Phospholipid Phase Transition Research Articles

Insights into Molecular Interactions between a GAPDH-Related Fish Antimicrobial Peptide, Analogs Thereof, and Bacterial Membranes.

Interactions between SJGAP (skipjack tuna GAPDH-related antimicrobial peptide) and four analogs thereof with model bacterial membranes were studied using Fourier-transform infrared spectroscopy (FTIR) and molecular dynamics (MD) simulations. MD trajectory analyses showed that the N-terminal segment of the peptide analogs has many contacts with the polar heads of membrane phospholipids, while the central α helix interacts strongly with the hydrophobic core of the membranes. The peptides also had a marked influence on the wave numbers associated with the phase transition of phospholipids organized as liposomes in both the interface and aliphatic chain regions of the infrared spectra, supporting the interactions observed in the MD trajectories. In addition, interesting links were found between peptide interactions with the aliphatic chains of membrane phospholipids, as determined by FTIR and from the MD trajectories, and the membrane permeabilization capacity of these peptide analogs, as previously demonstrated. To summarize, the combined experimental and computational efforts have provided insights into crucial aspects of the interactions between the investigated peptides and bacterial membranes. This work thus makes an original contribution to our understanding of the molecular interactions underlying the antimicrobial activity of these GAPDH-related antimicrobial peptides from Scombridae.

Effect of perfluorotetradecanoic acid on the morphology of a photopolymerizable phospholipid monolayer at the air‐water interface

AbstractFundamental physical chemical properties of a monolayer film comprised of the chiral, photopolymerizable phospholipid 1,2‐bis(10,12‐tricosadiynoyl)‐sn‐glycero‐3‐phosphocholine (Diyne PC) and the impact of a model perfluorocarbon, perfluorotetradecanoic acid (PF), on these film properties have been investigated, both with and without UV photopolymerization. Enantiomerically pure Diyne PC formed compact, stable monolayers at the air‐water interface, exhibited typical phospholipid phase transitions in surface pressure‐area isotherms and yielded micron‐scale, linear domains upon film compression. Photopolymerized films were significantly more expanded in comparison with the unpolymerized films and the resulting domains had a spiral morphology with a strongly preferred spiral direction. Mixing Diyne PC with PF altered the isotherm behavior, with the principal effects being a significant reduction in the plateau region associated with the characteristic LE‐LC phase transition and no larger‐scale spiral domain formation in the monolayers of the mixed films. Results are discussed in the context of interactions between the two different film components and the tendency of perfluorinated surfactants to disperse condensed phase regions of phospholipid‐based monolayer films. Overall, the tendency of PF to disperse condensed regions of the film makes patterning of films in mixed phospholipid‐perfluorocarbon monolayers particularly difficult.

Phase transitions of the pulmonary surfactant film at the perfluorocarbon-water interface

Pulmonary surfactant is a lipid-protein complex that forms a thin film at the air-water surface of the lungs. This surfactant film defines the elastic recoil and respiratory mechanics of the lungs. One generally accepted rationale of using oxygenated perfluorocarbon (PFC) as a respiratory medium in liquid ventilation is to take advantage of its low surface tensions (14–18 mN/m), which was believed to make PFC an ideal replacement of the exogenous surfactant. Compared with the extensive studies of the phospholipid phase behavior of the pulmonary surfactant film at the air-water surface, its phase behavior at the PFC-water interface is essentially unknown. Here, we reported the first detailed biophysical study of phospholipid phase transitions in two animal-derived natural pulmonary surfactant films, Infasurf and Survanta, at the PFC-water interface using constrained drop surfactometry. Constrained drop surfactometry allows in situ Langmuir-Blodgett transfer from the PFC-water interface, thus permitting direct visualization of lipid polymorphism in pulmonary surfactant films using atomic force microscopy. Our data suggested that regardless of its low surface tension, the PFC cannot be used as a replacement of pulmonary surfactant in liquid ventilation where the air-water surface of the lungs is replaced with the PFC-water interface that features an intrinsically high interfacial tension. The pulmonary surfactant film at the PFC-water interface undergoes continuous phase transitions at surface pressures less than the equilibrium spreading pressure of 50 mN/m and a monolayer-to-multilayer transition above this critical pressure. These results provided not only novel biophysical insight into the phase behavior of natural pulmonary surfactant at the oil-water interface but also translational implications into the further development of liquid ventilation and liquid breathing techniques.

New Meloxicam Derivatives-Synthesis and Interaction with Phospholipid Bilayers Measured by Differential Scanning Calorimetry and Fluorescence Spectroscopy.

The purpose of the present paper was to assess the ability of five newly designed and synthesized meloxicam analogues to interact with phospholipid bilayers. Calorimetric and fluorescence spectroscopic measurements revealed that, depending on the details of the chemical structure, the studied compounds penetrated bilayers and affected mainly their polar/apolar regions, closer to the surface of the model membrane. The influence of meloxicam analogues on the thermotropic properties of DPPC bilayers was clearly visible because these compounds reduced the temperature and cooperativity of the main phospholipid phase transition. Additionally, the studied compounds quenched the fluorescence of prodan to a higher extent than laurdan, what pointed to a more pronounced interaction with membrane segments close to its surface. We presume that a more pronounced intercalation of the studied compounds into the phospholipid bilayer may be related to the presence of the molecule of a two-carbon aliphatic linker with a carbonyl group and fluorine substituent/trifluoromethyl group (compounds PR25 and PR49) or the three-carbon linker together with the trifluoromethyl group (PR50). Moreover, computational investigations of the ADMET properties have shown that the new meloxicam analogues are characterized by beneficial expected physicochemical parameters, so we may presume that they will have a good bioavailability after an oral administration.

Isobavachalcone as an Active Membrane Perturbing Agent and Inhibitor of ABCB1 Multidrug Transporter.

Isobavachalcone (IBC) is an active substance from the medicinal plant Psoralea corylifolia. This prenylated chalcone was reported to possess antioxidative, anti-inflammatory, antibacterial, and anticancer activities. Multidrug resistance (MDR) associated with the over-expression of the transporters of vast substrate specificity such as ABCB1 (P-glycoprotein) belongs to the main causes of cancer chemotherapy failure. The cytotoxic, MDR reversing, and ABCB1-inhibiting potency of isobavachalcone was studied in two cellular models: human colorectal adenocarcinoma HT29 cell line and its resistant counterpart HT29/Dx in which doxorubicin resistance was induced by prolonged drug treatment, and the variant of MDCK cells transfected with the human gene encoding ABCB1. Because MDR modulators are frequently membrane-active substances, the interaction of isobavachalcone with model phosphatidylcholine bilayers was studied by means of differential scanning calorimetry. Molecular modeling was employed to characterize the process of membrane permeation by isobavachalcone. IBC interacted with ABCB1 transporter, being a substrate and/or competitive inhibitor of ABCB1. Moreover, IBC intercalated into model membranes, significantly affecting the parameters of their main phospholipid phase transition. It was concluded that isobavachalcone interfered both with the lipid phase of cellular membrane and with ABCB1 transporter, and for this reason, its activity in MDR cancer cells was presumptively beneficial.

The role of temperature in the formation of human-mimetic artificial cell membranes using droplet interface bilayers (DIBs).

Droplet interface bilayers (DIBs) have recently started to be used as human-mimetic artificial cell membranes. DIBs are bilayer sections created at the interface of two aqueous droplets, such that one droplet can be used as a donor compartment and the other as an acceptor compartment for the quantification of molecular transport across the artificial cell membrane. However, synthetic phospholipids are overwhelmingly used to create DIBs instead of naturally derived phospholipids, even though the diverse distribution of phospholipids in the latter is more biomimetic. We present the first systematic study of the role of temperature in DIB formation, which shows that the temperature at which DIBs are formed is a key parameter for the formation of DIBs using naturally derived phospholipids in a microfluidic platform. The phospholipids that are most abundant in mammalian cell membranes (phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), and phosphatidylinositol (PI)) only form DIBs when the temperature is above the phase transition temperature (Tm). Similarly, DIB formation usually only occurs above the highest Tm of a single phospholipid in a bespoke formulation. In addition, we show a new phenomenon wherein the DIB "melts" without disintegrating for bilayers formed predominantly of phospholipids that occupy cylindrical spaces. We also demonstrate differences in DIB formation rates as well as permeability of these biomimetic membranes. Given the difficulties associated with making DIBs using naturally derived phospholipids, we anticipate this work will illuminate the role of phospholipid phase transition in mono- and bilayer formation and lay the foundation for DIBs to be used as human-mimetic artificial cell membranes.

Insight into the Lubrication Behavior of Phospholipids Pre-adsorbed on Silica Surfaces at Different Adsorption Temperatures.

Phospholipids, as essential components in joint synovial fluid, play a dominant role in joint lubrication. In this study, atomic force microscopy was used to evaluate the normal and shear forces between two surfaces bearing three types of phospholipids with different acyl chain lengths, which were pre-adsorbed onto silica surfaces at different temperatures (25, 45, and 60 °C). When the pre-adsorption temperature was below the phospholipid phase transition temperature (Tm), a super-low friction coefficient [1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC): 0.002; 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC): 0.007] between two opposing silica surfaces in water was achieved because of the super-low shear strength of the hydration shell and robustness of the vesicle when the load was less than the critical value (DSPC: 500 nN; DPPC: 85 nN). However, when the pre-adsorption temperature exceeded Tm, the silica surface was covered by a bilayer structure with many defects, which exhibited poor adsorption density and low bearing capacity, resulting in a relatively high friction coefficient. This study gains insights into the influence of structure and temperature on the lubrication mechanism of phospholipids as biolubricants, providing guidance for the application of artificial joint synovial fluid.

Determination of n-alkane partitioning within phosphatidylethanolamine Lα/HII phases

Alkanes are known to promote the fluid lamellar (Lα)-to-inverted hexagonal (HII) phase transition of different phospholipids. In this work, we studied the interaction of decane and tetradecane with self-assemblies formed of 1-palmitoyl-2-oleoyl-sn-glycero-phosphoethanolamine (POPE), using sequential 2H and 31P solid-state NMR spectroscopy. This technique allowed calculating the partitioning constant of the alkanes between the Lα and HII phases of POPE. Our results show that both alkanes are preferentially distributed in the HII phase compared to the Lα phase. In the HII phase, both alkanes display a very high conformational disorder, consistent with their location in the intercylinder voids. This preferential partitioning in the HII phase is more pronounced for tetradecane than for decane. This finding is proposed to be associated with a less energetically favored insertion (limited solubility) of tetradecane in the lamellar phase. This proposition is supported by the observation that tetradecane experiences more conformational freedom than its shorter analog in POPE bilayers, suggesting that it is located, on average, closer to the middle of the bilayers. It is also established that the increase in size of the intercylinder voids, by the addition of 1-palmitoyl-2-oleoyl-sn-glycero-phosphocholine (POPC) in the HII cylinders, enhances the partitioning of decane in the HII phase compared to the Lα phase. These findings propose handles to modulate the balance of the relative Lα/HII phase stability.

Evidence for ATP Interaction with Phosphatidylcholine Bilayers.

ATP is a fundamental intracellular molecule and is thought to diffuse freely throughout the cytosol. Evidence obtained from nucleotide-sensing sarcolemmal ion channels and red blood cells, however, suggest that ATP is compartmentalized or buffered, especially beneath the sarcolemma, but no definitive mechanism for restricted diffusion or potential buffering system has been postulated. In this study, we provide evidence from alterations to membrane dipole potential, membrane conductance, changes in enthalpy of phospholipid phase transition, and from free energy calculations that ATP associates with phospholipid bilayers. Furthermore, all-atom molecular dynamics simulations show that ATP can form aggregates in the aqueous phase at high concentrations. ATP interaction with membranes provides a new model to understand the diffusion of ATP through the cell. Coupled with previous reports of diffusion restriction in the subsarcolemmal space, these findings support the existence of compartmentalized or buffered pools of ATP.

Melting of the Dipalmitoylphosphatidylcholine Monolayer.

Langmuir monolayer self-assembled at the air-water interface represents an excellent model for studying phase transition and lipid polymorphism in two dimensions. Compared with numerous studies of phospholipid phase transitions induced by isothermal compression, there are very scarce reports on two-dimensional phase transitions induced by isobaric heating. This is mainly due to technical difficulties of continuously regulating temperature variations while maintaining a constant surface pressure in a classical Langmuir-type film balance. Here, with technological advances in constrained drop surfactometry and closed-loop axisymmetric drop shape analysis, we studied the isobaric heating process of the dipalmitoylphosphatidylcholine (DPPC) monolayer. It is found that temperature and surface pressure are two equally important intensive properties that jointly determine the phase behavior of the phospholipid monolayer. We have determined a critical point of the DPPC monolayer at a temperature of 44 °C and a surface pressure of 57 mN/m. Beyond this critical point, no phase transition can exist in the DPPC monolayer, either by isothermal compression or by isobaric heating. The melting process of the DPPC monolayer studied here provides novel insights into the understanding of a wide range of physicochemical and biophysical phenomena, such as surface thermodynamics, critical phenomena, and biophysical study of pulmonary surfactants.

4. Effects of sugars on phospholipid phase transitions : relevance to dehydration tolerance(Seminar : Roles of Sugars in the Preservation of Biological Materials)

4. Effects of sugars on phospholipid phase transitions : relevance to dehydration tolerance(Seminar : Roles of Sugars in the Preservation of Biological Materials)

Chitosan-coated microvesicles: Effect of polysaccharide-phospholipid affinity on decafluorobutane dissolution

The stability of perfluorinated microvesicles is mainly determined by the presence of interfacial materials and their ability to hinder the gas component diffusibility into the bloodstream. The goal of this study is to increase the persistence of the gaseous-core by introducing chitosan-coated 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) microvesicles, reducing gas diffusion from microvesicles, and increasing for a long time ultrasonic signals. Our hypothesis was based on the irreversible adhesion of chitosan towards DSPC head groups observed in thin-films models. This affinity enhanced the stabilization of gaseous-core microvesicles, in which the polysaccharide effectively reduced the phospholipid phase transition enthalpy from 383±5.5Jmg−1 for plain to 150±9.7Jmg−1 for chitosan-coated microvesicles, providing a more stable structure that diminished the gaseous component lost and provided the persistence of intense 19F-NMR signals after 48h, twice as long compared to plain samples. As a result, stronger and long-lasting ultrasonic signals were produced by the more stable chitosan-containing microvesicles, thus, presenting great potential to increase the diagnostic and therapeutic applications of perfluorocarbon carries.

Calorimetry-Derived Composition Vectors to Resolve Component Raman Spectra in Phospholipid Phase Transitions.

Multidimensional least squares analysis is a well-established technique for resolving component vibrational spectra from mixed samples or systems. Component resolution of temperature-dependent vibrational spectra is challenging, however, due to the lack of a suitable model for the variation in sample composition with temperature. In this work, analysis of temperature-dependent Raman spectra of lipid membranes is accomplished by using "concentration" vectors independently derived from enthalpy changes determined by differential scanning calorimetry. Specifically, the lipid-bilayer phase transitions of DMPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) are investigated through Raman spectra acquired from individual, optically trapped vesicles in suspension as a function of temperature. Heat capacity profiles of the same vesicle suspension are measured using differential scanning calorimetry and numerically integrated to generate enthalpy change curves of each phase transition, which are in turn used to construct composition vectors. Multidimensional least squares analysis optimized for a fit to these composition vectors allows resolution of the component spectra corresponding to gel, ripple, and liquid-crystalline phases of the DMPC. The quality of fit of the calorimetry-derived results is confirmed by unstructured residual differences between the data and the model, and a composition variation predicted by the resolved spectra that matches the calorimetry results. This approach to analysis of temperature-dependent spectral data could be readily applied in other areas of materials characterization, where one is seeking to learn about structural changes that occur through temperature-dependent phase transitions.

Insights into composition/structure/function relationships of Doxil® gained from “high-sensitivity” differential scanning calorimetry

Thermotropic behavior of Doxil® and its generic, Lipodox®, was characterized using “high-sensitivity” differential scanning calorimetry (DSC). This is the first report that two distinct endotherms were observed in Doxil and Lipodox upon heating. The first (Tm at 51±2°C) is broad and of low enthalpy, representing the membrane lipid phase transition, which occurs despite having high (38mole%) cholesterol. The second (Tm at ∼70°C) is narrow, representing melting of the intraliposomal doxorubicin-sulfate nanocrystals. The thermograms of Doxil and Lipodox are practically identical. The membrane phase transition is similar to that of drug-free nanoliposomes of the same size and lipid composition as Doxil, suggesting lack of significant drug-membrane interaction. The melting endotherm of the intraliposomal nanocrystals is 2.0–2.5-fold narrower than that of the crystals formed in a solution of 250mM ammonium sulfate and >60mg/ml doxorubicin. This suggests that nanovolume of liposomes improves doxorubicin-sulfate crystallinity. Moreover, both phase transitions are reversible in cycled DSC scanning (15–90–15°C). This indicates an unexpected “non-leaky” phospholipid phase transition and excellent physical and chemical stabilities of Doxil after short exposure to high temperature. Reducing mole% of cholesterol results in a “leaky” membrane phase transition of higher enthalpy. Namely, high mole% cholesterol is essential for the resistance to drug leakage during phase transition. Pegylated liposomal doxorubicin in which HSPC was replaced by DPPC shows the same non-leaky phase transition but at a lower temperature, indicating this type of phase transition is not unique to Doxil. The presence of DSPE-PEG2k increases the cooperativity of the phase transition. High-sensitivity DSC helps illuminate composition/structure/function relationships of Doxil, and is useful for the equivalence/similarity studies.

Influence of different surfactants on the physicochemical properties of elastic liposomes

Elastic liposomes are capable to improve drug transport through the skin by acting as penetration enhancers due to the high fluidity and elasticity of the liposome membranes. Therefore, elastic liposomes were prepared and characterized to facilitate the transdermal transport of bioactive molecules. Liposomes consisted of dimyristoylphosphatidylcholine (DMPC) as the structural component, with different surfactants derived from lauric acid as elastic components: C12E5 (polyoxyethylene-5-lauryl ether), PEG4L (polyethyleneglycol-4-lauryl ester), PEG4DL (polyethylene glycol-4-dilauryl ester), PEG8L (polyethylene glycol-8-lauryl ester) and PEG8DL (polyethylene glycol-8-dilauryl ester). The elastic liposomes were characterized in terms of their phospholipid content, mean diameter, size distribution, elasticity and stability during storage, as well as their ability to incorporate surfactant and permeate through 50 nm pore size membranes. The results showed that the phospholipid phase transition temperature, the fluidity of the lipid bilayer resulting from incorporation of the surfactant and the preservation of particle integrity were factors determining the performance of the elastic liposomes in permeating through nanoporous membranes. The best results were obtained using DMPC combined with the surfactants PEG8L or PEG8DL. The findings demonstrate the potential of using elastic liposomes for transdermal administration of drugs.

Anisotropic Self-Assembly of Citrate-Coated Gold Nanoparticles on Fluidic Liposomes.

The behavior of self-assembly processes of nanoscale particles on plasma membranes can reveal mechanisms of important biofunctions and/or intractable diseases. Self-assembly of citrate-coated gold nanoparticles (cAuNPs) on liposomes was investigated. The adsorbed cAuNPs were initially fixed on the liposome surfaces and did not self-assemble below the phospholipid phase transition temperature (Tm ). In contrast, anisotropic cAuNP self-assembly was observed upon heating of the composite above the Tm, where the phospholipids became fluid. The number of self-assembled NPs is conveniently controlled by the initial mixing ratio of cAuNPs and liposomes. Gold nanoparticle protecting agents strongly affected the self-assembly process on the fluidic membrane.

Preliminary investigation into the effects of two dietary fatty acids, 20:5n-3 and 22:6n-3, on mortality of juvenile Mercenaria mercenaria during the approach to winter

Changes in the molar percentage of two omega-3 fatty acids, 20:5n-3 (EPA) and 22:6n-3 (DHA), occur in juvenile northern quahogs, Mercenaria mercenaria, as descending autumn temperatures approach winter lows in the temperate zone. Incorporation of these highly unsaturated fatty acids compensates for the effect of low temperature on cell membrane viscosity by lowering the phase transition of membrane phospholipids. Prior studies have demonstrated that the ability of bivalve mollusks, like M. mercenaria, to manufacture these compounds de novo does not exist or may be limited at best. Insufficient exogenous supply of EPA and DHA, therefore, may restrict their ability to overcome the inhibiting effect of high viscosity on cell membrane phospholipids during this interval. In the present study, a matrix of cultured algae containing various quantities of these compounds was administered to four groups of juvenile M. mercenaria as they adapted to low temperature. Each group displayed a unique response to diet in relative fatty acid profile and mortality. The group receiving abundant EPA adapted well during the early-phase temperature decline, and the one receiving abundant DHA adapted well to the late-phase temperature decline. The group receiving abundant DHA also enjoyed best overall survival following a severe 5-day drop in temperature. The strain native to the local region suffered between 31 and 117 % higher mortality than the notata strain during the 72-day period of temperature decline from 18 to 6 °C.

Spontaneous Curvature Measurements for Zwitterionic and Anionic Phospholipids under Biomimetic Perturbations and their Implications to the Mechanisms of Lamellar-Nonlamellar Phase Transitions

It is believed that the structural rearrangements of lipid bilayers into curved structures are highly related to the fusion and fission of biomembranes. Local fluctuations in lipid composition and/or environmental ion concentration are considered to play important roles in this regard, through their influences on the local membrane curvatures. To understand the underlying mechanisms, we measure the monolayer spontaneous curvatures (C0) of three phospholipids, including one zwitterionic phospholipid and two anionic phospholipids, under the perturbations induced by changes in phospholipid composition and calcium ion concentration ([Ca2+]). With the X-ray diffraction techniques and the reconstruction of electron density profiles, the C0 of the phospholipids and their mixtures were determined when the samples were affected by the biomimetic perturbations and at different temperatures. It is not surprising to see that while the C0 of the zwitterionic phospholipid demonstrates nearly negligible dependence on [Ca2+], those of the anionic phospholipids react strongly to the changes in [Ca2+], with the strength of the response correlated with the charge density. Interestingly, the situation is completely reversed when the temperature perturbation is concerned: Whereas the C0 of the zwitterionic phospholipid increases with temperature, as seen for most phospholipids, the modulation in temperature causes visually no effect on the C0 of the anionic phospholipids. The discrepancies in the reactions to the two perturbations are expected to involve the differential binding affinities of Ca2+ and the contributions of the electrostatic interactions. Based on these observations and energetic considerations, we explore the likely mechanisms underlying the calcium-induced phase transitions of zwitterionic phospholipids and the thermal induced phase transitions of anionic phospholipids from the lamellar phase to the inverted nonlamellar structures. The biological implications of these proposed mechanisms are discussed.

Hydrophobic nanoparticles promote lamellar to inverted hexagonal transition in phospholipid mesophases.

This study focuses on how the mesophase transition behaviour of the phospholipid dioleoyl phosphatidylethanolamine (DOPE) is altered by the presence of 10 nm hydrophobic and 14 nm hydrophilic silica nanoparticles (NPs) at different concentrations. The lamellar to inverted hexagonal phase transition (Lα-HII) of phospholipids is energetically analogous to the membrane fusion process, therefore understanding the Lα-HII transition with nanoparticulate additives is relevant to how membrane fusion may be affected by these additives, in this case the silica NPs. The overriding observation is that the HII/Lα boundaries in the DOPE p-T phase diagram were shifted by the presence of NPs: the hydrophobic NPs enlarged the HII phase region and thus encouraged the inverted hexagonal (HII) phase to occur at lower temperatures, whilst hydrophilic NPs appeared to stabilise the Lα phase region. This effect was also NP-concentration dependent, with a more pronounced effect for higher concentration of the hydrophobic NPs, but the trend was less clear cut for the hydrophilic NPs. There was no evidence that the NPs were intercalated into the mesophases, and as such it was likely that they might have undergone microphase separation and resided at the mesophase domain boundaries. Whilst the loci and exact roles of the NPs invite further investigation, we tentatively discuss these results in terms of both the surface chemistry of the NPs and the effect of their curvature on the elastic bending energy considerations during the mesophase transition.

From molecular modelling to photophysics of neutral oligo- and polyfluorenes incorporated into phospholipid bilayers.

The combination of various experimental techniques with theoretical simulations has allowed elucidation of the mode of incorporation of fluorene based derivatives into phospholipid bilayers. Molecular dynamics (MD) simulations on a fully hydrated 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) bilayer, with benzene (B), biphenyl (BP), fluorene (F) and tri-(9,9-di-n-octylfluorenyl-2,7-diyl), TF, have provided insights into the topography of these molecules when they are present in the phospholipid bilayer, and suggest marked differences between the behavior of the small molecules and the oligomer. Further information on the interaction of neutral fluorenes within the phospholipid bilayer was obtained by an infrared (IR) spectroscopic study of films of DMPC and of the phospholipid with PFO deuterated specifically on its alkyl chains (DMPC-PFO-d34). This was complemented by measurements of the effect of F, TF and two neutral polymers: polyfluorene poly(9,9-di-n-octylfluorenyl-2,7-diyl), PFO, and poly(9,9-di-n-dodecylfluorenyl-2,7-diyl), PFD, on the phospholipid phase transition temperature using differential scanning calorimetry (DSC). Changes in liposome size upon addition of F and PFO were followed by dynamic light scattering. In addition, the spectroscopic properties of F, TF, PFO and PFD solubilised in DMPC liposomes (absorption, steady-state and time-resolved fluorescence) were compared with those of the same probes in typical organic solvents (chloroform, cyclohexane and ethanol). Combining the insight from MD simulations with the results at the molecular level from the various experimental techniques suggests that while the small molecules have a tendency to be located in the phospholipid head group region, the polymers are incorporated within the lipid bilayers, with the backbone predominantly orthogonal to the phospholipid alkyl chains and with interdigitation of them and the PFO alkyl chains.