Hybrid bilayer membranes on metallurgical polished aluminum

Abstract

In this work we describe the functionalization of metallurgically polished aluminum surfaces yielding biomimetic electrodes suitable for probing protein/phospholipid interactions. The functionalization involves two simple steps: silanization of the aluminum and subsequent fusion of multilamellar vesicles which leads to the formation of a hybrid bilayer lipid membrane (hBLM). The vesicle fusion was followed in real-time by fast Fourier transform electrochemical impedance spectroscopy (FFT EIS). The impedance-derived complex capacitance of the hBLMs was approximately 0.61 µF cm^-2, a value typical for intact phospholipid bilayers. We found that the hBLMs can be readily disrupted if exposed to > 400 nM solutions of the pore-forming peptide melittin. However, the presence of cholesterol at 40% (mol) in hBLMs exhibited an inhibitory effect on the membrane-damaging capacity of the peptide. The melittin-membrane interaction was concentration dependent decreasing with concentration. The hBLMs on Al surface can be regenerated multiple times, retaining their dielectric and functional properties essentially intact.

Figure 1

Cyclic voltammograms of the metallurgical aluminum surface before and after silanization at the scan rate of 10 mV s⁻¹ in phosphate buffer, pH 7.1.

Figure 2

C–H stretching region of ATR-FTIR spectra of OTS self-assembled monolayer formed on the metallurgical polished aluminum surface.

Figure 3

Electrochemical impedance spectra in Cole–Cole plot of Al, OTS and DOPC:Chol (molar % ratio 6:4) hBLM, formed by vesicle fusion method after 30 min incubation time. The applied potential: − 0.7 V versus Ag/AgCl/NaCl_(sat.). Mean complex capacitance values with standard deviations are displayed as well.

Figure 4

Cole–Cole plots of FFT impedance spectra of the DOPC:Chol (molar % ratio 6:4) vesicle fusion process on silanized aluminum surface. The applied potential: − 0.7 V versus Ag/AgCl/NaCl_(sat.). Inset—surface coverage, θ, of the lipid layer over time.

Figure 5

Cole–Cole plots of electrochemical impedance spectroscopy of the OTS SAM regeneration for DOPC:Chol (molar % ratio 6:4) hybrid bilayer formation on the metallurgical aluminum surface. The applied potential for EI measurements: − 0.7 V versus Ag/AgCl/NaCl_(sat.). Inset shows an enlarged part of the complex capacitance plot of the hBLM formations.

Figure 6

Electrochemical impedance spectra in Cole–Cole plot of melittin interaction: (a) and (c)—with DOPC:Chol (molar % ratio 6:4), (b) and (d)—with DOPC hybrid bilayer lipid membranes. The applied potential: − 0.7 V versus Ag/AgCl/NaCl_(sat.).

Figure 7

(a)—Dependence of the membrane damage, MD, on melittin concentration, (b)—same data depicted in logMD versus inverse of melittin concentration.