Patrycja Dynarowicz-Łątka

Molecular interaction in mixed monolayers at the air/water interface

Abstract Different approaches used to characterize molecular interaction in mixed monolayers, formed either by adsorption or by spreading at the air/water interface, are reviewed in this paper.

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.

X-ray grazing incidence diffraction and Langmuir monolayer studies of the interaction of β-cyclodextrin with model lipid membranes

The interactions of beta-CD with one component monolayers of cholesterol (chol), 1-stearoyl-sn-glycero-3-phosphocholine (lyso-PC), 1,2-dipalmitpyl-sn-phosphocholine (DPPC), sphingomyelin (SM) and the SM/chol and DPPC/chol mixtures have been investigated by the Langmuir monolayer technique and the synchrotron grazing incidence X-ray diffraction (GIXD). The investigated lipid monolayers have been studied with and without the 10(-3) M solution of beta-CD in the aqueous subphase. The surface pressure-area (pi-A) isotherms and the relaxation of the monolayers (surface pressure-time curves) were monitored. Our experiments reveal that there is not impact of beta-CD on the packing properties of the DPPC monolayers, while the presence of beta-CD in subphase changes the in-plane organization of SM molecules. Monolayers composed of pure chol molecules have been rapidly affected by the presence of the beta-CD in the subphase. Our data show that beta-CD can complex and desorb one-chain phospholipid (lyso-PC) but this process is relatively slow and, as indicated by the GIXD data, beta-CD molecules are present at the air/water interface. Subtraction of cholesterol by the beta-CD from mixed binary systems containing SM/chol (70/30, 50/50 and 30/70 mol ratio) and DPPC/chol (70/30 and 50/50 mol ratio) has also been investigated. Our experiments proved that cholesterol can be removed from the mixed monolayers only when it is unbound. The beta-CD was not capable to distract the monolayers of the SM/chol, forming a stable complex of the 2:1 stoichiometry (as observed in the model lipid raft). Interestingly, at the surface pressure of 30 mN/m also at the molar proportion of 50/50 no cholesterol removal was observed. This was interpreted by relatively strong SM/chol interactions and the tight packing of the mixed monolayer. For model membranes, in which cholesterol was in large excess (SM/chol, 30/70) the beta-CD extraction of cholesterol was observed, and the membrane composition evolves towards the lipid proportion corresponding to the stable complex stoichiometry (SM/chol 2:1).

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.

Cholesterol and phytosterols effect on sphingomyelin/phosphatidylcholine model membranes—Thermodynamic analysis of the interactions in ternary monolayers

In this work thermodynamic analysis of the interactions between lipids in ternary sphingomyelin/DPPC/sterol Langmuir films were performed to compare the effect of cholesterol, beta-sitosterol and stigmasterol on a model membrane. The condensing effect of the respective sterols and the interactions between molecules in ternary mixtures were analyzed on the basis of the excess area per molecule and the excess free energy of mixing values. The stability of the mixed monolayers was verified with the free energy of mixing values. The conclusions on the ordering effect of sterols were drawn from the analysis of the compression modulus values. It was found that the stoichiometry of the mixed films of the highest thermodynamic stability and of the strongest interactions is the same for all the sterols investigated. The results obtained prove that the mammalian sterol induces the strongest contraction of the area and reveals the strongest stabilizing and ordering effect among the investigated sterol. Stigmasterol was found to condense a model membrane in a weaker extent as compared to beta-sitosterol, however, the differences in ordering properties of both phytosterols are less pronounced. The magnitude of the influence of the investigated sterols on a model membrane was thoroughly discussed from the point of view of the structure of their side chain, which determines the geometry of a sterol molecule.

Effect of saturation degree on the interactions between fatty acids and phosphatidylcholines in binary and ternary Langmuir monolayers

Fatty acids molecules have strong influence on membrane properties. In this work the interactions between fatty acids and phosphatidylcholines were studied in mixed binary and ternary Langmuir monolayers. The compounds investigated, both fatty acids and phospholipids were of the same hydrocarbon chain length and differed in their saturation degree (stearic acid (C18:0)), oleic acid (C18:1)), linoleic acid (C18:2) and distearoylphosphatidylcholine-DSPC and dioleoylphosphatidylcholine-DOPC). It was found that the investigated fatty acids interact more strongly with saturated phospholipid (DSPC) and more strongly affect the molecular organization of DSPC films as compared to DOPC monolayer. The saturated fatty acid (stearic acid) makes the phospholipid monolayers more rigid, while unsaturated fatty acids decrease the condensation of phospholipid films. The differences in the effect of the respective fatty acids on phosphatidylcholines monolayers were thoroughly analyzed from the point of view of geometry of the lipids molecules, which is determined by the saturation of the hydrocarbon chain.

Edelfosine disturbs the sphingomyelin–cholesterol model membrane system in a cholesterol-dependent way – The Langmuir monolayer study

Synthetic alkyl-lysophospholipids, represented by edelfosine (ED), reveal strong anticancer activity and therefore are promising drugs used in anticancer therapy. Primary target for edelfosine is cellular membrane, which is in contrast to traditional cytostatics affecting DNA. The mechanism of antitumor activity of edelfosine was hypothesized to be related to its accumulation in membrane rafts. Inspired by these findings, we have performed the Langmuir monolayer studies on the influence of edelfosine on systems composed of sphingomyelin (SM) and cholesterol (Chol), being the principal components of membrane rafts. Sphingomyelin-cholesterol proportion in monolayers was varied to reflect the composition of solely membrane rafts (SM/Chol=2:1) and contain excess of cholesterol (SM/Chol=1:1 and 1:2). Into these systems, edelfosine was added in various concentrations. The analysis of surface pressure-area isotherms, complemented with films visualization with Brewster angle microscopy (BAM) allowed us to compare the effect of edelfosine on condensation and ordering of SM/Chol monolayers. The results evidenced that the influence of ED on the interactions in model membranes and its fluidizing effect is highly cholesterol-dependent. The strongest decrease of monolayer ordering was observed for model raft system, while the excess of cholesterol present in the remaining mixtures was found to weaken the fluidizing effect of the drug.

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.

Effects of β-Cyclodextrin on the Structure of Sphingomyelin/Cholesterol Model Membranes

The interaction of beta-cyclodextrin (beta-CD) with mixed bilayers composed of sphingomylein and cholesterol (Chol) above and below the accepted stable complexation ratio (67:33) was investigated. Membranes with the same (symmetric) and different (asymmetric) compositions in their inner and outer leaflets were deposited at surface pressures of 20, 30, and 40 mN/m at the solid-liquid interface. Using neutron reflectometry, membranes of various global molar ratios (defined as the sum of the molar ratios of the inner and outer leaflets), were characterized before and after beta-CD was added to the subphase. The structure of bilayers with global molar ratios at or above the stable complexation ratio was unchanged by beta-CD, indicating that beta-CD is unable to remove sphingomyelin or complexed Chol. However, beta-CD removed all uncomplexed Chol from bilayers composed of global molar ratios below the stable complexation ratio. The removal of Chol by beta-CD was independent of the initial structure of the membranes as deposited, suggesting that asymmetric membranes homogenize by the exchange of molecules between leaflets. The interaction of beta-CD with the aforementioned membranes was independent of the deposition surface pressure except for a symmetric 50:50 membrane deposited at 40 mN/m. The scattering from 50:50 bilayers with higher packing densities (deposited at 40 mN/m) was unaffected by beta-CD, suggesting that the removal of Chol can depend on both the composition and packing density of the membrane.