HII Phase Research Articles

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.

An Investigation of Whether Vitamin E Preferentially Interacts with Polyunsaturated Lipids

Vitamin E (α-tocopherol) is a lipid-soluble antioxidant that has the role of protecting phospholipids from oxidation in membranes. A question that remains is how the low concentration of α-tocopherol found in whole cells can protect the relatively large concentration of polyunsaturated phospholipids found in membranes that are particularly vulnerable to oxidative attack. We hypothesize that α-tocopherol co-localizes with polyunsaturated phospholipids to optimize its role as an antioxidant. This project attempts to test this hypothesis by comparing the effect of α-tocopherol on the molecular organization of 1-palmitoyl-2-docosahexaenoyl-sn-glycerophosphatidylethanolamine (16:0-22:6PE, PDPE) and, as a monounsaturated control, 1-palmitoyl-2-oleoyl-sn-glycerophosphatidylethanolamine (16:0-18:1PE, POPE) in mixtures with sphingomyelin (SM). By solid-state 2H NMR spectroscopy, we directly observe order and phase behavior of POPE-d31 and PDPE-d31 (analogs of POPE and PDPE with a perdeuterated sn-1 chain) in the mixed membranes. In complementary X-ray diffraction and differential scanning calorimetry experiments we further probe phase behavior. The spectra observed for POPE-d31 in POPE/SM/α-tocopherol (2:2:1 mol) reveal that a transition from gel to liquid crystalline phase is no longer apparent. At higher temperatures there is a superposition of two spectral components that we ascribe to α-tocopherol promoting a transition from lamellar to inverted hexagonal (HII) phase. Analysis of depaked spectra shows that order is increased by about 8 % and that the amount of HII phase increases with temperature, ranging from 7 (31°C) to 41 % (65°C). In mixed membranes where POPE-d31 is replaced by PDPE-d31, we shall investigate whether there is a greater tendency for α-tocopherol to increase order and destabilize bilayer structure for the polyunsaturated phospholipid.

Initial step of pH-jump-induced lamellar to bicontinuous cubic phase transition in dioleoylphosphatidylserine/monoolein.

Electrostatic interactions (EI) are an important factor for phase transitions between lamellar liquid-crystalline (L(α)) and inverse bicontinuous cubic (Q(II)) phases. We investigated the low pH-induced L(α) to double-diamond cubic (Q(II)(D)) phase transition in dioleoylphosphatidylserine (DOPS)/monoolein (MO) using time-resolved small-angle X-ray scattering. Using a stopped-flow apparatus, a suspension of liposomes (multilamellar vesicles (MLVs) or large unilamellar vesicles (LUVs)) of 20%-DOPS/80%-MO membrane at neutral pH was rapidly mixed with a low pH buffer, and then the structural change of the membranes in the resultant suspension was observed as a function of time (i.e., pH-jump experiment). At the initial step, the L(α) phase was directly transformed into the hexagonal II (H(II)) phase, and subsequently, the H(II) phase slowly converted into the Q(II)(D) phase. We obtained the rate constants of the initial step (i.e., the L(α) to H(II) phase transition) and of the second step (i.e., the H(II) to Q(II)(D) phase transition) using the non-negative matrix factorization method. The rate constant of the initial step was independent of the MLV concentration, indicating that single MLVs can convert into the HII phase without any interaction with other MLVs. On the other hand, the rate constant of the initial step increased with a decrease in pH, 0.041 s(-1) at pH 2.6 and 0.013 s(-1) at pH 2.8, and also exhibited a size dependence; for smaller vesicles such as LUVs and smaller MLVs with diameters of ~1 μm, the rate constant was smaller. They were reasonably explained by the classical nucleation theory. These results provide the first experimental evidence of the total kinetics of EI-induced L(α)/Q(II) phase transitions.

Phase behavior of the DOPE + DOPC + alkanol system.

Small- and wide-angle X-ray diffraction was used to study the effect of 1-alkanols, as simple models of general anesthetics, (abbreviation CnOH, n = 8-18 is the even number of carbons in the aliphatic chain) on the lamellar to hexagonal Lα→ H(II) phase transition in the dioleoylphosphatidylethanolamine-dioleoylphosphatidylcholine = 3 : 1 mol/mol (DOPE + DOPC) system. All studied CnOHs were found to decrease the phase transition temperature of the DOPE + DOPC system in a CnOH chain length and concentration dependent manner and thus promote the formation of the HII phase. Anesthetically active C8OH and C10OH were found to decrease the lattice parameter d of the Lα phase, however longer non-anesthetic CnOHs increased the parameter d; this effect being more pronounced with increasing CnOH concentration. The lattice parameter of the HII phase was decreased in the presence of all CnOHs, even at the lowest concentrations studied. In the scope of the indirect mechanism of general anesthesia observed changes in the lattice parameter d (reflecting changes in the bilayer thickness) due to the intercalation of C8OH and C10OH might induce changes in the activity of integral membrane proteins engaged in neuronal pathways.

Formation of Inverse Topology Lyotropic Phases in Dioleoylphosphatidylcholine/Oleic Acid and Dioleoylphosphatidylethanolamine/Oleic Acid Binary Mixtures

The addition of saturated fatty acids (FA) to phosphatidylcholine lipids (PC) that have saturated acyl chains has been shown to promote the formation of lyotropic liquid-crystalline phases with negative mean curvature. PC/FA mixtures may exhibit inverse bicontinuous cubic phases (Im3m, Pn3m) or inverse topology hexagonal phases (HII), depending on the length of the acyl chains/fatty acid. Here we report a detailed study of the phase behavior of binary mixtures of dioleoylphosphatidylcholine (DOPC)/oleic acid (OA) and dioleoylphosphatidylethanolamine (DOPE)/oleic acid at limiting hydration, constructed using small-angle X-ray diffraction (SAXD) data. The phase diagrams of both systems show a succession of phases with increasing negative mean curvature with increasing OA content. At high OA concentrations, we have observed the occurrence of an inverse micellar Fd3m phase in both systems. Hitherto, this phase had not been reported for phosphatidylethanolamine/fatty acid mixtures, and as such it highlights an additional route through which fatty acids may increase the propensity of bilayer lipid membranes to curve. We also propose a method that uses the temperature dependence of the lattice parameters of the HII phases to estimate the spontaneous radii of curvature (R0) of the binary mixtures and of the component lipids. Using this method, we calculated the R0 values of the complexes comprising one phospholipid molecule and two fatty acid molecules, which have been postulated to drive the formation of inverse phases in PL/FA mixtures. These are -1.8 nm (±0.4 nm) for DOPC(OA)2 and -1.1 nm (±0.1 nm) for DOPE(OA)2. R0 values estimated in this way allow the quantification of the contribution that different lipid species make to membrane curvature elastic properties and hence of their effect on the function of membrane-bound proteins.

Optimizing Drug Release: Bilayer to Inverted Hexagonal Phase Transition of Cationic XTC2 and Anionic DSPS Lipid System is Influenced by pH, Temperature, and Salt Concentration

The effectiveness of a macromolecular drug delivered in lipid nanoparticles (LNP) depends upon the biophysical properties of the delivery vehicle. Recent research has shown that the design of the cationic lipid component of LNPs improves the intracellular delivery of therapeutic siRNA[1]. Even in these optimized LNPs, only 1% of the siRNA taken up by the cell via endocytosis is actually released into the cell cytosol. We proposed a mechanism of endosome disruption that relies on the formation of non-bilayer phases in the presence of anionic endosomal lipid and synthetic cationic lipids. A model system using prototypical anionic lipid 1,2-distearoyl(d70)-sn-glycero-3-[phospho-L-serine] (DSPS-d70) in 1:1 molar ratio to the cationic lipid DLin-KC2-DMA (XTC2) (pKa∼6.7) was characterized by 2H and 31P NMR spectroscopy. Through spectral analysis, we determined that at physiological pH (∼7.4) the XTC2/DSPS system exhibits a stable gel phase for temperatures below 45oC while an isotropic signal emerges at higher temperatures - no inverted hexagonal (HII) phase is observed. At low pH (∼4.75), the XTC2/DSPS system is principally in a bilayer gel phase at low temperatures with a non-bilayer HII phase predominating at higher temperatures. The transition from gel to HII phase is dependent on salt concentration and is most evident in the range of 15-25oC for 0.25M [Na+], 20-30oC for 0.5M [Na+] and 35-45oC for 1M [Na+]. Through depaking the spectra, order parameter profiles SCD have been obtained and compared for DSPS-d70 chains in bilayer and HII phases. These will be useful for computational simulation and eventually to design in vivo animal model experiments.[1] Semple, S.C., et al., Rational design of cationic lipids for siRNA delivery. Nat Biotechnol, 2010. 28(2): p. 172-6.

Molecular dynamics simulations of the lyotropic reverse hexagonal (HII) of Guerbet branched-chain β-d-glucoside

Through atomistic molecular dynamic simulations using a GROMOS53a6 force field for the carbohydrate, we studied the lyotropic reverse hexagonal phase HII from a glycolipid, namely the Guerbet branched-chain β-d-glucoside, at 14% and 22% water concentrations. Our simulations showed that at low water concentration (14%) the sugar head group overlapped extensively and protruded into the water channel. In contrast, in the 22% concentration system a water column free from the sugar headgroup ('free' water) was formed as expected for the system close to the limit of maximum hydration. In both concentrations, we found anomalous water diffusion in the xy-plane, i.e. the two-dimensional space confined by the surface of the cylinder. On the other hand, along the z-axis, the water diffusion obeyed the Einstein relation for the 22% system, while for the 14% system it was slightly anomalous. For the 22% system, the diffusion along the z-axis of the 'free' water obeyed the Einstein relation, while that of the 'bound' water is slightly anomalous. The xy-plane displacement of the 'bound' water was higher than that for the 'free' water at times longer than 200 ps, as a consequence of the exchange of water molecules between the two regions. Based on our findings, we proposed an alternative explanation to the observed spatial heterogeneity in the HII phase from probe diffusion by Penaloza et al. (Phys. Chem. Chem. Phys., 2012, 14(15), 5247-5250). We found the extent of contact with water was different at different oxygen atoms within the sugar ring. Generally, a higher probability of hydrogen bonding but a shorter lifetime was found in 22% water compared to the case of 14% water. Finally, we examined the extension and compression of the alkyl chain of a columnar.

Modelling of the Interaction between Cationic Lipid Dlin-Kc2-Dma (XTC2) and Anionic Lipid Distearoylphosphatidylserine (DSPS)

Short pieces of double stranded small interfering RNA (siRNA) could be used as a potential drug to cure cancer by binding to cancer gene's messenger RNA. However, the siRNA is not stable in the blood stream, and tends not to penetrate target cell membrane. So, it must be encapsulated in a lipid nanoparticle. LNPs have been shown to be strong candidates for drug delivery [1]. Lipid nanoparticles (LNPs) are a new tool for drug delivery systems. They are responsible for transferring short pieces of double stranded small interfering RNA (siRNA) into the cell. However, the underlying mechanism of action of lipid constituents of LNPs is not clearly known. Specifically, characteristics such as encapsulation efficiency, and the stability of LNPs are not predictable. Cationic lipids play essential roles in encapsulating negatively charged siRNA and also destabilizing the endosomal membrane. We have studied the bilayer and inverted hexagonal phase (HII) phase stability as it could potentially show the destabilizing effect of LNPs on the endosomal membrane for DLin-KC2-DMA (XTC2) at different temperatures. It has shown good destabilizing effects for bilayer structures. We also studied the interaction of the distearoylphosphatidylserine (DSPS) with XTC2 based on comparison with deuterium order parameters obtained from NMR experiments since DSPS has been used to optimize the structure of XTC2 [1]. pH, water content, temperature, and salt concentration effects on the bilayer HII phase stability is also studied.[1] S. de Youg, G. C. Chikh, L. L. Sekirov, S. Raney, S. C. Semple, S. K. Klimuk, Z. N. Yuan, M. Hope, P. R. Cullis, Y. K. Tam, Cancer Immunology and Immunotherapy, 56, 1251-1264, 2007.

A bacterial monorhamnolipid alters the biophysical properties of phosphatidylethanolamine model membranes

This work presents a biophysical study on the interactions of a monorhamnolipid (monoRL) produced by Pseudomonas aeruginosa MA01 with model dielaidoylphosphatidylethanolamine (DEPE) membranes. Incorporation of monoRL into DEPE shifts the onset temperature of the Lβ-to-Lα and the Lα-to-HII phase transitions toward lower values. Incorporation of monoRL into DEPE indicates the coexistence of lamellar and hexagonal–HII phases in rhamnolipid-containing samples at 60°C, at which pure DEPE is lamellar. Thus, both techniques show that monoRL facilitates formation of the hexagonal–HII phase in DEPE, i.e. it destabilizes the bilayer organization. The phase diagram for the phospholipid component indicates a near-ideal behavior, with better miscibility of monoRL into DEPE in the fluid phase than in the gel phase. The various vibrational mode bands of the acyl chains of DEPE were studied by FTIR spectroscopy, focusing on the CH2 symmetric stretching mode. Incorporation of monoRL into DEPE shifts the frequency of this band to higher wavenumbers, at temperatures both below and above the main gel to liquid–crystalline phase transition. Examination of the CO stretching band of DEPE indicates that monoRL/DEPE interactions result in an overall dehydration effect on the polar headgroup of DEPE. These results are discussed in light of the possible role of rhamnolipids as bilayer stabilizers/destabilizers during cell membrane fluctuation events.

Mapping Ion-Induced Mesophasic Transformation in Lyotropic In Situ Gelling System and its Correlation with Pharmaceutical Performance

To investigate influence of ion induced mesophasic transformation on pharmaceutical performance of in situ gelling system consisting of glyceryl monooleate. The prepared system showed mesophasic transformation during its conversion from sol to gel upon controlled hydration. The process of mesophasic transformation was studied by SAXS, DSC, rheology and plane polarized light microscopy. Further the influence of additives i.e. naproxen salts (sodium and potassium) and naproxen (base) on the process of mesophasic transformation was also elucidated. It was observed that addition of salt form of naproxen transformed W/O emulsions into cubic mesophase whereas addition of base form of naproxen formed reverse hexagonal (HII) phase upon controlled hydration. The cubic mesophase formed by naproxen salts retarded the drug release for initial 3h whereas HII phase showed sustained drug release characteristics for naproxen base following Higuchi drug release kinetics. The current work suggests that formulations with tailor made pharmaceutical performance can be developed by selecting proper additives in the system so as to obtain the desired mesophase 'on demand' thereby controlling drug release characteristics.

Fusion Peptides Promote Formation of Bilayer Cubic Phases in Lipid Dispersions. An X-Ray Diffraction Study

Small angle x-ray diffraction revealed a strong influence of the N-terminal influenza hemagglutinin fusion peptide on the formation of nonlamellar lipid phases. Comparative measurements were made on a series of three peptides, a 20-residue wild-type X-31 influenza virus fusion peptide, GLFGAIAGFIENGWEGMIDG, and its two point-mutant, fusion-incompetent peptides G1E and G13L, in mixtures with hydrated phospholipids, either dipalmitoleoylphosphatidylethanolamine (DPoPE), or monomethylated dioleoyl phosphatidylethanolamine (DOPE-Me), at lipid/peptide molar ratios of 200:1 and 50:1. All three peptides suppressed the HII phase and shifted the Lα–HII transition to higher temperatures, simultaneously promoting formation of inverted bicontinuous cubic phases, QII, which becomes inserted between the Lα and HII phases on the temperature scale. Peptide-induced QII had strongly reduced lattice constants in comparison to the QII phases that form in pure lipids. QII formation was favored at the expense of both Lα and HII phases. The wild-type fusion peptide, WT-20, was distinguished from G1E and G13L by the markedly greater magnitude of its effect. WT-20 disordered the Lα phase and completely abolished the HII phase in DOPE-Me/WT-20 50:1 dispersions, converted the QII phase type from Im3m to Pn3m and reduced the unit cell size from ∼38 nm for the Im3m phase of DOPE-Me dispersions to ∼15 nm for the Pn3m phase in DOPE-Me/WT-20 peptide mixtures. The strong reduction of the cubic phase lattice parameter suggests that the fusion-promoting WT-20 peptide may function by favoring bilayer states of more negative Gaussian curvature and promoting fusion along pathways involving Pn3m phase-like fusion pore intermediates rather than pathways involving HII phase-like intermediates.

Self-Assembly and UV-Curing Property of Polymerized Lyotropic Liquid Crystal Monomer of Sodium 3,4,5-tris(11-acryloxyundecyloxy)benzoate

A polymerized lyotropic liquid crystal monomer of sodium 3,4,5-tris(11-acryloxyundecyloxy)-benzoate was synthesized by a convenient route starting from 3,4,5-trihydroxybenzoic acid via esterification followed by etherification, acylation and finally neutralization. The chemical structure was confirmed by Fourier transform infrared (FT-IR) and H1 nuclear magnetic resonance spectral analysis. The self-organization behavior of the monomer with deionized water in methanol at room temperature was also demonstrated. The assemblies were characterized by polarized optical microscope and X-ray diffraction. The results show that a solution containing 80:20 of the monomer to water was found to be able to self-organize into Lamellar (La) phase and 92:8 with inverted hexagonal (HII) phase, which was in accordance with the theoretical calculation of critical packing parameter. It suggests that the concentration of the monomer was the key factor to influence assembly structure. Additionally, the acrylate conversion with different photoinitiators and nanostructure retention after polymerization were investigated. The research shows that the acrylate conversion of the monomer with Darocur2959 could reach up to 78% when irradiated by 30 mW/cm2 UV light of 365 nm for 30 min characterized by Real-time FT-IR as well as the sol-gel method. Meanwhile, the La and HII phase nanostructures were both retained after polymerization.

Lipophilicity of flavonoid complexes with iron(II) and their interaction with liposomes

We studied complex formation of flavonoids quercetin and taxifolin with iron(II) and the complex influence on phase transitions of phospholipid bilayer. UV–Vis spectroscopy revealed that the stoichiometry of flavonoid/iron complexes was equal to 3:2 and 2:1. Molecular modeling and experimental measurements demonstrated the increase of flavonoids lipophilicity after the complex formation. A considerable influence of quercetin–iron complex on Palmitoyl–Oleoyl–Phosphatidylethanolamine transitions from bilayer to hexagonal HII phase was detected by differential scanning calorimetry. The obtained data are related to flavonoid/iron complexes bioavailability, their influence on cell membrane functioning, and should be considered in designing liposomal vehicles for drug and gene delivery.

Anionic Phospholipids change the Effect of the Hydrophobic Surfactant Proteins on Structures of Hexagonal Lipids

Available evidence suggests that the hydrophobic surfactant proteins (SPs), SP-B and SP-C, accelerate adsorption of surfactant vesicles to an air/water interface by promoting formation of a negatively curved rate-limiting structure. In support of this model, the proteins induce several phosphatidylethanolamines to form inverse bicontinuous cubic phases, in which each leaflet has negative saddle-like curvature analogous to the hypothetical intermediate. The proteins could promote formation of cubic phases by changing spontaneous curvature (c0) of lipids, which would be reflected in the dimensions of the inverse hexagonal (HII) phase. With 1,2-dioleoyl phosphatidylethanolamine (DOPE), the SPs had no effect on the size of the HII phase, suggesting a constant c0. The study, however, lacked anionic phospholipids, which constitute ∼10% (mol:mol) of phospholipids in lung surfactant, and could engage in selective interactions with the cationic SPs. In this work, we used small-angle X-ray scattering to examine how SPs affect structures formed by DOPE mixed with 10% (mol:mol) anionic 1,2-dioleoyl phosphatidylglycerol (DOPG). With DOPG, the HII lattice-constant (a0) decreased in a dose-dependent manner with increasing levels of protein. This change could be caused by specific interactions between the SPs and DOPG, or nonspecific interactions between cationic SPs and the anionic phosphate group of DOPG. To determine whether the observed change in a0 was caused by nonspecific electrostatic effects, the measurements were repeated on samples prepared in buffered electrolyte. In the presence of counterions, the effect of SPs on a0 was significantly diminished. These results suggest that the change in the a0 for DOPE:DOPG in the absence of counterions is caused by nonspecific electrostatic interactions between the positively-charged proteins and anionic phospholipids, and are unlikely to play a major role in physiological media.Acknowledgements: Stanford Synchrotron Radiation Lightsource; NIH.

Formation of Ag Nanostrings Induced by Lyotropic Liquid–Crystalline Phospholipid Multilayer

Morphological variation of the Ag nanoparticles embedded in a lyotropic phospholipid (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, DOPE) membrane during hydration was investigated. Hydration at 5 °C resulted in transformation of the Ag nanoparticles into a bundle of Ag nanostrings as the Ag nanoparticles conformed to the H(II) phase of the DOPE molecules. Above 30 °C, the nanoparticles quickly coarsened into large polygonal-shaped particles since high mobility of the lipid molecules overwhelmed the tendency for the Ag nanoparticles to order. The result provided an insight into the long-term stability of nanoparticles trapped in different lipid membranes depending on the structural ordering of the molecules.

Thermotropic and lyotropic liquid crystalline phases of Guerbet branched-chain -D-glucosides

The effect of chain branching on glycolipid thermotropic and lyotropic phases was investigated for a series of synthetic β-D-glucosides derived from Guerbet alcohols, whose total hydrocarbon chain length ranged from C8 to C24. The compounds, which can be viewed as isosteric mimics for glycoglycerolipids, were synthesised in high purity and their liquid crystalline phases were studied using optical polarising microscopy (OPM), and small-angle X-ray diffraction. When dry, the shortest compound (total C8) exhibits a monotropic Lα phase while longer ones (C16 and C20) adopt inverse hexagonal HII phases. The C24 compound forms an ordered lamellar phase at room temperature, but exhibits a metastable HII phase upon cooling. Curiously the intermediate chain length homologue (C12) adopts an isotropic inverse micellar (L2) phase in the dry state over the range of temperatures studied. Upon hydration, the C8 compound dissolves, and the C12 compound forms a fluid lamellar Lα phase. The C16 Guerbet glucoside (i.e. β-Glc-C10C6) exhibits an inverse bicontinuous cubic phase of space group Ia3d in excess water, never previously observed in branched-chain lipids, and very seldom observed in excess water. The C20 compound remains in the HII phase upon hydrating, with the lattice parameter swelling substantially.

PH-Responsive Lyotropic Liquid Crystals for Controlled Drug Delivery

We present a food-grade lyotropic liquid crystal system, capable of responding to pH variations with a reversible switch in both the structure and physical properties. The system, which is composed by monolinolein and linoleic acid (97:3 wt % ratio) in the presence of excess water at 37 °C and 150 mM ionic strength, is specifically designed to reversibly change from a Im3m reverse bicontinuous cubic phase to a H(II) reverse columnar hexagonal phase, when changing the pH from neutral (pH 7) to acidic (pH 2) conditions, to simulate intestine and stomach conditions, respectively. The pH responsiveness is provided by the linoleic acid, which, being a weak acid (pK(a) ≈ 5), is essentially in the deprotonated charged state at pH 7 and mainly protonated and neutral at pH 2, imposing changes in the critical packing parameter (CPP) of the lyotropic liquid crystal. The use of this system as an efficient controlled-release delivery vehicle is demonstrated on the model hydrophilic drug phloroglucinol, by both release and diffusion studies at different pH, as followed by ultraviolet-visible (UV-vis) spectroscopy. The Im3m cubic phase at pH 7 is shown to release 4 times faster than the H(II) phase at pH 2, making this system an ideal candidate for oral administration of drugs for targeted delivery in intestine or colon tracts.

The Pivotal Plane of Phosphatidylethanolamine is Unaffected by the Hydrophobic Surfactant Proteins

Available evidence suggests that the hydrophobic surfactant proteins (SP), SP-B and SP-C, promote adsorption of the surfactant lipids to the alveolar air/water interface by facilitating formation of a rate-limiting negatively-curved structure. In support of this model, the proteins induce several phosphatidylethanolamines to form inverse bicontinuous cubic phases, in which each leaflet has the same saddle-shaped curvature as the hypothetical intermediate. Proteins could promote formation of the cubic phases by altering the spontaneous curvature (c0) of the phospholipids, which would change the dimensions of the inverse hexagonal (HII) phase. With 1,2-dioleoyl phosphatidylethanolamine (DOPE) in excess water, the SP had no effect on the HII phase, suggesting a constant c0. The SP, however, might still change c0 by shifting the location of the pivotal plane at which c0 is calculated. To determine if the proteins shift the pivotal plane, we measured X-ray diffraction from DOPE with 0 - 1% SP at 22 oC with different hydrations. Behavior at the different SP concentrations was indistinguishable. With increasing hydration the hexagonal lattice expanded until reaching the same limiting size at the same limiting hydration, indicating that the location of the Luzatti plane, which defines the boundary between the aqueous core and lipid, was unaffected. Detailed structural analysis of the lattice-dimensions at lower hydrations confirmed the existence of a pivotal plane at all levels of protein, and the derived location of the pivotal plane relative to the Luzatti plane (Vp/Vl) showed no response to protein concentration. Since neither Vp/Vl, nor the location of the Luzatti plane change with SP, the location of the pivotal plane remains constant. Unaffected position of the pivotal plane confirms that the SP induce cubic phases without changing the spontaneous curvature. (Studies conducted at the Stanford Synchrotron Radiation Lightsource).

Hydrostatic pressure effects on a hydrated lipid inverse micellar Fd3m cubic phase

Over a range of hydration, unsaturated diacylglycerol/phosphatidylcholine mixtures adopt an inverse micellar cubic phase, of crystallographic space group Fd3m. In this study hydrated DOPC:DOG mixtures with a molar ratio close to 1 : 2 were examined as a function of hydrostatic pressure, using synchrotron X-ray diffraction. The small-angle diffraction pattern at atmospheric pressure was used to calculate 2-D sections through the electron density map. Pressure initially has very little effect on the structure of the Fd3m cubic phase, in contrast to its effect on hydrated inverse bicontinuous cubic phases. At close to 2 kbar, a sharp transition occurs from the Fd3m phase to a pair of coexisting phases, an inverse hexagonal H(II) phase plus an (ordered) lamellar phase. Upon increasing the pressure to 3 kbar, a further sharp transition occurs from the H(II) phase to a (fluid) lamellar phase, in coexistence with the ordered lamellar phase. These transitions are fully reversible, but show hysteresis. Remarkably, the lattice parameter of the Fd3m phase is practically independent of pressure. These results show that these two lipids are miscible at low pressure, adopting a single lyotropic phase (Fd3m); they then become immiscible above a critical pressure, phase separating into DOPC-rich and DOG-rich phases.

Partitioning of ssRNA Molecules between Preformed Monolithic HII Liquid Crystalline Phases of Lipids and Supernatant Isotropic Phases

The interaction of nucleic acids with the nanoarchitectures formed by lipidic systems is a new area of research that may offer insights into the functioning of genetic materials in vivo. Here we report that ssRNA has a strong preference to reside in isotropic solution rather than in inverse hexagonal (HII) liquid crystalline phases. This is in contrast to dsDNA, which becomes localized in the pores of the HII phase. The RNA that does associate with the external surfaces of the HII phase appears to form an accretion layer, tens of molecules thick, but this layer still allows the transcription of dsDNA that resides within the pores of the phase.