Polar Head Group Of Phosphatidylcholine Research Articles

Effect of Oleic Acid, Cholesterol, and Octadecylamine on Membrane Stability of Freeze-Dried Liposomes Encapsulating Natural Antimicrobials

Liposomes have been broadly studied as delivery systems for bioactive compounds, although its relatively low stability remains a limitation for commercial application. In this study, phosphatidylcholine (PC) liposomes were prepared entrapping a mixture of garlic extract (GE) and nisin (Nis) using cholesterol (CHO), oleic acid (OA), or octadecylamine (ODA) as membrane stabilizers to evaluate their physical, chemical, bioactive, and stability properties, in fully hydrated state and after freeze-drying. GE/Nis-loaded liposomes presented hydrodynamic diameter below 200 nm and polydispersity index below 0.30, typical for small unilamellar vesicles produced by thin film method. Under induced oxidation, the PC-OA-GE/Nis liposomes presented 91% less lipid peroxidation compared with the unloaded PC liposomes. The Fourier transform infrared spectroscopy (FTIR) analysis revealed a high level of hydrogen bonds in the polar head group of PC after addition of GE/Nis in all liposome formulations, in agreement to the high values of water activity and hygroscopicity found in the samples after freeze-drying. During 5 months storage at 4 °C, fully hydrated and lyophilized liposomes showed an increment in their average size and polydispersity index, but these values were reduced by the trehalose addition as lyoprotector. All liposome preparations maintained 100% activity against Listeria monocytogenes; nevertheless, a gradual reduction of activity against Salmonella enterica serovar Enteritidis was observed, suggesting a partial loss of GE active compounds. Despite some physical modifications, freeze-dried liposomes containing OA as stabilizer showed best antimicrobial properties and high lipid oxidation resistance, constituting a promising approach to stabilize GE/Nis for long-term storage.

Direct Enzymatic Synthesis of 1‐Phosphatidyl‐β‐D‐glucose by Engineered Phospholipase D

AbstractA simple, direct method for the synthesis of 1‐phosphatidyl‐β‐D‐glucose (1‐PGlc) was established, being the first example of direct enzymatic synthesis of this recently discovered, highly important bioactive phospholipid. The method employs phospholipase D (PLD, E.C. 3.1.4.4)‐catalyzed transphosphatidylation, in which the polar head group of phosphatidylcholine is exchanged to glucose. Although wild‐type PLD can catalyze the transfer reaction, it provides only 6‐phosphatidyl‐glucose, a positional isomer of 1‐PGlc, due to its strong preference towards the primary hydroxyl group. To synthesize 1‐PGlc, engineered PLD variants, previously isolated as the ones active on secondary hydroxyls of inositol, were screened for the ability to catalyze the reaction. One of the variants, 187K/191W/385Y (KWY), was identified as the best‐performing in 1‐PGlc synthesis, however, in addition to 1‐PGlc the reaction with KWY also afforded undesired positional isomers as byproducts. To facilitate the isolation of 1‐PGlc, the isomers were converted into the corresponding amines by reductive amination. Following the column chromatography, the desired product was isolated with the overall yield of 12.5 %. The structure of the product was confirmed by 1H‐NMR analysis.

How Lipid Headgroups Sense the Membrane Environment: An Application of 14N NMR

The orientation of lipid headgroups may serve as a powerful sensor of electrostatic interactions in membranes. As shown previously by 2H NMR measurements, the headgroup of phosphatidylcholine (PC) behaves like an electrometer and varies its orientation according to the membrane surface charge. Here, we explored the use of solid-state 14N NMR as a relatively simple and label-free method to study the orientation of the PC headgroup in model membrane systems of varying composition. We found that 14N NMR is sufficiently sensitive to detect small changes in headgroup orientation upon introduction of positively and negatively charged lipids and we developed an approach to directly convert the 14N quadrupolar splittings into an average orientation of the PC polar headgroup. Our results show that inclusion of cholesterol or mixing of lipids with different length acyl chains does not significantly affect the orientation of the PC headgroup. In contrast, measurements with cationic (KALP), neutral (Ac-KALP), and pH-sensitive (HALP) transmembrane peptides show very systematic changes in headgroup orientation, depending on the amount of charge in the peptide side chains and on their precise localization at the interface, as modulated by varying the extent of hydrophobic peptide/lipid mismatch. Finally, our measurements suggest an unexpectedly strong preferential enrichment of the anionic lipid phosphatidylglycerol around the cationic KALP peptide in ternary mixtures with PC. We believe that these results are important for understanding protein/lipid interactions and that they may help parametrization of membrane properties in computational studies.

Palmitoleate Is a Mitogen, Formed upon Stimulation with Growth Factors, and Converted to Palmitoleoyl-phosphatidylinositol

Controversial correlations between biological activity and concentration of the novel lipokine palmitoleate (9Z-hexadecenoate, 16:1) might depend on the formation of an active 16:1 metabolite. For its identification, we analyzed the glycerophospholipid composition of mouse Swiss 3T3 fibroblasts in response to 16:1 using LC-MS/MS. 16:1 was either supplemented to the cell culture medium or endogenously formed when cells were stimulated with insulin or growth factors as suggested by the enhanced mRNA expression of 16:1-biosynthetic enzymes. The proportion of 1-acyl-2-16:1-sn-phosphatidylinositol (16:1-PI) was time-dependently and specifically increased relative to other glycerophospholipids under both conditions and correlated with the proliferation of fatty acid (16:1, palmitate, oleate, or arachidonate)-supplemented cells. Accordingly, cell proliferation was impaired by blocking 16:1 biosynthesis using the selective stearoyl-CoA desaturase-1 inhibitor CAY10566 and restored by supplementation of 16:1. The accumulation of 16:1-PI occurred throughout cellular compartments and within diverse mouse cell lines (Swiss 3T3, NIH-3T3, and 3T3-L1 cells). To elucidate further whether 16:1-PI is formed through the de novo or remodeling pathway of PI biosynthesis, phosphatidate levels and lyso-PI-acyltransferase activities were analyzed as respective markers. The proportion of 16:1-phosphatidate was significantly increased by insulin and growth factors, whereas lyso-PI-acyltransferases showed negligible activity for 16:1-coenzyme A. The relevance of the de novo pathway for 16:1-PI biosynthesis is supported further by the comparable incorporation rate of deuterium-labeled 16:1 and tritium-labeled inositol into PI for growth factor-stimulated cells. In conclusion, we identified 16:1 or 16:1-PI as mitogen whose biosynthesis is induced by growth factors.

Synthesis and hemocompatibity evaluation of segmented polyurethane end-capped with both a fluorine tail and phosphatidylcholine polar headgroups

To improve the hemocompatibility of polyurethanes, an amine monomer containing a long fluorine tail and phosphatidylcholine polar headgroups, 2-amino-3-oxo-3-(2-(2,2,3,3,4,4,5,5,6,6,7,7,8,8,8-pentadecafluorooctan amido) ethyl amino) propyl phosphorylcholine (FASPC) was firstly synthesized and characterized. Then four kinds of fluorinated phosphatidylcholine end-capped polyurethanes with different chemical structures were prepared. The surface properties of these prepared polyurethanes were characterized using X-ray photoelectron spectroscopic analysis (XPS) and water contact angle measurements. The results indicated that the phosphatidylcholine (PC) polar headgroups along with the fluorine tail could be easily enriched on the top surfaces, and the PC groups could be highly oriented on the outmost surface when the polymer film was in contact with water for only 30 s at room temperature. The evaluation of hemocompatibity was carried out via fibrinogen adsorption and platelet adhesion. Fibrinogen adsorption (37°C for 90 min) decreased by 98% to 87% compared to that on ordinary polyurethane surfaces, and almost no platelet adhesion and activation was observed at 37°C for 2 h.

CRP discriminates primary from secondary necrosis

Background and objectivesC-reactive protein (CRP) is an acute-phase protein whose serum concentration increases massively in response to tissue injury or inflammation. Beside membranes of bacteria CRP is known to bind...

Chemical Imaging of Cardiac Cell and Tissue by Using Secondary Ion Mass Spectrometry

Identification and localization of biomolecules in cells and tissue samples are important for understanding of subcellular structures and can be helpful in biomedical and pharmaceutical research. Isolated cardiac cells and tissue of rats are studied by using time-of-flight secondary ion mass spectrometry. This technique provides chemical composition of cardiac cell membrane and tissue surface in native form. The result is a spatially resolved chemical imaging of cell and tissue surfaces as a lateral distribution of biologically relevant molecules-phospholipids, along with fatty acids, and cholesterol. Phospholipids are represented by phosphatidylcholine and cardiolipin molecules and their fragments. Phosphatidylcholine polar head group at mass of 184.1 u has an origin in the cell membrane, and a two-dimensional distribution of this fragment provides clear chemical contours of the cell. The high-resolution contrast of the cell is observed within its environment represented with Na(+) ions. Images of PO(4)H(-) fragment and fatty acids with 16 or 18 C atoms are determined in cardiac tissue. Distributions of these 16 and 18 C fatty acids are the same within their groups, and interestingly, these two distribution groups are spatially complementary. Contours of phosphatidylcholine and cardiolipin fragments are also complementary, the distributions of 16 C fatty acids and phosphatidylcholine are identical, and the distributions of 18 C fatty acids and cardiolipin are also the same. This complementarity thus supports the chemical compositions of phosphatidylcholine and cardiolipin based on 16 C and 18 C fatty acids, respectively. The method provides information not only about cell and tissue morphology, shape, and condition but also about cellular membrane chemical composition and lateral distribution of biologically relevant molecules.

Synthesis and Hemocompatibility of Biomembrane Mimicing Poly(carbonate urethane)s Containing Fluorinated Alkyl Phosphatidylcholine Side Groups

In this article, we designed and synthesized biomembrane mimicing segmented poly(carbonate urethane)s containing fluorinated alkyl phosphatidylcholine (PC) side groups. To obtain these novel poly(carbonate urethane)s, a new diol with a long side chain fluorinated alkyl phosphatidylcholine polar headgroup (2-[2-2,2,3,3,4,4,5,5,6,6,7,7,8,8,9,9-hexadecafluoro-10-ethoxy-decyloxy-N-(2-hydroxy-1-hydroxymethyl-1-methyl-ethyl)-acetamide] phosphatidylcholine, HFDAPC) was first synthesized and characterized. Then a series of poly(carbonate urethane)s containing fluorinated alkyl phosphatidylcholine side groups were synthesized using methylenebis(phenylene isocyanate) (MDI), poly(1,6-hexyl-1,5-pentyl carbonate) diol (PHPCD), 1,4-butandiol (BDO), and HFDAPC. The obtained fluorinated phosphatidylcholine poly(carbonate urethane)s (FPCPCU) possessed high molecular weight, narrower molecular weight distribution, and good mechanical properties as characterized by GPC and Instron, showing an increased hydrophilicity and a possible arrangement of surface structure as characterized by water contact angle. XPS results indicated that the phosphatidylcholine polar headgroups have been indeed pulled out to the surface with the help of the migration of the fluorinated side chain that was directly connected with the phosphatidylcholine polar headgroup. A preliminary result by protein adsorption and platelet adhesion experiments suggested that only 5 approximately 12.5 mol % phosphatidylcholine could be enough for good hemocompatibility. The current work demonstrates a new synthetic approach that can be used to bring the bioactive PC groups to the surface of the PC-containing polyurethanes more effectively.

Novel Biomembrane‐Mimicking Polymer Surface with Environmental Responsiveness

AbstractSummary: In this article, we designed and synthesized novel segmented poly(carbonate urethane)s containing both hydrophobic fluorinated alkyl group and hydrophilic phosphatidylcholine polar head groups on the side chain. The contact angle measurement, XPS, together with ATR‐IR investigation indicated a reversible overturn of the phosphatidylcholine groups with the movement of the hydrophobic fluorinated alkyl groups when the samples were treated in dry air or water. The change in environment from air to water induced a reorganization of the surface in order to minimize the interfacial free energy, resulting in a macroscopic change of surface wettability. The good environmental responsiveness of such biomembrane‐mimicking films may find successful applications as biomaterials.Environmentally responsive surface using FPCPCU50 as an example; FPCPCU50 coated on aligned carbon nanotube film and dried in vacuum at 50 °C for 7 h and sample c treated in hot water at 80 °C for 1 h.imageEnvironmentally responsive surface using FPCPCU50 as an example; FPCPCU50 coated on aligned carbon nanotube film and dried in vacuum at 50 °C for 7 h and sample c treated in hot water at 80 °C for 1 h.

Phase behavior and hydrogen bonding in biomembrane mimicing polyurethanes with long side chain fluorinated alkyl phosphatidylcholine polar head groups attached to hard block

To achieve a good biocompatibility, two sets of novel segmented polyurethanes, namely, poly(ether urethane)s and poly(carbonate urethane)s, with long side chain fluorinated alkyl phosphatidylcholine polar head groups attached to hard block have been synthesized recently in our laboratory by using a new diol with a long side chain fluorinated alkyl phosphatidylcholine polar head group 2-[2-[2,2,3,3,4,4,5,5,6,6,7,7,8,8,9,9-hexadecafluoro-10-ethoxy-decyloxy]- N-(2-hydroxy-1-hydroxymethyl-1-methyl-ethyl)-acetamide] phosphatidylcholine, HFDAPC as an extender. These novel polyurethanes have shown a potential to be used as bio-membrane mimicry. In this article we investigated the phase behavior of these materials by Instron, DSC, DMA, and AFM because the phase behavior has a great effect on the surface properties thus the biological-related perspective. T g decreases first then increases for the poly(carbonate urethane)s, but increases first then decreases for the poly(ether urethane)s with an increasing in fluorinate phosphatidylcholine content. On the other hand, the tensile modulus was found decrease for the poly(carbonate urethane)s but increase for the poly(ether urethane)s with an increasing in fluorinate phosphatidylcholine content. It was found via AFM that the phase separation increases in poly(ether urethane)s but phase mixing increases in poly(carbonate urethane)s, with increasing content of fluorinated phosphatidylcholine side chain. The interaction between hard and soft segment, particularly, the hydrogen bonding was investigated by FTIR. The effect of fluorinated phosphatidylcholine side group on the phase separation of polyurethane was discussed and compared with that of fluorinated polyurethanes containing only fluorinated side chains. Our result demonstrated how the phase behavior of polyurethanes could be controlled by tailoring the interaction between hard and soft segment.

Interactions of Phosphatidylcholine Surface Monolayers with Triglyceride Cores and Enhanced ApoA-1 Binding in Lipid Emulsions

The binding maximum of apoA-1 (N) in triolein (TO)−egg yolk phosphatidylcholine (PC) emulsions was 10-fold larger than that in PC large unilamellar vesicles (LUV) of similar size (100 nm) with no significant difference in the affinity. Replacement of the long-chain triglyceride, TO, by medium-chain triglycerides or cholesteryl oleate in emulsion cores significantly decreased the N value. The 13C NMR chemical shifts of the PC carbonyl carbon at the surface layers indicated that PC polar headgroups are more separated and exposed to water molecules in emulsions than in vesicles. The N values were satisfactorily correlated with the chemical shift, that is, the degree of separation between the carbonyl groups at the surface. Although apoA-1 binding to the PC monolayers of emulsions brings about bending of the surface layers and creates local defects in the hydrocarbon regions in a similar manner as PC LUV, the surface−core interaction seems to fill the defects with the core neutral lipids, compensates for the ...

Surface Modification of Segmented Polyurethanes by Grafting Methacrylates and Phosphatidylcholine Polar Headgroups To Improve Hemocompatibility

A new type of phosphatidylcholine-containing segmented polyurethane (SPU) surface was produced by grafting various methacrylates and phosphatidylcholine polar headgroups to a vinyl-group-containing segmented polyurethane (V-SPU) using α,α‘-azobisisobutyronitrile (AIBN) as a radical initiator. 1,4-Butanediol (BD) as chain extender was used to synthesize the V-SPU, which is based on diphenylmethane diisocyanate (MDI) and vinyl-group-containing poly(butadiene) diol (PBD). Several methacrylates, such as methyl methacrylate (MMA), butyl methacrylate (BMA), stearyl methacrylate (SMA), and a phosphatidylcholine polar-headgroup-containing vinyl monomer, 2-(methacrylorloxy)ethyl-2-(trimethylammonium)ethyl phosphate (MTP), were grafted to the V-SPU. The bulk characteristics of the grafted V-SPUs were investigated by infrared (IR) spectroscopy, viscosity, and gel permeation chromatography (GPC) measurements. Mechanical properties of the typical SPU containing MMA were measured by dynamic viscoelasticity and tensile ...

Synthesis and Hemocompatibility Evaluation of Novel Segmented Polyurethanes with Phosphatidylcholine Polar Headgroups

A new diol with a phosphatidylcholine polar headgroup, [bis(2-hydroxymethyl)]propane-2-(trimethylammonio)ethyl phosphate (BTEP), was synthesized and characterized. The BTEP, together with 1,4-butanediol (BD) as a chain extender, was used to synthesize segmented polyurethanes (SPUs) based on diphenylmethane diisocyanate (MDI) and various types of soft segments such as polycarbonate, polyether, polyester, and hydrocarbon diols. The bulk characteristics of the resulting SPUs were investigated by infrared (IR) spectroscopy, viscosity, and gel-permeation chromatography (GPC) measurements. Good mechanical properties of the typical SPU containing poly(butadiene) glycol (PBD) were indicated by dynamic viscoelasticity and tensile measurements. The phosphatidylcholine polar groups were oriented on the surface of these materials as revealed by attenuated total reflectance−Fourier transform infrared spectroscopy (ATR−FTIR), electron spectroscopy for chemical analysis (ESCA), and contact angle measurements. The hemoco...

Poly(ethylene glycol)-induced and temperature-dependent phase separation in fluid binary phospholipid membranes

Exclusion of the strongly hygroscopic polymer, poly(ethylene glycol) (PEG), from the surface of phosphatidylcholine liposomes results in an osmotic imbalance between the hydration layer of the liposome surface and the bulk polymer solution, thus causing a partial dehydration of the phospholipid polar headgroups. PEG (average molecular weight of 6000 and in concentrations ranging from 5 to 20%, w/w) was added to the outside of large unilamellar liposomes (LUVs). This leads to, in addition to the dehydration of the outer monolayer, an osmotically driven water outflow and shrinkage of liposomes. Under these conditions phase separation of the fluorescent lipid 1-palmitoyl-2[6-(pyren-1-yl)]decanoyl-sn-glycero-3-phosphocholine (PPDPC) embedded in various phosphatidylcholine matrices was observed, evident as an increase in the excimer-to-monomer fluorescence intensity ratio (IE/IM). Enhanced segregation of the fluorescent lipid was seen upon increasing and equal concentrations of PEG both inside and outside of the LUVs, revealing that osmotic gradient across the membrane is not required, and phase separation results from the dehydration of the lipid. Importantly, phase separation of PPDPC could be induced by PEG also in binary mixtures with 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC), and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), for which temperature-induced phase segregation of the fluorescent lipid below Tm was otherwise not achieved. In the different lipid matrices the segregation of PPDPC caused by PEG was abolished above characteristic temperatures T0 well above their respective main phase transition temperatures Tm. For 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), DMPC, SOPC, and POPC, T0 was observed at approximately 50, 32, 24, and 20 degrees C, respectively. Notably, the observed phase separation of PPDPC cannot be accounted for the 1 degree C increase in Tm for DMPC or for the increase by 0.5 degrees C for DPPC observed in the presence of 20% (w/w) PEG. At a given PEG concentration maximal increase in IE/IM (correlating to the extent of segregation of PPDPC in the different lipid matrices) decreased in the sequence 1,2-dihexadecyl-sn-glycero-3-phosphocholine (DHPC) > DPPC > DMPC > SOPC > POPC, whereas no evidence for phase separation in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) LUV was observed (Lehtonen and Kinnunen, 1994, Biophys. J. 66: 1981-1990). Our results indicate that PEG-induced dehydration of liposomal membranes provides the driving force for the segregation of the pyrene lipid. In brief, phase separation of PPDPC from the matrix lipid could be attributed to the diminishing effective size of the phosphatidylcholine polar headgroup resulting from its partial dehydration by PEG. This in turn would allow for enhanced van der Waals interactions between the acyl chains of the matrix lipid, which then caused the exclusion of PPDPC due to the perturbing bulky pyrene moiety. Phase separation in DMPC/PPDPC liposomes was abolished by the inclusion of 25 mol % cholesterol and to a lesser extent by epicholesterol.

Calorimetric studies of freeze-induced dehydration of phospholipids

Differential scanning calorimetry (DSC) was used to determine the amount of water that freezes in an aqueous suspension of multilamellar dipalmitoylphosphatidylcholine (DPPC) liposomes. The studies were performed with dehydrated suspensions (12-20 wt% water) and suspensions containing an excess of water (30-70 wt% water). For suspensions that contained > or = 18 wt% water, two ice-formation events were observed during cooling. The first was attributed to heterogeneous nucleation of extraliposomal ice; the second was attributed to homogeneous nucleation of ice within the liposomes. In suspensions with an initial water concentration between 13 and 16 wt%, ice formation occurred only after homogeneous nucleation at temperatures below -40 degrees C. In suspensions containing < 13 wt% water, ice formation during cooling was undetectable by DSC, however, an endotherm resulting from ice melting during warming was observed in suspensions containing > or = 12 wt% water. In suspensions containing < 12 wt% water, an endotherm corresponding to the melting of ice was not observed during warming. The amount of ice that formed in the suspensions was determined by using an improved procedure to calculate the partial area of the endotherm resulting from the melting of ice during warming. The results show that a substantial proportion of water associated with the polar headgroup of phosphatidylcholine can be removed by freeze-induced dehydration, but the amount of ice depends on the thermal history of the samples. For example, after cooling to -100 degrees C at rates > or = 10 degrees C/min, a portion of water in the suspension remains supercooled because of a decrease in the diffusion rate of water with decreasing temperature. A portion of this supercooled water can be frozen during subsequent freeze-induced dehydration of the liposomes under isothermal conditions at subfreezing storage temperature Ts. During isothermal storage at Ts > or = -40 degrees C, the amount of unfrozen water decreased with decreasing Ts and increasing time of storage. After 30 min of storage at Ts = -40 degrees C and subsequent cooling to -100 degrees C, the amount of water associated with the polar headgroups was < 0.1 g/g of DPPC. At temperatures > -50 degrees C, the amount of unfrozen water associated with the polar headgroups of DPPC decreased with decreasing temperature in a manner predicted from the desorption isotherm of DPPC. However, at lower temperatures, the amount of unfrozen water remained constant, in large part, because the unfrozen water underwent a liquid-to-glass transformation at a temperature between -50 degrees and -140 degrees C.

Interaction of Carbohydrates with Phosphatidylcholine Inverse Micelles

AbstractSmall angle X‐ray scattering experiments indicate that egg phosphatidylcholine dissolved in benzene at a concentration of 12–50 mM organizes into approximately isometric inverse micelles with a size that depends upon the amount of water present. These inverse micelles can serve as self‐assembled hosts for monosaccharide derivatives. For a given amount of water, addition of carbohydrates causes an increase in the micelle size. In contrast, cyclohexane solutions of egg phosphatidylcholine contain highly anisotropic structures, possibly elongated rods. Monosaccharide derivatives can be extracted into these structures and are bound, at low water content, to the polar head groups. Upon addition of water the sugar is probably displaced from the polar head groups and gains some motional freedom in the water pool which is formed. This fact indicates that the polar head group of phosphatidylcholine interacts, probably by means of hydrogen bond formation, with monosaccharides and brings about their solubilization in nonpolar solvents.

Conformational analysis of the polar head group in phosphatidylcholine bilayers: a structural change induced by cations

The conformation of the polar head group of phosphatidylcholine in a bilayer in the liquid-crystalline state was deduced by analyzing the deuterium quadrupole splittings of the choline group and the phosphorus chemical shift anisotropy of the phosphate group in combination with the restriction of the choline conformation determined in laser Raman studies. The latter efficiently reduced the number of candidates for the actual conformation. A family of conformations was obtained for both the dynamic-structure and rigid-structure models, respectively. The polar head group is oriented roughly parallel to the membrane surface in both models. Furthermore, they are close to conformation A of the crystal structure of 1,2-dimyristoyl-sn-glycero-3-phosphocholine. The dynamic-structure model was concluded to be more reasonable in view of the fact that the polar head-group structures in most crystals comprise two conformations, which are nearly mirror images of each other. Conformational analysis was also carried out for the polar head group in the presence of multivalent cations. A possible conformational change of the polar head group induced by cations is discussed in the light of the present results.

Transbilayer phosphatidylcholine distributions in small unilamellar sphingomyelin-phosphatidylcholine vesicles: effect of altered polar head group

The effect of the altered polar head group of phosphatidylcholine (PC) on its transbilayer distributions in small unilamellar vesicles containing sphingomyelin (SM) was ascertained with phospholipase A2 as the external membrane probe. These vesicles were formed by sonication and fractionated by centrifugation. The vesicle size was determined by gel-permeation chromatography and solute entrapment. Experiments were done to confirm that phospholipase A2 treatments did not induce fusion, lyse the vesicles, or cause PC to migrate across the vesicle bilayer. The complete degradation of external PC in intact vesicles was assured by carrying out the enzyme reactions in the absence as well as in the presence of 9.2 X 10(-5) M bovine serum albumin. In small vesicles comprised of SM and 30 mol % 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), DPPC preferentially distributed in the inner monolayer. This preference of DPPC in these vesicles disappeared upon introducing one C2H5 group at the carbon atom adjacent to the quaternary ammonium residue in its polar head group and was reversed when the C2H5 group was replaced by C6H5 and C6H5CH2 substituents or when the P-N distance was increased. These results indicate that the effective polar head-group volume is an important factor in determining the phospholipid distributions across the small vesicle bilayer.

The interactions between monovalent ions and phosphatidyl cholines in aqueous bilayers.

The interactions of lithium and sodium ions and water with phosphatidylcholine bilayers have been studied by means of 7Li, 23Na and 2H NMR quadrupole splittings. The experimental results are interpreted in terms of a simple three-site model with two anisotropic sites ('binding' sites) and one isotropic site ('free' ions and water molecules). The findings obtained for the zwitterionic model membrane are compared with previous investigations of lamellar phases composed of ionic and nonionic amphiphiles. It is shown that the data obtained are compatible with our previous suggestion [Lindblom, G., Persson, N.-O., and Arvidson G. (1976) Adv. Chem. Ser. 152, 121] that an increase in the salt content in the water layer induces a conformational change in the polar head group of phosphatidylcholine. Thus at high salt concentration the phosphocholine head group tends to be oriented perpendicular to the lipid bilayer surface. The study also shows that increasing the amount of salt leads to a squeezing out of water between the bilayers. This is interpreted in terms of a reduction of the repulsion forces between the bilayers.

Short and long-range forces involved in cation-induced aggregation of chromaffin granule membranes

The kinetics and equilibria of cation-induced aggregation of chromaffin granule (CG) membranes are analyzed. The following mechanistic observations are made 1. (1) Cation-induced dimerization of CG membranes is brought about by reducing the membrane surface potential and hence by reducing the electrostatic repulsion between membranes. This can be accomplished by either monovalent or divalent cations. 2. (2) The dimerization reaction is nearly diffusion-controlled. The activation energy of the process is low. 3. (3) The kinetics are analyzed in terms of an “encounter complex” model formulated for comparison with the delay times involved in Ca 2+ -triggered exocytotic release. The half time for aggregation of vesicles with distances of separation between 26 Å and 126 .Å is calculated to be on the order of 3 msec. 4. (4) The role of van der Waals attraction, electrostatic repulsion and dipolar repulsion is considered. Calculations show that dipolar repulsion of phosphatidyl choline polar head groups should outweigh van der Waals attraction for short distances of separation. 5. (5) Phospholipid vesicles prepared from CG membrane lipids also show cation-induced aggregation. However, the behavior is qualitatively and quantitatively dissimilar to that of the CG membrane. The rates of aggregation are two orders of magnitude slower. This is shown to be due to a negative entropy of activation resulting from the necessity of matching the surfaces of the vesicles. Vesicle approaches involving high energies (enthalpies) of activation are excluded.