitance, Cm, can thus be estimated at a single frequency. Further, the capacitance of the tightly packed bilayer, Cbl, can be calculated from Cm if the coverage factor and the double-layer capacitance are known Cbl = (Cm — (1 — 0)Cdl)/0. 

By using the imaginary component of the measured impedance data for PtO-OTS and PtO-OTS-Rh electrodes (Table II) at a frequency of 1000 Hz (after subtracting Ru), the calculated Cm is 867 nF/cm2 and Cbl is thus 584 nF/cm2 using = 10 jiF/cm2 and 0 = 0.97, which are close to the theoretical values derived from the best curve fit simulation. We conclude that the simplified equivalent circuit may be adequate for the surface-bound membrane electrode. The thickness of the tightly packed membrane bilayer, d, can be calculated from d = e0e/Cbl, where e is the dielectric constant of... 

Surface-bound membranes formed on PtO electrodes were chemically and mechanically stable. The PtO-OTS-Rh electrodes were monitored by measuring the capacitance while the electrodes were kept in buffer at 4 °C for 11 days. Any dissolution of the surface-bound membrane would result in an increase in capacitance. Little change in capacitance was observed, which indicates that the membranes are stable. 

The simplified equivalent circuit in Figure 8b was used to evaluate surface-bound membranes on Si02, TiOa, and ITO electrodes. Figures 10 and 11 present the capacitance curves for n-Si-Si02 and TiOa electrodes with and without OTS- and rhodopsin-containing lipid membranes in KCl buffer. As with the PtO electrodes, the capacitance decreases upon formation of an OTS layer and the membrane on the oxide surface. Table II lists the... 

Electrochemical impedance spectroscopy provides a sensitive means for characterizing the structure and electrical properties of the surface-bound membranes. The results from impedance analysis are consistent with a single biomembrane-mimetic structure being assembled on metal and semiconductor electrode surfaces. The structures formed by detergent dialysis may consist of a hydrophobic alkyl layer as one leaflet of a bilayer and the lipid deposited by dialysis as the other. Proteins surrounded by a bound lipid layer may simultaneously incorporate into pores in the alkylsilane layer by hydrophobic interactions during deposition of the lipid layer. This model is further supported by the composition of the surface-bound membranes and by Fourier transform infrared analyses (9). 

J. Li, N.W. Downer, H.G. Smith, Evaluation of surface-bound membranes with electrochemical impedance spectroscopy, in Biomembrane Electrochemistry (American Chemical Society Advances in Chemistry Series 235), ed. by M. Blank, 1. Vodyanov (ACS, Washington, DC, 1994), pp. 491-510... 

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