Microcavity-Supported Lipid Bilayers; Evaluation of Drug–Lipid Membrane Interactions by Electrochemical Impedance and Fluorescence Correlation Spectroscopy

Abstract

Many drugs have intracellular or membrane-associated targets thus understanding their interaction with the cell membrane is of value in drug development. Cell-free tools used to predict membrane interactions should replicate the molecular organization of the membrane. Microcavity array supported lipid bilayer (MSLB) platform are versatile biophysical models of the cell membrane that combine liposome-like membrane fluidity with stability and addressability. We used an MSLB herein to interrogate drugmembrane interactions across seven drugs from different classes, including non-steroidal antiinflammatories; Ibuprofen (Ibu) and Diclofenac (Dic), antibiotics; Rifampicin (Rif), Levofloxacin (Levo) and Pefloxacin (Pef), and bisphosphonates; Alendronate (Ale) and Clodronate (Clo). Fluorescence lifetime correlation spectroscopy (FLCS) and electrochemical impedance spectroscopy (EIS) were used to evaluate the impact of drug on DOPC and binary bilayers over physiologically relevant drug concentrations. Whereas FLCS data revealed Ibu, Levo, Pef, Ale and Clo had no impact on lipid lateral

The Impact of Membrane Composition and Co‐Drug Synergistic Effects on Vancomycin Association with Model Membranes from Electrochemical Impedance Spectroscopy

“…Upon exposure to 1 μM Rif, the absolute resistance of E. coli membrane decreases dramatically from 5.52 � 0.01 to 2.36 � 0.02 MΩ and the absolute Q M of the bilayer increases from 6.22 � 0.002 to 6.36 � 0.001 μF • s mÀ 1 . As for our earlier report at DOPC-only bilayer, [44] Rif seems to penetrate the E. coli membrane (Inset, Figure 3a) and indeed the magnitude of decreased membrane resistance ( Figure 3b (blue and black symbols respectively). Note, the DR data (blue, Figure 3b) was normalized with respect to the resistance data of E. coli membrane when 1 μM Rif is present (cf.…”

“…[55] In our recent study, in contrast to vancomycin, passive permeation of Rif across the DOPC membrane was observed to occur and this resulted in the case of Rif alone, in large decreases in membrane resistance, as replicated below. [44] Here, we examined the interaction of vancomycin with E. coli membrane in the presence of 1 μM Rif in the contact solution and compared the results (Figure 3a) with earlier data, where no Rif was present ( Figure S5d, SI). Upon exposure to 1 μM Rif, the absolute resistance of E. coli membrane decreases dramatically from 5.52 � 0.01 to 2.36 � 0.02 MΩ and the absolute Q M of the bilayer increases from 6.22 � 0.002 to 6.36 � 0.001 μF • s mÀ 1 .…”

“…Fabrication of gold microcavity array electrodes with hexagonally packed pores of 1 μm diameter and depth 0.5 μm, and the assembly of pore spanning lipid bilayers span across the arrays were carried out as reported earlier by us. [44,45] The detailed methods are provided in SI ( Figures S1-S3, SI) and a schematic representation of such a biomimetic platform is shown in Scheme 1a. The impedance of the MSLBs of each composition was assessed in triplicate, as well as their temporal stability.…”

“…Gold microcavity array electrodes (~1 × 1 cm 2 ) prepared by polystyrene microsphere lithography technique as reported previously by us [44,45,64,65] and were selectively modified with a self-assembled monolayer (SAM) of 1 mM 6-Mercapto-1-hexanol (MH). The detailed fabrication and characterization methods are also presented in SI.…”

“…The MSLB functions as the working electrode enabling electrochemical measurements of drug-membrane interaction and protein-lipid detection. [44,45] The lipid bilayer is assembled over an aqueous reservoir by a combination of both Langmuir-Blodgett and vesicle disruption to permit asymmetric/symmetric lipid bilayer assembly in a robust, relatively quick and facile manner. [41,43] Herein, we exploit MSLB platforms, to study by, electrochemical impedance spectroscopy, the interactions of vancomycin with 5 symmetric biomembranes of increasing complexity; DOPC, DOPC : Chol and DOPC : SM : Chol, eukaryotic membranes, a mammalian plasma membrane mimetic fusogenic composition coined "Nature's own" by Lentz et al [46,47] (referred to forthwith as MPM) expected to promote membrane curvature and E. coli extract comprised of DOPE, DOPG and cardiolipin (CA) as bacterial membranes.…”