J. Miñones

Interactions between membrane sterols and phospholipids in model mammalian and fungi cellular membranes — A Langmuir monolayer study

This paper is aimed at investigating sterol/phospholipid interactions in the exact proportion that occurs in fungi/mammalian cells. We have performed a thorough analysis of surface pressure (pi)-area (A) isotherms with the Langmuir monolayer technique, complemented with Brewster angle microscopy (BAM) images. The following mixtures were analysed: cholesterol (Chol)-dipalmitoyl phosphatidylcholine (DPPC), Chol-dioleoyl phosphatidylcholine (DOPC), ergosterol (Erg)-DPPC, and Erg-DOPC. For each system, two different concentrations of the sterols were used, 13 and 30%, corresponding to the range of concentration found in various natural membranes. The obtained results show the existence of attractive interactions between phospholipids and cholesterol. Mixtures with ergosterol behave quite differently, i.e. either the interactions are repulsive (mixtures with DPPC) or the system is ideal (mixtures with DOPC). The obtained results have implications in the polyene antibiotics mode of action, i.e. the polyenes may interact easier with ergosterol, present in fungi cells, as compared to cholesterol--the main sterol of the mammalian cellular membranes.

Study of penetration of amphotericin B into cholesterol or ergosterol containing dipalmitoyl phosphatidylcholine Langmuir monolayers

Abstract The interactions of amphotericin B (AmB) with sterols and phospholipids have been studied by adsorption of AmB from aqueous solutions into Langmuir monolayers from dipalmitoyl phosphatidylcholine (DPPC), ergosterol, cholesterol and their mixtures. The results show that AmB exhibits stronger interaction with cholesterol than ergosterol in one-component monolayers. However, for DPPC–sterol monolayers, the effectiveness of AmB penetration depends on the proportion of both film components in the mixed film as well as on the strength of interaction between DPPC and particular sterol.

Orientational changes in dipalmitoyl phosphatidyl glycerol Langmuir monolayers.

Dipalmitoyl phosphatidyl glycerol (DPPG) as Langmuir monolayers at the air/water interface was investigated by means of surface pressure measurements in addition to Brewster angle microscopy (BAM) during film compression/expansion. A characteristic phase transition region appeared in the course of surface pressure-area (pi-A) isotherms for monolayers spread on alkaline water or buffer subphase, while on neutral or acidic water the plateau region was absent. This phase transition region was attributed to the ionization of DPPG monolayer. It has been postulated that the ionization of the phosphatidyl glycerol group leads to its increased solvation, which probably provokes both a change in the orientation of the polar group and its deeper penetration into bulk phase. Film compression along the transition region provokes the dehydration of polar groups and subsequent change of their conformation, thus causing the DPPG molecules to emerge up to the interface. Quantitative Brewster angle microscopy (BAM) measurements revealed that along the liquid-expanded to liquid-condensed phase transition the thickness of the ionized DPPG monolayer increases by 4.2 A as a result of the conformational changes of the ionized polar groups, which tend to emerge from the bulk subphase up to the surface.

Interactions of amphotericin B with saturated and unsaturated phosphatidylcholines at the air/water interface

Abstract Three synthetic phospholipids, differing in the degree of saturation of phosphatidylcholine (PC) acyl chains, namely: dipalmitoyl phosphatidylcholine (DPPC), dioleoyl phosphatidylcholine (DOPC) and palmitoyl-oleoyl phosphatidyl choline (POPC) were mixed with the antimycotic polyene antibiotic, amphotericin B (AmB), and investigated as monolayers at the air/water interface. The mixed films were spread on water (pH 6) at room temperature, and the surface pressure–area ( π / A ) isotherms were recorded upon compression. The interactions were examined by analysing the phase transitions of the monolayers and calculating the films compressibility. The obtained results indicate the existence of stronger interaction between AmB and the saturated phospholipid as compared with unsaturated ones, and reveal the influence of apolar structure of PC chains on the stoichiometry of AmB–PC complexes.

Interactions of lauryl gallate with phospholipid components of biological membranes.

The effect of different amounts of lauryl gallate (LG) on properties of the model membranes of phosphatidylcholines (PC), differing in the presence of double bonds in the hydrocarbon chains, and phosphatidylglycerol (PG) was described in terms of phase behaviour of mixtures, interactions between both components, monolayers stability and their organization. The Langmuir monolayer technique was used to monitor the surface thermodynamics (i.e. the excess area and excess Gibbs energy of mixing) on the basis of surface pressure-area per molecule (π-A) isotherms. Simultaneously, morphology of the studied monolayers was visualized by the Brewster angle microscopy (BAM). This allowed evaluating the kind and magnitude of interactions which influence on the phase behaviour and structural properties of the monolayers. The obtained results can be helpful to reveal the mechanism of phospholipid antioxidant protection and important pharmacological (antimicrobial) role of lauryl gallate for production of effective therapeutic substances.

Monolayer Properties of Uronic Acid Bicatenary Derivatives at the Air-Water Interface: Effect of Hydroxyl Group Stereochemistry Evidenced by Experimental and Computational Approaches

By screening uronic acid-based surfactant interfacial properties, the effect of the hydroxyl group stereochemistry (OH-4) on the conformation of bicatenary (disubstituted) derivatives at the air-water interface has been evidenced by experimental and computational approaches. Physical and optical properties of a monolayer characterized by Langmuir film balance, Brewster angle microscopy, and ellipsometry at 20 °C reveal that the derivative of glucuronate (C(14/14)-GlcA) forms a more expanded monolayer, and shows a transition state under compression, in the opposite to that of galacturonate (C(14/14)-GalA). Both films are very mechanically resistant (compression modulus > 300 mN m(-1)) and stable (collapse pressure exceeding 60 mN m(-1)), while that of C(14/14)-GalA exhibits a very high compression modulus up to 600 mN m(-1) like films in the solid state. Computational approaches provide single and assembly molecular models that corroborate the molecule expansion degree and interactions data from experimental results. Differences in the molecular conformation and film behaviours of uronic acid bicatenary derivatives at the air-water interface are attributed to the intra-H-bonding formation, which is more favourable with an OH-4 in the axial (C(14/14)-GalA) than in the equatorial position (C(14/14)-GlcA).

Study of the interaction of cholesterol with potentially hypocholesterolemic substances in a monolayer form

Abstract The behaviour of mixed monolayers of cholesterol (chol) and stigmastanil phosporylcholine (SPC) at the air–water interface was studied and compared with that of the cholesterol–arachidonic acid (Ara) system in order to examine the molecular interactions between the film components. The structural characteristics of the mixed films was determined from the π – A isotherms recorded on an automated Langmuir-type film balance. The studies confirm that the mixed films are homogeneous and their components exhibit negative deviations from ideality as the result of mutual interactions at the interface. The magnitude of these interactions depends on the surface pressure region, temperature, as well as the system composition. Values of the thermodynamic parameters such as Δ G exc , Δ S exc and Δ H exc provide evidence for the presence of interactions, which are stronger in the chol–SPC system than in the chol–Ara mixed films.

A study of the stability of amphotericin B monolayers

Amphotericin B (AmB) is a polyene antibiotic used as an antimycotic agent that forms relatively stable monolayers at low surface pressures (π) at the air/water interface. At increased π values, however, the monolayer structure undergoes a rearrangement, which is manifest as a constant surface pressure plateau in the corresponding π-A curve. Beyond such a plateau, when the monolayer is in a liquid condensed state, the area occupied by the molecules in the film is so small that this suggests the antibiotic is to some extent dissolved in the subphase. In order to confirm this hypothesis, the behaviour of AmB spread on various subphases was examined under variable conditions regarding the number of AmB molecules deposited, the rate at which the surface film was compressed and the temperature and ionic strength of the subphase. Relaxation experiments involving measurements of the rate of molecular area decrease at a constant surface pressure were also carried out. Based on the results, AmB desorption is diffusion controlled at low surface pressures and arises from its dissolution in two consecutive steps at surface pressures beyond those of plateau in the π -A curves, where AmB molecules are assumed to lie normal to the subphase surface.

Miscibility and Langmuir Studies of the Interaction of E2 (279–298) Peptide Sequence of Hepatitis G Virus/GB Virus-C with Dipalmitoylphosphatidyl Choline and Dimiristoylphosphatidyl Choline Phospholipids

Mixed monolayers of E2(279-298), a synthetic peptide belonging to the structural protein E2 of the GB virus C (GBV-C), formerly know as hepatitis G virus (HGV), and the phospholipids dipalmitoylphosphatidyl choline (DPPC) and dimiristoylphosphatidyl choline (DMPC),which differ in acyl chains length, were obtained at the A/W interface (monolayers of extension) in order to provide new insights on E2/phospholipids interaction. Analysis of the surface pressure-area isotherms, Brewster angle microscopy images, relative thickness, and mean areas per molecule has allowed us to establish the conditions under which the mixed components of the monolayer are miscible or immiscible and know how the level of the E2/phospholipid interaction varies with the composition of the mixed films, the surface pressure, and the hydrocarbon chains length of the phospholipids. The steric hindrance caused by the penetration of the polymer strands into the more or less ordered hydrocarbon chains of the phospholipids was suggested to explain the differences in the peptide interaction with the phospholipids studied. Therefore, the novelty of results obtained with the Langmuir film balance technique, supplemented with BAM images allow us to achieve a deeper understanding of the interaction.

Interactions in Monolayers: A Study of the Behavior of Poly(methyl methacrylate)—Lysozyme Mixed Films from Surface Pressure—Area and Ellipsometric Measurements

Mixed monolayers of PMMA-lysozyme show the existence of negative deviations from the additivity of the molecular areas (A(m)) when the composition of polymer mixtures is less than X(PMMA) 0.6, regardless of the surface pressure of the monolayers. The maximum deviation occurs in the mixed monolayer with composition X(PMMA) 0.25, which is attributed to the formation of a complex consisting of one polymer molecule and three protein molecules (1:3 stoichiometry), stabilized by hydrogen bonds between the NH groups of the protein and the CO groups of the polymer as well as by van der Waals attractive forces between the hydrocarbon chains of both components. When the relative proportion of the components in the mixed films significantly differs from the value corresponding to the stoichiometry of the complex (as in mixtures with X(PMMA) > 0.6), this complex cannot be formed, causing an immiscible system where the values of the experimental molecular areas coincide with those corresponding to ideal behavior. Measurements of monolayer thickness and BAM images allow confirmation on the microscopic level of the structural characteristics deduced from the π-A isotherms.