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A mixed monolayer of 1,2-dipalmitoyl-sn-glycero-3-(cytidine diphosphate) nucleolipid (DG-CDP) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) was transferred from the air–water interface onto an gold (1 1 1) electrode surface using the Langmuir-Schaeffer (horizontal touch) technique. Compression isotherms were recorded for the monolayer spread at the air–water interface. These measurements demonstrated that the mixture composition 7:3 of DG-CDP:DPPC displayed reduced repulsive interactions between polar heads and ideal packing of acyl chains. Chronocoulometric experiments were performed to demonstrate stability of this film at the gold–solution interface. Photon polarization modulation infrared reflection absorption spectroscopy showed that the tilt angle of the acyl chains is slightly higher, and the chains are less twisted in the mixed than in the pure nucleolipid monolayer. In the mixed film, the tilt of the cytosine moiety (the sensing element of the monolayer) assumes a large angle with respect to the surface normal, even when the monolayer becomes detached from the gold surface. The mixed monolayer has improved properties for future applications for detection of guanine, its complementary base. Spain Ministry of Science and Innovation and Junta de Andalucía. Natural Sciences and Engineering Research Council of Canada (NSERC)

In situ ATR-SEIRAS experiments were carried out to describe water structure at the surface of multilayers of Pd modified nanopatterned gold film electrodes in contact with aqueous 0.1 M HClO4 solution. The results show that the surface water becomes more ordered when the thickness of the Pd film increases. The “ice-like water” band becomes stronger and for the electrode with 5 ML Pd@Au, the ratio of “ice-like water” to “liquid-like water” becomes larger than that in bulk water. Concomitantly, the population of “monomers” or small water clusters diminishes. All surface water bands increase when hydrogen adsorption takes place. The bending band corresponding to “monomers” or “multimers” water is observed chiefly at the potentials of hydrogen adsorption. Natural Sciences and Engineering Research Council of Canada (NSERC)the National Natural Science Foundation of China (NSFC), the "111" Project (B17027), and the National Key Research and Development Program of China.

Chronocoulometry was applied to determine charge in a monolayer of nucleolipid (1,2-dipalmitoyl-sn‑glycero-3-(cytidine diphosphate)) deposited at a gold electrode surface. The immersion method was used to measure the potential of zero charge of the interface (Epzci), which is a sum of charge on the monolayer of the nucleolipid and charge on the gold surface. Photon polarization infrared reflection absorption spectroscopy (PM-IRRAS) was used to determine formation of the Watson-Crick complex between terminal cytidine moiety of the nucleolipid and guanine (its complementary base) added to the solution. The combination of electrochemical and spectroscopic studies allowed one to demonstrate that the Watson-Crick complex is formed when the interface is positively charged. The potential applied to the electrode affects not only the complex formation but also orientation of the cytosine moiety. The complex is formed when the cytosine moiety is oriented assuming a small angle with respect to the electrode surface. Natural Sciences and Engineering Research Council of Canada (NSERC), Spain Ministry of Science and Innovation and Junta de Andalucí

The role of the anion on the ionophore properties of valinomycin was studied in a model floating bilayer lipid membrane (fBLM) using supporting electrolytes containing K+ with four different counter anion species (ClO4–, H2PO4–, Cl–, and F–). The electrochemical impedance spectra indicate that the membrane resistance of the bilayer decreases with the decrease of Gibbs free energy of anion solvation. The IR spectra demonstrate that valinomycin does not readily bind to K+ in the KH2PO4, KCl, and KF electrolyte solutions, but in the presence of KClO4, valinomycin readily binds to K+, forming a valinomycin–K+ complex. The results in the present paper reveal the role of the counter anion on the transport of cations by valinomycin across the lipid bilayer. The valinomycin–cation complex creates an ion pair with the anion, and this ion pair can enter the hydrophobic region of the bilayer transporting the cation across the membrane. Anions with low solvation energies facilitate the formation of the ion pair improving the ion conductivity of valinomycin-incorporated bilayers. This paper sheds new light on the transport mechanism of valinomycin ionophores and provides new information about the bioactivity of this molecule. Natural Sciences and Engineering Research Council of Canada (NSERC)

The ion channel properties of gramicidin A (gA) in a model floating bilayer lipid membrane were studied by electrochemical impedance spectroscopy (EIS) and polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS). To determine if the transport of Na+ across the membrane is coupled with the transport of the counterion, fluoride and perchlorate anions were explored due to their large differences in hydration energy. The combined EIS and IR experiments demonstrated that in contrast to valinomycin, the mechanism of Na+ transfer through the gA channel is independent of the accompanying anion present in the supporting electrolyte. These studies also demonstrated that the conductivity of the gA sodium channel depends on the transmembrane potential and the IR measurements indicated that more gA molecules insert into the bilayer with the decrease of the transmembrane potential. The results of this present study demonstrate that the fBLM system is a suitable model bilayer for the in situ investigations of membrane-active peptides. Natural Sciences and Engineering Research Council of Canada (NSERC)

Reduction of oxygen catalyzed by laccase was studied by surface-enhanced infrared absorption spectroscopy (SEIRAS) combined with electrochemical techniques. Laccase molecules were adsorbed on modified carbon nanotubes (CNTs). SEIRAS provided spectra of Amide I band of adsorbed laccase. Fourier self-deconvolution (FSD) and two-dimensional correlation spectroscopy (2D COS) techniques were employed to reveal the sub-band structure of the Amide I band. The analysis demonstrated that laccase adsorbed on CNTs remains its native state both in the oxidized and reduced states. The detailed analysis showed that change of the oxidation state leads to a small change of the secondary structure of the protein. This change is comparable to the change observed for similar blue-copper oxidases in solution during redox titration. The results of this study demonstrated that CNT's immobilized laccase is an excellent catalyst of oxygen reduction. This study illustrates power of quantitative analysis of IR data to provide information about three-dimensional structure of surface immobilized proteins. Natural Sciences and Engineering Research Council of Canada (NSERC)

The electrochemical impedance spectroscopy (EIS) and polarization-modulation infrared reflection absorption spectroscopy (PM-IRRAS) techniques were employed to study the ionophore properties of valinomycin in a model floating bilayer lipid membrane (fBLM) in perchlorate supporting electrolytes with potassium and sodium cations. Valinomycin decreases the membrane resistance of the fBLM in KClO4 solution by about 180 times, while it has a negligible effect on the membrane resistivity in NaClO4 solution. The IR spectra indicate that valinomycin forms a complex with K+, but not with Na+. The valinomycin-K+ complex adopts a small tilt angle with respect to the electrode surface normal and is well interdigitated between the acyl chains of the bilayer. The EIS and PM-IRRAS results indicate that valinomycin forms complexes with K+ and transports K+ across the lipid bilayer. This transport is potential independent and hence has a passive character. Natural Sciences and Engineering Research Council of Canada (NSERC)

Neurodegeneration in Alzheimer's disease is associated with disruption of the neuronal cell membrane by the amyloid β (Aβ) peptide. However, the disruption mechanism and the resulting changes in membrane properties remain to be elucidated. To address this issue, herein the interaction of amyloid β monomers (AβMs) and amyloid β oligomers (AβOs) with a floating bilayer lipid membrane (fBLM) was studied using electrochemical and IR spectroscopy techniques. IR measurements showed that both Aβ forms interacted similarly with the hydrophobic membrane core (lipid acyl chains), causing conformational and orientational changes of the lipid acyl chains, thus decreasing acyl chain mobility and altering the lipid packing unit cell. In the presence of AβOs, these changes were more significant than those in the presence of AβMs. However, respective interactions of AβMs and AβOs with the membrane hydrophilic exterior (lipid heads) were quite different. AβMs dehydrated lipid heads without affecting their orientation while AβOs changed the orientation of lipid heads keeping their hydration level intact. Electrochemical measurements showed that only AβOs porated the fBLM, thus significantly changing the fBLM electrical properties. The present results provide new molecular-level insight into the mechanism of membrane destruction by AβOs and changes in the membrane properties. Polish National Science Centre, European Union Horizon 2020 Research and Innovation Programme under the Marie Skłodowska-Curie grant agreement, Polish Ministry of Science and Higher Education for the implementation of an international co-financed project in the years 2017−2021. Discovery grant from Natural Sciences and Engineering Research Council of Canada (NSERC).

This document is the Accepted Manuscript version of a Published Work that appeared in final form in Journal of Physical Chemistry C, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acs.jpcc.0c03453. The knowledge of the structure and properties of water adsorbed on metal surfaces is relevant for understanding electrocatalytic reactions. A monolayer of Pd deposited at a gold electrode surface is known to have enhanced electrocatalytic properties. To shed new light on these properties, the present study describes investigations of water structure at monolayer Pd film deposited at a nanostructured gold electrode consisting of ∼50 nm large nanoparticles supported at a Si hemisphere. Attenuated total reflection surface enhanced infrared reflection absorption spectroscopy (ATR-SEIRAS) was applied to measure vibrational spectra of water molecules at the electrode surface. The surface enhancement of the electric field of the photon allowed one to determine spectra of water molecules interacting with the Pd film. The results show that surface water is a mosaic of “ice-like”, “liquid-like”, and monomer or small clusters of water molecules. In addition, hydronium ions are present at the Pd surface in an acidic solution. The relative contribution of these structures depends on the pH and electrode potential. The surface water is more ordered at potentials corresponding to the adsorption of hydrogen atoms. Natural Sciences and Engineering Research Council of Canada (NSERC), National Natural Science Foundation of China (NSFC), "111" Project, National Key Research and Development Program of China

A hybrid bilayer lipid membrane (hBLM), constructed with a 1-hexadecanethiol self-assembled interior leaflet and a 1,2-dipalmitoyl-sn-glycero-3-cytidine nucleolipid exterior leaflet, was deposited at the surface of a gold (111) electrode. This system was used to investigate the molecular recognition reaction between the cytosine moieties of the lipid head group with guanine molecules in the bulk electrolyte solution. Electrochemical measurements and photon polarization modulation infrared reflection absorption spectroscopy (PMIRRAS) were employed to characterize the system and determine the extent of the molecular recognition reaction. The capacitance of the hBLM-covered gold electrode was very low (~1 μF cm−2), therefore the charge density at the gold surface was small. Changing the electrode potential had a minimal effect on the complexation between the cytosine moieties and guanine molecules due to small changes in the static electric field across the membrane. This behavior favored the formation of the guanine–cytosine complex. Spanish Ministry of Economy and Competitiveness, Andalusian government, Discovery grant from Natural Sciences and Engineering Council of Canada.

Monolayers of 1,2-dipalmitoyl-sn-glycero-3-cytidine were incubated with guanine in a 0.1 M NaF electrolyte at the surface of a Langmuir trough and transferred to gold (111) electrodes using the Langmuir–Schaefer technique. Chronocoulometry and photon polarization modulation infrared reflection absorption spectroscopy were employed to investigate the influence of the static electric field on the orientation and conformation of the cytidine nucleolipid molecules on the metal surface in the presence of guanine and to monitor the molecular recognition of guanine with the cytosine moiety. When the monolayer is exposed to guanine solutions, the cytosine moiety binds to the guanine residue in either a Watson–Crick complex at positively charged electrode surfaces or a noncomplexed state at negative surface charges. The positive electrostatic field causes the cytosine moiety and the cytosine–guanine complex to adopt a nearly parallel orientation with respect to the plane of the monolayer with a measured tilt angle of ∼10°. The parallel orientation is stabilized by the interactions between the permanent dipole of the cytosine moiety or the Watson–Crick complex and the static electric field. At negative charge densities, the tilt of the cytosine moiety increases by ∼15–20°, destabilizing the complex. Our results demonstrate that the static electric field has an influence on the molecular recognition reactions between nucleoside base pairs at the metal–solution interface and can be controlled by altering the surface charge at the metal. Spanish Ministry of Economy and Competitiveness and Andalusian government, Natural Sciences and Engineering Council of Canada (NSERC)

Colicin E1 is a channel-forming bacteriocin produced by certain Escherichia coli cells in an effort to reduce competition from other bacterial strains. The colicin E1 channel domain was incorporated into a 1,2-diphytanoyl-sn-glycero-3-phosphocholine floating bilayer situated on a 1-thio-?-d-glucose-modified gold (111) surface. The electrochemical properties of the colicin E1 channel in the floating bilayer were measured by electrochemical impedance spectroscopy; the configuration and orientation of colicin E1 in the bilayer were determined by polarization-modulation-infrared-reflection absorption spectroscopy. The EIS and IR results indicate that colicin E1 adopts a closed-channel state at the positive transmembrane potential, leading to high membrane resistance and a large tilt angle of ?-helices. When the transmembrane potential becomes negative, colicin E1 begins to insert into the lipid bilayer, corresponding to low membrane resistance and a low tilt angle of ?-helices. The insertion of colicin E1 into the lipid bilayer is driven by the negative transmembrane potential, and the ion-channel open and closed states are potential reversible. The data in this report provide new insights into the voltage-gated mechanism of colicin E1 ion channels in phospholipid bilayers and illustrate that the floating bilayer lipid membrane at the metal electrode surface is a robust platform to study membrane-active proteins and peptides in a quasi-natural environment. Natural Sciences and Engineering Research Council of Canada

The effects of amiloride on the structure and conductivity of alamethicin ion pore formation within negatively charged, gold-supported, 1,2-dimyristoyl-sn-glycero-3-phosphocholine/Egg-PG membranes were investigated with the help of electrochemical impedance spectroscopy (EIS), photon polarization modulation-infrared reflection spectroscopy (PM-IRRAS), and atomic force microscopy (AFM). The EIS results indicate that ion conductivity across negatively charged phospholipid bilayers containing alamethicin decreases by an order of magnitude when amiloride is introduced to the system. Despite the reduction in ion conductivity, the PM-IRRAS data shows that amiloride does not inhibit ion channel formation by alamethicin peptides. High-resolution AFM images revealed that amiloride enlarges and distorts the shape of alamethicin ion pores when introduced to the system, indicating that it is inserting itself into the mouth of the alamethicin pores. This effect is driven by electrostatic interactions between positively charged amiloride molecules and the negative charge on the membrane. Discovery grant from Natural Sciences and Engineering Council of Canada

The effect of the electrode potential on the orientation and conformation of the 1,2-dipalmitoyl-sn-glycero-3-cytidine monolayer deposited on a gold (111) electrode surface was described. The potential of zero free charge (Epzc) for the monolayer-covered electrode was determined to be −0.2 V vs SCE. The differential capacitance and charge density data indicated that the monolayer is stable at the electrode surface when (E – Epzc) > 0.0 V. At negative rational potentials, a progressive detachment (electrodewetting) of the monolayer occurs. The monolayer is fully detached from the electrode surface at (E – Epzc) < −0.6 V. The conformation and orientation of the acyl chains and the orientation of the cytosine moiety were determined with the help of photon polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS). The IR measurements demonstrate that the acyl chains are predominantly in the gel phase in the adsorbed state and tilted at an angle of ∼30° with respect to the electrode surface normal. The tilt angle of the acyl chains increases when the film is detached from the gold surface, indicating that the monolayer becomes more disordered. At (E – Epzc) > 0.0 V, the plane of the cytosine moiety assumes a small angle of ∼20° with respect to the surface. At negative potentials, the tilt angle of the cytosine fragment increases and rotates. With the help of DFT calculations, these changes were explained by the repulsion of the positive pole of the cytosine permanent dipole moment by the positively charged gold surface and its attraction to the metal surface at negative electrode potentials. This work provides unique information for the future development of sensors based on the molecular recognition of nucleoside targets. Spanish Ministry of Economy and Competitiveness and the Andalusian government, Spanish Ministry of Science and Technology. Natural Sciences and Engineering Council of Canada (NSERC)

Electrochemical and polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS) techniques were employed to study the ion channel properties of gramicidin A (gA) in tethered bilayer lipid membranes (tBLMs) with varying concentrations of gA added to the outer leaflet of the bilayer. The electrochemical impedance spectroscopy (EIS) results demonstrate that gA forms conductive ion channels in tBLM systems even at concentrations as low as 0.5 mol%. The impedance of the membrane depended on the gA concentration where a minimum was observed at 10 mol%. When the amount of gA in the bilayer is increased to a concentration above 10%, the ion channels appear to arrange into a non-conducting state. PM-IRRAS spectra suggest that the gA molecules adopt a tilt angle of ∼29 ± 4° in the conductive ion channel state. In the non-conducting state, the average tilt angle of the gA molecules is ∼52 ± 3°, which implies that gA peptides assume a random orientation in the tBLM. Natural Sciences and Engineering Research Council of Canada (NSERC)

The charge number per adsorbed molecule and surface concentration of 2,3-di-O-phytanyl-sn-glycero-1-octaethyleneglycol-D,L-α-lipoic acid ester (DPOL) at a Au(111) electrode surface was investigated by chronocoulometry using two different film deposition methods. First, a known amount of the thiolipid was transferred from the air-water interface of a Langmuir trough onto the gold electrode surface via a Langmuir-Blodgett (LB) deposition. The charge density measurements for this monolayer system were used to determine the charge number per adsorbed DPOL molecule (electrosorption valency). The observed electrosorption valency values of the adsorbed DPOL film were much lower than the expected number of transferred electrons for a simple reductive desorption process. In the second deposition method, charge densities were measured for the electrode covered by a self-assembled DPOL monolayer. The electrosorption valency values determined from the LB DPOL film were used to calculate the packing density of the DPOL molecules within the self-assembled monolayer. The surface concentration of the molecules within the thiol monolayer with octaethylene glycol chains gave similar results to a related thiolipid with tetraethylene glycol chains (DPTL). This new finding indicates that both of these molecules (DPTL and DPOL) assume a brush conformation in densely packed self-assembled monolayers. Natural Sciences and Engineering Research Council of Canada