The influence of terpinen-4-ol and eucalyptol - The essential oil components - on fungi and plant sterol monolayers.

Antifungal and herbicidal activity of terpenes, being the components of the essential oils, is directly related to the incorporation of these compounds into cellular membranes. Thus, the differences in the lipid composition of various pathogenic membranes may be the factor determining the activity of these molecules. One of the class of lipids, which form the membrane environment are sterols. The aim of this work was to compare the effect of two terpenes: terpinen-4-ol and eucalyptol on the monolayers formed by ergosterol and ß - sitosterol, which are the components of fungi and plant membranes, respectively. The modifications in the sterol monolayer properties were investigated in the surface pressure-area measurements and penetration studies as well as in a micrometer scale (Brewster angle microscopy experiments) and in nanoscale (GIXD technique). It was evidenced that although at higher surface pressure the terpene molecules are in part removed from the interface, they are able to substantially modify the condensation, morphology and molecular organization of the sterol film. It was also found that the incorporation of terpenes into sterol films is comparable for both sterols, however, ß - sitosterol monolayers properties are affected more strongly than ergosterol films. Finally, the analysis of the results of the studies performed on model membrane systems and the results of antimicrobial studies reported in literature, enabled us to suggest that the activity of terpenes depends on the membrane composition and that the sterol concentration may be important from the point of view of antifungal effect of terpinen-4-ol and eucalyptol.

Interactions of Long-Chain Perfluorotelomer Alcohol and Perfluorinated Hydrocarbons with Model Decomposer Membranes.

Perfluorinated hydrocarbons and their polar derivatives are produced annually in high quantities and find multiple industrial and technological applications due to their chemical and physical durability, significant hydro- and lipophobicity and excellent surface activity. Unfortunately, multiple perfluorinated compounds are recognized as persistent organic pollutants as they are completely nonbiodegradable and accumulate in soils and sediments. In our studies, we applied Langmuir monolayers formed by different structural phospholipids as models of soil bacteria and fungi membranes and investigated the effects exerted by long-chain perfluorinated pollutants, perfluorotelomer alcohol and two structurally different perfluorinated hydrocarbons, on the artificial membranes. Various mutually complemental methods such as surface pressure-mean molecular area isotherm registration, Brewster angle microscopy (BAM), and grazing incidence X-ray diffraction (GIXD) were applied. It turned out that the presence of the perfluorinated chemicals profoundly affected the phospholipid monolayers. The miscibility of the phospholipid with the perfluorotelomer alcohol depended strongly on the size and charge of the polar headgroup. Additionally, it was observed by BAM that the presence of the perfluorinated molecules significantly changed the texture of all the investigated phospholipid monolayers. On the basis of the BAM and GIXD results and other studies described in the scientific literature, we postulated that the perfluorinated hydrocarbons form an additional monolayer anchored on top of the phospholipid film. Our studies prove that both polar and nonpolar perfluorinated pollutants can be toxic to decomposer organisms and that their toxicity is strictly correlated with the phospholipid composition of the cellular membrane.

Influence of Cationic Phosphatidylcholine Derivative on Monolayer and Bilayer Artificial Bacterial Membranes.

An increasing number of bacterial infections and the rise in antibiotic resistance of a number of bacteria species forces one to search for new antibacterial compounds. The latter facts motivate the investigations presented herein and are aimed at studying the influence of a cationic lipid, 1-palmitoyl-2-oleoyl- sn-glycero-3-ethylphosphocholine (EPOPC), on model (mono- and bilayer) membranes. The monolayer experiments involved the analysis of the interactions of EPOPC with bacterial membrane lipids in one component and mixed systems as well as Brewster angle microcopy studies. The properties of liposomes were analyzed based on the results of dynamic light scattering (DLS) and zeta potential measurements as well as on the experiments concerning the release of calcein entrapped in liposomes after titration with surfactant solution and steady-state fluorescence anisotropy of DPH. The obtained results evidenced that EPOPC, even at low concentrations, strongly changes organization of model systems making them less condensed. Moreover, EPOPC decreases the hydrodynamic diameter of liposomes, increases their zeta potential, and destabilizes model membranes, increasing their fluidity and permeability. Also, the in vitro tests performed on Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) strains prove that EPOPC has some bacteriostatic properties which seem to be stronger toward Gram-negative than Gram-positive bacteria. All these findings allow one to conclude that EPOPC mode of action may be directly connected with the interactions of EPOPC molecules with bacterial membranes.

Influence of Parabens on Bacteria and Fungi Cellular Membranes: Studies in Model Two-Dimensional Lipid Systems.

Langmuir monolayers were used to study the influence of four commercially applied parabens on multicomponent systems composed of lipid species characteristic of the cellular membrane of microorganisms found in carbohydrates and proteins reaching products, including food and cosmetics. The aim of the undertaken studies was to shed new light on the problem of parabens' interactions with membrane lipids and their affinity for monolayers differing with regard to the composition, mutual lipid ratios, and physicochemical properties. The discussion is based on the π-A isotherm characteristics, surface morphology observation performed with BAM, and analysis of the diffraction data collected for the periodically ordered lipid domains present in the investigated multicomponent films. Our studies revealed that the selected parabens are capable of surface film modification and that the magnitude of this effect increases with the number of methylene groups in the ester part of paraben molecules. We found that the strongest destructive effect was observed for model 1 (Staphylococcus aureus), a lower effect was observed for model 2 (Pseudomonas aeruginosa), and the lowest effect was observed for model 3 (Candida albicans). It was inferred that such a trend appears due to the composition of the artificial membranes, i.e., above all, in the presence or lack of sterol molecules and the content of negatively charged lipids.

Polycyclic aromatic hydrocarbons in model bacterial membranes - Langmuir monolayer studies.

High molecular weight polycyclic aromatic hydrocarbons (HMW-PAHs) are persistent organic pollutants which due to their limited biodegradability accumulate in soils where their increased presence can lead to the impoverishment of the decomposer organisms. As very hydrophobic PAHs easily penetrate cellular membranes of soil bacteria and can be incorporated therein, changing the membrane fluidity and other functions which in consequence can lead to the death of the organism. The structure and size of PAH molecule can be crucial for its membrane activity; however the correlation between PAH structure and its interaction with phospholipids have not been investigated so far. In our studies we applied phospholipid Langmuir monolayers as model bacterial membranes and investigated how the incorporation of six structurally different PAH molecules change the membrane texture and physical properties. In our studies we registered surface pressure and surface potential isotherms upon the monolayer compression, visualized the monolayer texture with the application of Brewster angle microscopy and searched the ordering of the film-forming molecules with molecular resolution with the application of grazing incidence X-ray diffraction (GIXD) method. It turned out that the phospholipid-PAH interactions are strictly structure dependent. Four and five-ring PAHs of the angular or cluster geometry can be incorporated into the model membranes changing profoundly their textures and fluidity; whereas linear or large cluster PAHs cannot be incorporated and separate from the lipid matrix. The observed phenomena were explained based on structural similarities of the applied PAHs with membrane steroids and hopanoids.

Studies on the Behavior of Eucalyptol and Terpinen-4-ol-Natural Food Additives and Ecological Pesticides-in Model Lipid Membranes.

Effective application of the essential oils requires detailed exploration of their mechanism of action and the origin of diverse activity of their components. In this work, the influence of eucalyptol and terpinen-4-ol on artificial membranes was studied to verify whether the differences in the activity of these compounds are related to their effect on membranes. The properties of monolayers formed from structurally different lipids in the presence of terpenes were examined based on the results of the surface pressure-area measurements, penetration studies, and Brewster angle microscopy experiments. Both compounds were able to incorporate into the membrane and alter lipid/lipid interactions, making the monolayer less stable and more fluid. These effects were determined by monolayer composition (but not by its condensation per se) and the resulting rheological properties and were stronger in the presence of terpinen-4-ol. These findings confirm the hypothesis that differences in the antimicrobial potency of these terpenes are membrane-related, and membrane composition may determine their selectivity.

Phytohormone Behavior in the Model Environment of Plant and Human Lipid Membranes.

Interactions between three auxins (indole-3-acetic acid (IAA), 2-naphthoxyacetic acid (BNOA), and 2,4-dichlorophenoxyacetic acid (2,4-D)) and model two-dimensional lipid systems mimicking plant and human cell membranes were investigated in monolayers formed at the air/water solution interface. The analysis was based on the recorded π-A isotherm characteristics complemented with Brewster angle microscopy. The influence of auxins on model membranes was discussed on the basis of condensation changes, modification of mutual lipid-lipid interactions in the mixed films, and morphological alteration of the surface domains on the microscopic scale. It was demonstrated that the lipid composition and mutual proportion of the artificial membranes together with sterol to main the phospholipid ratio play a crucial role in the context of auxin behavior in the membrane-mimicking environment. Apart from specific molecular interactions between studied phytohormones represented by auxins and lipids, the condensation of the investigated monolayers was found to be a regulative factor of model systems' susceptibility toward auxin action. Two effects were recognized: fluidizing of monolayers being in the liquid state (model membranes) and initialization of the three-dimensional structure formation in ordered sterol films at high surface pressure. The influence of auxin molecules on lipid interactions in the monolayer and diminishing of the film condensation was the largest for BNOA, due to the presence of the most bulky nonpolar, aromatic fragment in the molecule. It was also demonstrated that auxins interact with model plant membranes more selectively, stronger, and at markedly lower concentration than with the human membrane models.

Essential oils as food eco-preservatives: Model system studies on the effect of temperature on limonene antibacterial activity.

Antimicrobial properties of essential oils predestine these substances to be used as ecological food preservatives. However, their activity is determined by variety of factors among which external conditions and food properties are highly important. Herein the influence of limonene on artificial membranes was studied to verify the effect of temperature on the incorporation of this compound into model bacterial membrane. The investigations were done on lipid monolayers and the experiments involved the surface pressure-area measurements, penetration studies and Brewster Angle Microscopy analysis. It was found that limonene incorporates into lipid monolayers causing their fluidization. However, the magnitude of alterations depends on limonene concentration, model membrane composition and, for a given composition, on system condensation. Moreover, the influence of limonene is stronger at lower temperatures and, in the light of collected data, this may be a consequence of strong volatility and evaporation of limonene increasing with temperature.

Sterol-Phospholipid Hybrids at the Air/Water Interface: Studies on Properties and Interactions with Parent Lipid Molecules.

The two synthetic sterol-phospholipid hybrids DCholPC and PCholPC were investigated in monolayers at the air/water interface. Study was based on π-A isotherm analysis complemented with application of grazing incidence X-ray diffraction. It was found that both compounds are capable of forming stable, highly condensed monolayers of a surface characteristics typical for sterols. GIXD studies show that the crystallographic area for DCholPC monolayer is very similar to that found for cholesterol (37.1 vs 38.0 Å(2)), while for PCholPC (28.8 Å(2)) it is significantly smaller as compared to area for the mixed Chol/DPPC 2/1 monolayer (33.4 Å(2)). In our study the problem of interactions between investigated sterol-phospholipid hybrids and natural membrane lipid components was for the first time analyzed in planar lipid systems. Studies on mixed monolayers showed that both hybrids, similarly to cholesterol, reveal a condensing effect toward DPPC acyl chains; however, DCholPC having two steroid moieties in the molecule was found to be more efficient. On the other hand, the sterol moiety and the hydrocarbon chain of PCholPC molecule are packed in the 2D crystalline phase extremely tight. Our studies showed that the investigated compounds can be applied as biocompatible components of stable liposomes.

Studies on the interactions between parabens and lipid membrane components in monolayers at the air/aqueous solution interface.

The interactions between parabens (PBs) and lipid components of mammalian and bacterial cell membranes were investigated in model systems of Langmuir monolayers. Me-, Et-, Pr- and Bu-paraben studied in this paper are frequently applied as cosmetics and food preservatives, since they possess broad antimicrobial activity. The mode of PB action is connected with their incorporation into the membrane of bacterial organisms, however; it is not known what is the role of the respective lipid species in this mechanism. This problem is crucial to understand the differences in paraben activity toward individual microorganisms and to shed the light onto the problem of PB cytotoxicity reported in studies on mammalian cells. In this paper, the mentioned aspects were investigated with application of the Langmuir monolayer technique complemented with BAM and GIXD. Our experiments revealed that the influence of PBs depends on their chemical structure, solution concentration and on the class of lipid. The strongest modification of the monolayer characteristics, leading to its collapse at low surface pressure, occurred in the presence of BuPB, having the largest chain. PBs interact preferentially with the monolayers possessing low degree of condensation, whereas for LC state, the effect was weaker and observed only as modification of the 2D unit cells. In the model systems, PBs interact with phospholipids characteristic for mammalian membranes (phosphatidylcholine) stronger than with bacterial (phosphatidylglycerol and cardiolipin). This strong influence of parabens on the model systems composed of animal lipids may explain cytotoxic activity of these preservatives.

Studies on the interactions of bisphenols with anionic phospholipids of decomposer membranes in model systems.

Bisphenol A (BPA) and other bisphenols constitute a class of organic pollutants, which because of their estrogenic properties, low dose activity and bioaccumulation pose considerable risk for public health as well as for the environment. Accumulated in the sediment bisphenols can endanger the decomposers' populations being incorporated into their cellular membranes; however, the mechanism of their membrane activity is unknown. Therefore, to study these phenomena we applied anionic phospholipid Langmuir monolayers as simple but versatile models of decomposers biomembranes. Phosphatidylglycerols and cardiolipins are not only the main components of bacterial membranes but also of crucial importance in mitochondrial and thylakoid membranes in eukaryotic cells. In our investigations we applied five compounds of the bisphenol class most commonly detected in the environment. To characterize the bisphenols-model membrane interactions we applied multiple mutually independent methods of physical chemistry; namely: the Langmuir monolayer technique, surface potential measurements, Brewster angle microscopy for the visualization of the monolayers' texture and grazing incidence X-ray diffraction for the discussion of the phospholipids packing within the monolayers. Our studies indicated that all the investigated bisphenols interact with the model membrane, but the strength of the interactions is dependent on the bisphenol structure and hydrophobicity and the fluidity of the model membranes. We proved that bisphenol S often treated as the least toxic BPA analog can also be incorporated to the model membranes changing their structure and fluidity.

Characteristics of the influence of auxins on physicochemical properties of membrane phospholipids in monolayers at the air/aqueous solution interface.

Interactions between representatives of plant hormones and selected membrane lipids have been studied in monolayers at the air/aqueous solutions interface with π-A isotherm analysis, microscopic visualization and grazing incidence X-ray diffraction technique (GIXD). Four phytohormones: indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), 2-naphthoxyacetic acid (BNOA) and 2,4-dichlorophenoxyacetic acid (2,4-D), belonging to the class of auxins differ as regards the chemical structure of the aromatic molecular fragment. The studied phospholipids have been chosen since they are omnipresent in the biological membranes of plant and animal kingdom. Our results revealed that both natural (IAA and IBA) and synthetic (2,4-D and BNOA) phytohormones modify the physicochemical characteristics of the investigated lipid monolayers. Auxins caused strong diminishing of the monolayer condensation, especially for DPPC and SOPE, which may be attributed to the phase transition in these monolayers. In the performed experiments the key step of auxins action occurs when the molecules interact with monolayers in the expanded state-when the space in the lipid head-group region is large enough to accommodate the molecules of water soluble auxins. The application of GIXD technique confirmed that auxin molecules are also present at the interface at higher surface pressure (30 mN/m). The obtained results showed that among the investigated auxins, the largest influence on the lipid monolayers occurred in the case of BNOA, which molecule possesses the largest aromatic fragment. In contrast, 2,4-D, having the smallest aryl group affects the studied lipid systems to the smallest extent.

Crucial Role of the Double Bond Isomerism in the Steroid B-Ring on the Membrane Properties of Sterols. Grazing Incidence X-Ray Diffraction and Brewster Angle Microscopy Studies.

Three cholesterol precursors-desmosterol, zymosterol, and lanosterol-were comprehensively characterized in monolayers formed at the air/water interface. The studies were based on registration of the surface pressure (π)-area (A) isotherms complemented with in situ analysis performed with application of modern physicochemical techniques: grazing incidence X-ray diffraction (GIXD) and Brewster angle microscopy (BAM). In this approach we were interested in the correlation between molecular structures of the studied sterols found in the cholesterol biosynthetic pathway and their membrane properties. Our results revealed that only desmosterol behaves in Langmuir monolayers comparably to cholesterol, the molecules of which arrange in the monolayers into a hexagonal lattice, while the two remaining sterols possess extremely different properties. We found that molecules of both zymosterol and lanosterol are organized on the water surface in the two-dimensional oblique unit cells despite the fact that they are oriented perpendicular to the monolayer plane. The comparison of chemical structures of the investigated sterols leads to the conclusion that the only structural motive that can be responsible for such unusual behavior is the double bond in the B sterol ring, which is located in desmosterol in a different position from in the other two sterols. This issue, which was neglected in the scientific literature, seems to have crucial importance for sterol activity in biomembranes. We showed that this structural modification in sterol molecules is directly responsible for their adaptation to proper functioning in biomembranes.

The influence of cholesterol precursor--desmosterol--on artificial lipid membranes.

The disorders in cholesterol biosynthesis pathway and various diseases manifest in the accumulation of cholesterol precursors in the human tissues and cellular membranes. In this paper the effect of desmosterol--one of cholesterol precursors--on model lipid membranes was studied. The investigations were performed for binary SM/desmo and POPC/desmo and ternary SM/POPC/desmo monolayers. Moreover, the experiments based on the gradual substitution of cholesterol by desmosterol in SM/POPC/chol=1:1:1 system were done. The obtained results allowed one to conclude that desmosterol is of lower domains promoting and stabilizing properties and packs less tightly with the lipids in monolayers. Moreover, desmosterol probably could replace cholesterol in model membranes, but only at its low proportion in the system (2%), however, at a higher degree of cholesterol substitution a significant decrease of the monolayer stability and packing and alterations in the film morphology were detected. The results collected in this work together with those from previous experiments allowed one to analyze the effect of a double bond in the sterol side chain as well as its position in the ring system on membrane activity of the molecule and to verify Bloch hypothesis.

Antagonistic effects of α-tocopherol and ursolic acid on model bacterial membranes.

α-tocopherol (Toc), the most active component of vitamin E can exert antagonistic effects disabling the therapy of cancers and bacterial infections. Such antagonisms were observed also between Toc and bioactive pentacyclic triterpenes (PT) exhibiting anticancer and antibacterial properties. Both Toc and PT are water-insoluble membrane active substances. Thus, our idea was to emulate their interactions with model Escherichia coli membranes. E. coli inner membranes were selected for the experiments because their lipid composition is quite simple and well characterized and the two main components are phosphatidylethanolamine and phosphatidylglycerol. As a model of E. coli membranes we applied Langmuir monolayers formed by the E. coli total extract of polar lipids (Etotal) as well as by the main lipid components: phosphatidylethanolamine (POPE) and phosphatidylglycerol (ECPG). The antagonistic effects of ursolic acid (Urs) and Toc were investigated with the application of ternary Langmuir monolayers formed by Urs, Toc and one of the phospholipids POPE or ECPG. Our studies indicated that the affinities of Urs and Toc towards the POPE molecule are comparable; whereas there are profound differences in the interactions of Urs and Toc with ECPG. Thus, the model experiments prove that in the case of E. coli membrane, the differences in the interactions between Urs and Toc with the anionic bacterial phosphatidylglycerol can be the key factor responsible for the antagonistic effects observed between PT and Toc in vivo.

The studies on the toxicity mechanism of environmentally hazardous natural (IAA) and synthetic (NAA) auxin--The experiments on model Arabidopsis thaliana and rat liver plasma membranes.

This paper concerns the studies towards membrane-damage effect of two auxins: indole-3-acetic acid - IAA and 1-naphthaleneacetic acid - NAA on plant (Arabidopsis thaliana) and animal (rat liver) model membranes. The foregoing auxins are plant growth regulators widely used in agriculture to control the quality of the crop. However, their accumulation in the environment makes them hazardous for the living organisms. The aim of our investigations was to compare the effect of natural (IAA) vs. synthetic (NAA) auxin on the organization of plant and animal model membranes and find a possible correlation between membrane-disturbing effect of these compounds and their toxicity. The collected data evidenced that auxins cause destabilization of membranes, decrease their condensation and weakens interactions of molecules. The alterations in the morphology of model systems were also noticed. The foregoing effects of auxins are concentration-dependent and additionally NAA was found to act on animal vs. plant membranes more selectively than IAA. Interestingly, both IAA and NAA induce the strongest disordering in model lipid system at the concentration, which is frequently reported as toxic to animal and plants. Based on the above findings it was proposed that membrane-damage effect induced by IAA and NAA may be important from the point of view of the mechanism of toxicity of these compounds and cannot be ignored in further investigations in this area.

Grazing incidence diffraction studies of the interactions between ursane-type antimicrobial triterpenes and bacterial anionic phospholipids.

α-Amyrin (AMalf) and ursolic acid (Urs) are ursane-type pentacyclic triterpenes which exhibit wide spectrum of antibacterial activity. These surface active compounds can be incorporated into bacterial membranes and alter their structure and function; however, the exact mechanism of their action still needs to be elucidated. Thus, we decided to study the interactions of these terpenes with specific anionic phospholipids:cardiolipins and phosphatidylglycerols extracted from Escherichia coli in the model environment of Langmuir monolayers. To characterize the ordering of the terpene molecules in one-component films as well as to study their interactions with the bacterial phospholipids in binary monolayers we applied grazing incidence X-ray diffraction (GIXD). It turned out that amyrins and ursolic acid molecules form crystalline hexagonal phases in Langmuir monolayers, in which the molecules are oriented uprightly. Regarding the mixtures, it was found that in the monolayers with Urs crystalline domains are present till moderate or even low Urs proportion. In contrast, in the mixtures with AMalf crystalline domains were observed only at the highest terpene concentration. In the interpretation of our results we underlined the significance of the interactions between the cyclopropane ring present in the hydrophobic part of the bacterial phospholipids and the terminal ring of the terpene structure. We proposed that the significant differences between the systems with AMalf and Urs are connected with the formation of hydrogen bonds between the Urs hydrophobic moieties. It can be inferred from the results that Urs is a more membrane-active agent than AMalf.

Studies of the interactions of ursane-type bioactive terpenes with the model of Escherichia coli inner membrane-Langmuir monolayer approach.

Pentacyclic triterpenes (PT), ursolic acid (Urs), and α-amyrin (AMalf) are natural products exhibiting broad spectrum of antibacterial activity. These compounds are membrane-active and can disorder bacterial membranes when incorporated; however, the exact mechanism of their membrane activity is unknown. In our studies, we applied Langmuir monolayer technique supported by Brewster angle microscopy to model the interactions of the selected PT with the lipid matrix of E. coli inner membrane. As the model membrane, we applied mixtures (75/25 mole/.mole %) of the representative Escherichia coli phosphatidylethanolamine (POPE), with the cardiolipin (ECCL) or phosphatidylglycerol (ECPG) extracted from the E. coli inner membrane. On the basis of the recorded isotherms, we performed thermodynamic analysis and calculated free energy of mixing ΔGexc. It turned out that the phospholipids forming the inner membrane of E. coli are ideally miscible, whereas in binary systems composed of PT and POPE, negative deviations from ideality indicating attractive interactions between the investigated PT and POPE molecules were observed. On the other hand, in ternary systems composed of PT, POPE and one of the E. coli anionic phospholipids large positive changes in ΔGexc were observed. Thus, both PT exhibit disorganizing effect on the model E. coli membrane. It was also proved that at low terpene proportion, AMalf can be more active than Urs. However, at higher proportion Urs incorporation can lead to the disintegration of cardiolipin-rich domains present in bacterial membrane.

The comparison of zymosterol vs cholesterol membrane properties--the effect of zymosterol on lipid monolayers.

In this work systematic investigations on the influence of zymosterol (zymo), which is one of cholesterol precursors, on lipid monolayers were done. The aim of these studies was to perform thorough comparison of zymosterol vs cholesterol membrane activity and fill the gap in the studies on the effect of sterols on membranes. The Langmuir monolayers experiments combined with Brewster angle microscopy studies were performed on binary (SM:zymo, POPC:zymo, GM3:zymo) and ternary (SM:POPC:zymo, SM:GM3:zymo) films differing in the sterol content. The obtained results evidenced differences in the influence of both sterols on lipid monolayers, which was manifested in the parameters calculated based on the isotherms as well is in monolayers morphology. It was found that zymosterol is of condensing, ordering and domain promoting abilities thus this molecule can be included to the group of membrane active sterols. However, zymosterol is much less effective than cholesterol as condensing and ordering agent. These findings were attributed to the differences in the structure of both sterols and their ability to pack tightly with other lipids in the mixed systems.

Incorporation of pentacyclic triterpenes into mitochondrial membrane--studies on the interactions in model 2D lipid systems.

Three representatives of naturally occurring pentacyclic triterpenes (PTs) were subjected to comprehensive studies aimed at the analysis of their interactions with phospholipids found naturally in mitochondrial membrane. To reach this goal, the selected compounds--α-amyrin (AMalf), betulinic acid (BAc), and ursolic acid (Urs)--were incorporated into two-component and multicomponent Langmuir monolayers acting as a model of mitochondrial membrane. As the lipids characteristic for mitochondria, phosphatidylcholine (POPC), phosphatidylethanolamine (POPE), and cardiolipin (BHCL) were chosen. Our studies were motivated by the fact that, according to the literature, the anticancer activity of PTs is correlated with their ability to incorporate into mitochondrial membrane and modify its properties. The undertaken studies were based on the surface pressure (π)-molecular area (A) isotherm registration complemented with the thermodynamic analysis and BAM visualization. It was found that all three terpenes with the exception of high betulinic acid proportion (30 and 50%) interact beneficially with POPC in two-component monolayers, while incorporation of BAc and Urs into POPE film is energetically unfavorable. As far as the model mitochondrial membrane composed of POPC/POPE/BHCL is concerned, the largest destructive influence (high positive values of ΔG(Exc) and decrease of the model monolayer condensation) was found in the case of terpene acids, while the effect of α-amyrin was energetically favorable. We postulated that the origin of the observed findings is connected with the specific interactions between bolaamphlilic terpene acids and POPE, known from its propensity to form intermolecular hydrogen bonds.