Bilayer Permeability Research Articles

Composition Dependence of Water Permeation Across Multicomponent Gel-Phase Bilayers.

The permeability of multicomponent phospholipid bilayers in the gel phase is investigated via molecular dynamics simulation. The physical role of the different molecules is probed by comparing multiple mixed-component bilayers containing distearylphosphatidylcholine (DSPC) with varying amounts of either the emollient isostearyl isostearate or long-chain alcohol (dodecanol, octadecanol, or tetracosanol) molecules. Permeability is found to depend on both the tail packing density and hydrogen bonding between lipid headgroups and water. Whereas the addition of emollient or alcohol molecules to a gel-phase DSPC bilayer can increase the tail packing density, it also disturbed the hydrogen-bonding network, which in turn can increase interfacial water dynamics. These phenomena have opposing effects on bilayer permeability, which is found to depend on the balance between enhanced tail packing and decreased hydrogen bonding.

Theoretical and Computational Investigations into Lipid Bilayer Permeation of Drugs

Transport phenomena across biomembranes are crucial processes in cellular biology, as well as in many medical, pharmaceutical, and environmental technologies. Therefore, determination of drug absorption is an important component of the drug discovery and development process in that it plays a key role in the decision to promote drug candidates to clinical trials. The path of a drug from the site of administration to its target cells or compartments implies the crossing of several semipermeable cell membranes, making it relevant to be able to predict whether and to what extent a molecule can pass through them. Lipid bilayers are an effective barrier to passive diffusion of ions and some hydrophilic small molecules, but many others can permeate bilayers through passive diffusion at rates that depend on bilayer composition and properties of the permeating solute. In this study, we consider the membrane deformation energy as the dominant factor in crossing the membrane into cells, as measured by in vitro cell-based experiments. We have investigated a new approach using the deformation free energy of a lipid bilayer based on the principle of a continuum theory. To gain atomistic insight into the passive permeability process, we have used physics-based methods, namely molecular dynamics simulations combined with the inhomogeneous solubility-diffusion model. The estimated permeabilities from our method are compared with other popular methods such as Quantitative Structure-Property Relationship (QSAR) analysis.

Rheological Droplet Interface Bilayers (rheo-DIBs): Probing the Unstirred Water Layer Effect on Membrane Permeability via Spinning Disk Induced Shear Stress

A new rheological droplet interface bilayer (rheo-DIB) device is presented as a tool to apply shear stress on biological lipid membranes. Despite their exciting potential for affecting high-throughput membrane translocation studies, permeability assays conducted using DIBs have neglected the effect of the unstirred water layer (UWL). However as demonstrated in this study, neglecting this phenomenon can cause significant underestimates in membrane permeability measurements which in turn limits their ability to predict key processes such as drug translocation rates across lipid membranes. With the use of the rheo-DIB chip, the effective bilayer permeability can be modulated by applying shear stress to the droplet interfaces, inducing flow parallel to the DIB membranes. By analysing the relation between the effective membrane permeability and the applied stress, both the intrinsic membrane permeability and UWL thickness can be determined for the first time using this model membrane approach, thereby unlocking the potential of DIBs for undertaking diffusion assays. The results are also validated with numerical simulations.

Erlang flow of hydrophilic pore formation and closure events in a lipid bilayer during phase transition resulting from diffusion in the radius space.

The Smoluchowski equation with an energy profile of a special type and an assumed hydrophobic ("half") pore source term is used to describe the process of hydrophilic pore formation in a lipid bilayer at the gel-liquid phase transition. The source term reflects the occurrence of molecule packing defects in a lipid bilayer at phase transition. The time sequences of the pore formation and closure events are treated as non-stationary, second-order Erlang flows whose characteristics depend on the equation solution. The computed distributions of the time intervals between hydrophilic pores, and pore lifetimes agree with the previously published experimental interpulse interval and pulse duration histograms for the current fluctuations through planar bilayer membranes of DPPC immersed in a LiCl aqueous solution containing polyethylene glycol. Thus, the statistical analysis of pore formation and closure times leads us to conclude that firstly, the increased permeability of a lipid bilayer during the gel-liquid phase transition is accounted for by the emergence of additional hydrophobic defects in the heterogeneous structure of the bilayer and secondly, that the non-exponential distributions of the lipid channel closed and open times observed in experiments are evidence that the process of hydrophilic pore formation is not a one-step process but involves at least two dependent events.

Comparing the action of HT61 and chlorhexidine on natural and model Staphylococcus aureus membranes.

HT61 and chlorhexidine (CHX) are both putative membrane-active antimicrobials, which non-specifically target the anionic lipids abundant in bacterial membranes. In model systems, the ability of these antimicrobials to partition into lipid monolayers and increase the permeability of lipid bilayers is dependent upon the presence and proportion of anionic lipids such as phosphatidylglycerol. Despite their apparent similarity in membrane affinity, we have found that HT61 and CHX differ in the extent to which they affect membrane integrity. HT61 was found to be capable of severely disrupting the lipid bilayer, resulting in lysis of Staphylococcus aureus membranes and the release of ATP from protoplasts. CHX, by contrast, does not disrupt the lipid bilayer to a sufficiently large degree to result in lysis of the membrane or release of ATP from S. aureus protoplasts. This suggests that although antimicrobials that interact with the membrane often have a common target, the action they have on the membrane may differ widely and may not be the primary mode of action of the antimicrobial.

Cationic amphiphiles bearing imidazole fragment: From aggregation properties to potential in biotechnologies

Fabrication of multifunctioning systems using noncovalent self-assembly principles are of great importance in biomimetic drug and gene delivery systems construction. Herein, supramolecular systems based on homologous series of amphiphiles bearing imidazolium fragment (CnH2n+1Im+Br−, where n=14, 16, 18) have been fabricated and studied. Their aggregation behavior, solubilization activity toward hydrophobic guest, interaction with DNA decamer as well as integration with lipid bilayer have been investigated. Elongation of hydrophobic fragment by two methylene groups results in 4-fold decrease of critical micelle concentration, however does not affect solubilization properties. Examination of capability of amphiphiles studied to interact with DNA decamer have revealed some unconventional features: size and charge characteristics of amphiphile/oligonucleotide complexes do not depend on the length of hydrophobic tail, but the latter affects the efficacy of component binding, which achieves 100% for all amphiphiles. Capability of amphiphiles studied to integrate with lipid bilayer strongly depends on the length of hydrophobic fragment: the lower homologue C14H29Im+Br− increases the permeability of lipid bilayer, while the higher homologues C16H33Im+Br− and C18H37Im+Br−, vice versa, stabilize it. Enhance of capability of the drug (indometacin) to penetrate through lipid bilayer has been shown in the presence of C14H29Im+Br−.

Permeability of a Fluid Lipid Bilayer to Short-Chain Alcohols from First Principles.

Computational prediction of membrane permeability to small molecules requires accurate description of both the thermodynamics and kinetics underlying translocation across the lipid bilayer. In this contribution, well-converged, microsecond-long free-energy calculations are combined with a recently developed subdiffusive kinetics framework to describe the membrane permeation of a homologous series of short-tail alcohols, from methanol to 1-butanol, with unprecedented fidelity to the underlying molecular models. While the free-energy profiles exhibit barriers for passage through the center of the bilayer in all cases, the height of these barriers decreases with the length of the aliphatic chain of the alcohol, in quantitative agreement with experimentally determined differential solvation free energies in water and oil. A unique aspect of the subdiffusive model employed herein, which was developed in a previous article, is the determination of a position-dependent fractional order which quantifies the degree to which the motion of the alcohol deviates from classical diffusion along the thickness of the membrane. In the aqueous medium far from the bilayer, this quantity approaches 1.0, the asymptotic limit for purely classical diffusion, whereas it dips below 0.75 near the center of the membrane irrespective of the permeant. Remarkably, the fractional diffusivity near the center of membrane, where its influence on the permeability is the greatest, is similar among the four permeants despite the large difference in molecular weight and lipophilicity between methanol and 1-butanol. The relative permeabilities, which are estimated from the free-energy and fractional diffusivity profiles, are therefore determined predominantly by differences in the former rather than the latter. The predicted relative permeabilities are highly correlated with existing experimental results, albeit they do not agree quantitatively with them. On the other hand, quite unexpectedly, the reported experimental values for the short-tail alcohols are nearly three orders of magnitude lower than the available experimental measurement for water. Plausible explanations for this apparent disagreement between theory and experiment are considered in detail.

Hydrophobic Nanoparticles Modify the Thermal Release Behavior of Liposomes.

Understanding the effect of embedded nanoparticles on the characteristics and behavior of lipid bilayers is critical to the development of lipid-nanoparticle assemblies (LNAs) for biomedical applications. In this work we investigate the effect of hydrophobic nanoparticle size and concentration on liposomal thermal release behavior. Decorated LNAs (D-LNAs) were formed by embedding 2 nm (GNP2) and 4 nm (GNP4) dodecanethiol-capped gold nanoparticles into DPPC liposomes at lipid to nanoparticle ratios (L:N) of 25,000:1, 10,000:1, and 5,000:1. D-LNA structure was investigated by cryogenic transmission electron microscopy, and lipid bilayer permeability and phase behavior were investigated based on the leakage of a model drug, carboxyfluorescein, and by differential scanning calorimetry, respectively. The presence of bilayer nanoparticles caused changes in the lipid bilayer release and phase behavior compared to pure lipid controls at very low nanoparticle to bilayer volume fractions (0.3%-4.6%). Arrhenius plots of the thermal leakage show that GNP2 led to greater increases in the leakage energy barrier compared to GNP4, consistent with GNP4 causing greater bilayer disruption due to their size relative to the bilayer thickness. Embedding hydrophobic nanoparticles as permeability modifiers is a unique approach to controlling liposomal leakage based on nanoparticle size and concentration.

Spectrophotometric Quantification of Peroxidase with p-Phenylene-diamine for Analyzing Peroxidase-Encapsulating Lipid Vesicles.

A spectrophotometric assay for the determination of horseradish peroxidase (HRP) in aqueous solution with p-phenylenediamine (PPD, benzene-1,4-diamine) as electron donor substrate and hydrogen peroxide (H2O2) as oxidant was developed. The oxidation of PPD by HRP/H2O2 leads to the formation of Bandrowski's base ((3E,6E)-3,6-bis[(4-aminophenyl)imino]cyclohexa-1,4-diene-1,4-diamine), which can be quantified by following the increase in absorbance at 500 nm. The assay was applied for monitoring the activity of HRP inside ≈180 nm-sized lipid vesicles (liposomes), prepared from POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) and purified by size exclusion chromatography. Because of the high POPC bilayer permeability of PPD and H2O2, the HRP-catalyzed oxidation of PPD occurs inside the vesicles once PPD and H2O2 are added to the vesicle suspension. In contrast, if instead of PPD the bilayer-impermeable substrate ABTS2- (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate)) is used, the oxidation of ABTS2- inside the vesicles does not occur. Therefore, using PPD and ABTS2- in separate assays allows distinguishing between vesicle-trapped HRP and HRP in the external bulk solution. In this way, the storage stability of HRP-containing POPC vesicles was investigated in terms of HRP leakage and activity of entrapped HRP. It was found that pH 7.0 suspensions of POPC vesicles (2.2 mM POPC) containing on average about 12 HRP molecules per vesicle are stable for at least 1 month without any significant HRP leakage, if stored at 4 °C. Such high stability is beneficial not only for bioanalytical applications but also for exploring the kinetic properties of vesicle-entrapped HRP through simple spectrophotometric absorption measurements with PPD as a sensitive and cheap substrate.

Kinetics of lipid bilayer permeation of a series of ionisable drugs and their correlation with human transporter-independent intestinal permeability

For low molecular weight drugs, lipid bilayer permeation is considered the major route for in vivo cell barrier passage. We recently introduced a fluorescence assay with liposomes to determine permeation kinetics of ionisable compounds across the lipid bilayer by monitoring drug-induced pH changes inside the liposomes. Here, we determined the permeability coefficients (PFLipP, FLipP for “Fluorescence Liposomal Permeability”) across 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayers of 35 ionisable drugs at pH6.0 and compared them to available in vivo human jejunal permeability (Peff) data. PFLipP values were furthermore compared with published Caco-2 cell permeability coefficients (PCaco-2), permeability coefficients determined with the parallel artificial membrane permeability assay (PAMPA) and with log D (pH6.0). The log PFLipP, corrected for predicted para-cellular diffusion, and log PCaco-2 correlated best with log Peff, with similar adjusted R2 (0.75 and 0.74, n=12). Our results suggest that transporter-independent intestinal drug absorption is predictable from liposomal permeability.

Engineering plant membranes using droplet interface bilayers.

Droplet interface bilayers (DIBs) have become widely recognised as a robust platform for constructing model membranes and are emerging as a key technology for the bottom-up assembly of synthetic cell-like and tissue-like structures. DIBs are formed when lipid-monolayer coated water droplets are brought together inside a well of oil, which is excluded from the interface as the DIB forms. The unique features of the system, compared to traditional approaches (e.g., supported lipid bilayers, black lipid membranes, and liposomes), is the ability to engineer multi-layered bilayer networks by connecting multiple droplets together in 3D, and the capability to impart bilayer asymmetry freely within these droplet architectures by supplying droplets with different lipids. Yet despite these achievements, one potential limitation of the technology is that DIBs formed from biologically relevant components have not been well studied. This could limit the reach of the platform to biological systems where bilayer composition and asymmetry are understood to play a key role. Herein, we address this issue by reporting the assembly of asymmetric DIBs designed to replicate the plasma membrane compositions of three different plant species; Arabidopsis thaliana, tobacco, and oats, by engineering vesicles with different amounts of plant phospholipids, sterols and cerebrosides for the first time. We show that vesicles made from our plant lipid formulations are stable and can be used to assemble asymmetric plant DIBs. We verify this using a bilayer permeation assay, from which we extract values for absolute effective bilayer permeation and bilayer stability. Our results confirm that stable DIBs can be assembled from our plant membrane mimics and could lead to new approaches for assembling model systems to study membrane translocation and to screen new agrochemicals in plants.

Role of Pore-Lining Residues in Defining the Rate of Water Conduction by Aquaporin-0

Compared to other aquaporins (AQPs), lens-specific AQP0 is a poor water channel, and its permeability was reported to be pH-dependent. To date, most water conduction studies on AQP0 were performed on protein expressed in Xenopus oocytes, and the results may therefore also reflect effects introduced by the oocytes themselves. Experiments with purified AQP0 reconstituted into liposomes are challenging because the water permeability of AQP0 is only slightly higher than that of pure lipid bilayers. By reconstituting high amounts of AQP0 and using high concentrations of cholesterol to reduce the permeability of the lipid bilayer, we improved the signal-to-noise ratio of water permeability measurements on AQP0 proteoliposomes. Our measurements show that mutation of two pore-lining tyrosine residues, Tyr-23 and Tyr-149 in sheep AQP0, to the corresponding residues in the high-permeability water channel AQP1 have additive effects and together increase the water permeability of AQP0 40-fold to a level comparable to that of AQP1. Molecular dynamics simulations qualitatively support these experimental findings and suggest that mutation of Tyr-23 changes the pore profile at the gate formed by residue Arg-187.

Voltage-Dependent Formation of Stable, Ion Conductive Pores in Suspended Lipid Bilayers from Oxidized Lipids

Oxidation of membrane lipids is a ubiquitous phenomenon in eukaryotic cells occurring as a consequence of the generation of reactive oxygen species by mitochondrial or peroxisomal activity and irradiation with photons or ions. Lipid hydroperoxidation, the primary step of the oxidative reactions, has been shown to induce molecular area increase, enhanced fluctuations and tube formation in giant unilamellar vesicles. Surprisingly, however, despite a 75% loss of elastic modulus, even fully hydroperoxidized membranes can remain effective barriers against small molecule diffusion1, contrary to membranes including other downstream oxidation products2,3,4. We were therefore interested to ascertain whether membrane permeability to ions of pure lipid bilayers is affected by lipid hydroperoxidation. Using a chip-based device for automated bilayer formation and parallel recording from suspended synthetic lipid bilayers (Orbit-16, Nanion), we compared responses to positive and negative voltages in the range of 50 to 200 mV of POPC- and 100% hydroperoxidized POPC (POPC-OOH)1 membranes formed on 50 µm-diam. microelectrode cavities of the MECA16 chip (Ionera). While control POPC membranes showed a stable conductance in the range of a few pS without evidence for pore-formation, POPC-OOH membranes responded with fluctuating conductance increases already at 50 mV. Conductance grew in a voltage and time-dependent manner, reaching values of several nS at +/-150 mV. Remarkably, the voltage induced conductance was stable over time and did not recover during a 1 s waiting period between pulses, showing cumulative behavior from one pulse to the next. Conductance fluctuations were characterized by multiple preferred levels separated by several tens of pS without evident equidistant periodicity. Taken together, these findings suggest that in response to voltage, hydroperoxidized POPC membranes form multiple, stable voltage-dependent, ion-permeant pores with conductances similar to protein ion channels.(1) Weber. et al. Lipid Oxidation Induces Structural Changes in Biomimetic Membranes. Soft Matter 2014, 10, 4241-4247.(2) Caetano et al. Photo-induced Destruction of Giant Vesicles in Methylene Blue Solutions. Langmuir, 2007, 23, 130-137.(3) Mertins et al. Physical Damage on Giant Vesicles Membrane as a Result of Methylene Blue Photoirradiation, Biophys. J. 2014, 106, 162-171.(4) Runas, K. A.; Malmstadt, N. Low Levels of Lipid Oxidation Radically Increase the Passive Permeability of Lipid Bilayers. Soft Matter 2015, 11, 499-505.

Modulation of lipid membrane structural and mechanical properties by a peptidomimetic derived from reduced amide scaffold

Understanding how antimicrobial peptidomimetics interact with lipid membranes is important in battling multidrug resistant bacterial pathogens. We study the effects of a recently reported peptidomimetic on lipid bilayer structural and mechanical properties. The compound referred to as E107-3 is synthesized based on the acylated reduced amide scaffold and has been shown to exhibit good antimicrobial potency. Our vesicle leakage assay indicates that the compound increases lipid bilayer permeability. We use micropipette aspiration to explore the kinetic response of giant unilamellar vesicles (GUVs). Exposure to the compound causes the GUV protrusion length LP to spontaneously increase and then decrease, followed by GUV rupture. Solution atomic force microscopy (AFM) is used to visualize lipid bilayer structural modulation within a nanoscopic regime. Unlike melittin, which produces pore-like structures, the peptidomimetic compound is found to induce nanoscopic heterogeneous structures. Finally, we use AFM-based force spectroscopy to study the impact of the compound on lipid bilayer mechanical properties. We find that incremental addition of the compound to planar lipid bilayers results in a moderate decrease of the bilayer puncture force FP and a 39% decrease of the bilayer area compressibility modulus KA. To explain our experimental data, we propose a membrane interaction model encompassing disruption of lipid chain packing and extraction of lipid molecules. The later action mode is supported by our observation of a double-bilayer structure in the presence of fusogenic calcium ions.

Bilayer permeability during phase transition as an Erlang flow of hydrophilic pores resulting from diffusion in the radius space

The formation of hydrophilic pores in a lipid bilayer during phase transition is described using the Smoluchowski equation with an additional term of the hydrophobic pore source. This term is added to account for defects in lipid packing during phase transition. We assume that the temporal sequence of the pores is a stochastic process, a non-stationary second-order Erlang flow. Flow characteristics depend on the equation solution and determine the formation times of the hydrophilic pores. The calculated distribution of the durations of intervals between hydrophilic pores is in a good agreement with experimental data published before. In the context of this model we describe the influence of poly(ethylene glycol) on the pore formation frequency.

Rapid Analysis of DOXIL Stability and Drug Release from DOXIL by HPLC Using a Glycidyl Methacrylate-Coated Monolithic Column.

In recent years, nanomedicines have received growing attention in a range of medical applications, including selective drug delivery technology. In this context, the analysis of liposome stability and drug release from liposomes is of particular importance, as the efficacy of a nanomedicine is determined by the release of the encapsulated drug. We investigated the influence of the surrounding environment on the stability and release of the encapsulated drug (i.e., doxorubicin) from DOXIL. Thus, for the purpose of this study, we selected the liposomal anticancer drug, DOXIL, as a typical nanomedicine, and investigated the influence of the surrounding environment on release of doxorubicin from DOXIL. We found that two pathways existed for doxorubicin release, namely the collapse of DOXIL, and an increase in the permeability of the lipid bilayer. DOXIL collapse occurred upon the addition of high concentrations (>60%) of a methanol solution, while an increase in permeability occurred at temperatures above the phase transition temperature of the DOXIL lipid bilayer, under basic conditions, and in the presence of membrane-permeable bases (e.g., Tris). As DOXIL is particularly stable and limited collapse of DOXIL occurred under physiological conditions, it is expected that doxorubicin release within the body took place through permeability changes in the lipid bilayer of the DOXIL structure.

Line Tension Assists Membrane Permeation at the Transition Temperature in Mixed-Phase Lipid Bilayers.

The umbrella sampling method has been used to evaluate the free energy profile for a large permeant moving through a lipid bilayer, represented using a coarse-grained simulation model, at and below its gel-fluid transition temperature. At the lipid transition temperature, determined to be 302 K for the MARTINI 2.0 model of DPPC, the permeation barrier for passage through an enclosed fluid domain embedded in a patch of gel was significantly lower than that for passage through a fluid stripe domain. In contrast, permeation through a fluid domain in a stripe geometry produced a free energy profile nearly identical to that of a gel-free fluid bilayer. In both cases, insertion of the permeant into a fluid domain coexisting with the gel phase led to a shift in phase composition, as lipids transitioned from fluid to gel to accommodate the area occupied by the permeant. In the case of the enclosed fluid domain, this transition produced a decrease in the length of the fluid-gel interface as the approximately circular fluid domain shrank. The observed decrease in the apparent permeation barrier, combined with an approximation for the change in interfacial length, enabled estimation of the interfacial line tension to be between 10 and 13 pN for this model. The permeation barrier was shown to drop even further in simulations performed at temperatures below the transition temperature. The results suggest a mechanism to explain the experimentally observed anomalous peak in the temperature-dependent permeability of lipid bilayers near their transition temperatures. The contribution of this mechanism toward the permeability of a gel phase containing a thermal distribution of fluid-phase domains is estimated using a simple statistical thermodynamic model.

Molecular Dynamics Simulations Predict the Pathways via Which Pristine Fullerenes Penetrate Bacterial Membranes.

Carbon fullerenes are emerging as effective devices for different biomedical applications, including the transportation of nanosized drugs and extraction of harmful oxidants and radicals. It has been proposed that fullerenes could be used as novel antibacterial agents, given the realization that the nanoparticles can kill pathogenic Gram-negative bacteria. To explore this at the molecular level, we simulated C60 fullerenes with bacterial membranes using the coarse-grain molecular dynamics Martini force field. We found that pristine C60 has a limited tendency to penetrate (incomplete core) Re mutant lipopolysaccharide (LPS) leaflets, but the translocation of C60 fullerenes into (complete core) Ra mutant LPS leaflets is not thermodynamically favored. Moreover, we showed that the permeability of the Re LPS bilayers depends sensitively on the system temperature, charge of ambient ions, and prevalence of palmitoyloleoylphosphoethanolamine (POPE) defect domains. The different permeabilities are rationalized in terms of transitory head group pore formation, which underpins the translocation of C60 into the lipid core. The Re LPS lipids readily form transient micropores when they are linked with monovalent cations or when they are heated to a high temperature. POPE lipids are shown to be particularly adept at forming these transient surface cavities, and their inclusion into Re LPS membranes facilitates the formation of particularly large pores that are tunneled by C60 aggregates of a significant size (∼5 nm wide). After insertion into the lipid core, the aggregates dissociate, and the disbanded nanoparticles migrate to the interface between separate POPE and LPS domains, where they weaken the boundaries between the coexisting lipid fractions and thereby promote lipid mixing.

A comparative study on the effect of Curcumin and Chlorin-p6 on the transport of the LDS cation across a negatively charged POPG bilayer: Effect of pH

We report the use of interface selective Second Harmonic generation technique to investigate the transport of the LDS cation across POPG liposomes in the pH range of 4.0 to 8.0 in the presence and absence of two amphiphilic drugs, Curcumin and Chlorin-p6 (Cp6). Our results show that bilayer permeability of liposomes is significantly affected by the presence of the drugs and pH of the medium as evidenced by significant changes in the transport kinetics of the LDS. Studies carried out in the pH range 4.0–8.0 show that while Cp6 significantly enhanced the transport of LDS at pH4.0, the transport of the cation was seen to increase with increasing pH, with maximum effect at pH7.4 for Curcumin. The pH dependent bilayer localization of both the drugs was investigated by conducting steady state FRET studies using DPH labeled lipids as donors. The FRET results and the relative population of the various ionic/nonionic species of the drugs at different pH suggest that distance dependent interaction between the various ionic species of the drugs and polar head groups of the lipid is responsible for the observed pH dependence enhancement of the drug induced membrane permeability. Another interesting observation was that the stability of Curcumin in presence of POPG liposomes was observed to degrade significantly near physiological pH (7.4 and 8.0). Although this degradation did not affect the liposome integrity, interestingly this was observed to enhance the transport of the LDS cation across the bilayer. That the degradation products of Curcumin are equally effective as the drug itself in enhancing the membrane permeability lends additional support to the current opinion that the bioactive degradation products of the drug may have a significant contribution to its observed pharmacological effects.

Investigating the Structure of Multicomponent Gel-Phase Lipid Bilayers

Single- and multicomponent lipid bilayers of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC), 1,2-distearoyl-sn-glycero-3-phosphatidylcholine (DSPC), isostearyl isostearate, and heptadecanoyl heptadecanoate in the gel phase are studied via molecular dynamics simulations. It is shown that the structural properties of multicomponent bilayers can deviate strongly from the structures of their single-component counterparts. Specifically, the lipid mixtures are shown to adopt a compact packing by offsetting the positioning depths at which different lipid species are located in the bilayer. This packing mechanism affects the area per lipid, the bilayer height, and the chain tilt angles and has important consequences for other bilayer properties, such as interfacial hydrogen bonding and bilayer permeability. In particular, the simulations suggest that bilayers containing isostearyl isostearate or heptadecanoyl heptadecanoate are less permeable than pure 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine or DSPC bilayers. Furthermore, hydrogen-bond analysis shows that the residence times of lipid-water hydrogen bonds depend strongly on the bilayer composition, with longer residence times for bilayers that have a higher DSPC content. The findings illustrate and explain the fundamental differences between the properties of single- and multicomponent bilayers.