Monika Naumowicz

Impedance analysis of phosphatidylcholine membranes modified with gramicidin D.

Electrochemical impedance spectroscopy (EIS) was used to the study of gramicidin D (GD) dimerization and to transport of monovalent cations across lipid bilayers by the dimers. Phosphatidylcholine (PC) membranes were studied, unmodified and modified with very low GD concentrations in the presence of various potassium ion concentrations. A new method was proposed to determine the parameters used to describe the gramicidin dimer: gramicidin surface concentration (c(GG)), area occupied by individual channel (A(G)) and gramicidin dimerization equilibrium constant (K(GG)). It was shown that electrochemical impedance spectroscopy measurements of lipid bilayer membranes yielded the K(GG) and A(G) values of the same order of magnitude as other measurement techniques.

Impedance analysis of phosphatidylcholine membranes modified with valinomycin

The effect of the ion carrier valinomycin on the electrochemical features of the phosphatidylcholine membrane was investigated by electrochemical impedance spectroscopy. Phosphatidylcholine and valinomycin were chosen for the study because they fulfil essential functions in lively organisms. The experimental impedance values obtained in the presence of different amounts of carrier, studied with several potassium ion concentrations, were used for the research ability of valinomycin to form a 1:1 potassium ion complex on the lipid bilayer/electrolyte solution interface. Based on derived mathematical equations, the heterogeneous equilibrium constant (Kh), association rate constant of the complex (kR) and dissociation rate constant of the complex (kD) were calculated. The result of the investigation is the proposal of a new method for the determination of the parameters used to describe the chemical reaction at the interface between a carrier molecule from the membrane and a monovalent ion from the aqueous phase.

Impedance analysis of phosphatidylcholine/α-tocopherol system in bilayer lipid membranes

The effect of α-tocopherol on the electrochemical features of the phosphatidylcholine membrane was investigated by impedance spectroscopy. Phosphatidylcholine and α-tocopherol were chosen for the study because they are present in biological membranes and they fulfill essential functions in living organisms. The experimental impedance values obtained in the presence of different amounts of α-tocopherol showed evidence of domain structures within the bilayer containing less than 0.048 molar fraction of α-tocopherol. Based on derived mathematical equations, the surface area of phospholipid/α-tocopherol domain was calculated; it amounts to 832 Å2. This value is consistent, taking into consideration ordering and condensation effects of α-tocopherol, with the acknowledged, well documented, stoichiometry of such a domain of 10:1. The result of the investigation is the proposal of a new method for the determination of the surface area and description of the stoichiometry of domains formed in any two-component system.

The interfacial tension of the lipid membrane formed from lipid-cholesterol and lipid-lipid systems

Interfacial tension has been determined for phosphatidylcholine-cholesterol, phosphatidylcholine-phosphatidylethanolamine, and phosphatidylethanolamine-cholesterol membranes. Phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cholesterol (Ch) were to be investigated, because of their presence in biological membranes. Interfacial tension values of pure components are 0.81×10−3 N/m, 1.67×10−3 N/m, and 2.36×10−3 N/m, respectively. The 1∶1 complexes were formed during formation of the PC-Ch, PC-PE, and PE-Ch lipid membranes. The following parameters describing the complexes were determined: A3−1, the surface concentrations of the lipid membranes formed from these complexes; γ3, the interfacial tensions of such membranes and K, the stability constants of these complexes.

Impedance Spectroscopic Investigation of the Bilayer Lipid Membranes Formed from the Phosphatidylserine–Ceramide Mixture

Electrochemical impedance spectroscopy was used for the study of two-component lipid membranes. Phosphatidylserine and ceramide were to be investigated because they play an important biochemical role in cell membranes. The research on biolipid interaction was focused on a quantitative description of processes that take part in a bilayer. Assumed models of interaction between amphiphilic molecules and the equilibria that take place there were described by mathematical equations for the studied system. The possibility of complex formation for a two-component system forming bilayers was assumed, which could explain the deviation from the additivity rule. The molecular area and the equilibrium constant of the complex were determined.

Chronopotentiometric Technique as a Method for Electrical Characterization of Bilayer Lipid Membranes

The basic electrical parameters of bilayer lipid membranes are capacitance and resistance. This article describes the application of chronopotentiometry to the research of lipid bilayers. Membranes were made from egg yolk phosphatidylcholine. The chronopotentiometric characteristic of the membranes depends on the current value. For low current values, no electroporation takes place and the voltage rises exponentially to a constant value. Based on these kinds of chronopotentiometric curves, a method of the membrane capacitance and the membrane resistance calculations are presented.

Chapter 5 Physicochemical Insights into Equilibria in Bilayer Lipid Membranes

Abstract This chapter concerns the equilibria of some biomolecules in bilayer lipid membranes. The presented research of biolipid interaction was concentrated on quantitative description of experiments that take part within the bilayer as well as on its surface. Assumed models of interaction between amphiphilic molecules, between transport proteins and monovalent cations and the equilibria that take place there as well as acid–base equilibria were described by mathematical equations for the systems studied. These theoretical models were verified experimentally using interfacial tension and electrochemical impedance spectroscopy techniques. For systems of two kinds of lipids, the possibility of a 1:1 complex formation was assumed what would explain the deviation from the additivity rule. Calculated values of parameters (equilibrium constants, molecular areas of complexes, interfacial tensions, capacitances and resistances of molecules and complexes) were used for calculation model curves. The comparison of model curves and experimental points verified assumed models. The complex formation between the gramicidin molecule and K + ion was investigated by the interfacial tension method. Electrochemical impedance spectroscopy was used for the study of gramicidin D dimerization and transport of monovalent cations across lipid bilayers by the dimers. Both experimental methods mentioned were used for the research ability of valinomycin to form a 1:1 potassium-ion complex on the lipid bilayer/electrolyte solution interface. Very simple methods were proposed to determine the parameters used to describe the gramicidin dimer and ionophore-K + complexes. It could be demonstrated that determined parameters values were of a similar order of magnitude to those obtained by other measuring techniques. The effect of pH of electrolyte solution on the bilayer lipid membrane built from different lipids was also determined. The obtained curves demonstrate the maximal interfacial tension values at the isoelectric point. A course of these curves was well characterized by the simplified description based on the Gibbs isotherm, but only in proximity of the isoelectric point. While, using the exact definition of surface excess within the Gibbs equation (taking into account volumes of adsorbed ions at the membrane surface) permits us to explain the run of experimental curves in the whole pH range. Also in this chapter the models derived to describe adsorption of the H + and OH − ions at lipid surfaces formed from phospholipids were proposed, which would reproduce changes in interfacial tension more correctly, particularly in the ranges distant from the isolelectric point. In models, contribution of the individual forms of lipid molecule to interfacial tension of the bilayer was assumed to be additive. This chapter concentrated especially on phospholipids because they are major fractions of lipids found in biological membranes: phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylserine (PS) were chosen. PC was the basic component of the formed bilayers because it has been widely examined and described in the literature, but also because it creates permanent monolayers and bilayers, of which one can easily build in the other components. Cholesterol, gramicidin and valinomycin were also studied since they play an important biochemical role in cell membranes. Cholesterol is an important factor in controlling physical properties of biological membranes and their functions. Gramicidin is a model of more complicated biological ionic channels. For this reason, many studies have been done using this simple channel-forming polypeptide. It has been reported that valinomycin acts as a selective carrier of K + ions in a variety of cell membranes as well as on liposomes and lipid bilayers. The interactions between membrane lipids are nowadays intensively developed. This results from the suitability of this research for understandings of phenomena proceeding in cellular membranes. However, there is still the lack of the quantitative description of the systems. It is required for a better understanding of the processes that take place in biological membranes with the aim of forming the artificial membrane that would very closely resemble the properties of the natural membrane. Therefore, the knowledge of molecular structure and organization of phospholipids is necessary. Data presented in this work, received from mathematical derivation and confirmed experimentally are of great importance for interpretation of phenomena occurring in lipid monolayers and bilayers. These results can help in a better understanding of biological membranes and in their biophysical studies. Simple and very interesting methods proposed in this chapter can be used with success for the determination of the equilibrium constant value of any 1:1 lipid–other lipid complex and any 1:1 ionophore–monovalent ion complex and for acid–base equilibria between any phospholipids and ions from electrolyte solution (H + and OH − ).

Impedance Analysis of Complex Formation Equilibria in Phosphatidylcholine Bilayers Containing Decanoic Acid or Decylamine

Bilayer lipid membranes composed of phosphatidylcholine and decanoic acid or phosphatidylcholine and decylamine were investigated using electrochemical impedance spectroscopy. Interaction between membrane components causes significant deviations from the additivity rule. Area, capacitance, and stability constant values for the complexes were calculated based on the model assuming 1:1 stoichiometry, and the model was validated by comparison of these values to experimental results. We established that phosphatidylcholine and decylamine form highly stable 1:1 complexes. In the case of decanoic acid-modified phosphatidylcholine membranes, complexes with stoichiometries other than 1:1 should be taken into consideration.

The Influence of pH on Phosphatidylethanolamine Monolayer at the Air/Aqueous Solution Interface

The dependence of the interfacial tension of a phosphatidylethanolamine (PE) monolayer on the pH of the aqueous solution has been studied. A theoretical equation is derived to describe this dependence. A simple model of the influence of pH on the phosphatidylethanolamine monolayer at the air/hydrophobic chains of PE is presented. The contributions of additive phosphatidylethanolamine forms (both interfacial tension values and molecular area values) depend on pH. The interfacial tension values and the molecular area values for PEH+ and PEOH− forms of phosphatidylethanolamine were calculated. The assumed model was verified experimentally. The experimental results agreed with those derived from the theoretical equation in a whole range of pH values.

Influence of pH on Sphingomyelin Monolayer at Air/Aqueous Solution Interface

The pH dependence of the interfacial tension is an important factor in the behavior of sphingomyelin (SM) monolayers. We developed a theoretical model to describe this dependence in which the interfacial tension and molecular area contributions of each sphingomyelin form were additive and dependent on pH. The interfacial tension values and the molecular areas values for the SMH(+) and SMOH(-) forms of sphingomyelin were calculated and the proposed model was experimentally verified. The theoretical predictions agreed with the experimental results for pH values ranging from 2 to 12.