Lipid Fatty Acid Chain Research Articles

Взаимодействие катионных антисептиков с кардиолипинсодержащей модельной бактериальной мембраной

Plasma membrane is one of the major targets for cationic antiseptics (CA). The study was aimed to assess molecular effects of CAs of different chemical classes on cardiolipin-containing regions of bacterial plasma membranes. The study was carried out using coarse-grained molecular modeling. Interaction of CAs, such as miramistin, chlorhexidine, picloxidine, and octenidine, with cardiolipin-containing bilayer was assessed based on the CA coarse-grained models. CAs reduced lipid lateral diffusion coefficients and increased the membrane area per lipid. All CAs, except miramistin, reduced the lipid fatty acid chain order parameters. Adding octenidine at a CA : lipid ratio of 1 : 4 resulted in cardiolipin clustering with subsequent pulling the neutral phosphatidylethanolamine molecules out of the model bilayer. It was found that CАs have the potential for sorption to lipid bilayer, causing clustering of negatively charged lipids. Antiseptic octenidine causes formation of cardiolipin microdomains. Abnormal lateral lipid distribution together with pulling out phosphatidylethanolamine molecules can result in increased lipid bilayer permeability. The most significant reduction of cardiolipin lateral diffusion coefficient by 2.8 ± 0.4 times was observed in the presence of CA chlorhexidine at an antiseptic : lipid ratio of 1 : 4.

Optimization of nanostructured lipid carriers loaded with retinoids by central composite design

The aim of this study was to design and optimize a new formulation of lipid nanoparticles for the topical administration of retinoids. The standard procedure involves nanostructured lipid carriers (NLCs) synthesized by the miniemulsion methodology, using sunflower oil as a bioactive ingredient and retinyl palmitate (RP) as model retinoid.A 5-factor, 3-level central composite design was used to predict responses and construct 3D-response contour plots. The independent variables were the concentration of total lipids, solid lipid, surfactant, encapsulated retinoid and number of carbons on the solid lipid fatty acid chain length, in function of the selected responses as the particle size, surface charge of the nanoparticles and the encapsulation efficiency (EE %) at 3 levels. Experimental trials were performed at all 31 possible combinations.An optimized NLC composition of 2.5% total lipids, 2.0% myristic acid and 0.5% sunflower oil and 1.5% of surfactant Tween 80 was reached. The type of surfactant was adjusted from Tween 80 to Span 80 to improve the electrostatic stability of the lipid nanoparticles and the resultant optimized NLC formulation was evaluated to incorporate another two retinoids as tretinoin (TRT) and adapalene (ADP).The resultant retinoid delivery systems were characterized in terms of particle size and electrostatic stability, morphology, crystallinity of the lipid matrix, EE (%) and drug loading capacity and release profile. Particle sizes of 134.5 ± 5.4 nm and zeta potential values of −57.0 ± 2.8 mV were obtained for optimized ADP-NLCs and high entrapment efficiency values were obtained for RP-NLC, TRT-NLC and ADP-NLC as 84.4 ± 3.0%, 84.1 ± 7.8% and 73.7 ± 3.3%, respectively. DSC analysis indicated that the melting temperatures of the three optimized NLCs loaded with the retinoids were above 40 °C, proving to be suitable for use in topical administration. The in vitro drug release studies were conducted for 48 h, using a dialysis regenerated cellulose membrane. The results demonstrated a very well-controlled release of RP and TRT, which improves the stability of these active compounds and confirms a high encapsulation efficiency.

Structure of the monomeric outer-membrane porin OmpG in the open and closed conformation

OmpG, a monomeric pore-forming protein from Escherichia coli outer membranes, was refolded from inclusion bodies and crystallized in two different conformations. The OmpG channel is a 14-stranded beta-barrel, with short periplasmic turns and seven extracellular loops. Crystals grown at neutral pH show the channel in the open state at 2.3 A resolution. In the 2.7 A structure of crystals grown at pH 5.6, the pore is blocked by loop 6, which folds across the channel. The rearrangement of loop 6 appears to be triggered by a pair of histidine residues, which repel one another at acidic pH, resulting in the breakage of neighbouring H-bonds and a lengthening of loop 6 from 10 to 17 residues. A total of 151 ordered LDAO detergent molecules were found in the 2.3 A structure, mostly on the hydrophobic outer surface of OmpG, mimicking the outer membrane lipid bilayer, with three LDAO molecules in the open pore. In the 2.7 A structure, OmpG binds one OG and one glucose molecule as sugar substrates in the closed pore.

Investigation on the flow behavior of dispersions of solid triglyceride nanoparticles

The flow behavior of low concentrated dispersions of solid lipid nanoparticles consisting of either trimyristin, tripalmitin or tristearin and different ionic and nonionic stabilizer blends were investigated using a rheometer with cone and plate apparatus and an Ubbelohde type capillary viscometer. The data demonstrate a remarkable influence of the matrix material, the stabilizer composition and the presence of small amounts of sodium chloride on the formulations’ rheological properties. A significant increase in dispersion viscosity was found in the triglyceride sequence trimyristin < tripalmitin < tristearin. This effect can be clearly attributed to an increase in particle shape anisometry with increasing length of the lipid's fatty acid chains. Surfactants, which are present during the crystallization of the dispersed lipid seem to have an additional effect on the nanoparticle shape. Beyond this, ionic surfactants and other salts affect the dispersion viscosity by altering the dimension of the electrical double layer (“second electroviscous effect”).

Phase transition of a single lipid bilayer measured by sum-frequency vibrational spectroscopy.

In this communication, we demonstrate the first use of sum-frequency generation (SFG) vibrational spectroscopy to measure directly the phase transition temperature (Tm) of a single planar supported lipid bilayer (PSLB). Three saturated phospholipids, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-diheptadecanoyl-sn-glycero-3-phosphocholine (DHPC), and 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), were studied. Lipid bilayer films were prepared by the the Langmuir-Blodgett method at a surface pressure of 30 nN/m. The symmetric nature of the bilayer was used to determine the Tm of bilayers by measuring the intensity of the symmetric methyl stretch at 2875 cm-1 from the lipid fatty acid chains as a function of temperature. A maximum in the CH3 symmetric stretch transition was observed at the Tm of the lipid film due to the reduction of symmetry in the bilayer. The SFG measured Tm for DPPC, DHPC, and DSPC were 41.0 +/- 0.4, 52.4 +/- 0.7, and 57.9 +/- 0.5 degrees C, respectively. These values correlate well with the literature values of 41.3 +/- 1.8, 49 +/- 3, and 54.5 +/- 1.5 degrees C for DPPC, DHPC, and DSPC, respectively obtained by differential scanning calorimetry (DSC) of lipid vesicles in solution. The high degree of correlation between the SFG spectroscopic measurements and the DSC results suggests the Tm of these lipids is not significantly altered upon immobilization on a surface.

Location of ubiquinone homologues in liposome membranes studied by fluorescence anisotropy of diphenyl-hexatriene and trimethylammonium-diphenyl-hexatriene

The measurements of diphenyl-hexatriene (DPH) and trimethylammonium-diphenyl-hexatriene (TMA-DPH) fluorescence anisotropy in dipalmitoylphosphatidylcholine (DPPC) and egg yolk lecithin (EYL) liposomes containing different concentrations of various ubiquinone (UQ) homologues have been performed. UQ-4 induced the highest DPH anisotropy increase in DPPC liposomes, whereas for higher UQ homologues the anisotropy was lowered with the increase of UQ side-chain length. These differences were less pronounced in EYL liposomes. It was concluded that at a higher content in the membranes (3–4 mol%), the short-chain ubiquinones are arranged parallel to lipid fatty acid chains, whereas long-chain homologues are progressively removed from the lipid acyl chains into the midplane region of the membrane. At the lower (1–2 mol%) concentrations, long-chain quinones seem to be evenly distributed within the membrane, especially in EYL membranes. UQ-10 in EYL liposomes perturbed TMA-DPH to a similar extend as the short-chain ubiquinones indicating that UQ-10 penetrates the interface regions of the membrane where its redox reactions occur. The localization and physical state of UQ-10 in native membranes is discussed.

Localization of adriamycin in model and natural membranes. Influence of lipid molecular packing.

The interaction of adriamycin with lipids was studied in model (monolayers, small unilamellar vesicles, large multilamellar vesicles) and natural (chinese hamster ovary cell) membranes by measurement of fluorescence energy transfer and fluorescence quenching. 2-APam, 7-ASte, 12-ASte and anthracene-phosphatidylcholine were used as fluorescent probes in which the anthracene group is well located at graded depths in the membrane. Egg-yolk phosphatidylcholine and a 1/1 mixture of it with bovine brain phosphatidylserine were used in model membrane systems. Large fluorescence energy transfer was observed between these molecules as donors and the drug as acceptor. With liposomes, at pH 7.4 and over an adriamycin concentration range of 0-100 microM, the efficiency of energy transfer was 12-ASte greater than 7-ASte greater than 2-APam, with 100% energy transfer for 12-ASte above a drug concentration of 30 microM. At pH 5, where the fatty acids are buried deeper (0.45 nm) in the lipid bilayer due to protonation of the carboxyl group, the order of energy transfer 7-ASTe greater than 12-ASte = 2-APam was observed. Measurements of fluorescence quenching using the non-permeant Cu2+ ion as quencher and spectrophotometric assays indicated that around 40% of the adriamycin molecules were deeply embedded in the lipid bilayer. Adriamycin molecules thus appear to penetrate the lipid bilayer, with the aminoglycosyl group interacting with the lipid phosphate groups and the dihydroanthraquinone residue in contact with the lipid fatty acid chains. In contrast, fluorescence energy transfer and quenching studies on CHO cells showed that adriamycin penetrated the plasma membrane of these cells to a much more limited extent than in the model membrane systems. This can be related to the squeezing out of the drug from a film of phosphatidylcholine which was observed in monolayers by means of surface pressure, potential and fluorescence experiments. These observations indicated that the penetration of adriamycin into lipid bilayers strongly depends on the molecular packing of the lipid.

Dependence of boundary lipid on fatty acid chain length in phosphatidylcholine vesicles containing a hydrophobic protein from myelin proteolipid

Lipophilin, a hydrophobic protein fraction, purified and delipidated from the proteolipid of human myelin, possesses a layer of boundary lipid surrounding it when incorporated into lipid vesicles. The protein reduces the energy absorbed during the lipid phase transition, indicating that the boundary lipid does not go through the phase transition. The amount of boundary lipid was estimated by plotting the enthalpy of the transition against the protein to lipid mole ratio and extrapolating to deltaH = 0 for a number of synthetic phosphatidylcholines, to determine the ability of fatty acid chains of varying length to interact with the protein. The amount of boundary lipid was found to be similar, 21-25 molecules per molecule of lipophilin, for fatty acid chains of length 14-18 carbons but somewhat less, 16 molecules of lipid per molecule of protein, for a fatty acid chain length of 12 or for one with a trans double bond (18:1tr). No preferential interaction was observed with a lipid containing a particular fatty acid chain length when the protein was incorporated into a mixture of these lipids. These results suggest that the binding of lipids to the boundary layer of other membrane proteins and enzymes may not depend significantly on lipid fatty acid chain length.

Effect of basic protein from human central nervous system myelin on lipid bilayer structure

The effect of myelin basic protein from normal human central nervous system on lipid organization has been investigated by studying model membranes containing the protein by differential scanning calorimetry or electron spin resonance spectroscopy. Basic protein was found to decrease the phase transition temperature of dipalmitoyl phosphatidylglycerol, phosphatidic acid, and phosphatidylserine. The protein had a greater effect on the freezing temperature, measured from the cooling scan, than on the melting temperature, measured from the heating scan. These results are consistent with partial penetration of parts of the protein into the hydrocarbon region of the bilayer in the liquid crystalline state and partial freezing out when the lipid has been cooled below its phase transition temperature. The effect of the protein on fatty acid chain packing was investigated by using a series of fatty acid spin labels with the nitroxide group located at different positions along the chain. If the protein has not yet penetrated, it increases the order throughout the bilayer in the gel phase, probably by decreasing the repulsion between the lipid polar head groups. Above the phase transition temperature, when parts of it are able to pentrate, it decreases the motion of the lipid fatty acid chains greatly near the polar head group region, but has little or no effect near the interior of the bilayer. Upon cooling again the protein still decreases the motion near the polar head group region but increases it greatly in the interior. Thus, the protein penetrates partway into the bilayer, distorts the packing of the lipid fatty acid chains, and prevents recrystallization, thus decreasing the phase transition temperature. The magnitude of the effect varied with the lipid and was greatest for phosphatidic acid and phosphatidylglycerol. It could be reversed upon cooling for phosphatidylglycerol but not phosphatidic acid. The protein was only observed to decrease the phase transition temperature of phosphatidylserine upon cooling. It had only a small effect on phosphatidylethanolamine and no effect on phosphatidylcholine. Thus, the protein may penetrate to a different extent into different lipids even if it binds to the polar head group region by electrostatic interactions.

Proton nuclear magnetic resonance studies of sarcoplasmic reticulum membranes. Correlation of the temperature-dependent Ca 2+ efflux with a reversible structural transition

Proton nuclear magnetic resonance spectra of ATP-dependent Ca 2+-accumulating sarcoplasmic reticulum vesicles isolated from rabbit leg white muscle show a reversible, temperature dependent structural transition. This transition involves a transformation of the choline methyl groups of lecithin from a rigid to a mobile state. The energy of the transformation is approx. 35 kcal/mole and involves nearly 60% of the lecithin molecules in the membrane. The fraction of mobile methyl groups as a function of temperature correlates with the temperature-dependent Ca 2+ efflux measured in these preparations. As determined by nuclear magnetic resonance, other agents which alter protein structure and simultaneously increase Ca 2+ efflux also increases the fraction of mobile choline methyl groups. Likewsise, irreversible heat denaturation of the membrane protein alters the temperature dependence of the transition. The proton magnetic resonance spectra are consistent with the organization of approx. 80% of the lipid fatty acid chains into a lamellar phase which permits only limited motion. On the other hand approx. 20% of the lipid molecules are in a fluid region of the membrane in which nearly isotropic motion occurs ( τ c ≈ 10 −10 sec ).