Sample records for solubilizes phospholipid vesicles

  1. The aminosterol antibiotic squalamine permeabilizes large unilamellar phospholipid vesicles.

    Selinsky, B S; Zhou, Z; Fojtik, K G; Jones, S R; Dollahon, N R; Shinnar, A E


    The ability of the shark antimicrobial aminosterol squalamine to induce the leakage of polar fluorescent dyes from large unilamellar phospholipid vesicles (LUVs) has been measured. Micromolar squalamine causes leakage of carboxyfluorescein (CF) from vesicles prepared from the anionic phospholipids phosphatidylglycerol (PG), phosphatidylserine (PS), and cardiolipin. Binding analyses based on the leakage data show that squalamine has its highest affinity to phosphatidylglycerol membranes, followed by phosphatidylserine and cardiolipin membranes. Squalamine will also induce the leakage of CF from phosphatidylcholine (PC) LUVs at low phospholipid concentrations. At high phospholipid concentrations, the leakage of CF from PC LUVs deviates from a simple dose-response relationship, and it appears that some of the squalamine can no longer cause leakage. Fluorescent dye leakage generated by squalamine is graded, suggesting the formation of a discrete membrane pore rather than a generalized disruption of vesicular membranes. By using fluorescently labeled dextrans of different molecular weight, material with molecular weight squalamine, but material with molecular weight >/=10,000 is retained. Negative stain electron microscopy of squalamine-treated LUVs shows that squalamine decreases the average vesicular size in a concentration-dependent manner. Squalamine decreases the size of vesicles containing anionic phospholipid at a lower squalamine/lipid molar ratio than pure PC LUVs. In a centrifugation assay, squalamine solubilizes phospholipid, but only at significantly higher squalamine/phospholipid ratios than required for either dye leakage or vesicle size reduction. Squalamine solubilizes PC at lower squalamine/phospholipid ratios than PG. We suggest that squalamine complexes with phospholipid to form a discrete structure within the bilayers of LUVs, resulting in the transient leakage of small encapsulated molecules. At higher squalamine/phospholipid ratios, these

  2. Two distinct mechanisms of vesicle-to-micelle and micelle-to-vesicle transition are mediated by the packing parameter of phospholipid-detergent systems

    Stuart, Marc C. A.; Boekema, Egbert J.


    The detergent solubilization and reformation of phospholipid vesicles was studied for various detergents. Two distinct mechanisms of vesicle-to-micelle and micelle-to-vesicle transition were observed by turbidimetry and cryo-electron microscopy. The first mechanism involves fast solubilization of ph

  3. Phospholipid Vesicles in Materials Science

    Granick, Steve [Univ. of Illinois, Champaign, IL (United States)


    The objective of this research was to develop the science basis needed to deploy phospholipid vesicles as functional materials in energy contexts. Specifically, we sought to: (1) Develop an integrated molecular-level understanding of what determines their dynamical shape, spatial organization, and responsiveness to complex, time-varying environments; and (2) Develop understanding of their active transportation in crowded environments, which our preliminary measurements in cells suggest may hold design principles for targeting improved energy efficiency in new materials systems. The methods to do this largely involved fluorescence imaging and other spectroscopy involving single particles, vesicles, particles, DNA, and endosomes. An unexpected importance outcome was a new method to image light-emitting diodes during actual operation using super-resolution spectroscopy.

  4. Effect of Vesicle-to-Micelle Transition on the Interactions of Phospholipid/Sodium Cholate Mixed Systems with Curcumin in Aqueous Solution.

    Zhang, Sha; Wang, Xiaoyong


    The role of vesicle-to-micelle transition has been investigated in the interactions of phospholipid vesicles, phospholipid/sodium cholate (NaC) mixed vesicles, and phospholipid/NaC mixed micelles with curcumin in aqueous solution. The addition of NaC causes phospholipid vesicles to transit into phospholipid/NaC mixed vesicles and phospholipid/NaC mixed micelles. Turbidity measurement reveals that the presence of curcumin increases the NaC concentration for the solubilization of phospholipid vesicles, which indicates that the bound curcumin tends to suppress the vesicle-to-micelle transition. The pyrene polarity index and curcumin fluorescence anisotropy measurements suggest that phospholipid/NaC mixed micelles have a more compact structure than that of phospholipid vesicles and phospholipid/NaC mixed vesicles. Curcumin associated with phospholipid vesicles, phospholipid/NaC mixed vesicles, and phospholipid/NaC mixed micelles often results in higher intensities of absorption and fluorescence than those of free curcumin. However, phospholipid/NaC mixed vesicles lead to the highest values of absorption and fluorescence intensities, binding constant, and radical-scavenging capacity with curcumin. The different structures in the phospholipid bilayer of phospholipid/NaC mixed vesicles and the hydrophobic part of phospholipid/NaC mixed micelles where curcumin located are discussed to explain the interaction behaviors of phospholipid/NaC mixed systems with curcumin.

  5. Hybrid, Nanoscale Phospholipid/Block Copolymer Vesicles

    Bo Liedberg


    Full Text Available Hybrid phospholipid/block copolymer vesicles, in which the polymeric membrane is blended with phospholipids, display interesting self-assembly behavior, incorporating the robustness and chemical versatility of polymersomes with the softness and biocompatibility of liposomes. Such structures can be conveniently characterized by preparing giant unilamellar vesicles (GUVs via electroformation. Here, we are interested in exploring the self-assembly and properties of the analogous nanoscale hybrid vesicles (ca. 100 nm in diameter of the same composition prepared by film-hydration and extrusion. We show that the self-assembly and content-release behavior of nanoscale polybutadiene-b-poly(ethylene oxide (PB-PEO/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC hybrid phospholipid/block copolymer vesicles can be tuned by the mixing ratio of the amphiphiles. In brief, these hybrids may provide alternative tools for drug delivery purposes and molecular imaging/sensing applications and clearly open up new avenues for further investigation.

  6. Determination of some solubilization parameters with surfactants of egg-yolk lecithin multilamellar vesicles by static light-scattering measurements

    Hobai, S; Hobai, St.; Fazakas, Zita


    Effective surfactant:phospholipid ratios (i.e. molar ratios in the mixed aggregates, vesicles or micelles) have been determined by static light-scattering for the interaction of egg-yolk lecithin (EYL) multilamellar vesicles (MLV) with Triton X-100 (TX-100), sodium deoxycholate (DOCNa) and cetyltrimethylammonium bromide (CTMB). The suspension of MLV-EYL was mixed with appropriate volumes of surfactant solution and was left overnight to reaches thermodynamic equilibrium. Rectan-gular optic diffusion data were used to compute the solubilization parameters: total surfactant concentrations, at saturation and solubilization Dtsat and Dtsol respectively, and effective molar ratios, Resat and Resol respectively. From the Resat value ob-tained graphically for interaction of vesicles with TX-100 resulted that in vesicle bilayers a surfactant molecule is surrounded with seven phospholipid molecules and the Resol value suggests that in mixed micelles ten lipid molecules with about fifteen surfactant molecules coexist. T...

  7. Slow Phospholipid Exchange between a Detergent-Solubilized Membrane Protein and Lipid-Detergent Mixed Micelles Brominated Phospholipids as Tools to Follow Its Kinetics

    Montigny, Cédric; Dieudonné, Thibaud; Orlowski, Stéphane


    Membrane proteins are largely dependent for their function on the phospholipids present in their immediate environment, and when they are solubilized by detergent for further study, residual phospholipids are critical, too. Here, brominated phosphatidylcholine, a phospholipid which behaves as an ...

  8. Magnetic field alignable domains in phospholipid vesicle membranes containing lanthanides.

    Beck, Paul; Liebi, Marianne; Kohlbrecher, Joachim; Ishikawa, Takashi; Rüegger, Heinz; Zepik, Helmut; Fischer, Peter; Walde, Peter; Windhab, Erich


    Magnetic fields were applied as a structuring force on phospholipid-based vesicular systems, using paramagnetic lanthanide ions as magnetic handles anchored to the vesicle membrane. Different vesicle formulations were investigated using small angle neutron scattering (SANS) in a magnetic field of up to 8 T, cryo-transmission electron microscopy (cryo-TEM), (31)P NMR spectroscopy, dynamic light scattering (DLS), and permeability measurements with a fluorescent water-soluble marker (calcein). The investigated vesicle formulations consisted usually of 80 mol % of the phospholipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 20 mol % of a chelator lipid (DMPE-DTPA; 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-diethylenetriaminepentaacetate) with complexed lanthanide ions (Tm(3+), Dy(3+), or La(3+)), and the total lipid concentration was 15 mM. Vesicles containing the paramagnetic lanthanide Tm(3+) or Dy(3+) exhibited a temperature-dependent response to magnetic fields, which can be explained by considering the formation of lipid domains, which upon reaching a critical size become alignable in a magnetic field. The features of this "magnetic field alignable domain model" are as follows: with decreasing temperature (from 30 to 2.5 degrees C) solid domains, consisting mainly of the higher melting phospholipid (DMPE-DTPA.lanthanide), begin to form and grow in size. The domains assemble the large magnetic moments conferred by the lanthanides and orient in magnetic fields. The direction of alignment depends on the type of lanthanide used. The domains orient with their normal parallel to the magnetic field with thulium (Tm(3+)) and perpendicular with dysprosium (Dy(3+)). No magnetic field alignable domains were observed if DMPE-DTPA is replaced either by POPE-DTPA (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine-diethylenetriamine-pentaacetate) or by DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine).

  9. Entrapment in phospholipid vesicles quenches photoactivity of quantum dots.

    Generalov, Roman; Kavaliauskiene, Simona; Westrøm, Sara; Chen, Wei; Kristensen, Solveig; Juzenas, Petras


    Quantum dots have emerged with great promise for biological applications as fluorescent markers for immunostaining, labels for intracellular trafficking, and photosensitizers for photodynamic therapy. However, upon entry into a cell, quantum dots are trapped and their fluorescence is quenched in endocytic vesicles such as endosomes and lysosomes. In this study, the photophysical properties of quantum dots were investigated in liposomes as an in vitro vesicle model. Entrapment of quantum dots in liposomes decreases their fluorescence lifetime and intensity. Generation of free radicals by liposomal quantum dots is inhibited compared to that of free quantum dots. Nevertheless, quantum dot fluorescence lifetime and intensity increases due to photolysis of liposomes during irradiation. In addition, protein adsorption on the quantum dot surface and the acidic environment of vesicles also lead to quenching of quantum dot fluorescence, which reappears during irradiation. In conclusion, the in vitro model of phospholipid vesicles has demonstrated that those quantum dots that are fated to be entrapped in endocytic vesicles lose their fluorescence and ability to act as photosensitizers.

  10. Entrapment in phospholipid vesicles quenches photoactivity of quantum dots

    Generalov R


    Full Text Available Roman Generalov1,2, Simona Kavaliauskiene1, Sara Westrøm1, Wei Chen3, Solveig Kristensen2, Petras Juzenas11Department of Radiation Biology, Institute for Cancer Research, Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway; 2School of Pharmacy, University of Oslo, Oslo, Norway; 3Department of Physics, The University of Texas at Arlington, Arlington, TX, USAAbstract: Quantum dots have emerged with great promise for biological applications as fluorescent markers for immunostaining, labels for intracellular trafficking, and photosensitizers for photodynamic therapy. However, upon entry into a cell, quantum dots are trapped and their fluorescence is quenched in endocytic vesicles such as endosomes and lysosomes. In this study, the photophysical properties of quantum dots were investigated in liposomes as an in vitro vesicle model. Entrapment of quantum dots in liposomes decreases their fluorescence lifetime and intensity. Generation of free radicals by liposomal quantum dots is inhibited compared to that of free quantum dots. Nevertheless, quantum dot fluorescence lifetime and intensity increases due to photolysis of liposomes during irradiation. In addition, protein adsorption on the quantum dot surface and the acidic environment of vesicles also lead to quenching of quantum dot fluorescence, which reappears during irradiation. In conclusion, the in vitro model of phospholipid vesicles has demonstrated that those quantum dots that are fated to be entrapped in endocytic vesicles lose their fluorescence and ability to act as photosensitizers.Keywords: fluorescence lifetime, free radicals, liposomes, lipodots, reactive oxygen species

  11. Inositol depletion restores vesicle transport in yeast phospholipid flippase mutants.

    Yamagami, Kanako; Yamamoto, Takaharu; Sakai, Shota; Mioka, Tetsuo; Sano, Takamitsu; Igarashi, Yasuyuki; Tanaka, Kazuma


    In eukaryotic cells, type 4 P-type ATPases function as phospholipid flippases, which translocate phospholipids from the exoplasmic leaflet to the cytoplasmic leaflet of the lipid bilayer. Flippases function in the formation of transport vesicles, but the mechanism remains unknown. Here, we isolate an arrestin-related trafficking adaptor, ART5, as a multicopy suppressor of the growth and endocytic recycling defects of flippase mutants in budding yeast. Consistent with a previous report that Art5p downregulates the inositol transporter Itr1p by endocytosis, we found that flippase mutations were also suppressed by the disruption of ITR1, as well as by depletion of inositol from the culture medium. Interestingly, inositol depletion suppressed the defects in all five flippase mutants. Inositol depletion also partially restored the formation of secretory vesicles in a flippase mutant. Inositol depletion caused changes in lipid composition, including a decrease in phosphatidylinositol and an increase in phosphatidylserine. A reduction in phosphatidylinositol levels caused by partially depleting the phosphatidylinositol synthase Pis1p also suppressed a flippase mutation. These results suggest that inositol depletion changes the lipid composition of the endosomal/TGN membranes, which results in vesicle formation from these membranes in the absence of flippases.

  12. Slow Phospholipid Exchange between a Detergent-Solubilized Membrane Protein and Lipid-Detergent Mixed Micelles: Brominated Phospholipids as Tools to Follow Its Kinetics

    Montigny, Cédric; Dieudonné, Thibaud; Orlowski, Stéphane; Vázquez-Ibar, José Luis; Gauron, Carole; Georgin, Dominique; Lund, Sten; le Maire, Marc; Møller, Jesper V.; Champeil, Philippe


    Membrane proteins are largely dependent for their function on the phospholipids present in their immediate environment, and when they are solubilized by detergent for further study, residual phospholipids are critical, too. Here, brominated phosphatidylcholine, a phospholipid which behaves as an unsaturated phosphatidylcholine, was used to reveal the kinetics of phospholipid exchange or transfer from detergent mixed micelles to the environment of a detergent-solubilized membrane protein, the paradigmatic P-type ATPase SERCA1a, in which Trp residues can experience fluorescence quenching by bromine atoms present on phospholipid alkyl chains in their immediate environment. Using dodecylmaltoside as the detergent, exchange of (brominated) phospholipid was found to be much slower than exchange of detergent under the same conditions, and also much slower than membrane solubilization, the latter being evidenced by light scattering changes. The kinetics of this exchange was strongly dependent on temperature. It was also dependent on the total concentration of the mixed micelles, revealing the major role for such exchange of the collision of detergent micelles with the detergent-solubilized protein. Back-transfer of the brominated phospholipid from the solubilized protein to the detergent micelle was much faster if lipid-free DDM micelles instead of mixed micelles were added for triggering dissociation of brominated phosphatidylcholine from the solubilized protein, or in the additional presence of C12E8 detergent during exchange, also emphasizing the role of the chemical nature of the micelle/protein interface. This protocol using brominated lipids appears to be valuable for revealing the possibly slow kinetics of phospholipid transfer to or from detergent-solubilized membrane proteins. Independently, continuous recording of the activity of the protein can also be used in some cases to correlate changes in activity with the exchange of a specific phospholipid, as shown here

  13. Permeability of phospholipid membrane for small polar molecules determined from osmotic swelling of giant phospholipid vesicles

    Peterlin, Primoz; Diamant, Haim; Haleva, Emir


    A method for determining permeability of phospholipid bilayer based on the osmotic swelling of micrometer-sized giant unilamellar vesicles (GUVs) is presented as an alternative to the two established techniques, dynamic light scattering on liposome suspension, and electrical measurements on planar lipid bilayers. In the described technique, an individual GUV is transferred using a micropipette from a sucrose/glucose solution into an isomolar solution containing the solute under investigation. Throughout the experiment, vesicle cross-section is monitored and recorded using a digital camera mounted on a phase-contrast microscope. Using a least-squares procedure for circle fitting, vesicle radius R is computed from the recorded images of vesicle cross-section. Two methods for determining membrane permeability from the obtained R(t) dependence are described: the first one uses the slope of R(t) for a spherical GUV, and the second one the R(t) dependence around the transition point at which a flaccid vesicle trans...

  14. Enhanced solubilization of curcumin in mixed surfactant vesicles.

    Kumar, Arun; Kaur, Gurpreet; Kansal, S K; Chaudhary, Ganga Ram; Mehta, S K


    Self-assemblies of equimolar double and single chain mixed ionic surfactants, with increasing numbers of carbon atoms of double chain surfactant, were analyzed on the basis of fluorescence and conductivity results. Attempts were also made to enhance the solubilization of curcumin in aqueous equimolar mixed surfactant systems. Mixed surfactant assembly was successful in retarding the degradation of curcumin in alkaline media (only 25-28 40% degraded in 10h at pH 13). Fluorescence spectroscopy and fluorescence quenching methods were employed to predict the binding position and mechanism of curcumin with self-assemblies. Results indicate that the interactions take place according to both dynamic and static quenching mechanisms and curcumin was distributed in a palisade layer of mixed aggregates. Antioxidant activity (using DPPH radical) and biocompatibility (using calf-thymus DNA) of curcumin-loaded mixed surfactant formulations were also evaluated. The prepared systems improved the stability, solubility and antioxidant activity of curcumin and additionally are biocompatible.

  15. Optimization of the Electroformation of Giant Unilamellar Vesicles (GUVs) with Unsaturated Phospholipids.

    Breton, Marie; Amirkavei, Mooud; Mir, Lluis M


    Giant unilamellar vesicles (GUV) are widely used cell membrane models. GUVs have a cell-like diameter and contain the same phospholipids that constitute cell membranes. The most frequently used protocol to obtain these vesicles is termed electroformation, since key steps of this protocol consist in the application of an electric field to a phospholipid deposit. The potential oxidation of unsaturated phospholipids due to the application of an electric field has not yet been considered even though the presence of oxidized lipids in the membrane of GUVs could impact their permeability and their mechanical properties. Thanks to mass spectrometry analyses, we demonstrated that the electroformation technique can cause the oxidation of polyunsaturated phospholipids constituting the vesicles. Then, using flow cytometry, we showed that the amplitude and the duration of the electric field impact the number and the size of the vesicles. According to our results, the oxidation level of the phospholipids increases with their level of unsaturation as well as with the amplitude and the duration of the electric field. However, when the level of lipid oxidation exceeds 25 %, the diameter of the vesicles is decreased and when the level of lipid oxidation reaches 40 %, the vesicles burst or reorganize and their rate of production is reduced. In conclusion, the classical electroformation method should always be optimized, as a function of the phospholipid used, especially for producing giant liposomes of polyunsaturated phospholipids to be used as a cell membrane model.

  16. Transporting of a Cell-Sized Phospholipid Vesicle Across Water/Oil Interface

    Hase, M; Hamada, T; Yoshikawa, K; Hase, Masahiko; Yamada, Ayako; Hamada, Tsutomu; Yoshikawa, Kenichi


    When a cell-sized water droplet, with a diameter of several tens of micro meter, is placed in oil containing phospholipids, a stable cell-sized vesicle is spontaneously formed as a water-in-oil phospholipid emulsion (W/O CE) with a phospholipid monolayer. We transferred the lipid vesicle thus formed in the oil phase to the water phase across the water/oil interface by micromanipulation, which suggests that the vesicle is transformed from a phospholipid monolayer as W/O CE into a bilayer. The lipid vesicle can then be transported back into the oil phase. This novel experimental procedure may be a useful tool for creating a model cellular system, which, together with a microreactor, is applicable as a micrometer-scale biochemical reaction field.

  17. Dynamic assembly of MinD on phospholipid vesicles regulated by ATP and MinE

    Hu, Zonglin; Gogol, Edward P.; Lutkenhaus, Joe


    Selection of the division site in Escherichia coli is regulated by the min system and requires the rapid oscillation of MinD between the two halves of the cell under the control of MinE. In this study we have further investigated the molecular basis for this oscillation by examining the interaction of MinD with phospholipid vesicles. We found that MinD bound to phospholipid vesicles in the presence of ATP and, upon binding, assembled into a well-ordered helical array that deformed the vesicle...

  18. Aggregation of phospholipid vesicles induced by the ribosome inactivating protein saporin.

    Hao, Q; Yan, L; Yang, H; Zhang, Y; Gao, G; Yao, Q; Li, Q


    Saporin-S6(SO-6) is a single chain ribosome inactivating protein, which can inhibit protein synthesis by inactivating eukaryotic ribosomes. The interaction of SO-6 with phospholipid model systems was described. SO-6 can specifically interact with negatively-charged phospholipid vesicles and it induces the aggregation of the lipid vesicles. The kinetics of the vesicle aggregation induced by SO-6 was studied. The saturating protein/lipid molar ratio was determined to be 1:100 based on titration experiments. The aggregation is dependent on the temperature in a range that was many times higher than the phase transition temperature of the phospholipid. The effect of pH on the aggregation of the vesicles can not be explained by simple deprotonation of side chain amino groups of the protein, and may be related to conformational changes of the protein. The maintenance of physiological ionic strength was required for the aggregation of SO-6 with vesicles. Finally, the interaction was prompted by Ca2+ ions, and was totally inhibited by EDTA, which suggests that SO-6 may interact with phospholipid vesicles in a Ca(2+)-dependent manner.

  19. Giant vesicles from 72-membered macrocyclic archaeal phospholipid analogues: initiation of vesicle formation by molecular recognition between membrane components

    Eguchi; Arakawa; Kakinuma; Rapp; Ghosh; Nakatani; Ourisson


    Stereochemically pure archeal acyclic bola-amphiphilic diphosphates 4 and 5, with the basic structure of the phospholipids found in Sulfolobus, have been synthesized for the first time. The self-assembly properties have been compared with those of the nearly identical 72-membered macrocyclic tetraether phosphates 3a and 3b, analogues of the major phospholipid components of Sulfolobus, Thermoplasma, and methanogenic Archea, which were also synthesized. Phase contrast and fluorescence microscopies have shown that the dipolar lipids 1 and 2 spontaneously formed vesicles. Whereas the macrocyclic dipolar phosphates 3 spontaneously formed vesicles (phase contrast and fluorescence microscopies), the bolaform phosphate 4 gave only a lamellar structure (synchrotron diffraction pattern: repeat distance of about 4.25 nm but with only a few layers). However, upon addition of the unphosphorylated precursors phytanol, phytol, or geranylgeraniol to the acyclic lipids 4 and 5, giant vesicles were rapidly formed. Addition of n-hexadecanol or cholesterol did not lead to vesicle formation. Therefore it was concluded that this vesicle formation occurs only when the added molecule is closely compatible with the constituents of the lipid layer and can be inserted into the double layer. A slight mismatch (cholesterol or n-hexadecanol/polyprenyl chains) is therefore enough to block the insertion process presumably required for vesicle formation.

  20. Bile Salt Micelles and Phospholipid Vesicles Present in Simulated and Human Intestinal Fluids

    Elvang, Philipp A; Hinna, Askell H; Brouwers, Joachim


    Knowledge about colloidal assemblies present in human intestinal fluids (HIFs), such as bile salt micelles and phospholipid vesicles, is regarded of importance for a better understanding of the in vivo dissolution and absorption behavior of poorly soluble drugs (Biopharmaceutics Classification...... distinct size fraction of colloidal assemblies, whereas FeSSIF contained 2 fractions of colloidal species with significantly different sizes. These size fractions likely represent (1) mixed taurocholate-phospholipid-micelles, as indicated by a size range up to 70 nm (in diameter) and a strong UV absorption...... sizes of approximately 50 and 200 nm, respectively (intensity-weighted mean diameter, Dz), likely representing mixed cholate/phospholipid micelles and phospholipid vesicles, respectively. The sizes of the smaller 2 fractions being below the size range of multiangle laser light scattering analysis (




    Lysozyme-induced fusion of phosphatidylserine (PS) vesicles was studied as a function of pH. Fusion, monitored by lipid-mixing, was measured by following the dilution of pyrene-labelled phosphatidylcholine, incorporated in PS vesicles, into unlabelled bilayers. It is demonstrated that

  2. Frequency-dependent electrodeformation of giant phospholipid vesicles in AC electric field

    Peterlin, Primoz


    A model of vesicle electrodeformation is described which obtains a parametrized vesicle shape by minimizing the sum of the membrane bending energy and the energy due to the electric field. Both the vesicle membrane and the aqueous media inside and outside the vesicle are treated as leaky dielectrics, and the vesicle itself is modelled as a nearly spherical shape enclosed within a thin membrane. It is demonstrated (a) that the model achieves a good quantitative agreement with the experimentally determined prolate-to-oblate transition frequencies in the kHz range, and (b) that the model can explain a phase diagram of shapes of giant phospholipid vesicles with respect to two parameters: the frequency of the applied AC electric field and the ratio of the electrical conductivities of the aqueous media inside and outside the vesicle, explored in a recent paper (S. Aranda et al., Biophys. J. 95:L19--L21, 2008). A possible use of the frequency-dependent shape transitions of phospholipid vesicles in conductometry of m...

  3. Ionic strength dependent vesicle adsorption and phase behavior of anionic phospholipids on a gold substrate.

    Pramanik, Sumit Kumar; Seneca, Senne; Ethirajan, Anitha; Neupane, Shova; Renner, Frank Uwe; Losada-Pérez, Patricia


    The authors report on the effect of ionic strength on the formation of supported vesicle layers of anionic phospholipids 1,2-dimyristoyl-sn-glycero-3-phospho-rac-glycerol (DMPG) and dimyristoylphosphatidylserine (DMPS) onto gold. Using quartz crystal microbalance with dissipation monitoring the authors show that vesicle adsorption is mainly governed by NaCl concentration, reflecting the importance of electrostatic interactions in anionic lipids, as compared to zwitterionic 1,2-dimyristoyl-sn-glycero-3-phosphocholine. At low ionic strength, low or no adsorption is observed as a result of vesicle-vesicle electrostatic repulsion. At medium ionic strength, the negative charges of DMPG and DMPS are screened resulting in larger adsorption and a highly dissipative intact vesicle layer. In addition, DMPS exhibits a peculiar behavior at high ionic strength that depends on the temperature of the process.

  4. Determining membrane permeability of giant phospholipid vesicles from a series of videomicroscopy images

    Peterlin, Primoz; Pisanski, Tomaz


    A technique for determining the permeability of a phospholipid membrane from a sequence of videomicrographs is described. A single giant unilamellar vesicle (GUV) is transferred using a micropipette from a solution of an impermeable solute (e.g., glucose or sucrose) into an iso-osmolar solution of a solute with a higher membrane permeability (e.g., glycerol). Upon the transfer, the vesicle swells until it reaches the tensile strength of the membrane, when the membrane breaks and a fraction of the vesicle volume is ejected, sufficient for the membrane to return to its relaxed value. The swelling-burst cycle repeats itself until the composition of the solution in the vesicle interior equilibrates with the external solution. A sequence of ~10.000 image frames is obtained from a CCD camera mounted on the optical microscope, documenting the process. On each frame, the vesicle radius is determined, and from the rate of swelling the membrane permeability can be obtained.

  5. Fusion of phospholipid vesicles induced by muscle glyceraldehyde-3-phosphate dehydrogenase in the absence of calcium.

    Morero, R D; Viñals, A L; Bloj, B; Farías, R N


    Ca2+-induced fusion of phospholipid vesicles (phosphatidylcholine/phosphatidic acid, 9:1 mol/mol) prepared by ethanolic injection was followed by five different procedures: resonance energy transfer, light scattering, electron microscopy, intermixing of aqueous content, and gel filtration through Sepharose 4-B. The five methods gave concordant results, showing that vesicles containing only 10% phosphatidic acid can be induced to fuse by millimolar concentrations of Ca2+. When the fusing capability of several soluble proteins was assayed, it was found that concanavalin A, bovine serum albumin, ribonuclease, and protease were inactive. On the other hand, lysozyme, L-lactic dehydrogenase, and muscle and yeast glyceraldehyde-3-phosphate dehydrogenase were capable of inducing vesicle fusion. Glyceraldehyde-3-phosphate dehydrogenase from rabbit muscle, the most extensively studied protein, proved to be very effective: 0.1 microM was enough to induce complete intermixing of bilayer phospholipid vesicles. Under conditions used in this work, fusion was accompanied by leakage of internal contents. The fusing capability of glyceraldehyde-3-phosphate dehydrogenase was not affected by 5 mM ethylenediaminetetraacetic acid. The Ca2+ concentration in the medium, as determined by atomic absorption spectroscopy, was 5 ppm. Heat-denatured enzyme was incapable of inducing fusion. We conclude that glyceraldehyde-3-phosphate dehydrogenase is a soluble protein inherently endowed with the capability of fusing phospholipid vesicles.

  6. Characterization of phospholipid+semifluorinated alkane vesicle system.

    Sabín, Juan; Ruso, Juan M; González-Pérez, Alfredo; Prieto, Gerardo; Sarmiento, Félix


    The aim of this study is to characterize vesicles obtained by the incorporation of the semifluorinated alkane, (perfluoro-n-hexyl)ethane (diblock F6H2) to a standard lipid, egg yolk phosphatidylcholine (PC). Large unilamellar vesicles (LUVs), prepared by extrusion, were characterized by fluorescence spectroscopy, zeta potential (zeta-potential) and light scattering. By using the fluorescence spectroscopy technique, the anisotropy of l,6-diphenyl-l,3,5-hexatriene (DPH) probe at different temperatures was determined. It was demonstrated that F6H2 is placed inside of the lipid bilayer and that the hydrocarbon acyl chain in the bilayers has higher viscosity in the presence of fluoroalkane. The zeta-potential of the PC-F6H2 system is negative and increases (in absolute value) from -10 to -19 mV when the temperature rises from 10 to 25 degrees C, this last value keeping practically constant with a further increase of temperature. The adsorption of K+ ions on the liposome surface was measured by zeta-potential. This adsorption originates a sudden increase of the initial zeta-potential followed by a slight decrease with K+ concentration. The application of the DLVO theory of colloidal stability showed a growing dependence of the DLVO potential with K+ concentration and consequently a increasing stability.




    The influence of a transmembrane pH gradient on the Ca2+-induced fusion of phospholipid vesicles, containing free fatty acids, has been investigated. Large unilamellar vesicles composed of an equimolar mixture of cardiolipin, dioleoylphosphatidylcholine, and cholesterol, containing 20 mol % oleic ac




    The interaction of clathrin with large unilamellar vesicles of various lipid compositions has been examined at neutral pH. Clathrin induces leakage of contents of vesicles that contain the acidic phospholipid phosphatidylserine. Leakage is greatly enhanced by the presence of a relatively minor

  9. Coupling of the fusion and budding of giant phospholipid vesicles containing macromolecules.

    Terasawa, Hidetoshi; Nishimura, Kazuya; Suzuki, Hiroaki; Matsuura, Tomoaki; Yomo, Tetsuya


    Mechanisms that enabled primitive cell membranes to self-reproduce have been discussed based on the physicochemical properties of fatty acids; however, there must be a transition to modern cell membranes composed of phospholipids [Budin I, Szostak JW (2011) Proc Natl Acad Sci USA 108:5249-5254]. Thus, a growth-division mechanism of membranes that does not depend on the chemical nature of amphiphilic molecules must have existed. Here, we show that giant unilamellar vesicles composed of phospholipids can undergo the coupled process of fusion and budding transformation, which mimics cell growth and division. After gaining excess membrane by electrofusion, giant vesicles spontaneously transform into the budded shape only when they contain macromolecules (polymers) inside their aqueous core. This process is a result of the vesicle maximizing the translational entropy of the encapsulated polymers (depletion volume effect). Because the cell is a lipid membrane bag containing highly concentrated biopolymers, this coupling process that is induced by physical and nonspecific interactions may have a general importance in the self-reproduction of the early cellular compartments.

  10. Solubilization of poorly water-soluble compounds using amphiphilic phospholipid polymers with different molecular architectures.

    Mu, Mingwei; Konno, Tomohiro; Inoue, Yuuki; Ishihara, Kazuhiko


    To achieve stable and effective solubilization of poorly water-soluble bioactive compounds, water-soluble and amphiphilic polymers composed of hydrophilic 2-methacryloyloxyethyl phosphorylcholine (MPC) units and hydrophobic n-butyl methacrylate (BMA) units were prepared. MPC polymers having different molecular architectures, such as random-type monomer unit sequences and block-type sequences, formed polymer aggregates when they were dissolved in aqueous media. The structure of the random-type polymer aggregate was loose and flexible. On the other hand, the block-type polymer formed polymeric micelles, which were composed of very stable hydrophobic poly(BMA) cores and hydrophilic poly(MPC) shells. The solubilization of a poorly water-soluble bioactive compound, paclitaxel (PTX), in the polymer aggregates was observed, however, solubilizing efficiency and stability were strongly depended on the polymer architecture; in other words, PTX stayed in the poly(BMA) core of the polymer micelle formed by the block-type polymer even when plasma protein was present in the aqueous medium. On the other hand, when the random-type polymer was used, PTX was transferred from the polymer aggregate to the protein. We conclude that water-soluble and amphiphilic MPC polymers are good candidates as solubilizers for poorly water-soluble bioactive compounds. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Hydration process of multi-stacked phospholipid bilayers to form giant vesicles

    Hishida, M; Yamada, N L; Yoshikawa, K


    A hydration process of multi-stacked phospholipid bilayers to form giant vesicles was investigated by means of time-resolved small angle X-ray scattering. It is found that the hydration of lipids in the liquid-crystalline phase proceeds with two stages: In the early stage until about 150 s after the hydration, lipid bilayers on a solid substrate swell about 20 % and arrive at a quasi-stable state. In the late stage, after several hundreds seconds, lipid bilayers gradually peel off from the stack. On the other hand, in case of a lipid in the gel phase, only the early stage is observed. These behaviors correspond to the ability of the giant vesicle formation depending on the lipid phases. The kinetics of the peeling-off process is discussed in terms of the Kramers' formulas by considering the effect of the unbinding transition.

  12. Chemical characterization of Citrus limon var. pompia and incorporation in phospholipid vesicles for skin delivery.

    Manconi, Maria; Manca, Maria Letizia; Marongiu, Francesca; Caddeo, Carla; Castangia, Ines; Petretto, Giacomo Luigi; Pintore, Giorgio; Sarais, Giorgia; D'hallewin, Guy; Zaru, Marco; Bacchetta, Gianluigi; Fadda, Anna Maria


    The components of pompia, a hybrid Citrus species cultivated only in Sardinia (Italy), were extracted using an environmentally-friendly method and food-grade solvents. Taking into account that only few data are available on pompia composition, the phytochemical fingerprint of its rind extract was obtained by accurate component separation and identification, combining HPLC and mass spectrometry. Different flavones such as naringin (23.77μg/mg), neoeriocitrin (46.53μg/mg) and neohesperidin (44.57μg/mg) were identified. Additionally, the antioxidant activity and phenolic content were confirmed by DPPH and Folin-Ciocalteu assays. The whole extract was incorporated in innovative phospholipid vesicles, namely glycerosomes, hyalurosomes and glycerol containing hyalurosomes, which were prepared using a high ratio of extract/phospholipid (1/3.5w/w). The in vitro biocompatibility of the nanoincorporated extract and its ability to potentiate the aptitude of the extract to counteract oxidative stress in skin cells were evaluated. The vesicles, especially glycerol containing hyalurosomes, were able to prevent oxidative damage and death of both keratinocytes and fibroblasts, promoting their viability. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Development of polyherbal antidiabetic formulation encapsulated in the phospholipids vesicle system

    Vinod Kumar Gauttam


    Full Text Available Multifactorial metabolic diseases, for instance diabetes develop several complications like hyperlipidemia, hepatic toxicity, immunodeficiency etc., Hence, instead of mono-drug therapy the management of the disease requires the combination of herbs. Marketed herbal drugs comprise of irrational combinations, which makes their quality control more difficult. Phytoconstituents, despite having excellent bioactivity in vitro demonstrate less or no in vivo actions due to their poor lipid solubility, resulting in high therapeutic dose regimen; phospholipids encapsulation can overcome this problem. Hence, present study was designed to develop a phospholipids encapsulated polyherbal anti-diabetic formulation. In the present study, polyherbal formulation comprises of lyophilized hydro-alcoholic (50% v/v extracts of Momordica charantia, Trigonella foenum-graecum and Withania somnifera 2:2:1, respectively, named HA, optimized based on oral glucose tolerance test model in normal Wistar rats. The optimized formulation (HA entrapped in the phosphatidylcholine and cholesterol (8:2 vesicle system is named HA lipids (HAL. The vesicles were characterized for shape, morphology, entrapment efficiency, polar-dispersity index and release profile in the gastric pH. The antidiabetic potential of HA, marketed polyherbal formulation (D-fit and HAL was compared in streptozotocin-induced diabetic rat model of 21 days study. The parameters evaluated were behavioral changes, body weight, serum glucose level, lipid profile and oxidative stress. The antidiabetic potential of HA (1000 mg/kg was at par with the D-fit (1000 mg/kg. However, the potential was enhanced by phospholipids encapsulation; as HAL (500 mg/kg has shown more significant (P < 0.05 potential in comparison to HA (1000 mg/kg and at par with metformin (500 mg/kg.

  14. Immunogenicity of recombinant class 1 protein from Neisseria meningitidis refolded into phospholipid vesicles and detergent.

    Niebla, O; Alvarez, A; Martín, A; Rodríguez, A; Delgado, M; Falcón, V; Guillén, G


    The possibility of eliciting bactericidal antibodies against a recombinant class 1 protein (P1) from Neisseria meningitidis, joined to the first 45 amino acids of the neisserial LpdA protein (PM82), was examined. P1 was produced in Escherichia coli as intracellular inclusion bodies, from which it was purified and reconstituted by (a) inclusion into phospholipid vesicles and detergent and (b) refolding in 0.1% SDS. When Balb/c mice were immunised, high titres of subtype-specific bactericidal antibodies against P1 were obtained in both cases. These results suggest that in spite of being a denaturing agent, it is possible to use SDS to reconstitute the P1 protein in a conformation that exposes the immunodominat regions.

  15. Interactions of Aqueous Imidazolium-Based Ionic Liquid Mixtures with Solid-Supported Phospholipid Vesicles

    Losada-Pérez, Patricia; Khorshid, Mehran; Renner, Frank Uwe


    Despite the environmentally friendly reputation of ionic liquids (ILs), their safety has been recently questioned given their potential as cytotoxic agents. The fundamental mechanisms underlying the interactions between ILs and cells are less studied and by far not completely understood. Biomimetic films are here important biophysical model systems to elucidate fundamental aspects and mechanisms relevant for a large range of biological interaction ranging from signaling to drug reception or toxicity. Here we use dissipative quartz crystal microbalance QCM-D to examine the effect of aqueous imidazolium-based ionic liquid mixtures on solid-supported biomimetic membranes. Specifically, we assess in real time the effect of the cation chain length and the anion nature on a supported vesicle layer of the model phospholipid DMPC. Results indicate that interactions are mainly driven by the hydrophobic components of the IL, which significantly distort the layer and promote vesicle rupture. Our analyses evidence the gradual decrease of the main phase transition temperature upon increasing IL concentration, reflecting increased disorder by weakening of lipid chain interactions. The degree of rupture is significant for ILs with long hydrophobic cation chains and large hydrophobic anions whose behavior is reminiscent of that of antimicrobial peptides. PMID:27684947

  16. Binding of acylated peptides and fatty acids to phospholipid vesicles: pertinence to myristoylated proteins.

    Peitzsch, R M; McLaughlin, S


    We studied the binding of fatty acids and acylated peptides to phospholipid vesicles by making electrophoretic mobility and equilibrium dialysis measurements. The binding energies of the anionic form of the fatty acids and the corresponding acylated glycines were identical; the energies increased by 0.8 kcal/mol per number of carbons in the acyl chain (Ncarbon = 10, 12, 14, 16), a value identical to that for the classical entropy-driven hydrophobic effect discussed by Tanford [The Hydrophobic Effect (1980) Wiley, New York]. The unitary Gibbs free binding energy, delta Gou, of myristoylated glycine, 8 kcal/mol, is independent of the nature of the electrically neutral lipids used to form the vesicles. Similar binding energies were obtained with other myristoylated peptides (e.g., Gly-Ala, Gly-Ala-Ala). The 8 kcal/mol, which corresponds to an effective dissociation constant of 10(-4) M for myristoylated peptides with lipids, provides barely enough energy to attach a myristoylated protein in the cytoplasm to the plasma membrane. Thus, other factors that reduce (e.g., hydrophobic interaction of myristate with the covalently attached protein) or enhance (e.g., electrostatic interactions of basic residues with acidic lipids; protein-protein interactions with intrinsic receptor proteins) the interaction of myristoylated proteins with membranes are likely to be important and may cause reversible translocation of these proteins to the membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

  17. Solubilization of beclomethasone dipropionate in sterically stabilized phospholipid nanomicelles (SSMs: physicochemical and in vitro evaluations

    Peh KK


    Full Text Available Mohanad Naji Sahib, Shaymaa Abdalwahed Abdulameer, Yusrida Darwis, Kok Khiang Peh, Yvonne Tze Fung TanSchool of Pharmaceutical Sciences, Universiti Sains Malaysia, Penang, MalaysiaBackground: The local treatment of lung disorders such as asthma and chronic obstructive pulmonary disease via pulmonary drug delivery offers many advantages over oral or intravenous routes of administration. This is because direct deposition of a drug at the diseased site increases local drug concentrations, which improves the pulmonary receptor occupancy and reduces the overall dose required, therefore reducing the side effects that result from high drug doses. From a clinical point of view, although jet nebulizers have been used for aerosol delivery of water-soluble compounds and micronized suspensions, their use with hydrophobic drugs has been inadequate.Aim: To evaluate the feasibility of sterically stabilized phospholipid nanomicelles (SSMs loaded with beclomethasone dipropionate (BDP as a carrier for pulmonary delivery.Methods: 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-methoxy-poly(ethylene glycol 5000 polymeric micelles containing BDP (BDP-SSMs were prepared by the coprecipitation and reconstitution method, and the physicochemical and in vitro characteristics of BDP-SSMs were investigated.Results: BDP-SSMs were successfully prepared with a content uniformity and reproducibility suitable for pulmonary administration. The maximum solubility of BDP in SSMs was approximately 1300 times its actual solubility. The particle size and zeta potential of BDP-SSMs were 19.89 ± 0.67 nm and -28.03 ± 2.05 mV, respectively. The SSMs system slowed down the release of BDP and all of the aerodynamic values of the aerosolized rehydrated BDP-SSMs were not only acceptable but indicated a significant level of deposition in the lungs.Conclusion: The SSM system might be an effective way of improving the therapeutic index of nebulized, poorly soluble corticosteroids

  18. Molecular interactions between bile salts, phospholipids and cholesterol : relevance to bile formation, cholesterol crystallization and bile salt toxicity

    Moschetta, Antonio


    Cholesterol is a nonpolar lipid dietary constituent, absorbed from the small intestine, transported in blood and taken up by the liver. In bile, the sterol is solubilized in mixed micelles by bile salts and phospholipids. In case of supersaturation, cholesterol is kept in vesicles with phospholipid

  19. Cholesterol affects the interaction between an ionic liquid and phospholipid vesicles. A study by differential scanning calorimetry and nanoplasmonic sensing.

    Russo, Giacomo; Witos, Joanna; Rantamäki, Antti H; Wiedmer, Susanne K


    The present work aims at studying the interactions between cholesterol-rich phosphatidylcholine-based lipid vesicles and trioctylmethylphosphonium acetate ([P8881][OAc]), a biomass dissolving ionic liquid (IL). The effect of cholesterol was assayed by using differential scanning calorimetry (DSC) and nanoplasmonic sensing (NPS) measurement techniques. Cholesterol-enriched dipalmitoyl-phosphatidylcholine vesicles were exposed to different concentrations of the IL, and the derived membrane perturbation was monitored by DSC. The calorimetric data could suggest that the binding and infiltration of the IL are delayed in the vesicles containing cholesterol. To clarify our findings, NPS was applied to quantitatively follow the resistance of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine incorporating 0, 10, and 50mol% of cholesterol toward the IL exposure over time. The membrane perturbation induced by different concentrations of IL was found to be a concentration dependent process on cholesterol-free lipid vesicles. Moreover, our results showed that lipid depletion in cholesterol-enriched lipid vesicles is inversely proportional to the increasing amount of cholesterol in the vesicles. These findings support that cholesterol-rich lipid bilayers are less susceptible toward membrane disrupting agents as compared to membranes that do not incorporate any sterols. This probably occurs because cholesterol tightens the phospholipid acyl chain packing of the plasma membranes, increasing their resistance and reducing their permeability. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Longitudinal diffusion behavior of hemicyanine dyes across phospholipid vesicle membranes as studied by second-harmonic generation and fluorescence spectroscopies.

    Yamaguchi, Akira; Nakano, Masaki; Nochi, Kimihisa; Yamashita, Tomohisa; Morita, Kotaro; Teramae, Norio


    The adsorption and longitudinal diffusion behaviors of a series of hemicyanine dyes to phospholipid vesicle membranes were studied by second-harmonic generation (SHG) and fluorescence spectroscopies. It was observed that the longitudinal diffusion of cationic hemicyanine dyes takes place immediately after the initial adsorption of these dyes to the outer surface of the vesicle membrane. In contrast, hardly any amount of a zwitterionic hemicyanine dye with a sulfonate group diffused across the vesicle membrane within the measurement time (<2000 s). Based on the difference in the time-course responses of SHG and fluorescence spectroscopies for all of the hemicyanine dyes tested, we propose that hydration of the sulfonate group is mainly responsible for the low diffusivity of the zwitterionic hemicyanine dye.

  1. Effect of carbon black nanomaterial on biological membranes revealed by shape of human erythrocytes, platelets and phospholipid vesicles.

    Pajnič, Manca; Drašler, Barbara; Šuštar, Vid; Krek, Judita Lea; Štukelj, Roman; Šimundić, Metka; Kononenko, Veno; Makovec, Darko; Hägerstrand, Henry; Drobne, Damjana; Kralj-Iglič, Veronika


    We studied the effect of carbon black (CB) agglomerated nanomaterial on biological membranes as revealed by shapes of human erythrocytes, platelets and giant phospholipid vesicles. Diluted human blood was incubated with CB nanomaterial and observed by different microscopic techniques. Giant unilamellar phospholipid vesicles (GUVs) created by electroformation were incubated with CB nanomaterial and observed by optical microscopy. Populations of erythrocytes and GUVs were analyzed: the effect of CB nanomaterial was assessed by the average number and distribution of erythrocyte shape types (discocytes, echinocytes, stomatocytes) and of vesicles in test suspensions, with respect to control suspensions. Ensembles of representative images were created and analyzed using computer aided image processing and statistical methods. In a population study, blood of 14 healthy human donors was incubated with CB nanomaterial. Blood cell parameters (concentration of different cell types, their volumes and distributions) were assessed. We found that CB nanomaterial formed micrometer-sized agglomerates in citrated and phosphate buffered saline, in diluted blood and in blood plasma. These agglomerates interacted with erythrocyte membranes but did not affect erythrocyte shape locally or globally. CB nanomaterial agglomerates were found to mediate attractive interaction between blood cells and to present seeds for formation of agglomerate - blood cells complexes. Distortion of disc shape of resting platelets due to incubation with CB nanomaterial was not observed. CB nanomaterial induced bursting of GUVs while the shape of the remaining vesicles was on the average more elongated than in control suspension, indicating indirect osmotic effects of CB nanomaterial. CB nanomaterial interacts with membranes of blood cells but does not have a direct effect on local or global membrane shape in physiological in vitro conditions. Blood cells and GUVs are convenient and ethically acceptable

  2. Smooth/rough layering in liquid-crystalline/gel state of dry phospholipid film, in relation to its ability to generate giant vesicles

    Hishida, M; Yoshikawa, K; Hishida, Mafumi; Seto, Hideki; Yoshikawa, Kenichi


    Morphological changes in a dry phospholipid film on a solid substrate were studied below and above the main transition temperature, between the gel and liquid-crystalline phases by phase-contrast microscopy and AFM. A Phospholipid film in the liquid-crystalline phase exhibits flat, smooth layering, whereas that in the gel phase shows rough, random layering. These film morphologies are discussed in relation to the ability to form giant vesicles through the natural swelling method.

  3. The Regulation of Vesicle Trafficking by Small GTPases and Phospholipids during Pollen Tube Growth

    Polarized and directional growth of pollen tubes is the only means by which immotile sperm of flowering plants reach the deeply embedded female gametes for fertilization. Vesicle trafficking is among the most critical cellular activities for pollen tube growth. Vesicle trafficking maintains membrane...

  4. Faceted phospholipid vesicles tailored for the delivery of Santolina insularis essential oil to the skin.

    Castangia, Ines; Manca, Maria Letizia; Caddeo, Carla; Maxia, Andrea; Murgia, Sergio; Pons, Ramon; Demurtas, Davide; Pando, Daniel; Falconieri, Danilo; Peris, José E; Fadda, Anna Maria; Manconi, Maria


    The aim of this work was to formulate Santolina insularis essential oil-loaded nanocarriers, namely Penetration Enhancer containing Vesicles (PEVs), evaluate the physico-chemical features and stability, and gain insights into their ability to deliver the oil to the skin. S. insularis essential oil was obtained by steam distillation, and was predominantly composed of terpenes, the most abundant being β-phellandrene (22.6%), myrcene (11.4%) and curcumenes (12.1%). Vesicles were prepared using phosphatidylcholine, and ethylene or propylene glycol were added to the water phase (10% (v/v)) to improve vesicle performances as delivery systems. Vesicles were deeply characterized by light scattering, cryogenic transmission electron microscopy and small/wide-angle X-ray scattering, the results showing polyhedral, faceted, unilamellar vesicles of ∼115 nm in size. The presence of the glycols improved vesicle stability under accelerated ageing conditions, without changes in size or migration phenomena (e.g. sedimentation and creaming). Confocal laser scanning microscopy images of pig skin treated with S. insularis formulations displayed a penetration ability of PEVs greater than that of control liposomes. Moreover, all formulations showed a marked in vitro biocompatibility in human keratinocytes. These findings suggest that the nanoformulation may be of value in enhancing the delivery of S. insularis essential oil to the skin, where it can exert its biological activities.

  5. Enthalpy/entropy driven activation of the first interquinone electron transfer in bacterial photosynthetic reaction centers embedded in vesicles of physiologically important phospholipids.

    Milano, Francesco; Dorogi, Márta; Szebényi, Kornélia; Nagy, László; Maróti, Péter; Váró, György; Giotta, Livia; Agostiano, Angela; Trotta, Massimo


    The thermodynamics and kinetics of light-induced electron transfer in bacterial photosynthetic RCs are sensitive to physiologically important lipids (phosphatidylcholine, cardiolipin and phosphatidylglycerol) in the environment. The analysis of the temperature-dependence of the rate of the P(+)Q(A)(-)Q(B)-->P(+)Q(A)Q(B)(-) interquinone electron transfer revealed high enthalpy change of activation in zwitterionic or neutral micelles and vesicles and low enthalpy change of activation in vesicles constituted of negatively charged phospholipids. The entropy change of activation was compensated by the changes of enthalpy, thus the free energy change of activation ( approximately 500 meV) did not show large variation in vesicles of different lipids.

  6. Formation of supported phospholipid bilayers from unilamellar vesicles investigated by atomic force microscopy

    Reviakine, [No Value; Brisson, A


    Since their introduction through the work of McConnell et al. in the early 80s, supported phospholipid bilayers (SPBs) have proven to be a versatile model system for investigating a wide variety of phenomena. Despite their continuous application in fundamental as well as applied research fields, the

  7. Growth and shape transformations of giant phospholipid vesicles upon interaction with an aqueous oleic acid suspension

    Peterlin, Primoz; Kogej, Ksenija; Svetina, Sasa; Walde, Peter


    The interaction of two types of vesicle systems was investigated: micrometer-sized, giant unilamellar vesicles (GUVs) formed from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and submicrometer-sized, large unilamellar vesicles (LUVs) formed from oleic acid and oleate, both in a buffered aqueous solution (pH=8.8). Individual POPC GUVs were transferred with a micropipette into a suspension of oleic acid/oleate LUVs, and the shape changes of the GUVs were monitored using optical microscopy. The behavior of POPC GUVs upon transfer into a 0.8 mM suspension of oleic acid, in which oleic acid/oleate forms vesicular bilayer structures, was qualitatively different from the behavior upon transfer into a 0.3 mM suspension of oleic acid/oleate, in which oleic acid/oleate is predominantly present in the form of monomers and possibly non-vesicular aggregates. In both cases, changes in vesicle morphology were observed within tens of seconds after the transfer. Vesicle initially started to evaginate. In 60% of the...

  8. Inhibition of Sendai virus fusion with phospholipid vesicles and human erythrocyte membranes by hydrophobic peptides

    Kelsey, D.R.; Flanagan, T.D.; Young, J.E.; Yeagle, P.L. (State Univ. of New York, Buffalo (USA))


    Hydrophobic di- and tripeptides which are capable of inhibiting enveloped virus infection of cells are also capable of inhibiting at least three different types of membrane fusion events. Large unilamellar vesicles (LUV) of N-methyl dioleoylphosphatidylethanolamine (N-methyl DOPE), containing encapsulated 1-aminonaphthalene-3,6,8-trisulfonic acid (ANTS) and/or p-xylene bis(pyridinium bromide) (DPX), were formed by extrusion. Vesicle fusion and leakage were then monitored with the ANTS/DPX fluorescence assay. Sendai virus fusion with lipid vesicles and Sendai virus fusion with human erythrocyte membranes were measured by following the relief of fluorescence quenching of virus labeled with octadecylrhodamine B chloride (R18). This study found that the effectiveness of the peptides carbobenzoxy-L-Phe-L-Phe (Z-L-Phe-L-Phe), Z-L-Phe, Z-D-Phe, and Z-Gly-L-Phe-L-Phe in inhibiting N-methyl DOPE LUV fusion or fusion of virus with N-methyl DOPE LUV also paralleled their reported ability to block viral infectivity. Furthermore, Z-D-Phe-L-PheGly and Z-Gly-L-Phe inhibited Sendai virus fusion with human erythrocyte membranes with the same relative potency with which they inhibited vesicle-vesicle and virus-vesicle fusion. The evidence suggests a mechanism by which these peptides exert their inhibition of plaque formation by enveloped viruses. This class of inhibitors apparently acts by inhibiting fusion of the viral envelope with the target cell membrane, thereby preventing viral infection. The physical pathway by which these peptides inhibit membrane fusion was investigated. {sup 31}P nuclear magnetic resonance (NMR) of proposed intermediates in the pathway for membrane fusion in LUV revealed that the potent fusion inhibitor Z-D-Phe-L-PheGly selectively altered the structure (or dynamics) of the hypothesized fusion intermediates and that the poor inhibitor Z-Gly-L-Phe did not.

  9. Effect of engineered TiO2 and ZnO nanoparticles on erythrocytes, platelet-rich plasma and giant unilamelar phospholipid vesicles

    Šimundić Metka


    Full Text Available Abstract Background Massive industrial production of engineered nanoparticles poses questions about health risks to living beings. In order to understand the underlying mechanisms, we studied the effects of TiO2 and ZnO agglomerated engineered nanoparticles (EPs on erythrocytes, platelet-rich plasma and on suspensions of giant unilamelar phospholipid vesicles. Results Washed erythrocytes, platelet-rich plasma and suspensions of giant unilamelar phospholipid vesicles were incubated with samples of EPs. These samples were observed by different microscopic techniques. We found that TiO2 and ZnO EPs adhered to the membrane of washed human and canine erythrocytes. TiO2 and ZnO EPs induced coalescence of human erythrocytes. Addition of TiO2 and ZnO EPs to platelet-rich plasma caused activation of human platelets after 24 hours and 3 hours, respectively, while in canine erythrocytes, activation of platelets due to ZnO EPs occurred already after 1 hour. To assess the effect of EPs on a representative sample of giant unilamelar phospholipid vesicles, analysis of the recorded populations was improved by applying the principles of statistical physics. TiO2 EPs did not induce any notable effect on giant unilamelar phospholipid vesicles within 50 minutes of incubation, while ZnO EPs induced a decrease in the number of giant unilamelar phospholipid vesicles that was statistically significant (p  Conclusions These results indicate that TiO2 and ZnO EPs cause erythrocyte aggregation and could be potentially prothrombogenic, while ZnO could also cause membrane rupture.

  10. A heteronuclear and homonuclear filtering strategy for studying the structure of membrane peptides in non-deuterated phospholipid vesicles

    Doan, B. T.; Nezry, C.; Rene, L.; Badet, B.; Beloeil, J. C.


    NMR study of membrane biomolecules comes up against a poor solubility in classical solvents. A strategy was elaborated to obtain structural information of peptides in non deuterated phospholipids vesicles. It is based on isotopic (HSQC-NOESY) and homonuclear selective filters, both using a fine water suppression. The method is illustrated with the substance P, a 11-residue membrane neuropeptide. L'étude par RMN des biomolécules membranaires est délicate en raison de leur faible solubilité. Une stratégie d'étude a été élaborée pour obtenir des informations structurales de peptides dans un environnement de vésicules de phospholipides non deutérés. Elle repose sur des filtres isotopiques de type HSQC-NOESY et des filtres homonucléaires avec impulsion sélective, avec une suppression fine de l'eau. Un exemple est donné sur un neuropeptide membranaire de 11 résidus : la substance P.

  11. A simple approach to predict the stability of phospholipid vesicles to nebulization without performing aerosolization studies.

    Subramanian, Sneha; Khan, Iftikhar; Korale, Oshadie; Alhnan, Mohamed Albed; Ahmed, Waqar; Najlah, Mohammad; Taylor, Kevin M G; Elhissi, Abdelbary


    Membrane extrusion was investigated for predicting the stability of soya phosphatidylcholine liposomes and surfactosomes (Tween 80-enriched liposomes) to nebulization. Formulations were prepared with or without cholesterol, and salbutamol sulfate (SBS) or beclometasone dipropionate (BDP) were incorporated as model hydrophilic or hydrophobic drugs respectively. Formulations were extruded through 5, 2, 1 and 0.4 μm polycarbonate membrane filters to study the influence of membrane pore size on drug retention by the vesicles. Surfactosomes were found to be very leaky to SBS, such that even without extrusion greater than 50% of the originally entrapped drug was lost; these losses were minimized by the inclusion of cholesterol. The smaller the membrane pore size, the greater the leakage of SBS; hence only around 10% were retained in cholesterol-free surfactosomes extruded through 0.4 μm filters. To study the influence of vesicle size on SBS retained entrapment, an excessive extrusion protocol was proposed (51 extrusion cycles through 1 μm filters) to compare the stability of freshly prepared vesicles (i.e. unextruded; median size approx. 4.5-6.5 μm) with those previously extruded through 1 μm pores. Cholesterol was essential for minimizing losses from liposomes, whilst for surfactosomes size reduction prior to extrusion was the only way to minimize SBS losses which reached up to 93.40% of the originally entrapped drug when no cholesterol was included. When extrusion was applied to BDP-loaded vesicles, greater proportions of the drug were retained in the vesicles compared to SBS. Even with extrusion through 0.4 μm, BDP retention was around 50-60% with little effect of formulation. Excessive extrusion showed BDP retention using small liposomes (1 μm) to be as high as 71-87%, compared to 50-66% for freshly prepared vesicles. The findings, based on extrusion, were compared to studies of vesicle stability to nebulization, published by a range of investigators. It was

  12. Matrix Vesicle Enzyme Activity and Phospholipid Content in Endosteal Bone Following Implantation of Osseointegrating and Non-Osseointegrating Implant Materials.


    matrix vesicles (Schwartz et al., 1990a; Boyan et al., 1988; Peress et al., 1974) and is intimately associated with mineral formation as the principle...directly related to the process of primary mineral formation in tissues adjacent to osseointegrating and non- integrating materials. Of particular...Evans, L. 1981. Mechanisms of mineral formation by matrix vesicles. In: Matrix Vesicles, Proc. 3rd Int. Conf. Matrix Vesicles. Ascenci, A., Bonucci, D

  13. The Lantibiotic Nisin Induces Transmembrane Movement of a Fluorescent Phospholipid

    Moll, Gert N.; Konings, Wil N.; Driessen, Arnold J.M.


    Nisin is a pore-forming antimicrobial peptide. The capacity of nisin to induce transmembrane movement of a fluorescent phospholipid in lipid vesicles was investigated. Unilamellar phospholipid vesicles that contained a fluorescent phospholipid

  14. Size-dependent ultrafast structural dynamics inside phospholipid vesicle bilayers measured with 2D IR vibrational echoes

    Kel, Oksana; Tamimi, Amr; Fayer, Michael D.


    The ultrafast structural dynamics inside the bilayers of dilauroylphosphatidylcholine (DLPC) and dipalmitoylphosphatidylcholine vesicles with 70, 90, and 125 nm diameters were directly measured with 2D IR vibrational echo spectroscopy. The antisymmetric CO stretch of tungsten hexacarbonyl was used as a vibrational probe and provided information on spectral diffusion (structural dynamics) in the alkyl region of the bilayers. Although the CO stretch absorption spectra remain the same, the interior structural dynamics become faster as the size of the vesicles decrease, with the size dependence greater for dipalmitoylphosphatidylcholine than for DLPC. As DLPC vesicles become larger, the interior dynamics approach those of the planar bilayer. PMID:24395796

  15. The Arabidopsis P4-ATPase ALA3 requires a ß-subunit to function in phospholipid translocation and secretory vesicle formation

    Lopez Marques, Rosa Laura

    The Arabidopsis P4-ATPase ALA3 requires a ß-subunit to function in phospholipid translocation and secretory vesicle formation   Lisbeth R. Poulsen1, Rosa L. López-Marqués1, Stephen C. McDowell2, Juha Okkeri3, Dirk Licht3, Alexander Schulz1, Thomas Pomorski3,  Jeffrey F. Harper2, and Michael G...... and in inducing membrane curvature, which is a requirement for vesicle formation. We show that Aminophospholipid ATPase3 (ALA3), a member of the P4-ATPase subfamily in the plant Arabidopsis thaliana, localizes to the Golgi apparatus and that genetic lesions of ALA3 result in impaired growth of roots and shoots....... The root growth defect is accompanied by the failure of the root to release border cells of the root cap. Electron micrograph data suggest that functioning and shedding of border cells are dependent on ALA3, as ala3 mutants are devoid of the characteristic proliferation of slime vesicles at the trans...




    This paper presents a study on the membrane fusion activity of a 23-residue synthetic peptide, representing the N-terminus of gp41 of the human immunodeficiency virus type I (HIV-1; LAV(1a) strain), in a model system involving large unilamellar vesicles (LUV) composed of the negatively charged 1-pal

  17. Thermodynamics and Structural Evolution during a Reversible Vesicle-Micelle Transition of a Vitamin-Derived Bolaamphiphile Induced by Sodium Cholate.

    Tian, Jun-Nan; Ge, Bing-Qiang; Shen, Yun-Feng; He, Yu-Xuan; Chen, Zhong-Xiu


    Interaction of endogenous sodium cholate (SC) with dietary amphiphiles would induce structural evolution of the self-assembled aggregates, which inevitably affects the hydrolysis of fat in the gut. Current work mainly focused on the interaction of bile salts with classical double-layered phospholipid vesicles. In this paper, the thermodynamics and structural evolution during the interaction of SC with novel unilamellar vesicles formed from vitamin-derived zwitterionic bolaamphiphile (DDO) were characterized. It was revealed that an increased temperature and the presence of NaCl resulted in narrowed micelle-vesicle coexistence and enlarged the vesicle region. The coexistence of micelles and vesicles mainly came from the interaction of monomeric SC with DDO vesicles, whereas micellar SC contributed to the total solubilization of DDO vesicles. This research may enrich the thermodynamic mechanism behind the structure transition of the microaggregates formed by amphiphiles in the gut. It will also contribute to the design of food formulation and drug delivery system.

  18. Measurement of the glucose permeation rate across phospholipid bilayers using small unilamellar vesicles. Effect of membrane composition and temperature.

    Bresseleers, G J; Goderis, H L; Tobback, P P


    Small unilamellar vesicles were used to measure the permeability of saturated phosphatidylcholine bilayers to glucose. The presented method circumvents most of the common restriction of classical permeability experiments. Increasing the fatty acid chain length of the lipids reduced the permeation rate significantly. Raising the temperature above that of the lipid phase transition drastically increased membrane permeability. Arrhenius plots demonstrated the activation energy to be independent of membrane composition and the phase-state of the lipids. The permeation process is discussed in terms of a constant energy to disrupt all hydrogen bonds between permeant and aqueous solvent prior to penetrating the membrane. The magnitude of the permeability coefficient is partly determined by a unfavourable change in entropy of activation on crossing the water/lipid interface. All results indicate that the penetration of the dehydrated permeant into the hydrophobic barrier is the rate-limiting step in the permeation of glucose.

  19. First-leaflet phase effect on properties of phospholipid bilayer formed through vesicle adsorption on LB monolayer.

    Park, Jin-Won


    Phospholipid bilayers were formed on mica using the Langmuir-Blodgett technique and liposome fusion, as a model system for biomembranes. Nanometer-scale surface physical properties of the bilayers were quantitatively characterized upon the different phases of the first leaflets. Lower hydration/steric forces on the bilayers were observed at the liquid phase of the first leaflet than at the solid phase. The forces appear to be related to the low mechanical stability of the lipid bilayer, which was affected by the first leaflet phase. The first leaflet phase also influenced the long-range repulsive forces over the second leaflet. Surface forces, measured using a modified probe with an atomic force microscope, showed that lower long-range repulsive forces were also found at the liquid phase of the first leaflet. Force measurements were performed at 300 mM sodium chloride solution so that the effect of the phase on the long-range repulsive forces could be investigated by reducing the effect of the repulsion between the second-leaflet lipid headgroups on the long-range repulsive forces. Forces were analyzed using the Derjaguin-Landau-Verwey-Overbeek theory so that the surface potential and surface charge density of the lipid bilayers were quantitatively acquired for each phase of the first leaflet.

  20. Continuous monitoring of phospholipid vesicle hydrolysis by phospholipase D (PLD) reveals differences in hydrolysis by PLDs from 2 Streptomyces species.

    Hirano, Satomi; Sekine, Kazuhisa; Handa, Tetsurou; Nakano, Minoru


    Phospholipase D (PLD)-mediated hydrolysis of phosphatidylcholine (PC) in large unilamellar vesicles (LUVs) consisting of PC and either glycerol monooleate (GMO) or methyl oleate (MeO) were monitored in situ and in real time by using a choline oxidase-immobilized oxygen electrode. This technique revealed reaction differences between 2 bacterial PLDs. PLD from Streptomyces chromofuscus, which is closely homologous to bacterial alkaline phosphatase, hydrolyzed only 6% of surface PC owing to product inhibition. The catalytic activity of this enzyme was not sensitive to the addition of GMO. On the other hand, typical bacterial PLD from Streptomyces sp. was found to hydrolyze all the PC molecules at the outer surface of LUVs suggesting that this enzyme is free from product inhibition. Introduction of GMO or MeO into the bilayer increased exposure of the PC headgroup and facilitated PC hydrolysis mediated by PLD from Streptomyces sp. GMO and MeO have the same lipophilic tail but the latter lacks hydroxyl groups on its polar head. From kinetic analysis by using the Michaelis-Menten model extended to the reaction at the interface, these compounds were found to activate PLD from Streptomyces sp. in different ways, i.e., MeO increased the protein binding to membranes and GMO stimulated the enzyme-substrate complex formation at membrane surface. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. Staphylococcus aureus interaction with phospholipid vesicles--a new method to accurately determine accessory gene regulator (agr activity.

    Maisem Laabei

    Full Text Available The staphylococcal accessory gene regulatory (agr operon is a well-characterised global regulatory element that is important in the control of virulence gene expression for Staphylococcus aureus, a major human pathogen. Hence, accurate and sensitive measurement of Agr activity is central in understanding the virulence potential of Staphylococcus aureus, especially in the context of Agr dysfunction, which has been linked with persistent bacteraemia and reduced susceptibility to glycopeptide antibiotics. Agr function is typically measured using a synergistic haemolysis CAMP assay, which is believe to report on the level of expression of one of the translated products of the agr locus, delta toxin. In this study we develop a vesicle lysis test (VLT that is specific to small amphipathic peptides, most notably delta and Phenol Soluble Modulin (PSM toxins. To determine the accuracy of this VLT method in assaying Agr activity, we compared it to the CAMP assay using 89 clinical Staphylococcus aureus isolates. Of the 89 isolates, 16 were designated as having dysfunctional Agr systems by the CAMP assay, whereas only three were designated as such by VLT. Molecular analysis demonstrated that of these 16 isolates, the 13 designated as having a functional Agr system by VLT transcribed rnaIII and secreted delta toxin, demonstrating they have a functional Agr system despite the results of the CAMP assay. The agr locus of all 16 isolates was sequenced, and only the 3 designated as having a dysfunctional Agr system contained mutations, explaining their Agr dysfunction. Given the potentially important link between Agr dysfunction and clinical outcome, we have developed an assay that determines this more accurately than the conventional CAMP assay.

  2. The Lantibiotic Nisin Induces Transmembrane Movement of a Fluorescent Phospholipid

    Moll, Gert N.; Konings, Wil N.; Driessen, Arnold J.M.


    Nisin is a pore-forming antimicrobial peptide. The capacity of nisin to induce transmembrane movement of a fluorescent phospholipid in lipid vesicles was investigated. Unilamellar phospholipid vesicles that contained a fluorescent phospholipid (1-acyl-2-{6-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]ca

  3. Vesicle-cholesterol interactions : Effects of added cholesterol on gel-to-liquid crystal transitions in a phospholipid membrane and five dialkyl-based vesicles as monitored using DSC

    Blandamer, Michael J.; Briggs, B; Cullis, PM; Rawlings, BJ; Engberts, Jan B. F. N.; Cullis, Paul M.; Rawlings, Bernard J.


    Differential scanning calorimetry (DSC) results are reported characterising the effects of added cholesterol (CHOL) on vesicle bilayer systems formed in aqueous systems by three sodium dialkylphosphates, (RO)(2)PO2--Na+ where R = dodecyl (DDP), tetradecyl (DTP) and octadecyl (DOP), (ii) two

  4. Implications of surface charge and curvature for the binding orientation of Thermomyces lanuginosus lipase on negatively charged or zwitterionic phospholipid vesicles as studied by ESR spectroscopy

    Hedin, E.M.K.; Høyrup, Lise Pernille Kristine; Patkar, S.A.;


    , Y., et al. (2000) Biochemistry 39, 413-423]. The productive-mode binding orientation of TLL at the lipid-water interface of small unilamellar vesicles (SUV) consisting of 1-palmitoyl-2-oleoyi-sn-glycero-3-phosphati-dylglycerol (POPG) was previously determined using electron spin resonance (ESR......) spectroscopy in combination with site-directed spin-labeling [Hedin, E. M. K., et al. (2002) Biochemistry 41, 1418514196]. In our investigation, we have studied the interfacial orientation of TLL when bound to large unilamellar vesicles (LUV) consisting of POPG, and bound to SUV consisting of 1-palmitoyl-2...... fluorescence quenching efficiency between each spin-label positioned on TLL, and the lipid membrane. ESR exposure and fluorescence quenching data show that TILL associates closer to the negatively charged PG surface than the zwitterionic PC surface, and binds to both POPG LUV and POPC SUV predominantly through...

  5. Characteristics of redox-linked proton ejection in cytochrome c oxidase reconstituted in phospholipid vesicles. New observations support mechanisms different from proton pumping.

    Papa, S; Lorusso, M; Capitanio, N; De Nitto, E


    Experimental observations reveal a number of characteristics of the redox-linked proton ejection from cytochrome c oxidase vesicles, which apparently cannot be explained by a proton pumping activity of the oxidase. These observations seem, on the other hand, to provide useful elements for alternative explanation(s) of the proton ejection. It is proposed here that the process is scalar and not vectorial and can derive from redox-linked rupture of protonated salt-bridges in the oxidase-lipid complex.

  6. Interaction of styrene with DODAB bilayer vesicles. influence on vesicle morphology and bilayer properties

    Jung, M.; Hubert, D.H.W.; Veldhoven, van E.; Frederik, P.M.; Blandamer, M.J.; Briggs, B.; Visser, A.J.W.G.; Herk, van A.M.; German, A.L.


    The solubilization of styrene in large unilamellar DODAB vesicles is investigated at a styrene to DODAB molar ratio of 2:1. The combination of various vesicle characterization methods allows a simultaneous look at vesicle morphology (cryo-TEM, DLS) and molecular interactions (micro-DSC, various fluo

  7. Solubilization of microsomal-associated phosphatidylinositol synthase from germinating soybeans.

    Robinson, M L; Carman, G M


    CDP-1,2-diacyl-sn-glycerol (CDP-diacylglycerol):myo-inositol phosphatidyltransferase (EC, phosphatidylinositol synthase) catalyzes the final step in the de novo synthesis of phosphatidylinositol in the endoplasmic reticulum fraction of germinating soybeans (Glycine max L. var Cutler 71). A variety of solubilization agents were examined for their ability to release phosphatidylinositol synthase activity from the microsome fraction. The most effective agent to solubilize the enzyme was the nonionic detergent Brij W-1. A 2.1-fold increase in specific activity was achieved using 1% Brij W-1 with 69% activity solubilized.Maximal solubilization of phosphatidylinositol synthase was completely dependent on Brij W-1 (1%), potassium ions (0.3 m), and manganese ions (0.5 mm). Solubilization of the enzyme was not affected by the protein concentration of microsomes between 3 to 20 milligrams per milliliter. Solubilization was not affected by the pH of solubilization buffer between 6.5 to 8.5. To our knowledge, this is the first phospholipid biosynthetic enzyme solubilized from plant membranes. The Brij W-1-solubilized phosphatidylinositol synthase remained at the top of a glycerol gradient, whereas the membrane-associated enzyme sedimented to the bottom of the gradient. Maximal activity of the Brij W-1-solubilized phosphatidylinositol synthase was dependent on manganese (5 mm) or magnesium (30 mm) ions, and Triton X-100 (3.6 mm) at pH 8.0 with Tris-HCl buffer. The apparent K(m) values for CDP-diacylglycerol and myo-inositol for the solubilized enzyme was 0.1 mm and 46 mum, respectively. Solubilized phosphatidylinositol synthase activity was thermally inactivated at temperatures above 30 degrees C.

  8. 拉曼光谱研究脉冲电场作用下磷脂脂质体构象的变化%Effect of pulsed electric fields on phospholipid vesicle conformation by Raman spectra

    贾晓; 曾新安


    In order to study the sterilization mechanism of pulsed electric fields(PEF) on microorganisms and the interaction between PEF conditions and cell membrane composed of phospholipids,the phospholipid vesicle conformation changing under various electric field strength treatments was analyzed by Raman spectra.It was demonstrated from the Raman spectra that the intensity ratio of I1085/I1064 increased while I1085/I1123 increased firstly and then decreased later with increasing electric field strength,but the ratio was still higher than the control sample.It indicated a little increase of the gauche/trans population compared with control sample.On the other hand,the ratio of I2885/I2850 decreased with increasing electric field strength.The result showed that the intermolecular disordering in hydrocarbon chain of the lipid lattice increased and the membrane fluidity enhanced after being subjected PEF treatment.%为深入研究脉冲电场的灭菌机理,探讨电场与细胞膜的作用机制,利用拉曼光谱谱图分析了不同电场场强下磷脂脂质体构象的变化。结果表明,拉曼峰值比I1085/I1064随着电场场强的增加而相应增大,而I1085/I1123随着电场场强的增加而先增加后减少,但比值仍高于空白样,说明经过电场处理后,脂质体C-C扭曲构象的数目增多;而I2885/I2850随着场强的增加呈现递减的趋势。由此可以说明经电场处理后的脂质体磷脂分子间烃链的无序度增加,导致细胞膜流动性增强。

  9. Interactions of amelogenin with phospholipids.

    Lokappa, Sowmya Bekshe; Chandrababu, Karthik Balakrishna; Dutta, Kaushik; Perovic, Iva; Evans, John Spencer; Moradian-Oldak, Janet


    Amelogenin protein has the potential to interact with other enamel matrix proteins, mineral, and cell surfaces. We investigated the interactions of recombinant amelogenin rP172 with small unilamellar vesicles as model membranes, toward the goal of understanding the mechanisms of amelogenin-cell interactions during amelogenesis. Dynamic light scattering (DLS), fluorescence spectroscopy, circular dichroism (CD), and nuclear magnetic resonance (NMR) were used. In the presence of phospholipid vesicles, a blue shift in the Trp fluorescence emission maxima of rP172 was observed (∼334 nm) and the Trp residues of rP172 were inaccessible to the aqueous quencher acrylamide. DLS studies indicated complexation of rP172 and phospholipids, although the possibility of fusion of phospholipids following amelogenin addition cannot be ruled out. NMR and CD studies revealed a disorder-order transition of rP172 in a model membrane environment. Strong fluorescence resonance energy transfer from Trp in rP172 to DNS-bound-phospholipid was observed, and fluorescence polarization studies indicated that rP172 interacted with the hydrophobic core region of model membranes. Our data suggest that amelogenin has ability to interact with phospholipids and that such interactions may play key roles in enamel biomineralization as well as reported amelogenin signaling activities.

  10. Computational prediction of solubilizers' effect on partitioning.

    Hoest, Jan; Christensen, Inge T; Jørgensen, Flemming S; Hovgaard, Lars; Frokjaer, Sven


    A computational model for the prediction of solubilizers' effect on drug partitioning has been developed. Membrane/water partitioning was evaluated by means of immobilized artificial membrane (IAM) chromatography. Four solubilizers were used to alter the partitioning in the IAM column. Two types of molecular descriptors were calculated: 2D descriptors using the MOE software and 3D descriptors using the Volsurf software. Structure-property relationships between each of the two types of descriptors and partitioning were established using partial least squares, projection to latent structures (PLS) statistics. Statistically significant relationships between the molecular descriptors and the IAM data were identified. Based on the 2D descriptors structure-property relationships R(2)Y=0. 99 and Q(2)=0.82-0.83 were obtained for some of the solubilizers. The most important descriptor was related to logP. For the Volsurf 3D descriptors models with R(2)Y=0.53-0.64 and Q(2)=0.40-0.54 were obtained using five descriptors. The present study showed that it is possible to predict partitioning of substances in an artificial phospholipid membrane, with or without the use of solubilizers.

  11. Oxidative and other posttranslational modifications in extracellular vesicle biology.

    Szabó-Taylor, Katalin; Ryan, Brent; Osteikoetxea, Xabier; Szabó, Tamás G; Sódar, Barbara; Holub, Marcsilla; Németh, Andrea; Pálóczi, Krisztina; Pállinger, Éva; Winyard, Paul; Buzás, Edit I


    Extracellular vesicles including exosomes, microvesicles and apoptotic vesicles, are phospholipid bilayer surrounded structures secreted by cells universally, in an evolutionarily conserved fashion. Posttranslational modifications such as oxidation, citrullination, phosphorylation and glycosylation play diverse roles in extracellular vesicle biology. Posttranslational modifications orchestrate the biogenesis of extracellular vesicles. The signals extracellular vesicles transmit between cells also often function via modulating posttranslational modifications of target molecules, given that extracellular vesicles are carriers of several active enzymes catalysing posttranslational modifications. Posttranslational modifications of extracellular vesicles can also contribute to disease pathology by e.g. amplifying inflammation, generating neoepitopes or carrying neoepitopes themselves.

  12. Phospholipid bilayer formation at a bare Si surface

    Gutberlet, T.; Steitz, R.; Fragneto, G.


    Neutron reflectivity was applied to monitor in situ the adsorption of small unilamellar phospholipid vesicles on a solid bare hydrophilic Si interface. The obtained reflectivity curves are consistent with the rupture and fusion model for the adsorption of phosphatidylcholine vesicles to solid...

  13. Digestion of phospholipids after secretion of bile into the duodenum changes the phase behavior of bile components.

    Birru, Woldeamanuel A; Warren, Dallas B; Ibrahim, Ahmed; Williams, Hywel D; Benameur, Hassan; Porter, Christopher J H; Chalmers, David K; Pouton, Colin W


    Bile components play a significant role in the absorption of dietary fat, by solubilizing the products of fat digestion. The absorption of poorly water-soluble drugs from the gastrointestinal tract is often enhanced by interaction with the pathways of fat digestion and absorption. These processes can enhance drug absorption. Thus, the phase behavior of bile components and digested lipids is of great interest to pharmaceutical scientists who seek to optimize drug solubilization in the gut lumen. This can be achieved by dosing drugs after food or preferably by formulating the drug in a lipid-based delivery system. Phase diagrams of bile salts, lecithin, and water have been available for many years, but here we investigate the association structures that occur in dilute aqueous solution, in concentrations that are present in the gut lumen. More importantly, we have compared these structures with those that would be expected to be present in the intestine soon after secretion of bile. Phosphatidylcholines are rapidly hydrolyzed by pancreatic enzymes to yield equimolar mixtures of their monoacyl equivalents and fatty acids. We constructed phase diagrams that model the association structures formed by the products of digestion of biliary phospholipids. The micelle-vesicle phase boundary was clearly identifiable by dynamic light scattering and nephelometry. These data indicate that a significantly higher molar ratio of lipid to bile salt is required to cause a transition to lamellar phase (i.e., liposomes in dilute solution). Mixed micelles of digested bile have a higher capacity for solubilization of lipids and fat digestion products and can be expected to have a different capacity to solubilize lipophilic drugs. We suggest that mixtures of lysolecithin, fatty acid, and bile salts are a better model of molecular associations in the gut lumen, and such mixtures could be used to better understand the interaction of drugs with the fat digestion and absorption pathway.

  14. The mechanism of detergent solubilization of liposomes and protein-containing membranes.

    Kragh-Hansen, U; le Maire, M; Møller, J V


    The present study explores intermediate stages in detergent solubilization of liposomes and Ca2+-ATPase membranes by sodium dodecyl sulfate (SDS) and medium-sized ( approximately C12) nonionic detergents. In all cases detergent partitioning in the membranes precedes cooperative binding and solubilization, which is facilitated by exposure to detergent micelles. Nonionic detergents predominantly interact with the lipid component of Ca2+-ATPase membranes below the CMC (critical micellar concentration), whereas SDS extracts Ca2+-ATPase before solubilization of lipid. At the transition to cooperative binding, n-dodecyl octaethylene glycol monoether (C12E8), Triton X-100, and dodecyldimethylamine oxide induce fusion of small unilamellar liposomes to larger vesicles before solubilization. Solubilization of Ca2+-ATPase membranes is accompanied by membrane fragmentation and aggregation rather than vesicle fusion. Detergents with strongly hydrophilic heads (SDS and beta-D-dodecylmaltoside) only very slowly solubilize liposomal membranes and do not cause liposome fusion. These properties are correlated with a slow bilayer flip-flop. Our data suggest that detergent solubilization proceeds by a combination of 1) a transbilayer attack, following flip-flop of detergent molecules across the lipid bilayer, and 2) extraction of membrane components directly by detergent micelles. The present study should help in the design of efficient solubilization protocols, accomplishing the often delicate balance between preserving functional properties of detergent sensitive membrane proteins and minimizing secondary aggregation and lipid content.

  15. Kinetics of solubilization with Triton X-100 of egg-yolk lecithin bilayers containing cholesterol

    Hobai, S


    The titration solubilization of multilamellar egg-yolk lecithin liposomes (MLV-EYL) with Triton X-100 was studied by rectangular optical diffusimetric measurements as a function of cholesterol (Chol) concentration. It was determinated the variation of optic percentage diffu-sion (per mmol surfactant), DDif%/mmol TX-100, in the course of solubilization of MLV-EYL-Chol system with TX-100 10mM. The statistical analysis of the titration curves can reveal the contribution of cholesterol to the stability of phospholipid bilayer membranes. The solubilization of the lecithin-cholesterol mixtures, with a high cholesterol content, much more bile salt requires.

  16. Concentration-Independent Spontaneously Forming Biomimetric Vesicles

    Nieh, M.-P.; Harroun, T. A.; Raghunathan, V. A.; Glinka, C. J.; Katsaras, J.


    In this Letter we present small-angle neutron scattering data from a biomimetic system composed of the phospholipids dimyristoyl and dihexanoyl phosphorylcholine (DMPC and DHPC, respectively). Doping DMPC-DHPC multilamellar vesicles with either the negatively charged lipid dimyristoyl phosphorylglycerol (DMPG, net charge -1) or the divalent cation, calcium (Ca2+), leads to the spontaneous formation of energetically stabilized monodisperse unilamellar vesicles whose radii are concentration independent and in contrast with previous experimental observations.

  17. Solubilizing excipients in oral and injectable formulations.

    Strickley, Robert G


    A review of commercially available oral and injectable solution formulations reveals that the solubilizing excipients include water-soluble organic solvents (polyethylene glycol 300, polyethylene glycol 400, ethanol, propylene glycol, glycerin, N-methyl-2-pyrrolidone, dimethylacetamide, and dimethylsulfoxide), non-ionic surfactants (Cremophor EL, Cremophor RH 40, Cremophor RH 60, d-alpha-tocopherol polyethylene glycol 1000 succinate, polysorbate 20, polysorbate 80, Solutol HS 15, sorbitan monooleate, poloxamer 407, Labrafil M-1944CS, Labrafil M-2125CS, Labrasol, Gellucire 44/14, Softigen 767, and mono- and di-fatty acid esters of PEG 300, 400, or 1750), water-insoluble lipids (castor oil, corn oil, cottonseed oil, olive oil, peanut oil, peppermint oil, safflower oil, sesame oil, soybean oil, hydrogenated vegetable oils, hydrogenated soybean oil, and medium-chain triglycerides of coconut oil and palm seed oil), organic liquids/semi-solids (beeswax, d-alpha-tocopherol, oleic acid, medium-chain mono- and diglycerides), various cyclodextrins (alpha-cyclodextrin, beta-cyclodextrin, hydroxypropyl-beta-cyclodextrin, and sulfobutylether-beta-cyclodextrin), and phospholipids (hydrogenated soy phosphatidylcholine, distearoylphosphatidylglycerol, L-alpha-dimyristoylphosphatidylcholine, L-alpha-dimyristoylphosphatidylglycerol). The chemical techniques to solubilize water-insoluble drugs for oral and injection administration include pH adjustment, cosolvents, complexation, microemulsions, self-emulsifying drug delivery systems, micelles, liposomes, and emulsions.

  18. Loading of Vesicles into Soft Amphiphilic Nanotubes using Osmosis

    Erne, Petra M.; van Bezouwen, Laura S.; Stacko, Peter; van Dtjken, Derk Jan; Chen, Jiawen; Stuart, Marc C. A.; Boekema, Eghert J.; Feringa, Ben L.


    The facile assembly of higher-order nanoarchitectures from simple building blocks is demonstrated by the loading of vesicles into soft amphiphilic nanotubes using osmosis. The nanotubes are constructed from rigid interdigitated bilayers which are capped with vesicles comprising phospholipid-based fl

  19. Degradation of cholesterol crystals in phospholipids

    Koren, Eugen; Koscec, Mirna; Fugate, Robert D.


    Based on previous studies from the laboratory that demonstrated degradation of cholesterol crystals ingested by macrophages in a cell culture system and indicated that intracellular phospholipids could play an important role in mobilization of crystalline cholesterol, the role of each of the three major intracellular phospholipid species in degradation of crystals is further explored. Fluorescently labeled cholesterol crystals are incubated with phospholipids over a period of 5 d. Morphological changes in crystals are monitored using digital imaging fluorescence microscopy, fluorescence redistribution after photobleaching, confocal microscopy, and epifluorescent and phase contrast microscopy. Results clearly demonstrate that all three phospholipids are able to mobilize crystalline cholesterol. However, the mechanisms by which they exert mobilization are different. Sphingomyelin and phosphatidylchloline are found to cause gradual and uniform dissolution of crystals, more or less preserving their original shape. Phosphatidylethanolamine appear to penetrate into the crystal, causing its fragmentation and solubilization. In the mixture of all three phospholipids representing the composition found in macrophages, both of the described mechanisms are working simultaneously.


    N. B. Gupta et al.


    Ethosomes are as novel vesicles in transdermal drug delivery show significant effects of drug penetration through the biological membrane with slight modification of well established drug carrier liposomes. Ethosomes are soft, malleable vesicles composed mainly of phospholipids, ethanol and water. The size of ethosome vesicles can be modulated from tens of nanometer to microns. The ethosomes can be prepared by Hot as well as Cold method. The evaluation parameters of ethosomes include visualiz...

  1. Composition and physical state of phospholipids in calanoid copepods from India and Norway

    Farkas, T.; Storebakken, T.; Bhosle, N.B.

    the adaptation of membrane lipids with seawater temperatures Phospholipid vesicles obtained from the tropic copepods proved more rigid than those from C finmarchicus, as assessed by diphenylhexatriene fluorescence polarization techniques In each case, there were...

  2. Role of phospholipids in endocytosis, phagocytosis, and macropinocytosis.

    Bohdanowicz, Michal; Grinstein, Sergio


    Endocytosis, phagocytosis, and macropinocytosis are fundamental processes that enable cells to sample their environment, eliminate pathogens and apoptotic bodies, and regulate the expression of surface components. While a great deal of effort has been devoted over many years to understanding the proteins involved in these processes, the important contribution of phospholipids has only recently been appreciated. This review is an attempt to collate and analyze the rapidly emerging evidence documenting the role of phospholipids in clathrin-mediated endocytosis, phagocytosis, and macropinocytosis. A primer on phospholipid biosynthesis, catabolism, subcellular distribution, and transport is presented initially, for reference, together with general considerations of the effects of phospholipids on membrane curvature and charge. This is followed by a detailed analysis of the critical functions of phospholipids in the internalization processes and in the maturation of the resulting vesicles and vacuoles as they progress along the endo-lysosomal pathway.

  3. Studying Factors Involved in Biogenesis of Lysobacter sp. XL1 Outer Membrane Vesicles.

    Kudryakova, I V; Suzina, N E; Vinokurova, N G; Shishkova, N A; Vasilyeva, N V


    The Gram-negative bacterium Lysobacter sp. XL1 produces outer membrane vesicles that are heterogeneous in size, density, and protein composition. One of the subpopulations is secretory vesicles for lytic protease L5 of Lysobacter sp. XL1 (Kudryakova et al. (2015) FEMS Microbiol. Lett., 362, fnv137). Protein L5 was assumed to influence biogenesis of these secretory vesicles that contain it. Using a Pseudomonas fluorescens Q2-87/B expression system, it was shown that the recombinant L5 protein may act as a factor of vesicle biogenesis. This points to a possible involvement of L5 protein in Lysobacter sp. XL1 vesicle biogenesis. Furthermore, it was established that the main phospholipid of Lysobacter sp. XL1 vesicles is cardiolipin, and vesicles are formed predominantly of outer membrane regions enriched with this phospholipid. This indicates that cardiolipin participates in biogenesis of all vesicle subpopulations in Lysobacter sp. XL1.

  4. Interaction of Phenol-Soluble Modulins with Phosphatidylcholine Vesicles

    Anthony C. Duong


    Full Text Available Several members of the staphylococcal phenol-soluble modulin (PSM peptide family exhibit pronounced capacities to lyse eukaryotic cells, such as neutrophils, monocytes, and erythrocytes. This is commonly assumed to be due to the amphipathic, α-helical structure of PSMs, giving PSMs detergent-like characteristics and allowing for a relatively non-specific destruction of biological membranes. However, the capacities of PSMs to lyse synthetic phospholipid vesicles have not been investigated. Here, we analyzed lysis of synthetic phosphatidylcholine (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, POPC vesicles by all Staphylococcus aureus and S. epidermidis PSMs. In addition, we investigated the lytic capacities of culture filtrates obtained from different S. aureus PSM deletion mutants toward POPC vesicles. Our results show that all staphylococcal PSMs have phospholipid vesicle-lysing activity and the capacity of S. aureus culture filtrate to lyse POPC vesicles is exclusively dependent on PSMs. Notably, we observed largely differing capacities among PSM peptides to lyse POPC vesicles. Interestingly, POPC vesicle-lytic capacities did not correlate with those previously seen for the lysis of eukaryotic cells. For example, the β-type PSMs were strongly lytic for POPC vesicles, but are known to exhibit only very low lytic capacities toward neutrophils and erythrocytes. Thus our results also suggest that the interaction between PSMs and eukaryotic membranes is more specific than previously assumed, potentially depending on additional structural features of those membranes, such as phospholipid composition or yet unidentified docking molecules.

  5. Interaction of fluorescent phospholipids with cyclodextrins.

    Denz, Manuela; Haralampiev, Ivan; Schiller, Sabine; Szente, Lajos; Herrmann, Andreas; Huster, Daniel; Müller, Peter


    Fluorescent analogs of phospholipids are often employed to investigate the structure and dynamics of lipids in membranes. Some of those studies have used cyclodextrins e.g., to modulate the lipid phase. However, the role of the fluorescence moiety of analogs for the interaction between cyclodextrins and fluorescent lipids has not been investigated so far in detail. Therefore, in the present study the interaction of various fluorescent phospholipid analogs with methylated α-, β- and γ- cyclodextrins was investigated. The analogs differed in their structure, in the length of the fatty acyl chain, in the position of the fluorescence group, and in the attached fluorescence moiety (7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) or dipyrrometheneboron difluoride (BODIPY)). In aqueous buffer, cyclodextrins bind fluorescent lipids disturbing the organization of the analogs. When incorporated into lipid vesicles, analogs are selectively extracted from the membrane upon addition of cyclodextrins. The results show that the interaction of cyclodextrins with fluorescent phospholipids depends on the cyclodextrin species, the fluorescence moiety and the phospholipid structure. The presented data should be of interest for studies using fluorescent phospholipids and cyclodextrins, since the interaction between the fluorescence group and the cyclodextrin may interfere with the process(es) under study.

  6. From Vesicles to Protocells: The Roles of Amphiphilic Molecules

    Yuka Sakuma


    Full Text Available It is very challenging to construct protocells from molecular assemblies. An important step in this challenge is the achievement of vesicle dynamics that are relevant to cellular functions, such as membrane trafficking and self-reproduction, using amphiphilic molecules. Soft matter physics will play an important role in the development of vesicles that have these functions. Here, we show that simple binary phospholipid vesicles have the potential to reproduce the relevant functions of adhesion, pore formation and self-reproduction of vesicles, by coupling the lipid geometries (spontaneous curvatures and the phase separation. This achievement will elucidate the pathway from molecular assembly to cellular life.

  7. Solubilization, purification, and reconstitution of alpha 2 beta 1 isozyme of Na+/K+ -ATPase from caveolae of pulmonary smooth muscle plasma membrane: comparative studies with DHPC, C12E8, and Triton X-100.

    Ghosh, Biswarup; Chakraborti, Tapati; Kar, Pulak; Dey, Kuntal; Chakraborti, Sajal


    We identified alpha(2), alpha(1), and beta(1) isoforms of Na(+)/K(+)-ATPase in caveolae vesicles of bovine pulmonary smooth muscle plasma membrane. The biochemical and biophysical characteristics of the alpha(2)beta(1) isozyme of Na(+)/K(+)-ATPase from caveolae vesicles were studied during solubilization and purification using the detergents 1,2-heptanoyl-sn-phosphatidylcholine (DHPC), poly(oxy-ethylene)8-lauryl ether (C(12)E(8)), and Triton X-100, and reconstitution with the phospholipid dioleoyl-phosphatidylcholine (DOPC). DHPC was determined to be superior to C(12)E(8), whereas C(12)E(8) was better than Triton X-100 in the active enzyme yields and specific activity. Fluorescence studies with DHPC-purified alpha(2)beta(1) isozyme of Na(+)/K(+)-ATPase elicited higher E1Na-E2 K transition compared with that of the C(12)E(8)- and Triton X-100-purified enzyme. The rate of Na(+) efflux in DHPC-DOPC-reconstituted isozyme was higher compared to the C(12)E(8)-DOPC- and Triton X100-DOPC-reconstituted enzyme. Circular dichroism analysis suggests that the DHPC-purified alpha(2)beta(1) isozyme of Na(+)/K(+)-ATPase possessed more organized secondary structure compared to the C(12)E(8)- and Triton X-100-purified isozyme.

  8. Structural analysis of bacteriorhodopsin solubilized by lipid-like phosphocholine biosurfactants with varying micelle concentrations.

    Wang, Xiaoqiang; Huang, Haihong; Sun, Chenghao; Huang, Fang


    Surfactants that can provide a more natural substitute for lipid bilayers are important in the purification and in vitro study of membrane proteins. Here we investigate the structural response of a model membrane protein, bacteriorhodopsin (BR), to phosphocholine biosurfactants. Phosphocholine biosurfactants are a type of biomimetic amphiphile that are similar to phospholipids, in which membrane proteins are commonly embedded. Multiple spectroscopic and zeta potential measurements are employed to characterize the conformational change, secondary and tertiary structure, oligomeric status, surface charge distribution and the structural stability of BR solubilized with phosphocholine biosurfactants of varying tail length. The process of phosphocholine micelle formation is found to facilitate the solubilization of BR, and for long-chain phosphocholines, concentrations much higher than their critical micelle concentrations achieve good solubilization. Phosphocholine biosurfactants are shown to be mild compared with the ionic surfactant SDS or CTAB, and tend to preserve membrane protein structure during solubilization, especially at low micelle concentrations, by virtue of their phospholipid-like zwitterionic head groups. The increase of alkyl chain length is shown to obviously enhance the capability of phosphocholine biosurfactants to stabilize BR. The underlying mechanism for the favorable actions of phosphocholine biosurfactant is also discussed.

  9. Compartmentalization and Transport in Synthetic Vesicles

    Christine eSchmitt


    Full Text Available Nano-scale vesicles have become a popular tool in life sciences. Besides liposomes that are generated from phospholipids of natural origin, polymersomes fabricated of synthetic block copolymers enjoy increasing popularity, as they represent more versatile membrane building blocks that can be selected based on their specific physicochemical properties, like permeability, stability or chemical reactivity.In this review, we focus on the application of simple and nested artificial vesicles in synthetic biology. First, we provide an introduction into the utilization of multi-compartmented vesosomes as compartmentalized nano-scale bioreactors. In the bottom-up development of protocells from vesicular nano-reactors, the specific exchange of pathway intermediates across compartment boundaries represents a bottleneck for future studies. To date, most compartmented bioreactors rely on unspecific exchange of substrates and products. This is either based on changes in permeability of the coblock polymer shell by physicochemical triggers or by the incorporation of unspecific porin proteins into the vesicle membrane. Since the incorporation of membrane transport proteins into simple and nested artificial vesicles offers the potential for specific exchange of substances between subcompartments, it opens new vistas in the design of protocells. Therefore we devote the main part of the review to summarize the technical advances in the use of phospholipids and block copolymers for the reconstitution of membrane proteins.

  10. Focus on Extracellular Vesicles: Development of Extracellular Vesicle-Based Therapeutic Systems

    Shin-ichiro Ohno


    Full Text Available Many types of cells release phospholipid membrane vesicles thought to play key roles in cell-cell communication, antigen presentation, and the spread of infectious agents. Extracellular vesicles (EVs carry various proteins, messenger RNAs (mRNAs, and microRNAs (miRNAs, like a “message in a bottle” to cells in remote locations. The encapsulated molecules are protected from multiple types of degradative enzymes in body fluids, making EVs ideal for delivering drugs. This review presents an overview of the potential roles of EVs as natural drugs and novel drug-delivery systems.

  11. Focus on Extracellular Vesicles: Development of Extracellular Vesicle-Based Therapeutic Systems.

    Ohno, Shin-Ichiro; Drummen, Gregor P C; Kuroda, Masahiko


    Many types of cells release phospholipid membrane vesicles thought to play key roles in cell-cell communication, antigen presentation, and the spread of infectious agents. Extracellular vesicles (EVs) carry various proteins, messenger RNAs (mRNAs), and microRNAs (miRNAs), like a "message in a bottle" to cells in remote locations. The encapsulated molecules are protected from multiple types of degradative enzymes in body fluids, making EVs ideal for delivering drugs. This review presents an overview of the potential roles of EVs as natural drugs and novel drug-delivery systems.

  12. Degradation of cholesterol crystals in macrophages: the role of phospholipids

    Koren, Eugen; Koscec, Mirna; Fugate, Robert D.


    Previous studies from this laboratory demonstrated degradation of cholesterol crystals ingested by macrophages in a cell culture system. Those studies also indicated that intracellular phospholipids could play an important role in mobilization of crystalline cholesterol. The purpose of this study was to further explore the role of each of the three major intracellular phospholipid species in degradation of crystals. Fluorescently labeled cholesterol crystals were incubated with phospholipids over a period of 5 days. Morphological changes in crystals were monitored by the use of digital imaging fluorescence microscopy, fluorescence redistribution after photobleaching, confocal microscopy, as well as epifluorescent and phase contrast microscopy. Results clearly demonstrated that all three phospholipids were able to mobilize crystalline cholesterol; however, mechanisms by which they exerted mobilization were different. Sphingomyelin and phosphatidylcholine were found to cause gradual and uniform dissolution of crystals, more or less preserving their original shape. Phosphatidylethanolamine appeared to penetrate into the crystal, causing its fragmentation and solubilization. In the mixture of all three phospholipids representing the composition found in macrophages, both of the described mechanisms were working simultaneously.

  13. Phospholipid bilayer formation at a bare Si surface

    Gutberlet, T.; Steitz, R.; Fragneto, G.;


    Neutron reflectivity was applied to monitor in situ the adsorption of small unilamellar phospholipid vesicles on a solid bare hydrophilic Si interface. The obtained reflectivity curves are consistent with the rupture and fusion model for the adsorption of phosphatidylcholine vesicles to solid...... interfaces. The results show details of the adsorbed bilayer system at ångström resolution and indicate the presence of a thin ∼6 Å thick water leaflet that separates the bilayer from the Si surface. The resolved structural details provide the basis for further investigation of processes such as adsorption...

  14. Sucrose Solutions as Prospective Medium to Study the Vesicle Structure SAXS and SANS study

    Kiselev, M A; Kisselev, A M; Lombardo, D; Killany, M; Lesieur, S


    The possibility to use sucrose solutions as medium for X-ray and neutron small-angle scattering experiments has been explored for dimyristoylphosphatidylcholine (DMPC) vesicles and mixed DMPC/C_(12)E_(8) aggregates. The influence of sucrose concentration on phospholipid vesicles size and polydispersity has been investigated by complimentary X-ray and neutron scattering. Sucrose solutions decreased vesicle size and polydispersity and increased a contrast between phospholipid membrane and bulk solvent sufficiently for X-rays. 40% sucrose in H2O increased X-ray contrast by up to 10 times compared to pure H2O. The range of sucrose concentration 30%-40% created the best experimental conditions for the X-ray small-angle experiment with phospholipid vesicles.


    N. B. Gupta et al.


    Full Text Available Ethosomes are as novel vesicles in transdermal drug delivery show significant effects of drug penetration through the biological membrane with slight modification of well established drug carrier liposomes. Ethosomes are soft, malleable vesicles composed mainly of phospholipids, ethanol and water. The size of ethosome vesicles can be modulated from tens of nanometer to microns. The ethosomes can be prepared by Hot as well as Cold method. The evaluation parameters of ethosomes include visualization, vesicle size and zeta potential, transition temperature, surface tension activity measurement, vesicle stability, drug content, penetration and permeation studies. Ethosomes have been found to be much more efficient at delivering drug to the skin than either liposomes or hydroalcoholic solution. Thus, it can be a logical conclusion that ethosomal formulation possesses promising future in effective dermal/transdermal delivery of bioactive agents.

  16. EKSTRAKSI DAN FRAKSINASI FOSFOLIPID DARI LIMBAH PENGOLAHAN MINYAK SAWIT [Extraction and Fractionation of Phospholipids from the Waste of Palm Oil Processing

    Teti Estiasih1*


    Full Text Available This research aimed to extract and fractionate phospholipids from the most potential source of phospholipids of the waste of palm oil processing i.e. palm pressed fiber, sludge, and spent bleaching earth (adsorbent. The extraction process was performed by using chloroform-methanol, followed by polar lipid separation by solubilization in choloroform (to eliminate neutral lipid and methanol (to extract phospholipids. Fractionation was performed based on the solubility in ethanol and acetone. There were 5 phospholipd fractions obtained, i.e. crude phospholipids (without fractionation, ethanol soluble, ethanol insloluble, ethanol soluble-acetone soluble, and ethanol soluble-acetone insoluble fractions. Each fraction was characterized for phospholipid composition and fatty acid profile. The result showed that palm pressed fiber had the highest content of phospholipids of 10,222.19 ppm. Extraction and separation of phospholipids from palm pressed fiber produced phospholipids with a purity of 61.67%. Phospholipids found in palm pressed fiber were phosphatidylinositol (PI, phosphatidylcholine (PC, phosphatidyletanolamin (PE, phosphatidylglycerol (PG, diphosphatidylglyerol (DPG, and phosphatidic acid (PA, while the impurities consisted of fatty acids and neutral lipids. Oleic acid is the most dominant fatty acid ini crude palm pressed fiber phospholipids. Fractionation increased the purity of phospholipids. Different phospholipids fractions showed different phospholipid composition and fatty acid profile. Ethanol soluble fraction had more PI, PC, PG, and PA, but less PE and DPG as compared to crude phospholipids. The ratio of PC to PE also increased after ethanol fractionation. The amount of more non polar phospholipids (PG, DPG, PA were less in ethanol soluble-acetone insoluble fractions compared to ethanol soluble-acetone soluble fraction. The solubilization in ethanol increased total unsaturated fatty acids in ethanol soluble fractions, meanwhile the

  17. Anionic phospholipids modulate peptide insertion into membranes.

    Liu, L P; Deber, C M


    While the insertion of a hydrophobic peptide or membrane protein segment into the bilayer can be spontaneous and driven mainly by the hydrophobic effect, anionic lipids, which comprise ca. 20% of biological membranes, provide a source of electrostatic attractions for binding of proteins/peptides into membranes. To unravel the interplay of hydrophobicity and electrostatics in the binding of peptides into membranes, we designed peptides de novo which possess the typical sequence Lys-Lys-Ala-Ala-Ala-X-Ala-Ala-Ala-Ala-Ala-X-Ala-Ala-Trp-Ala-Ala-X-Ala-Al a-Ala-Lys-Lys-Lys-Lys-amide, where X residues correspond to "guest" residues which encompass a range of hydrophobicity (Leu, Ile, Gly, and Ser). Circular dichroism spectra demonstrated that peptides were partially (40-90%) random in aqueous buffer but were promoted to form 100% alpha-helical structures by anionic lipid micelles. In neutral lipid micelles, only the relatively hydrophobic peptides (X = L and I) spontaneously adopted the alpha-helical conformation, but when 25% of negatively charged lipids were mixed in to mimic the content of anionic lipids in biomembranes, the less hydrophobic (X = S and G) peptides then formed alpha-helical conformations. Consistent with these findings, fluorescence quenching by the aqueous-phase quencher iodide indicated that in anionic (dimyristoylphosphatidylglycerol) vesicles, the peptide Trp residue was buried in the lipid vesicle hydrophobic core, while in neutral (dimyristoylphosphatidylcholine) vesicles, only hydrophobic (X = L and I) peptides were shielded from the aqueous solution. Trp emission spectra of peptides in the presence of phospholipids doxyl-labeled at the 5-, 7-, 10-, 12-, and 16-fatty acid positions implied not only a transbilayer orientation for inserted peptides but also that mixed peptide populations (transbilayer + surface-associated) may arise. Overall results suggest that for hydrophobic peptides with segmental threshold hydrophobicity below that which

  18. Nonionic Microemulsions as Solubilizers of Hydrophobic Drugs: Solubilization of Paclitaxel

    Jen-Ting Lo


    Full Text Available The strategy using nonionic microemulsion as a solubilizer for hydrophobic drugs was studied and is demonstrated in this work. The aqueous phase behaviors of mixed nonionic surfactants with various oils at 37 °C are firstly constructed to give the optimal formulations of nonionic microemulsions with applications in the enhanced solubilization of the model hydrophobic drug, paclitaxel, at 37 °C. Briefly, the suitable oil phase with paclitaxel significantly dissolved is microemulsified with appropriate surfactants. Surfactants utilized include Tween 80, Cremophor EL, and polyethylene glycol (4.3 cocoyl ether, while various kinds of edible oils and fatty esters are used as the oil phase. On average, the apparent solubility of paclitaxel is increased to ca. 70–100 ppm in the prepared microemulsions at 37 °C using tributyrin or ethyl caproate as the oil phases. The sizes of the microemulsions attained are mostly from ca. 60 nm to ca. 200 nm. The cytotoxicity of the microemulsion formulations is assessed with the cellular viability of 3T3 cells. In general, the cell viability is above 55% after 24 h of cultivation in media containing these microemulsion formulations diluted to a concentration of total surfactants equal to 50 ppm and 200 ppm.

  19. Magainin 2a - Induced Permeabilization of Phospholipid Vesicles


    Dr. Hao- Chia Chen Dr. Robert W. Williams Dr. Noreen Williams for his guidance, advice, positive approach to research, and detailed review the interaction of magainin peptides with membranes. An assay which measures the release of the fluores - cent dye, 6CF from 6CF-loaded SUVs

  20. Entrapment in phospholipid vesicles quenches photoactivity of quantum dots

    Generalov R; Kavaliauskiene S; Westrøm S; Chen W; Kristensen S; Juzenas P


    Roman Generalov1,2, Simona Kavaliauskiene1, Sara Westrøm1, Wei Chen3, Solveig Kristensen2, Petras Juzenas11Department of Radiation Biology, Institute for Cancer Research, Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway; 2School of Pharmacy, University of Oslo, Oslo, Norway; 3Department of Physics, The University of Texas at Arlington, Arlington, TX, USAAbstract: Quantum dots have emerged with great promise for biological applications as fluorescent markers for imm...

  1. A novel isolation strategy for obtaining crude membrane vesicles from bovine skim milk

    Blans, Kristine; Larsen, Lotte Bach; Wiking, Lars


    Bovine milks content of phospholipid membranes have largely been explored in the cream fraction, and known as the milk fat globule membrane that surrounds fat droplets. In skim milk, the population of phospholipid membranes is reported to constitute membrane vesicles with a soluble content known...... components. Here we present a novel strategy for a short, gentle and non-denaturing isolation of skim milk-derived membrane vesicles. Methods: Untreated fresh bovine milk was defatted to remove milk fat globules. The resulting skim milk was subjected to ultracentrifugation. The resulting ochre...... fraction can be obtained from skim milk by ultracentrifugation. Casein micelle remnants as well as smaller protein components in the crude vesicle fraction can be successfully removed by size chromatography. Electron microscopy of the vesicle isolate reveals circular structures with membrane vesicle...

  2. A novel isolation strategy for obtaining crude membrane vesicles from bovine skim milk

    Blans, Kristine; Larsen, Lotte Bach; Wiking, Lars

    Bovine milks content of phospholipid membranes have largely been explored in the cream fraction, and known as the milk fat globule membrane that surrounds fat droplets. In skim milk, the population of phospholipid membranes is reported to constitute membrane vesicles with a soluble content known...... components. Here we present a novel strategy for a short, gentle and non-denaturing isolation of skim milk-derived membrane vesicles. Methods: Untreated fresh bovine milk was defatted to remove milk fat globules. The resulting skim milk was subjected to ultracentrifugation. The resulting ochre...... fraction can be obtained from skim milk by ultracentrifugation. Casein micelle remnants as well as smaller protein components in the crude vesicle fraction can be successfully removed by size chromatography. Electron microscopy of the vesicle isolate reveals circular structures with membrane vesicle...

  3. Structure and organization of phospholipid/polysaccharide nanoparticles

    Gerelli, Y; Bari, M T Di; Deriu, A [Dipartimento di Fisica and CNISM, Universita degli Studi di Parma and CRS SOFT, INFM-CNR (Italy); Cantu, L [Dipartimento di Chimica, Biochimica e Biotecnologie per la Medicina-LITA, Universita di Milano (Italy); Colombo, P; Como, C; Motta, S; Sonvico, F [Dipartimento Farmaceutico, Universita degli Studi di Parma (Italy); May, R [Institut Laue-Langevin, Grenoble (France)], E-mail:


    In recent years nanoparticles and microparticles composed of polymeric or lipid material have been proposed as drug carriers for improving the efficacy of encapsulated drugs. For the production of these systems different materials have been proposed, among them phospholipids and polysaccharides due to their biocompatibility, biodegradability, low cost and safety. We report here a morphological and structural investigation, performed using cryo-TEM, static light scattering and small angle neutron and x-ray scattering, on phospholipid/saccharide nanoparticles loaded with a lipophilic positively charged drug (tamoxifen citrate) used in breast cancer therapy. The lipid component was soybean lecithin; the saccharide one was chitosan that usually acts as an outer coating increasing vesicle stability. The microscopy and scattering data indicate the presence of two distinct nanoparticle families: uni-lamellar vesicles with average radius 90 A and multi-lamellar vesicles with average radius 440 A. In both families the inner core is occupied by the solvent. The presence of tamoxifen gives rise to a multi-lamellar structure of the lipid outer shell. It also induces a positive surface charge into the vesicles, repelling the positively charged chitosan molecules which therefore do not take part in nanoparticle formation.

  4. Theoretical Assessment of Fluorinated Phospholipids in the Design of Liposomal Drug-Delivery Systems

    Madsen, Jesper J.united st; Fristrup, Peter; Peters, Günther H.J.


    Fluorinated phospholipid analogues are investigated as potential substrates for phospholipase A(2) (PLA(2)) using classical molecular dynamics simulations and quantum mechanics/density functional theory calculations. The fluorinated phospholipid analogues are a-fluoro (HF-ProAEL) and alpha......,alpha-difluoro (F-2-ProAEL) conjugates of (R)-1-O-hexadecyl-2-palmitoyl-sn-glycero-3-phoshocholineglycerol (ProAEL). Our results provide a theoretical assessment of the potential usefulness of these fluorinated lipids in the rational design of liposomal drug-delivery systems. The a-fluorine-substituted phospholipid...... at a progressively faster rate; the more electronegative substituent at the a-position effectively lowers the energy barrier for hydrolysis. We conclude that the partially fluorinated phospholipid analogues facilitate rational design of liposomal vesicles of phospholipid mixtures with desirable physicochemical...


    LI Zichen; XIE Ximng; FAN Qinghua; FANG Yifei


    α-Eleostearic acid and β-eleostearic acid formed vesicles in aqueous medium when an ethanol solutionofeleostearic acid was injected rapidly into a vigorously vortexed aqueous phase. Formation of the vesicles was demonstrated by electron microscopic observation and bromothymol blue encapsulation experiments. Polymerizations of the eleostearic acids in the formed vesicles carried out by UV irradiation produced poly-α-eleostearic acid and poly-β-eleostearic acid vesicles.

  6. Initiation of waves in BZ encapsulated vesicles using light - towards design of computing architectures

    Costello, Ben de Lacy; Ahearn, Matt; Holley, Julian; Bull, Larry; Adamatzky, Andrew


    A gas free analogue of the Belousov-Zhabotinsky reaction catalysed by ferroin and encapsulated in phospholipid stabilised vesicles is reported. A reaction mixture which exhibits spontaneous oscillation and excitation transfer between vesicles was formulated. By adjusting the reagent concentrations a quiescent state with fewer spontaneous oscillations was achieved. Using relatively low power laser sources of specific wavelengths (green 532nm and blue 405nm) it was shown that waves could be reproducibly initiated within the BZ vesicles. Furthermore, despite the reduced excitability of the system overall the initiated waves exhibited vesicle to vesicle transfer. It was possible to manipulate single vesicles and design simple circuits based on a 2D validation of collision based circuits. Therefore, we conclude that this BZ system exhibits promise for computing applications based on 3D networks of vesicles.

  7. Phospholipid Complex Technique for Superior Bioavailability of Phytoconstituents

    Kattamanchi Gnananath


    Full Text Available Phytoconstituents have been utilized as medicines for thousands of years, yet their application is limited owing to major hurdles like deficit lipid solubility, large molecular size and degradation in the gastric environment of gut. Recently, phospholipid-complex technique has unveiled in addressing these stumbling blocks either by enhancing the solubilizing capacity or its potentiating ability to pass through the biological membranes and it also protects the active herbal components from degradation. Hence, this phospholipid-complex-technique can enable researchers to deliver the phytoconstituents into systemic circulation by using certain conventional dosage forms like tablets and capsules. This review highlights the unique property of phospholipids in drug delivery, their role as adjuvant in health benefits, and their application in the herbal medicine systems to improve the bioavailability of active herbal components. Also we summarize the prerequisites for phytosomes preparation like the selection of type of phytoconstituents, solvents used, various methods employed in phytosomal preparation and its characterization. Further we discuss the key findings of recent research work conducted on phospholipid-based delivery systems which can enable new directions and advancements to the development of herbal dosage forms.

  8. Maize endophytic bacteria as mineral phosphate solubilizers.

    de Abreu, C S; Figueiredo, J E F; Oliveira, C A; Dos Santos, V L; Gomes, E A; Ribeiro, V P; Barros, B A; Lana, U G P; Marriel, I E


    In the present study, we demonstrated the in vitro activity of endophytic phosphate-solubilizing bacteria (PSB). Fifty-five endophytic PSB that were isolated from sap, leaves, and roots of maize were tested for their ability to solubilize tricalcium phosphate and produce organic acid. Partial sequencing of the 16S rRNA-encoding gene showed that the isolates were from the genus Bacillus and different species of Enterobacteriaceae. The phosphate solubilization index on solid medium and phosphate solubilization in liquid medium varied significantly among the isolates. There was a statistically significant difference (P ≤ 0.05) for both, the values of phosphate-solubilizing activity and pH of the growth medium, among the isolates. Pearson correlation was statistically significant (P ≤ 0.05) between P-solubilization and pH (R = -0.38), and between the gluconic acid production and the lowering of the pH of the liquid medium at 6 (R = 0.28) and 9 days (R = 0.39). Gluconic acid production was prevalent in all the PSB studied, and Bacillus species were most efficient in solubilizing phosphate. This is the first report on the characterization of bacterial endophytes from maize and their use as potential biofertilizers. In addition, this may provide an alternative strategy for improving the phosphorus acquisition efficiency of crop plants in tropical soils.

  9. Phospholipids and sports performance

    Purpura Martin


    Full Text Available Abstract Phospholipids are essential components of all biological membranes. Phosphatidylcholine (PC and Phosphatidylserine (PS are Phosphatidyl-phospholipids that are required for normal cellular structure and function. The participation in physical activity often challenges a variety of physiological systems; consequently, the ability to maintain normal cellular function during activity can determine sporting performance. The participation in prolonged intense exercise has been shown to reduce circulatory choline concentrations in some individuals. As choline is a pre-cursor to the neurotransmitter Acetylcholine, this finding has encouraged researchers to investigate the hypothesis that supplementation with PC (or choline salts could enhance sporting performance. Although the available data that evaluates the effects of PC supplementation on performance are equivocal, acute oral supplementation with PC (~0.2 g PC per kg body mass has been demonstrated to improve performance in a variety of sporting activities where exercise has depleted circulatory choline concentrations. Short term oral supplementation with soy-derived PS (S-PS has been reported to attenuate circulating cortisol concentrations, improve perceived well-being, and reduce perceived muscle soreness after exercise. More recently, short term oral supplementation (750 mg per day of S-PS for 10 days has been demonstrated to improve exercise capacity during high intensity cycling and tended to increase performance during intermittent running. Although more research is warranted to determine minimum dietary Phospholipid requirements for optimal sporting performance, these findings suggest that some participants might benefit from dietary interventions that increase the intakes of PC and PS.

  10. Investigating how vesicle size influences vesicle adsorption on titanium oxide: a competition between steric packing and shape deformation.

    Ferhan, Abdul Rahim; Jackman, Joshua A; Cho, Nam-Joon


    Understanding the adsorption behavior of lipid vesicles at solid-liquid interfaces is important for obtaining fundamental insights into soft matter adsorbates as well as for practical applications such as supported lipid bilayer (SLB) fabrication. While the process of SLB formation has been highly scrutinized, less understood are the details of vesicle adsorption without rupture, especially at high surface coverages. Herein, we tackle this problem by employing simultaneous quartz crystal microbalance-dissipation (QCM-D) and localized surface plasmon resonance (LSPR) measurements in order to investigate the effect of vesicle size (84-211 nm diameter) on vesicle adsorption onto a titanium oxide surface. Owing to fundamental differences in the measurement principles of the two techniques as well as a mismatch in probing volumes, it was possible to determine both the lipid mass adsorbed near the sensor surface as well as the total mass of adsorbed lipid and hydrodynamically coupled solvent in the adsorbed vesicle layer as a whole. With increasing vesicle size, the QCM-D frequency signal exhibited monotonic behavior reaching an asymptotic value, whereas the QCM-D energy dissipation signal continued to increase according to the vesicle size. In marked contrast, the LSPR-tracked lipid mass near the sensor surface followed a parabolic trend, with the greatest corresponding measurement response occurring for intermediate-size vesicles. The findings reveal that the maximum extent of adsorbed vesicles contacting a solid surface occurs at an intermediate vesicle size due to the competing influences of vesicle deformation and steric packing. Looking forward, such information can be applied to control the molecular self-assembly of phospholipid assemblies as well as provide the basis for investigating deformable, soft matter adsorbates.

  11. A bola-phospholipid bearing tetrafluorophenylazido chromophore as a promising lipid probe for biomembrane photolabeling studies.

    Xia, Yi; Sengupta, Kheya; Maggiani, Alain; Qu, Fanqi; Peng, Ling


    A bola-phospholipid probe, carrying a tetrafluorophenylazido chromophore in the middle of the transmembrane diacyl chain, was synthesized and characterized with a view to studying biomembranes by a photolabeling approach. This probe shows the advantageous stability of bola-lipids in giant vesicle formation alongside excellent photochemical properties conferred by the tetrafluorophenylazido chromophore, and thus constitutes a promising probe for biomembrane photolabeling studies.

  12. DNA-mediated self-assembly of artificial vesicles.

    Maik Hadorn

    Full Text Available BACKGROUND: Although multicompartment systems made of single unilamellar vesicles offer the potential to outperform single compartment systems widely used in analytic, synthetic, and medical applications, their use has remained marginal to date. On the one hand, this can be attributed to the binary character of the majority of the current tethering protocols that impedes the implementation of real multicomponent or multifunctional systems. On the other hand, the few tethering protocols theoretically providing multicompartment systems composed of several distinct vesicle populations suffer from the readjustment of the vesicle formation procedure as well as from the loss of specificity of the linking mechanism over time. METHODOLOGY/PRINCIPAL FINDINGS: In previous studies, we presented implementations of multicompartment systems and resolved the readjustment of the vesicle formation procedure as well as the loss of specificity by using linkers consisting of biotinylated DNA single strands that were anchored to phospholipid-grafted biotinylated PEG tethers via streptavidin as a connector. The systematic analysis presented herein provides evidences for the incorporation of phospholipid-grafted biotinylated PEG tethers to the vesicle membrane during vesicle formation, providing specific anchoring sites for the streptavidin loading of the vesicle membrane. Furthermore, DNA-mediated vesicle-vesicle self-assembly was found to be sequence-dependent and to depend on the presence of monovalent salts. CONCLUSIONS/SIGNIFICANCE: This study provides a solid basis for the implementation of multi-vesicle assemblies that may affect at least three distinct domains. (i Analysis. Starting with a minimal system, the complexity of a bottom-up system is increased gradually facilitating the understanding of the components and their interaction. (ii Synthesis. Consecutive reactions may be implemented in networks of vesicles that outperform current single compartment

  13. Characterization of phospholipids in insulin secretory granules and mitochondria in pancreatic beta cells and their changes with glucose stimulation.

    MacDonald, Michael J; Ade, Lacmbouh; Ntambi, James M; Ansari, Israr-Ul H; Stoker, Scott W


    The lipid composition of insulin secretory granules (ISG) has never previously been thoroughly characterized. We characterized the phospholipid composition of ISG and mitochondria in pancreatic beta cells without and with glucose stimulation. The phospholipid/protein ratios of most phospholipids containing unsaturated fatty acids were higher in ISG than in whole cells and in mitochondria. The concentrations of negatively charged phospholipids, phosphatidylserine, and phosphatidylinositol in ISG were 5-fold higher than in the whole cell. In ISG phosphatidylserine, phosphatidylinositol, phosphatidylethanolamine, and sphingomyelin, fatty acids 12:0 and 14:0 were high, as were phosphatidylserine and phosphatidylinositol containing 18-carbon unsaturated FA. With glucose stimulation, the concentration of many ISG phosphatidylserines and phosphatidylinositols increased; unsaturated fatty acids in phosphatidylserine increased; and most phosphatidylethanolamines, phosphatidylcholines, sphingomyelins, and lysophosphatidylcholines were unchanged. Unsaturation and shorter fatty acid length in phospholipids facilitate curvature and fluidity of membranes, which favors fusion of membranes. Recent evidence suggests that negatively charged phospholipids, such as phosphatidylserine, act as coupling factors enhancing the interaction of positively charged regions in SNARE proteins in synaptic or secretory vesicle membrane lipid bilayers with positively charged regions in SNARE proteins in the plasma membrane lipid bilayer to facilitate docking of vesicles to the plasma membrane during exocytosis. The results indicate that ISG phospholipids are in a dynamic state and are consistent with the idea that changes in ISG phospholipids facilitate fusion of ISG with the plasma membrane-enhancing glucose-stimulated insulin exocytosis.

  14. Mammalian P4-ATPases and ABC transporters and their role in phospholipid transport.

    Coleman, Jonathan A; Quazi, Faraz; Molday, Robert S


    Transport of phospholipids across cell membranes plays a key role in a wide variety of biological processes. These include membrane biosynthesis, generation and maintenance of membrane asymmetry, cell and organelle shape determination, phagocytosis, vesicle trafficking, blood coagulation, lipid homeostasis, regulation of membrane protein function, apoptosis, etc. P(4)-ATPases and ATP binding cassette (ABC) transporters are the two principal classes of membrane proteins that actively transport phospholipids across cellular membranes. P(4)-ATPases utilize the energy from ATP hydrolysis to flip aminophospholipids from the exocytoplasmic (extracellular/lumen) to the cytoplasmic leaflet of cell membranes generating membrane lipid asymmetry and lipid imbalance which can induce membrane curvature. Many ABC transporters play crucial roles in lipid homeostasis by actively transporting phospholipids from the cytoplasmic to the exocytoplasmic leaflet of cell membranes or exporting phospholipids to protein acceptors or micelles. Recent studies indicate that some ABC proteins can also transport phospholipids in the opposite direction. The importance of P(4)-ATPases and ABC transporters is evident from the findings that mutations in many of these transporters are responsible for severe human genetic diseases linked to defective phospholipid transport. This article is part of a Special Issue entitled Phospholipids and Phospholipid Metabolism.

  15. Nonenzymatic biomimetic remodeling of phospholipids in synthetic liposomes.

    Brea, Roberto J; Rudd, Andrew K; Devaraj, Neal K


    Cell membranes have a vast repertoire of phospholipid species whose structures can be dynamically modified by enzymatic remodeling of acyl chains and polar head groups. Lipid remodeling plays important roles in membrane biology and dysregulation can lead to disease. Although there have been tremendous advances in creating artificial membranes to model the properties of native membranes, a major obstacle has been developing straightforward methods to mimic lipid membrane remodeling. Stable liposomes are typically kinetically trapped and are not prone to exchanging diacylphospholipids. Here, we show that reversible chemoselective reactions can be harnessed to achieve nonenzymatic spontaneous remodeling of phospholipids in synthetic membranes. Our approach relies on transthioesterification/acyl shift reactions that occur spontaneously and reversibly between tertiary amides and thioesters. We demonstrate exchange and remodeling of both lipid acyl chains and head groups. Using our synthetic model system we demonstrate the ability of spontaneous phospholipid remodeling to trigger changes in vesicle spatial organization, composition, and morphology as well as recruit proteins that can affect vesicle curvature. Membranes capable of chemically exchanging lipid fragments could be used to help further understand the specific roles of lipid structure remodeling in biological membranes.

  16. Formation and size distribution of self-assembled vesicles

    Huang, Changjin; Quinn, David; Suresh, Subra


    When detergents and phospholipid membranes are dispersed in aqueous solutions, they tend to self-assemble into vesicles of various shapes and sizes by virtue of their hydrophobic and hydrophilic segments. A clearer understanding of such vesiculation processes holds promise for better elucidation of human physiology and disease, and paves the way to improved diagnostics, drug development, and drug delivery. Here we present a detailed analysis of the energetics and thermodynamics of vesiculation by recourse to nonlinear elasticity, taking into account large deformation that may arise during the vesiculation process. The effects of membrane size, spontaneous curvature, and membrane stiffness on vesiculation and vesicle size distribution were investigated, and the critical size for vesicle formation was determined and found to compare favorably with available experimental evidence. Our analysis also showed that the critical membrane size for spontaneous vesiculation was correlated with membrane thickness, and further illustrated how the combined effects of membrane thickness and physical properties influenced the size, shape, and distribution of vesicles. These findings shed light on the formation of physiological extracellular vesicles, such as exosomes. The findings also suggest pathways for manipulating the size, shape, distribution, and physical properties of synthetic vesicles, with potential applications in vesicle physiology, the pathobiology of cancer and other diseases, diagnostics using in vivo liquid biopsy, and drug delivery methods. PMID:28265065

  17. Fungal rock phosphate solubilization using sugarcane bagasse.

    Mendes, Gilberto O; Dias, Carla S; Silva, Ivo R; Júnior, José Ivo Ribeiro; Pereira, Olinto L; Costa, Maurício D


    The effects of different doses of rock phosphate (RP), sucrose, and (NH(4))(2)SO(4) on the solubilization of RP from Araxá and Catalão (Brazil) by Aspergillus niger, Penicillium canescens, Eupenicillium ludwigii, and Penicillium islandicum were evaluated in a solid-state fermentation (SSF) system with sugarcane bagasse. The factors evaluated were combined following a 2(3) + 1 factorial design to determine their optimum concentrations. The fitted response surfaces showed that higher doses of RP promoted higher phosphorus (P) solubilization. The addition of sucrose did not have effects on P solubilization in most treatments due to the presence of soluble sugars in the bagasse. Except for A. niger, all the fungi required high (NH(4))(2)SO(4) doses to achieve the highest level of P solubilization. Inversely, addition of (NH(4))(2)SO(4) was inhibitory to P solubilization by A. niger. Among the fungi tested, A. niger stood out, showing the highest solubilization capacity and for not requiring sucrose or (NH(4))(2)SO(4) supplementation. An additional experiment with A. niger showed that the content of soluble P can be increased by adding higher RP doses in the medium. However, P yield decreases with increasing RP doses. In this experiment, the maximal P yield (approximately 60 %) was achieved with the lower RP dose (3 g L(-1)). Our results show that SSF can be used to obtain a low cost biofertilizer rich in P combining RP, sugarcane bagasse, and A. niger. Moreover, sugarcane bagasse is a suitable substrate for SSF aiming at RP solubilization, since this residue can supply the C and N necessary for the metabolism of A. niger within a range that favors RP solubilization.

  18. Microencapsulation technology by nature: Cell derived extracellular vesicles with therapeutic potential.

    Kittel, A; Falus, A; Buzás, E


    Cell derived extracellular vesicles are submicron structures surrounded by phospholipid bilayer and released by both prokaryotic and eukaryotic cells. The sizes of these vesicles roughly fall into the size ranges of microbes, and they represent efficient delivery platforms targeting complex molecular information to professional antigen presenting cells. Critical roles of these naturally formulated units of information have been described in many physiological and pathological processes. Extracellular vesicles are not only potential biomarkers and possible pathogenic factors in numerous diseases, but they are also considered as emerging therapeutic targets and therapeutic vehicles. Strikingly, current drug delivery systems, designed to convey therapeutic proteins and peptides (such as liposomes), show many similarities to extracellular vesicles. Here we review some aspects of therapeutic implementation of natural, cell-derived extracellular vesicles in human diseases. Exploration of molecular and functional details of extracellular vesicle release and action may provide important lessons for the design of future drug delivery systems.

  19. Single-vesicle detection and analysis of peptide-induced membrane permeabilization

    Kristensen, Kasper; Ehrlich, Nicky; Henriksen, Jonas Rosager


    methods as alternatives to the quenching-based assays for studying peptide-induced leakage from large unilamellar lipid vesicles. Specifically, we use fluorescence correlation spectroscopy (FCS) to study the leakage of fluorescent molecules of different sizes from large unilamellar lipid vesicles......The capability of membrane-active peptides to disrupt phospholipid membranes is often studied by investigating peptide-induced leakage of quenched fluorescent molecules from large unilamellar lipid vesicles. In this article, we explore two fluorescence microscopy-based single-vesicle detection...... dispersed in aqueous solution, and we use confocal imaging of surface-immobilized large unilamellar lipid vesicles to investigate whether there are heterogeneities in leakage between individual vesicles. Of importance, we design an experimental protocol that allows us to quantitatively correlate the results...

  20. Fusion of Nonionic Vesicles

    Bulut, Sanja; Oskolkova, M. Z.; Schweins, R.


    We present an experimental study of vesicle fusion using light and neutron scattering to monitor fusion events. Vesicles are reproducibly formed with an extrusion procedure using an single amphiphile triethylene glycol mono-n-decyl ether in water. They show long-term stability for temperatures...... around 20 C, but at temperatures above 26 C we observe an increase in the scattered intensity due to fusion. The system is unusually well suited for the study of basic mechanisms of vesicle fusion. The vesicles are flexible with a bending rigidity of only a few k(H)T. The monolayer spontaneous curvature...

  1. Phospholipidic Monolayers on Formamide

    Graner, François; Perez-Oyarzun, Santiago; Saint-Jalmes, Arnaud; Flament, Cyrille; Gallet, François


    We report the first phase diagram of a Langmuir film at the air-formamide interface. Stable films of phospholipids such as DPPC or DSPC undergo phase transitions observed on isotherms or by fluorescence microscopy. They display bidimensional gas, liquid and solid phases, as well as two mesophases; the latter coexist with liquid on a sharp first-order transition plateau. We compare these observations with known results on films on water. Nous présentons le premier diagramme de phase d'un film de Langmuir à l'interface air-formamide. On observe, sur des isothermes et par microscopie de fluorescence, des transitions de phase dans des films stables de phospholipides comme le DPPC ou le DSPC. Cinq phases bidimensionnelles sont mises en évidence : gaz, liquide, solide, ainsi que deux mésophases ; ces deux dernières coexistent avec le liquide sur un plateau très marqué de transition du premier ordre. Nous comparons ces observations avec les résultats connus pour les films sur l'eau.

  2. Intermolecular crosslinking of fatty acyl chains in phospholipids: use of photoactivable carbene precursors.

    Gupta, C M; Radhakrishnan, R; Gerber, G E; Olsen, W L; Quay, S C; Khorana, H G


    Phospholipids containing photolysable carbene precursors (beta-trifluoro-alpha-diazopropionoxy and m-diazirinophenoxy groups) in omega-positions of sn-2 fatty acyl chains were prepared. Photolysis of their vesicles produced crosslinked products in 40-60% yields. Crosslinking was mostly intermolecular and occurred by carbene insertion into the C-H bonds of a second fatty acyl chain. Crosslinking products were characterized by (i) their gel permeation behavior, (ii) analysis of products formed by base-catalyzed transesterification, (iii) degradation with phospholipases A2 and C, (iv) gas chromatography/mass spectrometry, and (v) use of mixtures of phospholipids carrying the carbene precursors and a phospholipid containing radioactively labeled fatty acyl groups. Nitrenes generated from the aliphatic or aromatic azido groups in phospholipids were unsatisfactory for forming crosslinks by insertion in C-H bonds. PMID:288050

  3. Synthesis of Photoactivatable Phospholipidic Probes

    Qing PENG; Fan Qi QU; Yi XIA; Jie Hua ZHOU; Qiong You WU; Ling PENG


    We synthesized and characterized photoactivatable phospholipidic probes 1-3. These probes have the perfluorinated aryl azide function at the polar head of phospholipid. They are stable in dark and become highly reactive upon photoirradiation. The preliminary results suggest that they are promising tools to study the topology of membrane proteins and protein-lipid interactions using photolabeling approach.

  4. Interaction of SynaptotagminⅠ with Phospholipid Membrane: A Monolayer Study

    贺雨虹; 隋森芳


    Synaptotagmin Ⅰ(sytⅠ) is an abundant integral membrane protein of the synaptic vesicle and the C2A domain is an important functional domain in the cytoplasmic part of sytⅠ. C2A prefers to interact with plasmic membranes of neuron cells in vivo and such interaction is closely related to the sytⅠ physiological function as a Ca2+ sensor in the Ca2+-regulated neurotransmitter release. However, the interaction nature between C2A and phospholipids is not well understood. Monolayers at an air/water interface were used to study the interaction between C2A and a phospholipid membrane. The results show that C2A preferentially inserts into the negatively charged phosphatidylserine monolayer and Ca2+ ions are required for the interaction. Electrostatic force is mostly responsible for the insertion of C2A into dipalmitoyl phosphatidylserine monolayers.

  5. Complement-mediated solubilization of immune complexes. Solubilization inhibition and complement factor levels in SLE patients

    Baatrup, Gunnar; Petersen, Ivan; Kappelgaard, E;


    Thirty-two of 36 serum samples from 19 SLE patients showed reduced capacity to mediate complement-dependent solubilization of immune complexes (IC). SLE patients with nephritis exerted the lowest complement-mediated solubilization capacity (CMSC) whereas sera from patients with inactive disease g...

  6. Extracellular Vesicle (EV) Array

    Jørgensen, Malene; Bæk, Rikke; Pedersen, Shona


    their phenotype and determine their concentration in biological fluids. To identify circulating as well as cell culture-derived vesicles, the current standard is immunoblotting or a flow cytometrical analysis for specific proteins, both of which requires large amounts of purified vesicles....

  7. Synaptic vesicle endocytosis.

    Saheki, Yasunori; De Camilli, Pietro


    Neurons can sustain high rates of synaptic transmission without exhausting their supply of synaptic vesicles. This property relies on a highly efficient local endocytic recycling of synaptic vesicle membranes, which can be reused for hundreds, possibly thousands, of exo-endocytic cycles. Morphological, physiological, molecular, and genetic studies over the last four decades have provided insight into the membrane traffic reactions that govern this recycling and its regulation. These studies have shown that synaptic vesicle endocytosis capitalizes on fundamental and general endocytic mechanisms but also involves neuron-specific adaptations of such mechanisms. Thus, investigations of these processes have advanced not only the field of synaptic transmission but also, more generally, the field of endocytosis. This article summarizes current information on synaptic vesicle endocytosis with an emphasis on the underlying molecular mechanisms and with a special focus on clathrin-mediated endocytosis, the predominant pathway of synaptic vesicle protein internalization.

  8. Solubilization of Hydrophobic Dyes in Surfactant Solutions

    Ali Reza Tehrani-Bagha


    Full Text Available In this paper, the use of surfactants for solubilization of hydrophobic organic dyes (mainly solvent and disperse dyes has been reviewed. The effect of parameters such as the chemical structures of the surfactant and the dye, addition of salt and of polyelectrolytes, pH, and temperature on dye solubilization has been discussed. Surfactant self-assemble into micelles in aqueous solution and below the concentration where this occurs—the critical micelle concentration (CMC—there is no solubilization. Above the CMC, the amount of solubilized dye increases linearly with the increase in surfactant concentration. It is demonstrated that different surfactants work best for different dyes. In general, nonionic surfactants have higher solubilization power than anionic and cationic surfactants. It is likely that the reason for the good performance of nonionic surfactants is that they allow dyes to be accommodated not only in the inner, hydrocarbon part of the micelle but also in the headgroup shell. It is demonstrated that the location of a dye in a surfactant micelle can be assessed from the absorption spectrum of the dye-containing micellar solution.

  9. Biosurfactant-producing strains in enhancing solubilization and biodegradation of petroleum hydrocarbons in groundwater.

    Liu, Hong; Wang, Hang; Chen, Xuehua; Liu, Na; Bao, Suriguge


    Three biosurfactant-producing strains designated as BS-1, BS-3, and BS-4 were screened out from crude oil-contaminated soil using a combination of surface tension measurement and oil spreading method. Thin layer chromatography and infrared analysis indicated that the biosurfactants produced by the three strains were lipopeptide, glycolipid, and phospholipid. The enhancement of solubilization and biodegradation of petroleum hydrocarbons in groundwater employing biosurfactant-producing strains was investigated. The three strain mixtures led to more solubilization of petroleum hydrocarbons in groundwater, and the solubilization rate was 10.5 mg l−1. The combination of biosurfactant-producing strains and petroleum-degrading strains exhibited a higher biodegradation efficiency of 85.4 % than the petroleum-degrading strains (71.2 %). Biodegradation was enhanced the greatest with biosurfactant-producing strains and petroleum-degrading strains in a ratio of 1:1. Fluorescence microscopy images illustrate that the oil dispersed into smaller droplets and emulsified in the presence of biosurfactant-producing strains, which attached to the oil. Thus, the biodegradation of petroleum hydrocarbons in groundwater was enhanced.

  10. Elastic vesicles as topical/transdermal drug delivery systems.

    Choi, M J; Maibach, H I


    Skin acts a major target as well as a principle barrier for topical/transdermal drug delivery. Despite the many advantages of this system, the major obstacle is the low diffusion rate of drugs across the stratum corneum. Several methods have been assessed to increase the permeation rate of drugs temporarily. One simple and convenient approach is application of drugs in formulation with elastic vesicles or skin enhancers. Elastic vesicles are classified with phospholipid (Transfersomes((R)) and ethosomes) and detergent-based types. Elastic vesicles were more efficient at delivering a low and high molecular weight drug to the skin in terms of quantity and depth. Their effectiveness strongly depends on their physicochemical properties: composition, duration and application volume, and entrapment efficiency and application methods. This review focuses on the effect of elastic liposomes for enhancing the drug penetration and defines the action mechanism of penetration into deeper skin.

  11. Preparation of large monodisperse vesicles.

    Ting F Zhu

    Full Text Available Preparation of monodisperse vesicles is important both for research purposes and for practical applications. While the extrusion of vesicles through small pores (approximately 100 nm in diameter results in relatively uniform populations of vesicles, extrusion to larger sizes results in very heterogeneous populations of vesicles. Here we report a simple method for preparing large monodisperse multilamellar vesicles through a combination of extrusion and large-pore dialysis. For example, extrusion of polydisperse vesicles through 5-microm-diameter pores eliminates vesicles larger than 5 microm in diameter. Dialysis of extruded vesicles against 3-microm-pore-size polycarbonate membranes eliminates vesicles smaller than 3 microm in diameter, leaving behind a population of monodisperse vesicles with a mean diameter of approximately 4 microm. The simplicity of this method makes it an effective tool for laboratory vesicle preparation with potential applications in preparing large monodisperse liposomes for drug delivery.

  12. Shape deformations of giant unilamellar vesicles with a laser tweezer array

    Losert, Wolfgang; Poole, Cory; Bradford, Peter; English, Doug


    Vesicles are phospholipid bilayers that form a surface enclosing a volume of water or solution. They are of importance as model systems to study cells, as well as having practical applications such as containers for performing nanochemistry and facilitating drug delivery. Their properties have been studied for decades. Using a holographic laser tweezer array (LTA), which converts a single laser beam into many laser tweezer points, we stretch the vesicles in controlled ways from several points at once, measuring each force applied. Here, we present data on shape deformations of simple, spherical vesicles and on membrane fracture.

  13. Outer membrane vesicles of Lysobacter sp. XL1: biogenesis, functions, and applied prospects.

    Kudryakova, Irina V; Shishkova, Nina A; Vasilyeva, Natalia V


    Outer membrane vesicles (OMVs) produced by Gram-negative bacteria have been intensively investigated in recent times. Vesicle formation models have been proposed, some factors affecting the process were established, and important roles vesicles play in vital activities of their producing cells were determined. Studies of pathogenic bacterial vesicles contribute to understanding the causes of acute infection and developing drugs on their basis. Despite intensive research, issues associated with the understanding of vesicle biogenesis, the mechanisms of bacterium-bacterium and pathogen-host interactions with participation of vesicles, still remain unresolved. This review discusses some results obtained in the research into OMVs of Lysobacter sp. XL1 VKM B-1576. This bacterium secretes into the environment a spectrum of bacteriolytic enzymes that hydrolyze peptidoglycan of competing bacteria, thus leading to their lysis. One of these enzymes, lytic endopeptidase L5, has been shown not only to be secreted by means of vesicles but also to be involved in their formation. As part of vesicles, the antimicrobial potential of L5 enzyme has been found to be considerably expanded. Vesicles have been shown to have a therapeutic effect in respect of anthrax infection and staphylococcal sepsis modelled in mice. The scientific basis for constructing liposomal antimicrobial preparations from vesicle phospholipids and recombinant bacteriolytic enzyme L5 has been formed.

  14. The potential of phosphate solubilizing bacteria isolated from sugarcane wastes for solubilizing phosphate



    Full Text Available Most of P in agricultural soils is in unavailable forms for plant growth. Phosphate solubilizing bacteria can increase soil P availability. This study was aimed to isolate phosphate solubilizing bacteria from sugarcane waste compost and to test ability of the isolated bacterial to dissolve phosphate. The bacteria were isolated from three types of sugarcane waste, i.e. filter cake compost, bagasse compost, and a mixture of filter cake + bagasse + trash biomass compost. The potential colony was further purified by the Pikovskaya method on selective media. Eight isolates of phosphate solubilizing bacteria were obtained from all wasted studied. Amongst them, T-K5 and T-K6 isolates were superior in dissolving P from Ca3(PO42 in the media studied. The two isolates were able to solubilize P with solubilizing index of 1.75 and 1.67 for T-K5 and T-K6, respectively. Quantitatively, T-K6 isolate showed the highest P solubilization (0.74 mg / L, followed by T-K5 isolate (0.56 mg / L, while the lowest P solubilization (0.41 mg / L was observed for T-K4 isolate. The increase of soluble P was not always followed by the decrease in pH.

  15. Formation of self-assembled quantum dot-chlorin e6 complex: influence of nanoparticles phospholipid coating

    Karabanovas, V.; Skripka, A.; Valanciunaite, J.; Kubiliute, R.; Poderys, V.; Rotomskis, R.


    The clinical use of phospholipid-coated quantum dots (QDs)-photosensitizer complexes as therapeutic nanoagents depends on colloidal stability of these complexes and efficiency of Förster resonance energy transfer from QDs to bound photosensitizer molecules. In this study, we demonstrate modification of CdSe/ZnS QDs with different phospholipids such as 1,2-dipalmitoyl- sn-glycero-3-phosphoethanolamine- N-[methoxy(polyethylene glycol)-2000] (PEG-DPPE); 1,2-dioleoyl- sn-glycero-3-phosphoethanolamine- N-[methoxy(polyethylene glycol)-2000 (PEG-DOPE) and 1,2-dioleoyl- sn-glycero-3-phosphocholine (DOPC) and the complex formation with photosensitizer chlorin e6 (Ce6). QDs were successfully solubilized in water by coating QDs with PEG-DPPE and PEG-DOPE phospholipids. However, an attempt to solubilize QDs using PEG-free phospholipids (DOPC) was ineffective. While QDs modified with DOPC:PEG-DOPE mixtures at molar ratios of 1:1 and 2:1 showed long-term stability in aqueous solution, colloidal solution of QDs modified by DOPC:PEG-DPPE (molar ratio 2:1) was unstable. We showed that Ce6 forms a stable complex only with QDs coated with unsaturated phospholipids PEG-DOPE and DOPC:PEG-DOPE. Close localization of Ce6 molecules to the core of QDs ensures efficient energy transfer from QDs to bound Ce6 molecules that is crucial for its further application in photodynamic therapy of cancer.

  16. Pulling on adhered vesicles

    Smith, Ana-Suncana; Goennenwein, Stefanie; Lorz, Barbara; Seifert, Udo; Sackmann, Erich


    A theoretical model describing pulling of vesicles adhered in a contact potential has been developed. Two different regimes have been recognized. For weak to middle-strength adhesive potentials, locally stable shapes are found in a range of applied forces, separated from the free shape by an energy barrier. The phase diagram contains regions with either a unique bound shape or an additional meta-stable shape. Upon pulling, these shapes unbind discontinuously since the vesicle disengage from the surface while still possessing a finite adhesion area (Smith 2003a). In a strong adhesion regime, a competition between adhesion and tether formation is observed. A critical onset force is identified where a tether spontaneously appears as a part of a second order shape transition. Further growth of a tether is followed by a detachment process which terminates at a finite force when a vesicle continuously unbinds from the substrate (Smith 2003b). Both critical forces, as well as all shape parameters, are calculated as a function of the reduced volume and the strength of adhesive potential. Analogous experimental study has been performed where a vertical magnetic tweezers are used in combination with micro-interferometric and confocal techniques to reproduce the same symmetry as in the theoretical investigation. Giant vesicles are bound to the substrate by numerous specific bonds formed between ligands and receptors incorporated into the vesicle and the substrate, respectively. Application of a constant force is inducing a new thermodynamic equilibrium of the system where the vesicle is partially unbound from the substrate (Goennenwein 2003). The shapes of vesicles are compared prior and during application of the force. Very good agreement is obtained, particularly in the middle-strength adhesion regime (Smith 2003c). References: 1. A.-S. Smith, E. Sackmann, U. Seifert: Effects of a pulling force on the shape of a bound vesicle, Europhys. Lett., 64, 2 (2003). 2. A.-S. Smith

  17. Deformation analysis of vesicles in an alternating-current electric field.

    Tang, Yu-Gang; Liu, Ying; Feng, Xi-Qiao


    In this paper the shape equation for axisymmetric vesicles subjected to an ac electric field is derived on the basis of the liquid-crystal model. The equilibrium morphology of a lipid vesicle is determined by the minimization of its free energy in coupled mechanical and ac electric fields. Besides elastic bending, the effects of the osmotic pressure difference, surface tension, Maxwell pressure, and flexoelectric and dielectric properties of phospholipid membrane as well are taken into account. The influences of elastic bending, osmotic pressure difference, and surface tension on the frequency-dependent behavior of a vesicle membrane in an ac electric field are examined. The singularity of the ac electric field is also investigated. Our theoretical results of vesicle deformation agree well with previous experimental and numerical results. The present study provides insights into the physical mechanisms underpinning the frequency-dependent morphological evolution of vesicles in the electric and mechanical fields.

  18. Cell signalling and phospholipid metabolism

    Boss, W.F.


    These studies explored whether phosphoinositide (PI) has a role in plants analogous to its role in animal cells. Although no parallel activity of PI in signal transduction was found in plant cells, activity of inositol phospholipid kinase was found to be modulated by light and by cell wall degrading enzymes. These studies indicate a major role for inositol phospholipids in plant growth and development as membrane effectors but not as a source of second messengers.

  19. Solubilization of paraffinic deposits for microemulsions

    Gomes, Erika A.S.; Soares, Ranieri G.F.; Nascimento, Roseane E.S.; Dantas Neto, Afonso A.; Barros Neto, Eduardo L. [Universidade Federal do Rio Grande do Norte (UFRN), Natal, RN (Brazil)


    The oil company has been intensifying its efforts to find more efficient solutions for the problems related to the paraffin in wells and transport lines. When applied in the flow lines, the solvents dissolve the paraffin and they must be used hot, since the temperature increases the solubility of the wax and, consequently, its removal rate. The microemulsions appear as an alternative capable of acting in the solubilization and in the inhibition of the formation of deposits due to its great interfacial area, low superficial tension and high capacity of solubilization. They present some advantages in relation to the methods of use of chemical products due to its flexibility of composition in which they can be used, presenting low toxicity and inflammability, without any loss of its capacity of solubilization. The use of oil-in-water microemulsion aims to solubilized paraffin in the disperse phase, where one can find the apolar part of the molecule of the surfactant and the also apolar chain of paraffin, occurring, therefore the 'encapsulation' of the crystal, prohibiting the growth of the chain due to the affinity of paraffin and oil. In this in case, it is possible to transport the inserted paraffin in direct micelles, reducing the precipitation and optimizing the transport. (author)

  20. Multicomponent mixtures for cryoprotection and ligand solubilization

    Lidia Ciccone


    Full Text Available Mixed cryoprotectants have been developed for the solubilization of ligands for crystallization of protein–ligand complexes and for crystal soaking. Low affinity lead compounds with poor solubility are problematic for structural studies. Complete ligand solubilization is required for co-crystallization and crystal soaking experiments to obtain interpretable electron density maps for the ligand. Mixed cryo-preserving compounds are needed prior to X-ray data collection to reduce radiation damage at synchrotron sources. Here we present dual-use mixes that act as cryoprotectants and also promote the aqueous solubility of hydrophobic ligands. Unlike glycerol that increases protein solubility and can cause crystal melting the mixed solutions of cryo-preserving compounds that include precipitants and solubilizers, allow for worry-free crystal preservation while simultaneously solubilizing relatively hydrophobic ligands, typical of ligands obtained in high-throughput screening. The effectiveness of these mixture has been confirmed on a human transthyretin crystals both during crystallization and in flash freezing of crystals.

  1. Solubilization of coal in organic media

    Lahaye, P.; Decroocq, D.


    The use of solvent extraction to solubilize coal is discussed. Simple extractions are described which are conducted at moderate temperatures to exclude extraneous chemical reactions which would lead to uncontrolled changes in the components of the treated coal. Sample preparation, extraction apparatus, and the determination of extract yield for different experiments are also described.

  2. Solubilization and purification of the ATPase from the tonoplast of Hevea.

    Marin, B; Preisser, J; Komor, E


    The tonoplast-bound ATPase of Hevea brasiliensis (caoutchouc tree) was solubilized with dichloromethan and purified 100-fold with two ammonium sulfate precipitation steps and a G-200 gel filtration step. The resulting ATPase activity eluted according to a molecular mass of approximately 200 kDa and chromatographed at an isoelectric pH of 5.3. Subunits of molecular mass 110 kDa, 68 kDa, 24 kDa and 12 kDa appeared after treatment with 1% sodium dodecyl sulfate or spontaneously during storage of the solubilized ATPase. Dodecyl sulfate/polyacrylamide gel electrophoresis yielded four polypeptides of molecular mass 54 kDa, 66 kDa, 23 kDa and 13 kDa. From protein determination by ultraviolet absorption and Coomassie stain it appears that the 54-kDa and the 66-kDa polypeptides exist in multiple copies. No close resemblance to the membrane-bound ATPase of mitochondria, plastids, plasmalemma, chromaffin granules and synaptic vesicles is seen. No antibody cross-reaction to F1 of bacteria is observed. Therefore it is concluded that the vacuolar ATPase represents a novel type of ATPase. Many properties of the tonoplast-bound ATPase such as pH-dependence, substrate specificity, ion-dependence and inhibitor sensitivity did not change when the enzyme had been solubilized and purified. The phosphatase activity was lost during the purification procedure. The stimulation of ATP-hydrolysis in tonoplast vesicles by uncouplers and ionophores was absent in the solubilized ATPase, and also the stimulation by chloride was significantly reduced. Anion channel blockers, such as triphenyltin and 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene, which are strong inhibitors of membrane-bound ATPase, fully or partly lost their inhibiting effect after solubilization of the ATPase. These results are interpreted to indicate that ionophores do not directly affect the ATPase molecule, whereas chloride might have a small direct effect on the ATPase besides its effect as a permeating anion.

  3. Solution structure and phospholipid interactions of the isolated voltage-sensor domain from KvAP.

    Butterwick, Joel A; MacKinnon, Roderick


    Voltage-sensor domains (VSDs) are specialized transmembrane segments that confer voltage sensitivity to many proteins such as ion channels and enzymes. The activities of these domains are highly dependent on both the chemical properties and the physical properties of the surrounding membrane environment. To learn about VSD-lipid interactions, we used nuclear magnetic resonance spectroscopy to determine the structure and phospholipid interface of the VSD from the voltage-dependent K(+) channel KvAP (prokaryotic Kv from Aeropyrum pernix). The solution structure of the KvAP VSD solubilized within phospholipid micelles is similar to a previously determined crystal structure solubilized by a nonionic detergent and complexed with an antibody fragment. The differences observed include a previously unidentified short amphipathic α-helix that precedes the first transmembrane helix and a subtle rigid-body repositioning of the S3-S4 voltage-sensor paddle. Using (15)N relaxation experiments, we show that much of the VSD, including the pronounced kink in S3 and the S3-S4 paddle, is relatively rigid on the picosecond-to-nanosecond timescale. In contrast, the kink in S3 is mobile on the microsecond-to-millisecond timescale and may act as a hinge in the movement of the paddle during channel gating. We characterized the VSD-phospholipid micelle interactions using nuclear Overhauser effect spectroscopy and showed that the micelle uniformly coats the KvAP VSD and approximates the chemical environment of a phospholipid bilayer. Using paramagnetically labeled phospholipids, we show that bilayer-forming lipids interact with the S3 and S4 helices more strongly than with S1 and S2.


    Florencia, Henning María; Susana, Sanchez; Laura, Bakás


    Lipopolysaccharide (LPS) is an endotoxin released from the outer membrane of Gram negative bacteria during infections. It have been reported that LPS may play a rol in the outer membrane of bacteria similar to that of cholesterol in eukaryotic plasma membranes. In this article we compare the effect of introducing LPS or cholesterol in liposomes made of dipalmitoylphosphatidylcholine/dioleoylphosphatidylcholine on the solubilization process by Triton X-100. The results show that liposomes containing LPS or Cholesterol are more resistant to solubilization by Triton X-100 than the binary phospholipid mixtures at 4°C. The LPS distribution was analyzed on GUVs of DPPC:DOPC using FITC-LPS. Solid and liquid-crystalline domains were visualized labeling the GUVs with LAURDAN and GP images were acquired using a two-photon microscope. The images show a selective distribution of LPS in gel domains. Our results support the hypothesis that LPS could aggregate and concentrate selectively in biological membranes providing a mechanism to bring together several components of the LPS-sensing machinery. PMID:19324006

  5. Visualization and analysis of lipopolysaccharide distribution in binary phospholipid bilayers

    Henning, Maria Florencia [Instituto de Investigaciones Bioquimicas La Plata (INIBIOLP), CCT-La Plata, CONICET, Facultad de Ciencias Medicas, UNLP, Calles 60 y 120, 1900 La Plata (Argentina); Sanchez, Susana [Laboratory for Fluorescence Dynamics, University of California-Irvine, Irvine, CA (United States); Bakas, Laura, E-mail: [Instituto de Investigaciones Bioquimicas La Plata (INIBIOLP), CCT-La Plata, CONICET, Facultad de Ciencias Medicas, UNLP, Calles 60 y 120, 1900 La Plata (Argentina); Departamento de Ciencias Biologicas, Facultad de Ciencias Exactas, UNLP, Calles 47 y 115, 1900 La Plata (Argentina)


    Lipopolysaccharide (LPS) is an endotoxin released from the outer membrane of Gram-negative bacteria during infections. It have been reported that LPS may play a role in the outer membrane of bacteria similar to that of cholesterol in eukaryotic plasma membranes. In this article we compare the effect of introducing LPS or cholesterol in liposomes made of dipalmitoylphosphatidylcholine/dioleoylphosphatidylcholine on the solubilization process by Triton X-100. The results show that liposomes containing LPS or cholesterol are more resistant to solubilization by Triton X-100 than the binary phospholipid mixtures at 4 {sup o}C. The LPS distribution was analyzed on GUVs of DPPC:DOPC using FITC-LPS. Solid and liquid-crystalline domains were visualized labeling the GUVs with LAURDAN and GP images were acquired using a two-photon microscope. The images show a selective distribution of LPS in gel domains. Our results support the hypothesis that LPS could aggregate and concentrate selectively in biological membranes providing a mechanism to bring together several components of the LPS-sensing machinery.

  6. Phospholipid and Hydrocarbon Interactions with a Charged Electrode Interface.

    Levine, Zachary A; DeNardis, Nadica Ivošević; Vernier, P Thomas


    Using a combination of molecular dynamics simulations and experiments we examined the interactions of alkanes and phospholipids at charged interfaces in order to understand how interfacial charge densities affect the association of these two representative molecules with electrodes. Consistent with theory and experiment, these model systems reveal interfacial associations mediated through a combination of Coulombic and van der Waals forces. van der Waals forces, in particular, mediate rapid binding of decane to neutral electrodes. No decane binding was observed at high surface charge densities because of interfacial water polarization, which screens hydrophobic attractions. The positively charged choline moiety of the phospholipid palmitoyloleoylphosphatidylcholine (POPC) is primarily responsible for POPC attraction by a moderately negatively charged electrode. The hydrocarbon tails of POPC interact with the hydrophobic electrode interface similarly to decane. Previously reported electrochemical results confirm these findings by demonstrating bipolar displacement currents from PC vesicles adhering to moderately negatively charged interfaces, originating from the choline interactions observed in simulations. At more negatively charged interfaces, choline-to-surface binding was stronger. In both simulations and experiments the maximal interaction of anionic PS occurs with a positively charged interface, provided that the electrostatic forces outweigh local Lennard-Jones interactions. Direct comparisons between the binding affinities measured in experiments and those obtained in simulations reveal previously unobserved atomic interactions that facilitate lipid vesicle adhesion to charged interfaces. Moreover, the implementation of a charged interface in molecular dynamics simulations provides an alternative method for the generation of large electric fields across phospholipid bilayers, especially for systems with periodic boundary conditions, and may be useful for

  7. Mixed micelle formation between amino acid-based surfactants and phospholipids.

    Faustino, Célia M C; Calado, António R T; Garcia-Rio, Luís


    The mixed micelle formation in aqueous solutions between an anionic gemini surfactant derived from the amino acid cystine (C(8)Cys)(2), and the phospholipids 1,2-diheptanoyl-sn-glycero-3-phosphocholine (DHPC, a micelle-forming phospholipid) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC, a vesicle-forming phospholipid) has been studied by conductivity and the results compared with the ones obtained for the mixed systems with the single-chain surfactant derived from cysteine, C(8)Cys. Phospholipid-surfactant interactions were found to be synergistic in nature and dependent on the type of phospholipid and on surfactant hydrophobicity. Regular solution theory was used to analyse the gemini surfactant-DHPC binary mixtures and the interaction parameter, β(12), has been evaluated, as well as mixed micelle composition. The results have been interpreted in terms of the interplay between reduction of the electrostatic repulsions among the ionic head groups of the surfactants and steric hindrances arising from incorporation of the zwitterionic phospholipids in the mixed micelles.

  8. Molecular Dynamic Analysis of Hyaluronic Acid and Phospholipid Interaction in Tribological Surgical Adjuvant Design for Osteoarthritis.

    Siódmiak, Jacek; Bełdowski, Piotr; Augé, Wayne K; Ledziński, Damian; Śmigiel, Sandra; Gadomski, Adam


    Tribological surgical adjuvants constitute a therapeutic discipline made possible by surgical advances in the treatment of damaged articular cartilage beyond palliative care. The purpose of this study is to analyze interactions between hyaluronic acid and phospholipid molecules, and the formation of geometric forms, that play a role in the facilitated lubrication of synovial joint organ systems. The analysis includes an evaluation of the pathologic state to detail conditions that may be encountered by adjuvants during surgical convalescence. The synovial fluid changes in pH, hyaluronic acid polydispersity, and phospholipid concentration associated with osteoarthritis are presented as features that influence the lubricating properties of adjuvant candidates. Molecular dynamic simulation studies are presented, and the Rouse model is deployed, to rationalize low molecular weight hyaluronic acid behavior in an osteoarthritic environment of increased pH and phospholipid concentration. The results indicate that the hyaluronic acid radius of gyration time evolution is both pH- and phospholipid concentration-dependent. Specifically, dipalmitoylphosphatidylcholine induces hydrophobic interactions in the system, causing low molecular weight hyaluronic acid to shrink and at high concentration be absorbed into phospholipid vesicles. Low molecular weight hyaluronic acid appears to be insufficient for use as a tribological surgical adjuvant because an increased pH and phospholipid concentration induces decreased crosslinking that prevents the formation of supramolecular lubricating forms. Dipalmitoylphosphatidylcholine remains an adjuvant candidate for certain clinical situations. The need to reconcile osteoarthritic phenotypes is a prerequisite that should serve as a framework for future adjuvant design and subsequent tribological testing.

  9. An artificial cell based on gene expression in vesicle

    Noireaux, Vincent


    A new experimental approach is presented to build an artificial cell using the translation machinery of a cell-free expression system as the hardware and a DNA synthetic program as the software. Cytoplasmic extracts, encapsulated in phospholipid vesicles, are used to assemble custom-made genetic circuits to develop the functions of a minimal cell. The objective is to understand how a DNA algorithm can be designed to build an operating system that has some of the properties of life. We show how a long-lived bioreactor is built to carry out in vitro transcription and translation in cell-sized vesicles. To develop the synthetic membrane into an active interface, a few amphipathic peptides and an insertion mechanism of integral membrane proteins have been tested. With vesicles composed of different phospholipids, the fusion protein alpha-hemolysin-eGFP can be expressed to reveal patterns on the membrane. Finally, specific degradation mechanisms are introduced to create a sink for the synthesized messengers and proteins. Perspectives and limitations of this approach will be discussed.

  10. Sphingosine induces the aggregation of imine-containing peroxidized vesicles.

    Jiménez-Rojo, Noemi; Viguera, Ana R; Collado, M Isabel; Sims, Kacee H; Constance, Chad; Hill, Kasey; Shaw, Walt A; Goñi, Félix M; Alonso, Alicia


    Lipid peroxidation plays a central role in the pathogenesis of many diseases like atherosclerosis and multiple sclerosis. We have analyzed the interaction of sphingosine with peroxidized bilayers in model membranes. Cu(2+) induced peroxidation was checked following UV absorbance at 245nm, and also using the novel Avanti snoopers®. Mass spectrometry confirms the oxidation of phospholipid unsaturated chains. Our results show that sphingosine causes aggregation of Cu(2+)-peroxidized vesicles. We observed that aggregation is facilitated by the presence of negatively-charged phospholipids in the membrane, and inhibited by anti-oxidants e.g. BHT. Interestingly, long-chain alkylamines (C18, C16) but not their short-chain analogues (C10, C6, C1) can substitute sphingosine as promoters of vesicle aggregation. Furthermore, sphinganine but not sphingosine-1-phosphate can mimic this effect. Formation of imines in the membrane upon peroxidation was detected by (1)H-NMR and it appeared to be necessary for the aggregation effect. (31)P-NMR spectroscopy reveals that sphingosine facilitates formation of non-lamellar phase in parallel with vesicle aggregation. The data might suggest a role for sphingosine in the pathogenesis of atherosclerosis. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Solubilization isotherms of aromatic solutes in surfactant aggregates

    Gadelle, F.; Koros, W.J.; Schechter, R.S. (Univ. of Texas, Austin, TX (United States). Dept. of Chemical Engineering)


    Several factors affecting solubilization of aromatic solutes in surfactant micelles have been investigated. Solubilization isotherms of benzene, toluene, and chlorobenzene in various aqueous micellar solutions were determined using head space gas chromatography. Cationic surfactants such as cetylpyridinium chloride or cetyltrimethylammonium bromide present high solubilization capacities. Comparable anionic surfactants exhibit lower solubilization and a greater tendency to precipitate. It was observed that nonionic surfactants show high solubilization on a molar basis. Solubilization in mixed cationic-anionic micelles was also investigated. It also appears that the molecular size of the solute determines the extent of the solubilization. Finally, the shape of the different isotherms indicates that knowing the amount solubilized at saturation of the micellar solution is not sufficient to estimate solubilization at solute concentrations lower than the solute aqueous solubility. Solubilization of organics in surfactant micelles is of major importance in many applications. One new application is micellar-enhanced ultrafiltration. Another application of interest is the surfactant-enhanced aquifer remediation, a process in which a micellar aqueous solution is flushed into contaminated groundwaters to enhance recovery of pollutants by micellar solubilization.

  12. In vitro and in vivo anti-tumor effects of novel Span 80 vesicles containing immobilized Eucheuma serra agglutinin.

    Omokawa, Yousuke; Miyazaki, Tatsuhiko; Walde, Peter; Akiyama, Koichi; Sugahara, Takuya; Masuda, Seizo; Inada, Akihiro; Ohnishi, Yasuyuki; Saeki, Toshiaki; Kato, Keiichi


    The lectin Eucheuma serra agglutinin (ESA) is known from previous studies to specifically bind to high-mannose type N-glycans and to induce apoptotic cancer cell death in vitro. In this study, Span 80 vesicles, with an average diameter between about 200 and 400 nm, containing immobilized ESA were prepared from the nonionic surfactant Span 80, also known as sorbitan monooleate. The vesicles were investigated in vitro and in vivo to evaluate the vesicles's potential applicability as novel drug delivery system. The results obtained are promising since the following was observed: (i) vesicular ESA had the same hemagglutinating activity as free ESA, demonstrating its biological activity when bound to the vesicles; (ii) vesicles containing immobilized ESA decreased the viability of Colo201 cancer cells in vitro while the growth of normal cells was not affected; (iii) the vesicles showed binding to Colo201 cells in vitro and caused inhibition of cancer cell growth in nude mice to which the vesicle-treated cells were added; (iv) the vesicles diminished tumor growth after intravenous administration to nude mice which contained an implanted Colo201 tumor; (v) the vesicles showed a tendency to accumulate at the site of the tumor 6h after i.v. administration to nude mice. Thus, all measurements carried out indicate that this type of Span 80 vesicle can be considered as promising alternatives to conventional phospholipid-based vesicles.

  13. Phosphate solubilizing ability of two Arctic Aspergillus niger strains

    Shiv Mohan Singh,


    Full Text Available Many filamentous fungi were isolated from the soils of Ny-Ålesund, Spitsbergen, Svalbard, and were screened in vitro for their phosphate solubilizing ability. Two strains of Aspergillus niger showed good tricalcium phosphate (TCP solubilizing ability in Pikovskaya's medium. The TCP solubilization index was calculated at varying levels of pH and temperatures. The ability of Aspergillus niger strain-1 to solubilize and release inorganic-P was 285 µg ml–1, while Aspergillus niger strain-2 solubilized 262 µg ml–1 from 0.5% TCP after seven days. This is the first report of TCP solubilization by Arctic strains that may serve as very good phosphate solubilizers in the form of biofertilizer.

  14. Functional and structural stability of the epidermal growth factor receptor in detergent micelles and phospholipid nanodiscs

    Mi, Li-Zhi; Grey, Michael J; Nishida, Noritaka


    , recombinant EGFR constructs containing the extracellular, transmembrane, juxtamembrane, and kinase domains are overexpressed and purified from human embryonic kidney 293 cell cultures. The oligomerization state, overall structure, and functional stability of the purified EGF-bound receptor are characterized...... differential functional stability in Triton X-100 versus dodecyl maltoside. Furthermore, the kinase activity can be significantly stabilized by reconstituting purified EGF-bound EGFR dimers in phospholipid nanodiscs or vesicles, suggesting that the environment around the hydrophobic transmembrane...

  15. Stress-tolerant P-solubilizing microorganisms.

    Vassilev, N; Eichler-Löbermann, B; Vassileva, M


    Drought, high/low temperature, and salinity are abiotic stress factors accepted as the main reason for crop yield losses in a world with growing population and food price increases. Additional problems create nutrient limitations and particularly low P soil status. The problem of phosphate fertilizers, P plant nutrition, and existing phosphate bearing resources can also be related to the scarcity of rock phosphate. The modern agricultural systems are highly dependent on the existing fertilizer industry based exclusively of this natural, finite, non-renewable resource. Biotechnology offers a number of sustainable solutions that can mitigate these problems by using plant beneficial, including P-solubilizing, microorganisms. This short review paper summarizes the current and future trends in isolation, development, and application of P-solubilizing microorganisms in stress environmental conditions bearing also in mind the imbalanced cycling and unsustainable management of P. Special attention is devoted to the efforts on development of biotechnological strategies for formulation of P-solubilizing microorganisms in order to increase their protection against adverse abiotic factors.

  16. Review: P4-ATPases as Phospholipid Flippases-Structure, Function, and Enigmas

    Andersen, Jens P; Vestergaard, Anna L; Mikkelsen, Stine A


    P4-ATPases comprise a family of P-type ATPases that actively transport or flip phospholipids across cell membranes. This generates and maintains membrane lipid asymmetry, a property essential for a wide variety of cellular processes such as vesicle budding and trafficking, cell signaling, blood...... is highlighted by the finding that genetic defects in two P4-ATPases ATP8A2 and ATP8B1 are associated with severe human disorders. Recent studies have provided insight into how P4-ATPases translocate phospholipids across membranes. P4-ATPases form a phosphorylated intermediate at the aspartate of the P......-type ATPase signature sequence, and dephosphorylation is activated by the lipid substrate being flipped from the exoplasmic to the cytoplasmic leaflet similar to the activation of dephosphorylation of Na+/K+-ATPase by exoplasmic K+. How the phospholipid is translocated can be understood in terms...

  17. Phosphorylation of dynamin I on Ser-795 by protein kinase C blocks its association with phospholipids

    Powell, K A; Valova, V A; Malladi, C S


    Dynamin I is phosphorylated in nerve terminals exclusively in the cytosolic compartment and in vitro by protein kinase C (PKC). Dephosphorylation is required for synaptic vesicle retrieval, suggesting that its phosphorylation affects its subcellular localization. An in vitro phospholipid binding...... assay was established that prevents lipid vesiculation and dynamin lipid insertion into the lipid. Dynamin I bound the phospholipid in a concentration-dependent and saturable manner, with an apparent affinity of 230 +/- 51 nM. Optimal binding occurred with mixtures of phosphatidylserine...... and phosphatidylcholine of 1:3 with little binding to phosphatidylcholine or phosphatidylserine alone. Phospholipid binding was abolished after dynamin I phosphorylation by PKC and was restored after dephosphorylation by calcineurin. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry revealed...

  18. Lipid-Bilayer Dynamics Probed by a Carbon Dot-Phospholipid Conjugate.

    Nandi, Sukhendu; Malishev, Ravit; Bhunia, Susanta Kumar; Kolusheva, Sofiya; Jopp, Jürgen; Jelinek, Raz


    Elucidating the dynamic properties of membranes is important for understanding fundamental cellular processes and for shedding light on the interactions of proteins, drugs, and viruses with the cell surface. Dynamic studies of lipid bilayers have been constrained, however, by the relatively small number of pertinent molecular probes and the limited physicochemical properties of the probes. We show that a lipid conjugate comprised of a fluorescent carbon dot (C-dot) covalently attached to a phospholipid constitutes a versatile and effective vehicle for studying bilayer dynamics. The C-dot-modified phospholipids readily incorporated within biomimetic membranes, including solid-supported bilayers and small and giant vesicles, and inserted into actual cellular membranes. We employed the C-dot-phospholipid probe to elucidate the effects of polymyxin-B (a cytolytic peptide), valproic acid (a lipophilic drug), and amyloid-β (a peptide associated with Alzheimer's disease) upon bilayer fluidity and lipid dynamics through the application of various biophysical techniques.

  19. Isolation of phosphate-solubilizing fungus and its application in solubilization of rock phosphates.

    Wu, Yingben; He, Yuelin; Yin, Hongmei; Chen, Wei; Wang, Zhen; Xu, Lijuan; Zhang, Aiqun


    Microorganisms have been obtained to improve the agronomic value of rock phosphates (RPs), but the phosphorus solubilizing rate by these approaches is very slow. It is important to explore a high-efficient phosphate-solubilizing approach with a kind of microorganisms. This study aimed to isolate a high-efficient level of phosphate-solubilizing fungus from rhizosphere soil samples phosphate mines (Liuyang County, Hunan province, China) and apply it in solubilization of RPs. The experiments were carried out by the conventional methodology for morphological and biochemical fungus characterization and the analysis of 18s rRNA sequence. Then the effects of time, temperature, initial pH, phosphorus (P) sources, RPs concentration, shaking speed and silver ion on the content of soluble P released by this isolate were investigated. The results showed this isolate was identified as Galactomyces geotrichum P14 (P14) in GeneBank and the maximum amount of soluble P was 1252.13 mg L(-1) within 40 h in a modified phosphate growth agar's medium (without agar) where contained tricalcium phosphate (TCP) as sole phosphate source. At the same time, it could release phosphate and solubilize various rock phosphates. The isolated fungus can convert RPs from insoluble form into plant available form and therefore it hold great potential for biofertilizers to enhance soil fertility and promote plant growth.

  20. Extracellular vesicles for drug delivery

    Vader, Pieter; Mol, Emma A; Pasterkamp, Gerard; Schiffelers, Raymond M

    Extracellular vesicles (EVs) are cell-derived membrane vesicles, and represent an endogenous mechanism for intercellular communication. Since the discovery that EVs are capable of functionally transferring biological information, the potential use of EVs as drug delivery vehicles has gained

  1. Effects of phosphonium-based ionic liquids on phospholipid membranes studied by small-angle X-ray scattering.

    Kontro, Inkeri; Svedström, Kirsi; Duša, Filip; Ahvenainen, Patrik; Ruokonen, Suvi-Katriina; Witos, Joanna; Wiedmer, Susanne K


    The effects of ionic liquids on model phospholipid membranes were studied by small-angle X-ray scattering, dynamic light scattering (DLS) and zeta potential measurements. Multilamellar 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine liposomes and large unilamellar vesicles composed of l-α-phosphatidylcholine (eggPC) and l-α-phosphatidylglycerol (eggPG) (80:20mol%) or eggPC, eggPG, and cholesterol (60:20:20mol%) were used as biomimicking membrane models. The effects of the phosphonium-based ionic liquids: tributylmethylphosphonium acetate, trioctylmethylphosphonium acetate, tributyl(tetradecyl)-phosphonium acetate, and tributyl(tetradecyl)-phosphonium chloride, were compared to those of 1-ethyl-3-methyl-imidazolium acetate. With multilamellar vesicles, the ionic liquids that did not disrupt liposomes decreased the lamellar spacing as a function of concentration. The magnitude of the effect depended on concentration for all studied ionic liquids. Using large unilamellar vesicles, first a slight decrease in the vesicle size, then aggregation of vesicles was observed by DLS for increasing ionic liquid concentrations. At concentrations just below those that caused aggregation of liposomes, large unilamellar vesicles were coated by ionic liquid cations, evidenced by a change in their zeta potential. The ability of phosphonium-based ionic liquids to affect liposomes is related to the length of the hydrocarbon chains in the cation. Generally, the ability of ionic liquids to disrupt liposomes goes hand in hand with inducing disorder in the phospholipid membrane. However, trioctylmethylphosphonium acetate selectively extracted and induced a well-ordered lamellar structure in phospholipids from disrupted cholesterol-containing large unilamellar vesicles. This kind of effect was not seen with any other combination of ionic liquids and liposomes.

  2. Oxidative stability of marine phospholipids

    Lu, Henna Fung Sieng; Nielsen, Nina Skall; Baron, Caroline Pascale

    prepared in the form of emulsions by high pressure homogenizer. Then, the oxidative and hydrolytic stability of phospholipids was investigated by measurement of simple chemical analyses such as Peroxide Value and Free Fatty Acids, and 31PNMR after 32 days storage at 2ºC. The oxidative stability of MPL...

  3. Laser-induced europium(III) luminescence as a probe of the metal ion mediated association of human prothrombin with phospholipid.

    Rhee, M J; Horrocks, W D; Kosow, D P


    7F0 leads to 5D0 excitation spectroscopy of Eu(III) has been used to investigate the Eu(III) and phospholipid binding properties of human prothrombin. The results indicate that human prothrombin contains four high-affinity Eu(III) binding sites which are distributed into two classes of binding sites. When 4 equiv of Eu(III) is bound to prothrombin, the prothrombin is capable of binding to phospholipid vesicles. The deuterium isotope effect on the lifetime of the Eu(III)-prothrombin complex and the Eu(III)-prothrombin-phospholipid complex was used to determine the number of water molecules coordinated to the Eu(III). In both complexes, each of the Eu(III)'s coordinated to 2.5 +/- 0.5 water molecules. These results indicate that the binding of the Eu(III)-prothrombin complex to the phospholipid does not require the formation of a prothrombin-Eu(III)-phospholipid bridge.


    Sandeep Kumar Parihar*, Mithun Bhowmick, Rajeev Kumar and Balkrishna Dubey


    Full Text Available Ethosomes are noninvasive delivery carriers that enable drugs to reach the deep skin layers and/or the systemic circulation. These are soft, malleable vesicles tailored for enhanced delivery of active agents. They are composed mainly of phospholipids, high concentration of ethanol and water. The high concentration of ethanol makes the ethosomes unique, as ethanol is known for its disturbance of skin lipid bilayer organization; therefore, when integrated into a vesicle membrane, it gives that vesicle the ability to penetrate the stratum corneum. Also, because of their high ethanol concentration, the lipid membrane is packed less tightly than conventional vesicles but has equivalent stability, allowing a more malleable structure and improves drug distribution ability in stratum corneum lipids. The Ethosomes were found to be suitable for various applications within the pharmaceutical, biotechnology, veterinary, cosmetic, and nutraceutical markets. These “soft vesicles” represents novel vesicular carrier for enhanced delivery to/through skin.

  5. Phosphate solubilizing bacteria and their role in plant growth promotion.

    Rodríguez, H; Fraga, R


    The use of phosphate solubilizing bacteria as inoculants simultaneously increases P uptake by the plant and crop yield. Strains from the genera Pseudomonas, Bacillus and Rhizobium are among the most powerful phosphate solubilizers. The principal mechanism for mineral phosphate solubilization is the production of organic acids, and acid phosphatases play a major role in the mineralization of organic phosphorous in soil. Several phosphatase-encoding genes have been cloned and characterized and a few genes involved in mineral phosphate solubilization have been isolated. Therefore, genetic manipulation of phosphate-solubilizing bacteria to improve their ability to improve plant growth may include cloning genes involved in both mineral and organic phosphate solubilization, followed by their expression in selected rhizobacterial strains. Chromosomal insertion of these genes under appropriate promoters is an interesting approach.

  6. Characterization of the mechanisms for coal solubilization by filamentous fungi

    Fernandez, M.; Luna, N.; Monistrol, I.F.; Laborda, F. [Madrid Univ. (Spain). Dept. de Microbiologia y Parasitologia


    The mechanisms implicated on coal solubilization have been studied in this work. Two fungal strains (Trichoderma and Penicillium) has been isolated which were able to solubilize different kinds of Spanish coals (hard coal, subbituminous coal and lignite) when growing in liquid media in the presence of coal. Extracellular peroxidase, esterase and some times phenol oxidase enzymes were present in the culture supernatant when a high degree of coal solubilization was attained. Those enzymes could be involved in the coal attack. (orig.)

  7. Composition Effect of the Outer Layer on the Vesicle Fusion Catalyzed by Phospholipase D

    Park, Jin Won [Seoul National University, Seoul (Korea, Republic of)


    Phospholipase D (PLD) catalyzed the generation of phosphatidic acid (PA) from phosphatidylcholine (PC) at the outer layer of the vesicles prepared through layer by layer via a double emulsion technique. The generation induced a curvature change in the vesicles, which eventually led them to fuse each other. The ratio of two-fattyacid-tail ethanolamine (PE) to one-fatty-acid-tail ethanolamine (PE) was found to acquire the condition where the mixed-phospholipid vesicles were stable identically with pure two-fatty-acid-tail PC. The effect of the outer-layer mixture on the PLD-induced vesicle fusion was investigated using the fluorescence intensity change. 8-Aminonaph- thalene-1,3,6-trisulfonic acid disodium salt (ANTS) and p-Xylene-bis(N-pyridinium bromide) (DPX) were encapsulated in the vesicles, respectively, for the quantification of the fusion. The fluorescence scale was calibrated with the fluorescence of a 1/1 mixture of ANTS and DPX vesicles in NaCl buffer taken as 100% fluorescence (0% fusion) and the vesicles containing both ANTS and DPX as 0% fluorescence (100% fusion), considering the leakage into the medium studied directly in a separate experiment using vesicles containing both ANTS and DPX. The fusion data for each composition were acquired with the subtraction of the leakage from the quenching. From the monitoring, the vesicle fusion caused by the PLD reaction seems dominantly to occur rather than the vesicle lysis, because the composition effect on the fusion was observed identically with that on the change in the vesicle structure. Furthermore, the diameter measurements also support the fusion dominancy.

  8. Phosphate solubilizing bacteria around Indian peninsula

    DeSouza, M.J.B.D; Nair, S.; Chandramohan, D

    nitrophenylphosphate by the culture. One mole of nitrophenol is equivalent to 1 atom of phosphorus. The absorbance was read at 418 nm using DU-6 Beckmann spectrophotometer. The activity was expressed as ?mol.Pml-1d-1. The cultures which showed good phosphate... strengthening the fact that the isolates are of marine origin. Maximum activity of 5 ? was observed for strain P6 in 4.5% NaCl Location Beaches Islands Coasts Offshore Table 1?Phosphate solubilizing bacteria in various niches. No. of cultures...

  9. Bacterial phospholipide antigens and their taxonomic significance.

    Karalnik, B V; Razbash, M P; Akhmetova, E A


    The investigation of interrelationships between the phospholipides of various microorganisms (33 strains of corynebacteria, mycobacteria and staphylococci) using crossed antibody neutralization reactions with phospholipide antigenic erythrocyte diagnostic was used for the assessment of the degree of antigenic propinquity and antigenic differences between the phospholipides of bacteria of the same species, genus, and of different genera. The role of the determinants of the corresponding (their own) and "foreign" genera in the antigenic differences between the phospholipides of the microorganisms investigated was established. On the basis of the results obtained the conclusion has been drawn that the method of assessment of antigenic interrelationships between phospholipides can be used for the study of some taxonomic problems.

  10. Phospholipids in foods: prooxidants or antioxidants?

    Cui, Leqi; Decker, Eric A


    Lipid oxidation is one of the major causes of quality deterioration in natural and processed foods and thus a large economic concern in the food industry. Phospholipids, especially lecithins, are already widely used as natural emulsifiers and have been gaining increasing interest as natural antioxidants to control lipid oxidation. This review summarizes the fatty acid composition and content of phospholipids naturally occurring in several foods. The role of phospholipids as substrates for lipid oxidation is discussed, with a focus on meats and dairy products. Prooxidant and antioxidant mechanisms of phospholipids are also discussed to get a better understanding of the possible opportunities for using phospholipids as food antioxidants.

  11. Calcium transport in vesicles energized by cytochrome oxidase

    Rosier, Randy N. [Univ. of Rochester, NY (United States)


    Experiments on the reconstitution of cytochrome oxidase into phospholipid vesicles were carried out using techniques of selectivity energizing the suspensions with ascorbate and cytochrome c or ascorbate, PMS, and internally trapped cytochrome c. It was found that the K+ selective ionophore valinomycin stimulated the rate of respiration of cytochrome oxidase vesicles regardless of the direction of the K+ flux across the vesicle membranes. The stimulation occurred in the presence of protonophoric uncouplers and in the complete absence of potassium or in detergent-lysed suspensions. Gramicidin had similar effects and it was determined that the ionophores acted by specific interaction with cytochrome oxidase rather than by the previously assumed collapse of membrane potentials. When hydrophobic proteins and appropriate coupling factors were incorporated into the cytochrome oxidase, vesicles phosphorylation of ADP could be coupled to the oxidation reaction of cytochrome oxidase. Relatively low P:O, representing poor coupling of the system, were problematical and precluded measurements of protonmotive force. However the system was used to study ion translocation.

  12. Liquefaction/solubilization processes of Spanish coals by microorganisms

    Laborda, F.; Monistrol, I.F.; Luna, N.; Fernandez, M. [Madrid Univ. (Spain). Dept. de Microbiologia y Parasitologia


    Several fundamental aspects of microbial coal liquefaction/solubilization have been studied in this work. The first one is the mechanisms implicated on coal transformation. During coal solubilization, fungal cells produced extracellular peroxidase, esterase and some times phenol oxidase enzymes which appear to be involved in solubilization. Moreover, the analysis of liquefaction/solubilization products was done. In this regard, a reduction on the average size of humic acids derived from lignite was observed, probably due to depolymerization caused by microorganisms. Finally, microorganisms showed a specific adherence to the coal surface, that seems to promote the microbial attack to coal. (orig.)

  13. Purification of RNA by SDS solubilization and phenol extraction.

    Rio, Donald C; Ares, Manuel; Hannon, Gregory J; Nilsen, Timothy W


    This protocol describes a method for RNA purification by sodium dodecyl sulfate (SDS) solubilization and phenol extraction. It is of wide utility and is used routinely to deproteinize RNAs in biological material that has been solubilized in SDS, an ionic detergent that dissolves membranes, disrupts protein-nucleic acid interactions, and inactivates ribonucleases. Once solubilized, addition of phenol or phenol:chloroform:isoamyl alcohol (PCA) completely denatures the protein, and it becomes insoluble in aqueous solution. PCA extraction is the method of choice for preparing cytoplasmic RNA from tissue culture cells or in any other situation (e.g., enzyme reactions) where solubilization in SDS is easily achievable.

  14. Thermally-induced aggregation and fusion of protein-free lipid vesicles.

    Ibarguren, Maitane; Bomans, Paul H H; Ruiz-Mirazo, Kepa; Frederik, Peter M; Alonso, Alicia; Goñi, Félix M


    Membrane fusion is an important phenomenon in cell biology and pathology. This phenomenon can be modeled using vesicles of defined size and lipid composition. Up to now fusion models typically required the use of chemical (polyethyleneglycol, cations) or enzymatic catalysts (phospholipases). We present here a model of lipid vesicle fusion induced by heat. Large unilamellar vesicles consisting of a phospholipid (dioleoylphosphatidylcholine), cholesterol and diacylglycerol in a 43:57:3 mol ratio were employed. In this simple system, fusion was the result of thermal fluctuations, above 60 °C. A similar system containing phospholipid and cholesterol but no diacylglycerol was observed to aggregate at and above 60 °C, in the absence of fusion. Vesicle fusion occurred under our experimental conditions only when (31)P NMR and cryo-transmission electron microscopy of the lipid mixtures used in vesicle preparation showed non-lamellar lipid phase formation (hexagonal and cubic). Non-lamellar structures are probably the result of lipid reassembly of the products of individual fusion events, or of fusion intermediates. A temperature-triggered mechanism of lipid reassembly might have occurred at various stages of protocellular evolution.

  15. Pellet-free isolation of human and bovine milk extracellular vesicles by size-exclusion chromatography

    Blans, Kristine Ingrid Marie; Hansen, Maria Stenum; Sørensen, Laila V.


    -marker proteins in other relevant milk fractions such as milk fat globules. Nanoparticle tracking analysis and electron microscopy reveals the presence of heterogeneous sized vesicle structures in milk EV isolates. Lipid analysis by thin layer chromatography shows that EV isolates are devoid of triacylglycerides...... accomplished in three steps based on size-exclusion chromatography (SEC) resulting in effective and reproducible EV isolation from raw milk. The approaches do not require any EV pelleting and can be applied to both human and bovine milk. We show that SEC effectively separates phospholipid membrane vesicles...

  16. Pairing of cholesterol with oxidized phospholipid species in lipid bilayers

    Khandelia, Himanshu; Loubet, Bastien; Olzynska, Agnieszka


    We claim that (1) cholesterol protects bilayers from disruption caused by lipid oxidation by sequestering conical shaped oxidized lipid species such as 1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine (PZPC) away from phospholipid, because cholesterol and the oxidized lipid have complementary...... shapes and (2) mixtures of cholesterol and oxidized lipids can self-assemble into bilayers much like lysolipid–cholesterol mixtures. The evidence for bilayer protection comes from molecular dynamics (MD) simulations and dynamic light scattering (DLS) measurements. Unimodal size distributions of extruded...... vesicles (LUVETs) made up of a mixture of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and PZPC containing high amounts of PZPC are only obtained when cholesterol is present in high concentrations. In simulations, bilayers containing high amounts of PZPC become porous, unless cholesterol is also present...

  17. Nanomechanics of electrospun phospholipid fiber

    Mendes, Ana C., E-mail:, E-mail:; Chronakis, Ioannis S., E-mail:, E-mail: [Technical University of Denmark, DTU-Food, Søltofts Plads B227, DK-2800, Kgs. Lyngby (Denmark); Nikogeorgos, Nikolaos; Lee, Seunghwan [Department of Mechanical Engineering, Technical University of Denmark, DK-2800 Kgs. Lyngby (Denmark)


    Electrospun asolectin phospholipid fibers were prepared using isooctane as a solvent and had an average diameter of 6.1 ± 2.7 μm. Their mechanical properties were evaluated by nanoindentation using Atomic Force Microscopy, and their elastic modulus was found to be approximately 17.2 ± 1 MPa. At a cycle of piezo expansion-retraction (loading-unloading) of a silicon tip on a fiber, relatively high adhesion was observed during unloading. It is proposed that this was primarily due to molecular rearrangements at the utmost layers of the fiber caused by the indentation of the hydrophilic tip. The phospholipid fibers were shown to be stable in ambient conditions, preserving the modulus of elasticity up to 24 h.

  18. Modulating alignment of membrane proteins in liquid-crystalline and oriented gel media by changing the size and charge of phospholipid bicelles

    Lorieau, Justin L; Maltsev, Alexander S.; Louis, John M; Bax, Ad


    We demonstrate that alignment of a structured peptide or small protein solubilized in mixed phospholipid:detergent micelles or bicelles, when embedded in a compressed gel or liquid crystalline medium, can be altered by either changing the phospholipid aggregate shape, charge, or both together. For the hemagglutinin fusion peptide solubilized in bicelles, we show that bicelle shape and charge do not change its helical hairpin structure but impact its alignment relative to the alignment medium, both in charged compressed acrylamide gel and in liquid crystalline d (GpG). The method can be used to generate sets of residual dipolar couplings (RDCs) that correspond to orthogonal alignment tensors, and holds promise for high-resolution structural refinement and dynamic mapping of membrane proteins. PMID:23508769

  19. Modulating alignment of membrane proteins in liquid-crystalline and oriented gel media by changing the size and charge of phospholipid bicelles

    Lorieau, Justin L.; Maltsev, Alexander S.; Louis, John M.; Bax, Ad, E-mail: [National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health, Laboratory of Chemical Physics (United States)


    We demonstrate that alignment of a structured peptide or small protein solubilized in mixed phospholipid:detergent micelles or bicelles, when embedded in a compressed gel or liquid crystalline medium, can be altered by either changing the phospholipid aggregate shape, charge, or both together. For the hemagglutinin fusion peptide solubilized in bicelles, we show that bicelle shape and charge do not change its helical hairpin structure but impact its alignment relative to the alignment medium, both in charged compressed acrylamide gel and in liquid crystalline d(GpG). The method can be used to generate sets of residual dipolar couplings that correspond to orthogonal alignment tensors, and holds promise for high-resolution structural refinement and dynamic mapping of membrane proteins.

  20. [Bile phospholipids; function and significance].

    Salvioli, G; Salati, R


    The part played by phospholipides in the genesis of cholesterol gallstone considered. This is present in patients who frequently present a lecithin synthesis defect at hepatic level since precursors are used for forming triglycerides. Nevertheless polyunsaturated phosphatidicholine has a negative influence on the SB + PL/C ratio in the bile of T-tube subjects receiving 2 g of substance i.v. for 5 days.

  1. A vesicle bioreactor as a step toward an artificial cell assembly

    Noireaux, Vincent; Libchaber, Albert


    An Escherichia coli cell-free expression system is encapsulated in a phospholipid vesicle to build a cell-like bioreactor. Large unilamellar vesicles containing extracts are produced in an oil-extract emulsion. To form a bilayer the vesicles are transferred into a feeding solution that contains ribonucleotides and amino acids. Transcription-translation of plasmid genes is isolated in the vesicles. Whereas in bulk solution expression of enhanced GFP stops after 2 h, inside the vesicle permeability of the membrane to the feeding solution prolongs the expression for up to 5 h. To solve the energy and material limitations and increase the capacity of the reactor, the -hemolysin pore protein from Staphylococcus aureus is expressed inside the vesicle to create a selective permeability for nutrients. The reactor can then sustain expression for up to 4 days with a protein production of 30 µM after 4 days. Oxygen diffusion and osmotic pressure are critical parameters to maintain expression and avoid vesicle burst. -hemolysin | cell-free protein expression | membrane-anchoring polypeptide

  2. Haloarchaea Endowed with Phosphorus Solubilization Attribute Implicated in Phosphorus Cycle.

    Yadav, Ajar Nath; Sharma, Divya; Gulati, Sneha; Singh, Surender; Dey, Rinku; Pal, Kamal Krishna; Kaushik, Rajeev; Saxena, Anil Kumar


    Archaea are unique microorganisms that are present in ecological niches of high temperature, pH and salinity. A total of 157 archaea were obtained from thirteen sediment, water and rhizospheric soil samples collected from Rann of Kutch, Gujarat, India. With an aim to screen phosphate solubilizing archaea, a new medium was designed as Haloarchaea P Solubilization (HPS) medium. The medium supported the growth and P solubilization activity of archaea. Employing the HPS medium, twenty isolates showed the P-solubilization. Phosphate solubilizing archaea were identified as seventeen distinct species of eleven genera namely Haloarcula, Halobacterium, Halococcus, Haloferax, Halolamina, Halosarcina, Halostagnicola, Haloterrigena, Natrialba, Natrinema and Natronoarchaeum. Natrinema sp. strain IARI-WRAB2 was identified as the most efficient P-solubilizer (134.61 mg/L) followed by Halococcus hamelinensis strain IARI-SNS2 (112.56 mg/L). HPLC analysis detected seven different kinds of organic acids, namely: gluconic acid, citric acid, formic acid, fumaric acid succinic acid, propionic acid and tartaric acid from the cultures of these isolates. These phosphate solubilizing halophilic archaea may play a role in P nutrition to vegetation growing in these hypersaline soils. This is the first report for these haloarchaea to solubilize considerable amount of P by production of organic acids and lowering of pH.

  3. [The solubilization of four insoluble phosphates by some microorganisms].

    Zhao, Xiaorong; Lin, Qimei; Li, Baoguo


    Four insoluble phosphates of ferric phosphate (Fe-P), aluminum phosphate (Al-P), fluorapatite (FAP) and rock phosphate (RP) were used as a sole phosphorus resource for some phosphate-solubilizing microorganisms. It was found that there was significant difference in solubilizing these phosphates by the tested isolates. The fungi normally were more powerful than the bacteria in dissolving the phsophates. The microorganisms generally solubilized more phosphate when supplied with NO3- than with NH4+. However, the isolates of 2TCiF2 and 4TCiF6 had much higher capacity to solubilize FAP and Al-P respectively in NH4+ medium. Most of the isolates solubilized readily FAP and RP, and then Al-P. Ferric phosphate was the least soluble to these isolates. Only isolate 2TCiF2 showed strong ability to solubilize Fe-P. In particular, two Aspergillus sp. had much higher capacity of dissolving Fe-P when suppled with NO3-. The isolates of Evwinia sp. 4TCRi22 and Enterobacter sp. 1TCRi15 had higher capacity of solubilizing FAP. But two Arthrobacter sp. showed the highest activity in RP medium. It is supposed that complexion of organic acids with metals may be the main reason for these isolates to solubilize the phosphates. However, other chelant substances may be much more important for Enterobacter sp. and Erwinia sp. to release phosphorus from the phsphates.

  4. Association of endogenous substrate with solubilized bovine brain sialidase.

    Schengrund, C L; Repman, M A


    Nonidet P40 solubilized up to 90% of the sialidase, active towards added ganglioside substrate, that was associated with the total membrane fraction prepared from gray matter of bovine brains. Solubilized sialidase acted upon endogenous substrate (sialic acid containing compounds solubilized with the enzyme), hydrolyzing approximately 50% of the readily available sialosyl residues within 20 min. During a 2-hr reaction time 80% of the polysialylated gangliosides solubilized with the enzyme were acted upon. A 20-min lag was observed before sialidase acted upon added ganglioside substrate. The lag could be reduced to less than 2 min when the enzyme was allowed to act on endogenous substrate prior to exposure to exogenous substrate, suggesting that the solubilized enzyme acted preferentially on endogenous substrate. A protease inhibitor prevented much of the 86% loss of activity towards added substrate that was seen when the enzyme was stored at 4 degrees C for 6 days; activity towards endogenous substrate decreased only 34%.


    Sandeep Kumar Parihar*, Mithun Bhowmick, Rajeev Kumar and Balkrishna Dubey


    Ethosomes are noninvasive delivery carriers that enable drugs to reach the deep skin layers and/or the systemic circulation. These are soft, malleable vesicles tailored for enhanced delivery of active agents. They are composed mainly of phospholipids, high concentration of ethanol and water. The high concentration of ethanol makes the ethosomes unique, as ethanol is known for its disturbance of skin lipid bilayer organization; therefore, when integrated into ...

  6. Evolution of chloroplast vesicle transport.

    Westphal, Sabine; Soll, Jürgen; Vothknecht, Ute C


    Vesicle traffic plays a central role in eukaryotic transport. The presence of a vesicle transport system inside chloroplasts of spermatophytes raises the question of its phylogenetic origin. To elucidate the evolution of this transport system we analyzed organisms belonging to different lineages that arose from the first photosynthetic eukaryote, i.e. glaucocystophytes, chlorophytes, rhodophytes, and charophytes/embryophytes. Intriguingly, vesicle transport is not apparent in any group other than embryophytes. The transfer of this eukaryotic-type vesicle transport system from the cytosol into the chloroplast thus seems a late evolutionary development that was acquired by land plants in order to adapt to new environmental challenges.

  7. Phosphate Solubilizing Bacteria Adaptive to Vinasse

    Kahar Muzakhar


    Full Text Available Microorganisms identified as phosphate solubilizing bacteria (PSB adaptive to vinasse were successfully screened from sugarcane soil from an agriculatural estate in Jatiroto. By conducting a screening on Pikovskaya’s agar medium (PAM, we found that five different isolates were detected as PSB (pvk-5a, pvk-5b, pvk-6b, pvk-7a, and pvk-8a. Of the five isolates only three could be grown and were found to be adaptive to vinasse based medium without any nutrients added (pvk-5a, pvk-5b and pvk-7a. The three isolates were characterized as coccus and Gram negative with no endospores detected. We suggest that these three isolates can be used as biofertilizer agent to support organic farming.

  8. The toolbox of vesicle sidedness determination

    Meszaros, Peter; Hoekstra, Dick; Kok, Jan Willem


    Vesicles prepared from cellular plasma membranes are widely used in science for different purposes. The outer membrane leaflet differs from the inner membrane leaflet of the vesicle, and during vesicle preparation procedures two types of vesicles will be generated: right-side-out vesicles, of which

  9. Reduced complement-mediated immune complex solubilizing capacity and the presence of incompletely solubilized immune complexes in SLE sera

    Baatrup, Gunnar; Petersen, I; Jensenius, J C


    Reduced complement-mediated solubilization (CMS) of pre-formed immune complexes (IC) was demonstrated in sera from 11 out of 12 SLE patients. The presence of incompletely solubilized endogeneous IC in SLE sera was indicated by the following findings: (1) When IC positive SLE sera with reduced CMS...

  10. Softening of phospholipid membranes by the adhesion of silica nanoparticles - as seen by neutron spin-echo (NSE)

    Hoffmann, Ingo; Michel, Raphael; Sharp, Melissa; Holderer, Olaf; Appavou, Marie-Sousai; Polzer, Frank; Farago, Bela; Gradzielski, Michael


    The interactions between nanoparticles and vesicles are of significant interest both from a fundamental as well as from a practical point of view, as vesicles can serve as a model system for cell membranes. Accordingly the effect of nanoparticles that bind to the vesicle bilayer is very important with respect to understanding their biological impact and also may shed some light on the mechanisms behind the effect of nanotoxicity. In this study we have investigated the influence of small adsorbed silica nanoparticles (SiNPs) on the structure of zwitterionic DOPC vesicles. By a combination of SANS, cryo-TEM, and DLS, we observed that the SiNPs are bound to the outer vesicle surface without significantly affecting the vesicle structure. Most interestingly, by means of neutron spin-echo (NSE) local bilayer fluctuations were studied and one finds a small but marked decrease of the membrane rigidity upon binding of the nanoparticles. This surprising finding may be a relevant aspect for the further understanding of the effects that nanoparticles have on phospholipid bilayers.The interactions between nanoparticles and vesicles are of significant interest both from a fundamental as well as from a practical point of view, as vesicles can serve as a model system for cell membranes. Accordingly the effect of nanoparticles that bind to the vesicle bilayer is very important with respect to understanding their biological impact and also may shed some light on the mechanisms behind the effect of nanotoxicity. In this study we have investigated the influence of small adsorbed silica nanoparticles (SiNPs) on the structure of zwitterionic DOPC vesicles. By a combination of SANS, cryo-TEM, and DLS, we observed that the SiNPs are bound to the outer vesicle surface without significantly affecting the vesicle structure. Most interestingly, by means of neutron spin-echo (NSE) local bilayer fluctuations were studied and one finds a small but marked decrease of the membrane rigidity upon

  11. Interaction of caldesmon with endoplasmic reticulum membrane: effects on the mobility of phospholipids in the membrane and on the phosphatidylserine base-exchange reaction.

    Makowski, P; Makuch, R; Sikorski, A F; Jezierski, A; Pikula, S; Dabrowska, R


    We have previously demonstrated by tryptophan fluorescence the interaction of caldesmon with anionic phospholipid vesicles [Czurylo, Zborowski and Dabrowska (1993) Biochem. J. 291, 403-408]. In the present work we investigated the interaction of caldesmon with natural-membrane (rat liver endoplasmic reticulum) phospholipids by co-sedimentation assay. The results indicate that 1 mol of caldesmon binds approx. 170 mol of membrane phospholipids with a binding affinity constant of 7.3 x 10(6) M-1. The caldesmon-membrane phospholipid complex dissociates with increasing salt concentration and in the presence of Ca2+/calmodulin. As indicated by EPR measurements of membrane lipids labelled with 5-doxyl stearate and TEMPO-phosphatidylethanolamine, binding of caldesmon results in an increase in mobility of the acyl chains (in the region of carbon 5) and a decrease in polar headgroup mobility of phospholipids. Interaction of caldesmon with phospholipids is accompanied by inhibition of phosphatidylethanolamine synthesis via a phospholipid base-exchange reaction, with phosphatidylserine as substrate. This shows that, of the endoplasmic reticulum membrane phospholipids, the main target of caldesmon is phosphatidylserine. PMID:9371708

  12. Ether- versus ester-linked phospholipid bilayers containing either linear or branched apolar chains.

    Balleza, Daniel; Garcia-Arribas, Aritz B; Sot, Jesús; Ruiz-Mirazo, Kepa; Goñi, Félix M


    We studied the properties of bilayers formed by ether-and ester-containing phospholipids, whose hydrocarbon chains can be either linear or branched, using sn-1,2 dipalmitoyl, dihexadecyl, diphytanoyl, and diphytanyl phosphatidylcholines (DPPC, DHPC, DPhoPC, and DPhPC, respectively) either pure or in binary mixtures. Differential scanning calorimetry and confocal fluorescence microscopy of giant unilamellar vesicles concurred in showing that equimolar mixtures of linear and branched lipids gave rise to gel/fluid phase coexistence at room temperature. Mixtures containing DHPC evolved in time (0.5 h) from initial reticulated domains to extended solid ones when an equilibrium was achieved. The nanomechanical properties of supported planar bilayers formed by each of the four lipids studied by atomic force microscopy revealed average breakdown forces Fb decreasing in the order DHPC ≥ DPPC > DPhoPC > DPhPC. Moreover, except for DPPC, two different Fb values were found for each lipid. Atomic force microscopy imaging of DHPC was peculiar in showing two coexisting phases of different heights, probably corresponding to an interdigitated gel phase that gradually transformed, over a period of 0.5 h, into a regular tilted gel phase. Permeability to nonelectrolytes showed that linear-chain phospholipids allowed a higher rate of solute + water diffusion than branched-chain phospholipids, yet the former supported a smaller extent of swelling of the corresponding vesicles. Ether or ester bonds appeared to have only a minor effect on permeability.

  13. Incorporation of peptides in phospholipid aggregates using ultrasound.

    Silva, Raquel; Little, Collin; Ferreira, Helena; Cavaco-Paulo, Artur


    This study presents the highlights of ultrasonic effects on peptides incorporated on phospholipid aggregates (liposomes). These liposomes or vesicles are known as transport agents in skin drug delivery and for hair treatment. They might be a good model to deliver larger peptides into hair to restore fibre strength after hair coloration, modelling, permanent wave and/or straightening. The preparation of liposomes 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC) with peptides (LLLLK LLLLK LLLLK LLLLK; LLLLL LCLCL LLKAK AK) was made by the thin film hydration method. The LUVs (uni-lamellar vesicles) were obtained by sonication, applying different experimental conditions, such as depth (mm) and power intensity (%). Photon-correlation spectroscopy (PCS) and electronic microscopy (EM) results confirmed that the incorporation of these peptides, with different sequence of amino acids, presented differences on the diameter, zeta-potential of membrane surface and shape of liposomes. The liposomes that included peptide LLLLK LLLLK LLLLK LLLLK present an increased in zeta-potential values after using ultrasound and an "amorphous" aspect. Conversely, the liposomes that incorporated the peptide LLLLL LCLCL LLKAK AK presented a define shape (rod shape) and the potential surface of liposome did not change significantly by the use of ultrasound.

  14. Vesicles and vesicle fusion: coarse-grained simulations

    Shillcock, Julian C.


    of vesicles that is crucial for this transport is their ability to fuse to target membranes and release their contents to the distal side. In industry, some personal care products contain vesicles to help transport reagents across the skin, and research on drug formulation shows that packaging active...

  15. In vivo Stability of Ester- and Ether-Linked Phospholipid-Containing Liposomes as Measured by Perturbed Angular Correlation Spectroscopy

    Derksen, Johannes T.; Baldeschwieler, John D.; Scherphof, Gerrit L.


    To evaluate liposome formulations for use as intracellular sustained-release drug depots, we have compared the uptake and degradation in rat liver and spleen of liposomes of various compositions, containing as their bulk phospholipid an ether-linked phospholipid or one of several ester-linked phospholipids, by perturbed angular correlation spectroscopy. Multilamellar and small unilamellar vesicles (MLVs and SUVs), composed of egg phosphatidylcholine, sphingomyelin, distearoyl phosphatidylcholine (DSPC), dipalmitoyl phosphatidylcholine (DPPC) or its analog dihexadecylglycerophosphorylcholine (DHPC), and cholesterol plus phosphatidylserine, and containing 111In complexed to nitrilotriacetic acid, were injected intravenously in rats. Recovery of 111In-labeled liposomes in blood, liver, and spleen was assessed at specific time points after injection and the percentage of liposomes still intact in liver and spleen was determined by measurement of the time-integrated angular perturbation factor 111In of the [G22(∞ )] label. We found that MLVs but not SUVs, having DHPC as their bulk phospholipid, showed an increased resistance against lysosomal degradation as compared to other phospholipid-containing liposomes. The use of diacyl phospholipids with a high gel/liquid-crystalline phase-transition temperature, such as DPPC and DSPC, also retarded degradation of MLV, but not of SUV in the dose range tested, while the rate of uptake of these liposomes by the liver was lower.

  16. Vesicles and vesicle gels - structure and dynamics of formation

    Gradzielski, M


    Vesicles constitute an interesting morphology formed by self-aggregating amphiphilic molecules. They exhibit a rich structural variety and are of interest both from a fundamental point of view (for studying closed bilayer systems) and from a practical point of view (whenever one is interested in the encapsulation of active molecules). In many circumstances vesicular structures have to be formed by external forces, but of great interest are amphiphilic systems, where they form spontaneously. Here the question arises of whether this means that they are also thermodynamically stable structures, which at least in some systems appears to be the case. If such vesicles are well defined in size, it is possible to pack them densely and thereby form vesicle gels that possess highly elastic properties even for relatively low volume fractions of amphiphile. Conditions for the formation and the microstructure of such vesicle gels have been studied in some detail for the case of unilamellar vesicles. Another important and ...

  17. Extracellular vesicles for drug delivery

    Vader, Pieter; Mol, Emma A; Pasterkamp, Gerard; Schiffelers, Raymond M


    Extracellular vesicles (EVs) are cell-derived membrane vesicles, and represent an endogenous mechanism for intercellular communication. Since the discovery that EVs are capable of functionally transferring biological information, the potential use of EVs as drug delivery vehicles has gained consider

  18. Phosphate Solubilization Potentials of Rhizosphere Isolates from Central Anatolia (Turkey)

    Ogut, M.; Er, F.


    Plant available-phosphorus (P) is usually low in Anatolian soils due mainly to the precipitation as calcium (Ca) and magnesium (Mg) phosphates in alkaline conditions. Phosphate solubilizing microorganisms (PSM) can enhance plant P-availability by dissolving the hardly soluble-P within the rhizosphere, which is the zone that surrounds the plant roots. PSM's can be used as seed- or soil-inocula to increase plant P-uptake and the overall growth. A total of 162 PSM's were isolated from the rhizosphere of wheat plants excavated from different fields located along a 75 km part of a highway in Turkey. The mean, the standart deviation, and the median for solubilized-P (ppm) in a 24 h culture in a tricalcium phosphate broth were 681, 427, and 400 for glucose; 358, 266, and 236 for sucrose; and 102, 117, and 50 for starch, respectively. There was not a linear relationship between the phosphate solubilized in the liquid cultures and the solubilization index obtained in the Pikovskaya's agar. Nine isolates representing both weak and strong solubilizers [Bacillus megaterium (5), Bacillus pumilis (1), Pseudomonas syringae pv. phaseolica (1), Pseudomonas fluorescens (1), Arthrobacter aurescens (1) as determined by the 16S rRNA gene sequence analysis] were further studied in a five day incubation. Pseudomonas syringae pv. phaseolica solubilized statistically (P<0.05) higher phosphate (409 ppm) than all the other strains did. There was not a statistically significant (P<0.05) difference in solubilized-P among the Bacillus strains. The pH of the medium fell to the levels between 4 and 5 from the initial neutrality. The phosphate solubilizing strains variably produced gluconic, 2-keto-D-gluconic, glycolic, acetic and butyric acids. The organic acids produced by these microorganisms seem to be the major source of phosphate solubilization in vitro.

  19. Synaptic vesicle pools and dynamics.

    Alabi, AbdulRasheed A; Tsien, Richard W


    Synaptic vesicles release neurotransmitter at chemical synapses, thus initiating the flow of information in neural networks. To achieve this, vesicles undergo a dynamic cycle of fusion and retrieval to maintain the structural and functional integrity of the presynaptic terminals in which they reside. Moreover, compelling evidence indicates these vesicles differ in their availability for release and mobilization in response to stimuli, prompting classification into at least three different functional pools. Ongoing studies of the molecular and cellular bases for this heterogeneity attempt to link structure to physiology and clarify how regulation of vesicle pools influences synaptic strength and presynaptic plasticity. We discuss prevailing perspectives on vesicle pools, the role they play in shaping synaptic transmission, and the open questions that challenge current understanding.

  20. Processes of liquefaction/solubilization of Spanish coals by microorganisms

    Laborda, F.; Monistrol, I.F.; Luna, N.; Fernandez, M. [Universidad de Alcala de Henares, Madrid (Spain). Dept. de Microbiologia y Parasitologia


    Several fundamental aspects of microbial coal liquefaction/solubilization were studied. The liquefied/solubilized products from coal by microorganisms were analysed. The liquid products analysed by IR titration and UV/visible spectrometry showed some alterations with regard to the original coal. Humic acids extracted from the liquefied lignite showed a reduction in the average molecular weight and a increase in the condensation index, probably due to depolymerization caused by microorganisms. The mechanisms implicated in coal biosolubilization by two fungal strains, M2 (Trichoderma sp.) and M4 (Penicillium sp.) were also studied. Extracellular peroxidase, esterase and phenoloxidase enzymes appear to be involved in coal solubilization. (orig.)

  1. Solubilization of proteins: the importance of lysis buffer choice.

    Peach, Mandy; Marsh, Noelle; Miskiewicz, Ewa I; MacPhee, Daniel J


    The efficient extraction of proteins of interest from cells and tissues is not always straightforward. Here we demonstrate the differences in extraction of the focal adhesion protein Kindlin-2 from choriocarcinoma cells using NP-40 and RIPA lysis buffer. Furthermore, we demonstrate the use of a more denaturing urea/thiourea lysis buffer for solubilization, by comparing its effectiveness for solubilization of small heat-shock proteins from smooth muscle with the often utilized RIPA lysis buffer. Overall, the results demonstrate the importance of establishing the optimal lysis buffer for specific protein solubilization within the experimental workflow.

  2. Solubilization of benzene and cyclohexane by sodium deoxycholate micelles

    Christian, S.D.; Smith, L.S.; Bushong, D.S.; Tucker, E.E.


    Vapor pressure-solubility data were obtained for the aqueous systems benzene-sodium deoxycholate and cyclohexane- sodium deoxycholate at 25/sup 0/C. The results are consistent with a mass action model similar to the BET equation. Equilibrium constants are inferred to characterize interactions of hydrocarbons with solubilization sites assumed to consist of units of four deoxycholate anions. Although addition of sodium chloride increases the middle aggregation number, solubilization results are affected very little by variation in salt concentration. When pure liquid hydrocarbon standard states are employed, solubilization results for benzene and cyclohexane (at varying salt concentrations) are quite similar. 26 references.

  3. Phospholipid liposomes functionalized by protein

    Glukhova, O. E.; Savostyanov, G. V.; Grishina, O. A.


    Finding new ways to deliver neurotrophic drugs to the brain in newborns is one of the contemporary problems of medicine and pharmaceutical industry. Modern researches in this field indicate the promising prospects of supramolecular transport systems for targeted drug delivery to the brain which can overcome the blood-brain barrier (BBB). Thus, the solution of this problem is actual not only for medicine, but also for society as a whole because it determines the health of future generations. Phospholipid liposomes due to combination of lipo- and hydrophilic properties are considered as the main future objects in medicine for drug delivery through the BBB as well as increasing their bioavailability and toxicity. Liposomes functionalized by various proteins were used as transport systems for ease of liposomes use. Designing of modification oligosaccharide of liposomes surface is promising in the last decade because it enables the delivery of liposomes to specific receptor of human cells by selecting ligand and it is widely used in pharmacology for the treatment of several diseases. The purpose of this work is creation of a coarse-grained model of bilayer of phospholipid liposomes, functionalized by specific to the structural elements of the BBB proteins, as well as prediction of the most favorable orientation and position of the molecules in the generated complex by methods of molecular docking for the formation of the structure. Investigation of activity of the ligand molecule to protein receptor of human cells by the methods of molecular dynamics was carried out.

  4. Effect of sodium deoxycholate and sodium cholate on DPPC vesicles: A fluorescence anisotropy study with diphenylhexatriene

    Usharani Subuddhi; Ashok K Mishra


    Effects of two bile salts, namely sodium deoxycholate (NaDC) and sodium cholate (NaC), on DPPC small unilamellar vesicles have been investigated using the steady-state fluorescence anisotropy () of diphenylhexatriene (DPH) as a tool. It was found that the variation of is sensitive enough to monitor different stages of interaction of bile salts with DPPC vesicles. NaDC induced significant changes in the membrane well below its CMC (6 mM). Even at 4 mM, which is still lower than the CMC, the phospholipids were completely solubilised by the NaDC micelles. The effect of NaC on DPPC vesicles, however, was much less significant, especially in the sub-micellar concentration regime. Being more hydrophilic NaC does not interact with the membrane efficiently. Complete solubilisation of phospholipids took place only when the concentration of NaC was above its CMC (16 mM). The experiments also showed that the bile salt-induced changes of vesicle structure were strongly dependent on the concentration of the bile salt and not on the molar ratio of lipid and bile salt.

  5. Reduction-sensitive liposomes from a multifunctional lipid conjugate and natural phospholipids: reduction and release kinetics and cellular uptake.

    Goldenbogen, Björn; Brodersen, Nicolai; Gramatica, Andrea; Loew, Martin; Liebscher, Jürgen; Herrmann, Andreas; Egger, Holger; Budde, Bastian; Arbuzova, Anna


    The development of targeted and triggerable delivery systems is of high relevance for anticancer therapies. We report here on reduction-sensitive liposomes composed of a novel multifunctional lipidlike conjugate, containing a disulfide bond and a biotin moiety, and natural phospholipids. The incorporation of the disulfide conjugate into vesicles and the kinetics of their reduction were studied using dansyl-labeled conjugate 1 in using the dansyl fluorescence environmental sensitivity and the Förster resonance energy transfer from dansyl to rhodamine-labeled phospholipids. Cleavage of the disulfide bridge (e.g., by tris(2-carboxyethyl)phosphine (TCEP), dithiothreitol (DTT), l-cysteine, or glutathione (GSH)) removed the hydrophilic headgroup of the conjugate and thus changed the membrane organization leading to the release of entrapped molecules. Upon nonspecific uptake of vesicles by macrophages, calcein release from reduction-sensitive liposomes consisting of the disulfide conjugate and phospholipids was more efficient than from reduction-insensitive liposomes composed only of phospholipids. The binding of streptavidin to the conjugates did not interfere with either the subsequent reduction of the disulfide bond of the conjugate or the release of entrapped molecules. Breast cancer cell line BT-474, overexpressing the HER2 receptor, showed a high uptake of the reduction-sensitive doxorubicin-loaded liposomes functionalized with the biotin-tagged anti-HER2 antibody. The release of the entrapped cargo inside the cells was observed, implying the potential of using our system for active targeting and delivery. © 2011 American Chemical Society

  6. Oriented bilayers of a proteolipid complex, Annexin V phospholipids, for solid state NMR analyses

    Saurel, O.; Demange, P.; Milon, A.


    We were able to obtain oriented bilayers in the presence of annexin V (35 kDa) and to measure structural and dynamic parameters. NMR results obtained so far show that annexin does not affect the structure of the membrane but modifies the dynamics of the phospholipids (T2e decrease). Our new method to prepare oriented biological samples, based on the preparation of small unilamellar proteolipid vesicles in biological buffers, should be suitable for any membrane protein phospholipid complex and for solid state NMR or neutron diffraction experiments. La préparation de petites vésicules protéo-lipidiques dans un tampon physiologique, nous a permis d'obtenir des bicouches orientées en présence d'annexine V (35 kDa) et de mesurer les paramètres structuraux et dynamiques de ce complexe protéolipidique. Les résultats obtenus par RMN du solide montrent que l'annexine n'affecte pas la structure en bicouche de la membrane mais modifie la dynamique des phospholipides (diminution du temps de relaxation T2e). Cette nouvelle méthode de préparation d'échantillons orientés peut être applicable à tout complexe protéine-phospholipides pour des études par RMN à l'état solide ou par diffraction des neutrons.

  7. Allosteric regulation of G protein-coupled receptor activity by phospholipids.

    Dawaliby, Rosie; Trubbia, Cataldo; Delporte, Cédric; Masureel, Matthieu; Van Antwerpen, Pierre; Kobilka, Brian K; Govaerts, Cédric


    Lipids are emerging as key regulators of membrane protein structure and activity. These effects can be attributed either to the modification of bilayer properties (thickness, curvature and surface tension) or to the binding of specific lipids to the protein surface. For G protein-coupled receptors (GPCRs), the effects of phospholipids on receptor structure and activity remain poorly understood. Here we reconstituted purified β2-adrenergic receptor (β2R) in high-density lipoparticles to systematically characterize the effect of biologically relevant phospholipids on receptor activity. We observed that the lipid headgroup type affected ligand binding (agonist and antagonist) and receptor activation. Specifically, phosphatidylgycerol markedly favored agonist binding and facilitated receptor activation, whereas phosphatidylethanolamine favored antagonist binding and stabilized the inactive state of the receptor. We then showed that these effects could be recapitulated with detergent-solubilized lipids, demonstrating that the functional modulation occurred in the absence of a bilayer. Our data suggest that phospholipids act as direct allosteric modulators of GPCR activity.

  8. The use of natural and synthetic phospholipids as pharmaceutical excipients*

    van Hoogevest, Peter; Wendel, Armin


    In pharmaceutical formulations, phospholipids obtained from plant or animal sources and synthetic phospholipids are used. Natural phospholipids are purified from, e.g., soybeans or egg yolk using non-toxic solvent extraction and chromatographic procedures with low consumption of energy and minimum possible waste. Because of the use of validated purification procedures and sourcing of raw materials with consistent quality, the resulting products differing in phosphatidylcholine content possess an excellent batch to batch reproducibility with respect to phospholipid and fatty acid composition. The natural phospholipids are described in pharmacopeias and relevant regulatory guidance documentation of the Food and Drug Administration (FDA) and European Medicines Agency (EMA). Synthetic phospholipids with specific polar head group, fatty acid composition can be manufactured using various synthesis routes. Synthetic phospholipids with the natural stereochemical configuration are preferably synthesized from glycerophosphocholine (GPC), which is obtained from natural phospholipids, using acylation and enzyme catalyzed reactions. Synthetic phospholipids play compared to natural phospholipid (including hydrogenated phospholipids), as derived from the number of drug products containing synthetic phospholipids, a minor role. Only in a few pharmaceutical products synthetic phospholipids are used. Natural phospholipids are used in oral, dermal, and parenteral products including liposomes. Natural phospholipids instead of synthetic phospholipids should be selected as phospholipid excipients for formulation development, whenever possible, because natural phospholipids are derived from renewable sources and produced with more ecologically friendly processes and are available in larger scale at relatively low costs compared to synthetic phospholipids. Practical applications: For selection of phospholipid excipients for pharmaceutical formulations, natural phospholipids are preferred

  9. Effects of PEO-PPO-PEO triblock copolymers on phospholipid membrane integrity under osmotic stress.

    Wang, Jia-Yu; Chin, Jaemin; Marks, Jeremy D; Lee, Ka Yee C


    The effects of PEO-PPO-PEO triblock copolymers, mainly Poloxamer 188, on phospholipid membrane integrity under osmotic gradients were explored using giant unilamellar vesicles (GUVs). Fluorescence leakage assays showed two opposing effects of P188 on the structural integrity of GUVs depending on the duration of their incubation time. A two-state transition mechanism of interaction between the triblock copolymers and the phospholipid membrane is proposed: an adsorption (I) and an insertion (II) state. While the triblock copolymer in state I acts to moderately retard the leakage, their insertion in state II perturbs the lipid packing, thus increasing the membrane permeability. Our results suggest that the biomedical application of PEO-PPO-PEO triblock copolymers, either as cell membrane resealing agents or as accelerators for drug delivery, is directed by the delicate balance between these two states.

  10. Amphiphilic naproxen prodrugs: differential scanning calorimetry study on their interaction with phospholipid bilayers.

    Giuffrida, Maria Chiara; Pignatello, Rosario; Castelli, Francesco; Sarpietro, Maria Grazia


    Naproxen, a nonsteroid anti-inflammatory drug studied for Alzheimer's disease, was conjugated with lipoamino acids (LAA) directly or through a diethylamine (EDA) spacer to improve the drug lipophilicity and the interaction with phospholipid bilayers. The interaction of naproxen and its prodrugs with biomembrane models consisting of dimyristoylphosphatidylcholine multilamellar vesicles was studied by differential scanning calorimetry. The transfer of prodrugs from a lipophilic carrier to a biomembrane model was also studied. Naproxen conjugation to lipoamino acids improves its interaction with biomembrane models and affects the transfer from a lipophilic carrier to biomembrane model. LAA portion may localize between the phospholipid chains; the entity of the interaction depends not only on the presence of the spacer but also on the LAA chain length. Variation of LAA portion can modulate the naproxen prodrugs affinity towards the biological membrane as well as towards the lipophilic carrier. © 2017 Royal Pharmaceutical Society.

  11. Intracellular vesicles as reproduction elements in cell wall-deficient L-form bacteria.

    Yves Briers

    Full Text Available Cell wall-deficient bacteria, or L-forms, represent an extreme example of bacterial plasticity. Stable L-forms can multiply and propagate indefinitely in the absence of a cell wall. Data presented here are consistent with the model that intracellular vesicles in Listeria monocytogenes L-form cells represent the actual viable reproductive elements. First, small intracellular vesicles are formed along the mother cell cytoplasmic membrane, originating from local phospholipid accumulation. During growth, daughter vesicles incorporate a small volume of the cellular cytoplasm, and accumulate within volume-expanding mother cells. Confocal Raman microspectroscopy demonstrated the presence of nucleic acids and proteins in all intracellular vesicles, but only a fraction of which reveals metabolic activity. Following collapse of the mother cell and release of the daughter vesicles, they can establish their own membrane potential required for respiratory and metabolic processes. Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release. Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition. Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species. From a more general viewpoint, this type of multiplication mechanism seems reminiscent of the physicochemical self-reproducing properties of abiotic lipid vesicles used to study the primordial reproduction pathways of putative prokaryotic precursor cells.

  12. Secretory vesicle priming by CAPS is independent of its SNARE-binding MUN domain.

    Nguyen Truong, Cuc Quynh; Nestvogel, Dennis; Ratai, Olga; Schirra, Claudia; Stevens, David R; Brose, Nils; Rhee, JeongSeop; Rettig, Jens


    Priming of secretory vesicles is a prerequisite for their Ca(2+)-dependent fusion with the plasma membrane. The key vesicle priming proteins, Munc13s and CAPSs, are thought to mediate vesicle priming by regulating the conformation of the t-SNARE syntaxin, thereby facilitating SNARE complex assembly. Munc13s execute their priming function through their MUN domain. Given that the MUN domain of Ca(2+)-dependent activator protein for secretion (CAPS) also binds syntaxin, it was assumed that CAPSs prime vesicles through the same mechanism as Munc13s. We studied naturally occurring splice variants of CAPS2 in CAPS1/CAPS2-deficient cells and found that CAPS2 primes vesicles independently of its MUN domain. Instead, the pleckstrin homology domain of CAPS2 seemingly is essential for its priming function. Our findings indicate a priming mode for secretory vesicles. This process apparently requires membrane phospholipids, does not involve the binding or direct conformational regulation of syntaxin by MUN domains of CAPSs, and is therefore not redundant with Munc13 action.

  13. Secretory Vesicle Priming by CAPS Is Independent of Its SNARE-Binding MUN Domain

    Cuc Quynh Nguyen Truong


    Full Text Available Priming of secretory vesicles is a prerequisite for their Ca2+-dependent fusion with the plasma membrane. The key vesicle priming proteins, Munc13s and CAPSs, are thought to mediate vesicle priming by regulating the conformation of the t-SNARE syntaxin, thereby facilitating SNARE complex assembly. Munc13s execute their priming function through their MUN domain. Given that the MUN domain of Ca2+-dependent activator protein for secretion (CAPS also binds syntaxin, it was assumed that CAPSs prime vesicles through the same mechanism as Munc13s. We studied naturally occurring splice variants of CAPS2 in CAPS1/CAPS2-deficient cells and found that CAPS2 primes vesicles independently of its MUN domain. Instead, the pleckstrin homology domain of CAPS2 seemingly is essential for its priming function. Our findings indicate a priming mode for secretory vesicles. This process apparently requires membrane phospholipids, does not involve the binding or direct conformational regulation of syntaxin by MUN domains of CAPSs, and is therefore not redundant with Munc13 action.

  14. Self-assembly of mixed lipids into bicelles and vesicles: molecular dynamics simulations

    Sharma, Hari; Wang, Zilu; Dormidontova, Elena

    Formation of complex supramolecular nanostructures, such as micelles, bicelles, vesicles (liposomes) etc. via self-assembly of simple molecules has provided a new pathway for the design and development of effective drug carriers. Solid nanoparticles or functional biopolymers, such as RNA, DNA, peptides can be encapsulated into these carriers for controlled delivery or selective targeting. We performed coarse grained molecular dynamics simulation using the MARTINI force field to study the self-assembly of a binary surfactant mixture composed of long and short phospholipids, DPPC and DHPC, in the ratio 3:1. We found that at low temperature lipids self-assemble into a bicelle (nanodisc) with the longer lipid mainly forming the interior and short lipid the rim of the bicelle. At higher temperature the nanodisc transforms into a vesicle with homogeneously distributed lipids. The structural changes of these nanodiscs and vesicles imposed by gold nanoparticle encapsulation and pegylation will be addressed.

  15. Micro and nano liposome vesicles containing curcumin for a drug delivery system

    Nguyen, Tuan Anh; Duoc Tang, Quan; Chanh Tin Doan, Duc; Chien Dang, Mau


    Micro and nano liposome vesicles were prepared using a lipid film hydration method and a sonication method. Phospholipid, cholesterol and curcumin were used to form micro and nano liposomes containing curcumin. The size, structure and properties of the liposomes were characterized by using optical microscopy, transmission electron microscopy, and UV-vis and Raman spectroscopy. It was found that the size of the liposomes was dependent on their composition and the preparation method. The hydration method created micro multilamellars, whereas nano unilamellars were formed using the sonication method. By adding cholesterol, the vesicles of the liposome could be stabilized and stored at 4 °C for up to 9 months. The liposome vesicles containing curcumin with good biocompatibility and biodegradability could be used for drug delivery applications.

  16. Application of Potential Phosphate-Solubilizing Bacteria and Organic Acids on Phosphate Solubilization from Phosphate Rock in Aerobic Rice

    Qurban Ali Panhwar; Shamshuddin Jusop; Umme Aminun Naher; Radziah Othman; Mohd Ismail Razi


    A study was conducted at Universiti Putra Malaysia to determine the effect of phosphate-solubilizing bacteria (PSB) and organic acids (oxalic & malic) on phosphate (P) solubilization from phosphate rock (PR) and growth of aerobic rice. Four rates of each organic acid (0, 10, 20, and 30 mM), and PSB strain (Bacillus sp.) were applied to aerobic rice. Total bacterial populations, amount of P solubilization, P uptake, soil pH, and root morphology were determined. The results of the study showed ...

  17. Solubilization of silk protein by radiation

    Sudatis, Boonya; Pongpat, Suchada [Office of Atomic Energy of Peace, Bangkok (Thailand)


    Gamma irradiated silk fibroin at doses of 0, 5, 10, 20, 40, 60, 80, 100, 125, 250, 500, 750 and 1000 kGy were soaked in water for 1 hr. Silk fibroin solubilized percentage was investigated from lost weight of sample (dried at 105{sup 0}C), they were 0, 0, 0.7, 0, 0.11, 0.11, 0, 0.73, 0.77, 4.38, 8.32, 10.22 and 18.52 respectively. It showed that at the higher dose up to 250 kGy had direct effect to solubility, and increased with increasing dose. In addition, silk sericin dissolved 77.76, 82.22, 83.55, 84.31, 86.04, 86.67 and 87.37% after gamma irradiation at the doses of 0, 50, 100, 200, 500, 750 and 1000 kGy respectively. It presents that radiation can cause silk protein, fibroin and sericin dissolve because of their degradation. (author)

  18. Solubilization and Humanization of Paraoxonase-1

    Mohosin Sarkar


    Full Text Available Paraoxonase-1 (PON1 is a serum protein, the activity of which is related to susceptibility to cardiovascular disease and intoxication by organophosphorus (OP compounds. It may also be involved in innate immunity, and it is a possible lead molecule in the development of a catalytic bioscavenger of OP pesticides and nerve agents. Human PON1 expressed in E. coli is mostly found in the insoluble fraction, which motivated the engineering of soluble variants, such as G2E6, with more than 50 mutations from huPON1. We examined the effect on the solubility, activity, and stability of three sets of mutations designed to solubilize huPON1 with fewer overall changes: deletion of the N-terminal leader, polar mutations in the putative HDL binding site, and selection of the subset of residues that became more polar in going from huPON1 to G2E6. All three sets of mutations increase the solubility of huPON1; the HDL-binding mutant has the largest effect on solubility, but it also decreases the activity and stability the most. Based on the G2E6 polar mutations, we “humanized” an engineered variant of PON1 with high activity against cyclosarin (GF and found that it was still very active against GF with much greater similarity to the human sequence.

  19. Crude oil degradation by phosphate-solubilizing bacteria

    DeSouza, M.J.B.D.; Nair, S.; David, J.J.; Chandramohan, D.

    Phosphate-solubilizing bacteria were isolated from tropical areas around the Indian peninsula. Two of the isolates showed high phosphatase activity. The isolates were identified as Klebsiella pneumoniae and Bacillus pumilus, and they showed high...

  20. Moroccan rock phosphate solubilization during a thermo-anaerobic ...



    Dec 4, 2013 ... thermo-anaerobic grassland waste biodegradation process. Moussa S. .... plating the nutrient agar isolate on the solid NBRIP medium. (Nautiyal, 1999) ..... solubilizing fungi isolated from phosphate mines. Ecol. Eng. 33:187-.

  1. Rock Phosphate Solubilization Mechanisms of One Fungus and One Bacterium

    LIN Qi-mei; ZHAO Xiao-rong; ZHAO Zi-juan; LI Bao-guo


    Many microorganisms can dissolve the insoluble phosphates like apatite. However, the mechanisms are still not clear. This study was an attempt to investigate the mechanisms of rock phosphate solubilization by an Aspergillus 2TCiF2 and an Arthrobacter1TCRi7. The results indicated that the fungus produced a large amount of organic acids, mainly oxalic acid. The total quantity of the organic acids produced by the fungus was 550 times higher than that by the bacterium. Different organic acids had completely different capacities to solubilize the rock. Oxalic acid and citric acid had stronger capacity to dissolve the rock than malic acid, tartaric acid, lactic acid, acetic acid, malonic acid and succinic acid. The fungus solubilized the rock through excreting both proton and organic acids. The rock solubilization of the bacterium depended on only proton.

  2. Enriching vermicompost by nitrogen fixing and phosphate solubilizing bacteria.

    Kumar, V; Singh, K P


    The effect of inoculation of vermicompost with nitrogen-fixing Azotobacter chroococcum strains, Azospirillum lipoferum and the phosphate solubilizing Pseudomonas striata on N and P contents of the vermicompost was assessed. Inoculation of N2 fixing bacteria into vermicompost increased contents of N and P. Enriching vermicompost with rock phosphate improved significantly the available P when inoculated with P. striata. During the incubation period, the inoculated bacterial strains proliferated rapidly, fixed N and solubilized added and native phosphate.

  3. Al3+-induced vesicle formation


    Aluminium laurate [Al(OOCC11H23)3] was synthesized as a surfactant, which can dissolve in micellar solution of a zwitterionic surfactant, tetradecycldimethylamine oxide (C14DMAO). The phase behavior of the mixtures of Al(OOCC11H23)3 and C14DMAO in water was studied and birefringent Lα-phase was observed. The birefringent Lα-phase consists of vesicles that were demonstrated by Polarizer and Transmission Electron Microscope (TEM) micrographs. Al3+-coordinated vesicles could be used as templating-precursor, providing a vesicle-route for preparation of inorganic nanoscale particles.

  4. Composition, structure and properties of POPC–triolein mixtures. Evidence of triglyceride domains in phospholipid bilayers

    Duelund, Lars; Jensen, Grethe Vestergaard; Hannibal-Bach, Hans Kristian


    We have in this study investigated the composition, structure and spectroscopical properties of multilamellar vesicles composed of a phospholipid, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and up to 10mol% of triolein (TO), a triglyceride. We found in agreement with previous result......% TO contained pure TO domains. These observations are consistent with an earlier MD simulation study by us and our co-workers suggesting triglycerides to be located in lens shaped, blister-like domains between the two lipid bilayer leaflets (Khandelia et al. (2010) [26])....

  5. Destabilization and fusion of zwitterionic large unilamellar lipid vesicles induced by a beta-type structure of the HIV-1 fusion peptide

    Nieva, JL; Nir, S; Wilschut, J


    The peptide HIVarg, corresponding to a sequence of 23 amino acid residues at the N-terminus of HIV-1 gp41, has the capacity to induce fusion of large unilamellar vesicles (LUV) consisting of negatively charged or zwitterionic phospholipids. In the present study, we further characterize this destabil

  6. Research and engineering assessment of biological solubilization of phosphate

    Rogers, R.D.; McIlwain, M.E.; Losinski, S.J.; Taylor, D.D.


    This research and engineering assessment examined a microbial phosphate solubilization process as a method of recovering phosphate from phosphorus containing ore compared to the existing wet acid and electric arc methods. A total of 860 microbial isolates, collected from a range of natural environments were tested for their ability to solubilize phosphate from rock phosphate. A bacterium (Pseudomonas cepacia) was selected for extensive characterization and evaluation of the mechanism of phosphate solubilization and of process engineering parameters necessary to recover phosphate from rock phosphate. These studies found that concentration of hydrogen ion and production of organic acids arising from oxidation of the carbon source facilitated microbial solubilization of both pure chemical insoluble phosphate compounds and phosphate rock. Genetic studies found that phosphate solubilization was linked to an enzyme system (glucose dehydrogenase). Process-related studies found that a critical solids density of 1% by weight (ore to liquid) was necessary for optimal solubilization. An engineering analysis evaluated the cost and energy requirements for a 2 million ton per year sized plant, whose size was selected to be comparable to existing wet acid plants.

  7. Phosphate-solubilizing bacteria associated with runner bean rhizosphere

    Mihalache Gabriela


    Full Text Available Soil microorganisms, especially rhizobacteria, play a key role in soil phosphorus (P dynamics and the subsequent availability of phosphate to plants. Utilization of phosphate-solubilizing bacteria as biofertilizers instead of synthetic chemicals is known to improve plant growth through the supply of plant nutrients, and may help to sustain environmental health and soil productivity. The main purpose of this study was to identify new phosphate-solubilizing bacteria isolated from runner bean rhizosphere. Ten out of 25 isolated bacterial strains solubilized Ca3(PO42 in qualitative and quantitative P-solubilization. The strain that exhibited the highest potential to solubilize Ca3(PO42, was selected for further determination of the mechanisms involved in the process. The medium pH was measured, organic acids released in the culture medium were identified by HPLC analysis, and the acid and alkaline phosphatase activities were determined. Our results showed that strain R7 solubilized phosphorous through the production of various organic acids such as lactic, isocitric, tartaric and pyruvic acids, and that it can be used as a potential biofertilizer.

  8. Connexin channels and phospholipids: association and modulation

    Harris Andrew L


    Full Text Available Abstract Background For membrane proteins, lipids provide a structural framework and means to modulate function. Paired connexin hemichannels form the intercellular channels that compose gap junction plaques while unpaired hemichannels have regulated functions in non-junctional plasma membrane. The importance of interactions between connexin channels and phospholipids is poorly understood. Results Endogenous phospholipids most tightly associated with purified connexin26 or connexin32 hemichannels or with junctional plaques in cell membranes, those likely to have structural and/or modulatory effects, were identified by tandem electrospray ionization-mass spectrometry using class-specific interpretative methods. Phospholipids were characterized by headgroup class, charge, glycerol-alkyl chain linkage and by acyl chain length and saturation. The results indicate that specific endogenous phospholipids are uniquely associated with either connexin26 or connexin32 channels, and some phospholipids are associated with both. Functional effects of the major phospholipid classes on connexin channel activity were assessed by molecular permeability of hemichannels reconstituted into liposomes. Changes to phospholipid composition(s of the liposome membrane altered the activity of connexin channels in a manner reflecting changes to the surface charge/potential of the membrane and, secondarily, to cholesterol content. Together, the data show that connexin26 and connexin32 channels have a preference for tight association with unique anionic phospholipids, and that these, independent of headgroup, have a positive effect on the activity of both connexin26 and connexin32 channels. Additionally, the data suggest that the likely in vivo phospholipid modulators of connexin channel structure-function that are connexin isoform-specific are found in the cytoplasmic leaflet. A modulatory role for phospholipids that promote negative curvature is also inferred. Conclusion

  9. Effects of three stabilizing agents--proline, betaine, and trehalose--on membrane phospholipids.

    Rudolph, A S; Crowe, J H; Crowe, L M


    We have studied the interaction between three compounds which accumulate in organisms under hydration stress--proline, betaine, and trehalose--and the membrane phospholipids dimyristoylphosphatidylcholine (DMPC), palmitoyloleoylphosphatidylcholine (POPC), and dimyristoylphosphatidylethanolamine in bulk solution. Film balance studies reveal that these compounds increase the area/molecule of these lipids. Differential scanning calorimetry has been employed to investigate the effect these agents have on the gel-to-liquid crystalline phase transition of multilamellar and small unilamellar vesicles of DMPC, dipalmitoylphosphatidylcholine, and POPC:phosphatidylserine (90:10 mole ratio) in bulk solution. In the presence of 1 M proline, trehalose, or betaine, the midtransition temperature in small unilamellar vesicles is reduced (up to 7 degrees C in 1 M trehalose), and the transition broadened. In contrast, multilamellar vesicles of similar lipid composition show an increased transition temperature in the presence of the same concentration of these compounds. This result suggests that the inner lamellae in multilamellar vesicles may be dehydrated with only a few outer lamellae exposed to the protective compound. Finally, we have used stereomodels of phosphatidylcholine to investigate the mechanism of action of these agents. Hydrogen bonding of trehalose to the head group region results in an increase in the distance between head groups of 6.9 A. This amount of spreading compares well with data from the monolayer experiments which indicate that maximal spreading of DMPC monolayers by trehalose is 6.5 A. Molecular models of proline and betaine have also been constructed, and these models suggest potential interactions between these compounds and phosphatidylcholines. For the amphipath proline, this interaction may involve intercalation between phospholipid head groups.

  10. Solubilization and fractionation of paired helical filaments.

    González, P J; Correas, I; Avila, J


    Paired helical filaments isolated from brains of two different patients with Alzheimer's disease were extensively treated with the ionic detergent, sodium dodecyl sulphate. Filaments were solubilized at different extents, depending on the brain examined, thus suggesting the existence of two types of paired helical filaments: sodium dodecyl sulphate-soluble and insoluble filaments. In the first case, the number of structures resembling paired helical filaments greatly decreased after the detergent treatment, as observed by electron microscopy. Simultaneously, a decrease in the amount of sedimentable protein was also observed upon centrifugation of the sodium dodecyl sulfate-treated paired helical filaments. A sodium dodecyl sulphate-soluble fraction was isolated as a supernatant after low-speed centrifugation of the sodium dodecyl sulphate-treated paired helical filaments. The addition of the non-ionic detergent Nonidet-P40 to this fraction resulted in the formation of paired helical filament-like structures. When the sodium dodecyl sulphate-soluble fraction was further fractionated by high-speed centrifugation, three subfractions were observed: a supernatant, a pellet and a thin layer between these two subfractions. No paired helical filaments were observed in any of these subfractions, even after addition of Nonidet P-40. However, when they were mixed back together, the treatment with Nonidet P-40 resulted in the visualization of paired helical filament-like structures. These results suggest that at least two different components are needed for the reconstitution of paired helical filaments as determined by electron microscopy. The method described here may allow the study of the components involved in the formation of paired helical filaments and the identification of possible factors capable of blocking this process.

  11. Mitochondrial phospholipids: role in mitochondrial function.

    Mejia, Edgard M; Hatch, Grant M


    Mitochondria are essential components of eukaryotic cells and are involved in a diverse set of cellular processes that include ATP production, cellular signalling, apoptosis and cell growth. These organelles are thought to have originated from a symbiotic relationship between prokaryotic cells in an effort to provide a bioenergetic jump and thus, the greater complexity observed in eukaryotes (Lane and Martin 2010). Mitochondrial processes are required not only for the maintenance of cellular homeostasis, but also allow cell to cell and tissue to tissue communication (Nunnari and Suomalainen 2012). Mitochondrial phospholipids are important components of this system. Phospholipids make up the characteristic outer and inner membranes that give mitochondria their shape. In addition, these membranes house sterols, sphingolipids and a wide variety of proteins. It is the phospholipids that also give rise to other characteristic mitochondrial structures such as cristae (formed from the invaginations of the inner mitochondrial membrane), the matrix (area within cristae) and the intermembrane space (IMS) which separates the outer mitochondrial membrane (OMM) and inner mitochondrial membrane (IMM). Phospholipids are the building blocks that make up these structures. However, the phospholipid composition of the OMM and IMM is unique in each membrane. Mitochondria are able to synthesize some of the phospholipids it requires, but the majority of cellular lipid biosynthesis takes place in the endoplasmic reticulum (ER) in conjunction with the Golgi apparatus (Fagone and Jackowski 2009). In this review, we will focus on the role that mitochondrial phospholipids play in specific cellular functions and discuss their biosynthesis, metabolism and transport as well as the differences between the OMM and IMM phospholipid composition. Finally, we will focus on the human diseases that result from disturbances to mitochondrial phospholipids and the current research being performed to help

  12. Congenital agenesis of seminal vesicle

    Hong-Fei Wu; Di Qiao; Li-Xin Qian; Ning-Hong Song; Ning-Han Feng; Li-Xin Hua; Wei Zhang


    Congenital agenesis of the seminal vesicle (CASV) is frequently associated with congenital absence of the vas deferens (CAVD) or ipsilateral congenital vasoureteral communication. We reported two cases of a rare condition that the vas deferens open ectopically into Mullerian duct cyst associated with agenesis of the ipsilateral seminal vesicle. The diagnosis was confirmed by vasography. Transurethral unroofing of the Mullerian duct cyst was performed in both patients with favourable results, however, assisted reproductive technology (ART) was still necessary for them to father children.

  13. Characterization and solubilization of nucleotide-specific, Mg/sup 2 +/-ATPase and Mg/sup 2 +/-pyrophosphatase of tonoplast

    Wagner, G.J.; Mulready, P.


    Nucleotide-specific, Mg/sup 2 +/-dependent ATPase and Mg/sup 2 +/-dependent pyrophosphatase were recovered with purified tonoplast obtained from isolated Tulipa petal vacuoles. Relative Mg/sup 2 +/-dependent hydrolysis of ATP, GTP and pyrophosphate, the only substrates hydrolyzed to a substantial degree, was 1.0, 0.3, and 0.6, respectively. Tonoplast ATPase required Mg/sup 2 +/, and essentially no Mg/sup 2 +/-dependent or Mg/sup 2 +/-independent p-nitrophenylphosphatase (which was associated with intact vacuoles) occurred with the membrane. Tonoplast ATPase was stimulated 10 to 30% by KCl, but was little effected by other cations (other than NH/sub 4//sup +/) or anions. No activity was observed with CaATP as substrate. The enzyme was cold stable and was inhibited by DCCD and Dio-9, but not by oligomycin. Its pH optimum was 7.0 and its specific activity was about 50 P/sub i//mg protein per h at 37/sup 0/C. Properties of membrane-bound and Polidocanol (polyoxyethylene ether, 9 lauryl ether) solubilized enzyme were similar. Reduced activity of solubilized enzyme was partially restored with phospholipids. Tonoplast ATPase appears to be an integral membrane component which requires phospholipids for maximal activity. Tonoplast Mg/sup 2 +/-pyrophosphatase had a pH optimum of greater than or equal to 8.5, was stimulated 2.5-fold by 50 mM KCl, and was largely lost upon detergent treatment. Properties of tonoplast ATPase observed are consistent with the characteristics of proton transport exhibited by isolated, intact Tulipa vacuoles. These observations suggest that tonoplast ATPase functions in proton transport. 34 references, 7 figures, 6 tables.

  14. Bile salt/phospholipid mixed micelle precursor pellets prepared by fluid-bed coating.

    Dong, Fuxia; Xie, Yunchang; Qi, Jianping; Hu, Fuqiang; Lu, Yi; Li, Sanming; Wu, Wei


    Bile salt/phospholipid mixed micelles (MMs) are potent carriers used for oral absorption of drugs that are poorly soluble in water; however, there are many limitations associated with liquid formulations. In the current study, the feasibility of preparing bile salt/phospholipid MM precursor (preMM) pellets with high oral bioavailability, using fluid-bed coating technology, was examined. In this study, fenofibrate (FB) and sodium deoxycholate (SDC) were used as the model drug and the bile salt, respectively. To prepare the MMs and to serve as the micellular carrier, a weight ratio of 4:6 was selected for the sodium deoxycholate/phospholipids based on the ternary phase diagram. Polyethylene glycol (PEG) 6000 was selected as the dispersion matrix for precipitation of the MMs onto pellets, since it can enhance the solubilizing ability of the MMs. Coating of the MMs onto the pellets using the fluid-bed coating technology was efficient and the pellets were spherical and intact. MMs could be easily reconstituted from preMM pellets in water. Although they existed in a crystalline state in the preMM pellets, FB could be encapsulated into the reconstituted MMs, and the MMs were redispersed better than solid dispersion pellets (FB:PEG = 1:3) and Lipanthyl®. The redispersibility of the preMM pellets increased with the increase of the FB/PEG/micellar carrier. PreMM pellets with a FB:PEG:micellar carrier ratio of 1:1.5:1.5 showed 284% and 145% bioavailability relative to Lipanthyl® and solid dispersion pellets (FB:PEG = 1:3), respectively. Fluid-bed coating technology has considerable potential for use in preparing sodium deoxycholate/phospholipid preMM pellets, with enhanced oral bioavailability for poorly water-soluble drugs.

  15. Lipid peroxidation generates biologically active phospholipids including oxidatively N-modified phospholipids.

    Davies, Sean S; Guo, Lilu


    Peroxidation of membranes and lipoproteins converts "inert" phospholipids into a plethora of oxidatively modified phospholipids (oxPL) that can act as signaling molecules. In this review, we will discuss four major classes of oxPL: mildly oxygenated phospholipids, phospholipids with oxidatively truncated acyl chains, phospholipids with cyclized acyl chains, and phospholipids that have been oxidatively N-modified on their headgroups by reactive lipid species. For each class of oxPL we will review the chemical mechanisms of their formation, the evidence for their formation in biological samples, the biological activities and signaling pathways associated with them, and the catabolic pathways for their elimination. We will end by briefly highlighting some of the critical questions that remain about the role of oxPL in physiology and disease.

  16. Formation of substrate-supported membranes from mixtures of long- and short-chain phospholipids.

    Morigaki, Kenichi; Kimura, Shigeki; Okada, Keisuke; Kawasaki, Takashi; Kawasaki, Kazunori


    We studied the formation of substrate-supported planar phospholipid bilayers (SPBs) on glass and silica from mixtures of long- and short-chain phospholipids to assess the effects of detergent additives on SPB formation. 1,2-Hexyanoyl-sn-glycero-3-phosphocholine (DHPC-C6) and 1,2-heptanoyl-sn-glycero-3-phosphocholine (DHPC-C7) were chosen as short-chain phospholipids. 1-Palmitoyl-2-oleol-sn-glycero-3-phosphocholine (POPC) was used as a model long-chain phospholipid. Kinetic studies by quartz crystal microbalance with dissipation monitoring (QCM-D) showed that the presence of short-chain phospholipids significantly accelerated the formation of SPBs. Rapid rinsing with a buffer solution did not change the adsorbed mass on the surface if POPC/DHPC-C6 mixtures were used below the critical micelle concentration (cmc) of DHPC-C6, indicating that an SPB composed of POPC molecules remained on the surface. Fluorescence microscopy observation showed homogeneous SPBs, and the fluorescence recovery after photobleaching (FRAP) measurements gave a diffusion coefficient comparable to that for SPBs formed from POPC vesicles. However, mixtures of POPC/DHPC-C7 resulted in a smaller mass of lipid adsorption on the substrate. FRAP measurements also yielded significantly smaller diffusion coefficients, suggesting the presence of defects. The different behaviors for DHPC-C6 and DHPC-C7 point to the dual roles of detergents to enhance the formation of SPBs and to destabilize them, depending on their structures and aggregation properties.

  17. Liposomal solubilization of new 3-hydroxy-quinolinone derivatives with promising anticancer activity: a screening method to identify maximum incorporation capacity

    Di Cagno, Massimiliano; Styskala, Jakub; Hlaváč, Jan


    Four new 3-hydroxy-quinolinone derivatives with promising anticancer activity could be solubilized using liposomes as vehicle to an extent that allows their in vitro and in vivo testing without use of toxic solvent(s). A screening method to identify the maximum incorporation capacity of hydrophobic...... drugs within liposomes was successfully applied. The compounds and lipid(s) were dissolved in methanol, and the solvent was removed by rotary evaporation. The film was resuspended with phosphate buffer (pH 7.4), and the dispersion was sonicated to reduce vesicle size. Ultracentrifugation was used...

  18. Interaction of alpha-lactalbumin with dimyristoyl phosphatidylcholine vesicles. II. A fluorescence polarization study.

    Herreman, W; van Tornout, P; van Cauwelaert, F H; Hanssens, I


    The interaction of alpha-lactalbumin with dimyristoyl phosphatidylcholine vesicles was studied as a function of temperature, pH and the molar ratio of phospholipid to protein. The method consisted of measuring the fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene used as a probe embedded in the vesicles. After incubation of the protein with the phospholipid for 2 h at 23 degrees C, the polarization of the light emitted by this probe shifted to higher values; the shift was greater at acidic pH than at neutral pH. After incubation at 37 degrees C, no shift in polarization was found at pH 7, 6 and 5 while a strong increase occurred at pH 4. Lowering the temperature, after incubation at 37 degrees C, had little effect on the polarization at neutral pH. At pH 5, however, and in the transition range of the phospholipid, the polarization increased greatly. A kinetic study of the interaction carried out around the transition temperature of dimyristoyl phosphatidylcholine as a function of pH shows that the speed of complex formation between alpha-lactalbumin and the lipid increases from neutral to acidic pH. From the present results and in agreement with our earlier calorimetric and fluorescence data (Hanssens, I., Houthuys, C., Herreman, W. and van Cauwelaert, F.H. (1980) Biochim. Biophys, Acta 602, 539--557), it is concluded that at neutral pH the interaction mechanism is probably different from that at acidic pH. At neutral pH and at all temperatures, alpha-lactalbumin is mainly absorbed electrostatically to the outer surface of the vesicle with little or no influence on the transition temperature of the phospholipid. At this pH, only around the transition temperature is penetration possible. At pH 4, however, the protein is able to penetrate the vesicle at all temperatures and to interact hydrophobically with the phospholipid fatty acid chains. As a result of this interaction, the transition temperature is increased by about 4 degrees C. This different

  19. Cell signalling and phospholipid metabolism. Final report

    Boss, W.F.


    These studies explored whether phosphoinositide (PI) has a role in plants analogous to its role in animal cells. Although no parallel activity of PI in signal transduction was found in plant cells, activity of inositol phospholipid kinase was found to be modulated by light and by cell wall degrading enzymes. These studies indicate a major role for inositol phospholipids in plant growth and development as membrane effectors but not as a source of second messengers.

  20. Phospholipids as Biomarkers for Excessive Alcohol Use


    S.T., Bauman, K.E., & Foshee, V. A. (2005). Neighborhood Influences on Adolescent Cigarette and Alcohol Use: Mediating Effects through Parent and...AWARD NUMBER: W81XWH-12-1-0497 TITLE: Phospholipids as Biomarkers for Excessive Alcohol Use...NUMBER Phospholipids as Biomarkers for Excessive Alcohol Use 5b. GRANT NUMBER W81XWH-12-1-0497 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S

  1. Solubilization of trichloroethylene by polyelectrolyte/surfactant complexes

    Uchiyama, Hirotaka; Christian, S.D.; Tucker, E.E.; Scamehorn, J.F. (Univ. of Oklahoma, Norman, OK (United States))


    An automated vapor pressure method is used to obtain solubilization isotherms for trichloroethylene (TCE) in polyelectrolyte/surfactant complexes throughout a wide range of solute activities at 20 and 25 C. The polyelectrolyte chosen is sodium poly(styrenesulfonate), PSS< and the surfactant is cetylpyridinium chloride or N-hexadecylpyridinium chloride, CPC. Data are fitted to the quadratic equation K = K[sub 0](1[minus][alpha]X + [beta]X[sup 2]), which correlates the solubilization equilibrium constant (K) with the mole fraction of TCE (X) in the micelles or complexes at each temperature. Activity coefficients are also obtained for TCE in the PSS/CPC complexes as a function of X. The general solubilization of TCE in PSS/CPC complexes resembles that of TCE in CPC micelles, as well as that of benzene or toluene in CPC micelles, suggesting that TCE solubilizes in ionic micelles both within the hydrocarbon micellar interior and near the micellar surface. The presence of the polyelectrolyte causes a small decrease in the ability of the cationic surfactant to solubilize TCE, while greatly reducing the concentration of the surfactant present in monomeric form. PSS/CPC complexes may be useful in colloid-enhanced ultrafiltration processes to purify organic-contaminated water.

  2. Enzymatic modification of phospholipids forfunctional applications and human nutrition

    Guo, Zheng; Vikbjerg, Anders / Falk; Xu, Xuebing


    Rapid progress in biochemistry of phospholipids and evolution of modern bioengineering has brought forth a number of novel concepts and technical advancements in the modification of phospholipids for industrial applications and human nutrition. Highlights cover preparation of novel phospholipid...... analogs based on the latest understanding of pivotal role of phospholipids in manifold biological processes, exploration of remarkable application potentials of phospholipids in meliorating human health, as well as development of new chemical and biotechnological approaches applied to the modification...

  3. Gravimetric determination of phospholipid concentration.

    Tejera-Garcia, Roberto; Connell, Lisa; Shaw, Walter A; Kinnunen, Paavo K J


    Accurate determination of lipid concentrations is an obligatory routine in a research laboratory engaged in studies using this class of biomaterials. For phospholipids, this is frequently accomplished using the phosphate assay (Bartlett, G.R. Phosphorus Assay in Column Chromatography. J. Biol. Chem. 234, 466-468, 1959). Given the purity of the currently commercially available synthetic and isolated natural lipids, we have observed that determination of the dry weight of lipid stock solutions provides the fastest, most accurate, and generic method to assay their concentrations. The protocol described here takes advantage of the high resolution and accuracy obtained by modern weighing technology. We assayed by this technique the concentrations of a number of phosphatidylcholine samples, with different degrees of acyl chain saturation and length, and in different organic solvents. The results were compared with those from Bartlett assay, (31)P NMR, and Langmuir compression isotherms. The data obtained show that the gravimetric assay yields lipid concentrations with a resolution similar or better than obtained by the other techniques.

  4. Raman spectroscopy of single extracellular vesicles reveals subpopulations with varying membrane content (Conference Presentation)

    Smith, Zachary J.; Lee, Changwon; Rojalin, Tatu; Carney, Randy P.; Hazari, Sidhartha; Knudson, Alisha; Lam, Kit S.; Saari, Heikki; Lazaro Ibañez, Elisa; Viitala, Tapani; Laaksonen, Timo; Yliperttula, Marjo; Wachsmann-Hogiu, Sebastian


    Exosomes are small (~100nm) membrane bound vesicles excreted by cells as part of their normal biological processes. These extracellular vesicles are currently an area of intense research, since they were recently found to carry functional mRNA that allows transfer of proteins and other cellular instructions between cells. Exosomes have been implicated in a wide range of diseases, including cancer. Cancer cells are known to have increased exosome production, and may use those exosomes to prepare remote environments for metastasis. Therefore, there is a strong need to develop characterization methods to help understand the structure and function of these vesicles. However, current techniques, such as proteomics and genomics technologies, rely on aggregating a large amount of exosome material and reporting on chemical content that is averaged over many millions of exosomes. Here we report on the use of laser-tweezers Raman spectroscopy (LTRS) to probe individual vesicles, discovering distinct heterogeneity among exosomes both within a cell line, as well as between different cell lines. Through principal components analysis followed by hierarchical clustering, we have identified four "subpopulations" of exosomes shared across seven cell lines. The key chemical differences between these subpopulations, as determined by spectral analysis of the principal component loadings, are primarily related to membrane composition. Specifically, the differences can be ascribed to cholesterol content, cholesterol to phospholipid ratio, and surface protein expression. Thus, we have shown LTRS to be a powerful method to probe the chemical content of single extracellular vesicles.


    Thurapati Pandu Raju


    Full Text Available Advanced biochemical and pre-clinical studies have proved the potential of plant flavonoids, polyphenolics and other hydrophilic natural compounds for the treatment of skin disorders, different types of carcinoma, anti-aging and many other areas of therapeutics and preventive medicine. The hydrophilic nature and unique chemical structure of these compounds pose major challenge because of their poor bioavailability through the skin or gut. The bioavailability can be improved by the use of delivery systems, which can enhance the rate and the extent of drug solubilizing into aqueous intestinal fluids as well as the capacity to cross the lipid rich biomembranes. Phospholipid based drug delivery systems have been found promising for the effective and efficacious herbal drug delivery. Complexing the polyphenolic phytoconstituents in molar ratio with phosphatidylcholine results into a new herbal drug delivery system- "Phytosome". Phytosomes show better pharmacokinetic and therapeutic profile than conventional herbal extracts.

  6. Direct Visualization of the Action of Triton X-100 on Giant Vesicles of Erythrocyte Membrane Lipids

    Casadei, Bruna R.; Domingues, Cleyton C.; de Paula, Eneida; Riske, Karin A.


    The raft hypothesis proposes that microdomains enriched in sphingolipids, cholesterol, and specific proteins are transiently formed to accomplish important cellular tasks. Equivocally, detergent-resistant membranes were initially assumed to be identical to membrane rafts, because of similarities between their compositions. In fact, the impact of detergents in membrane organization is still controversial. Here, we use phase contrast and fluorescence microscopy to observe giant unilamellar vesicles (GUVs) made of erythrocyte membrane lipids (erythro-GUVs) when exposed to the detergent Triton X-100 (TX-100). We clearly show that TX-100 has a restructuring action on biomembranes. Contact with TX-100 readily induces domain formation on the previously homogeneous membrane of erythro-GUVs at physiological and room temperatures. The shape and dynamics of the formed domains point to liquid-ordered/liquid-disordered (Lo/Ld) phase separation, typically found in raft-like ternary lipid mixtures. The Ld domains are then separated from the original vesicle and completely solubilized by TX-100. The insoluble vesicle left, in the Lo phase, represents around 2/3 of the original vesicle surface at room temperature and decreases to almost 1/2 at physiological temperature. This chain of events could be entirely reproduced with biomimetic GUVs of a simple ternary lipid mixture, 2:1:2 POPC/SM/chol (phosphatidylcholine/sphyngomyelin/cholesterol), showing that this behavior will arise because of fundamental physicochemical properties of simple lipid mixtures. This work provides direct visualization of TX-100-induced domain formation followed by selective (Ld phase) solubilization in a model system with a complex biological lipid composition. PMID:24896120

  7. Expression, Solubilization, and Purification of Bacterial Membrane Proteins.

    Jeffery, Constance J


    Bacterial integral membrane proteins play many important roles, including sensing changes in the environment, transporting molecules into and out of the cell, and in the case of commensal or pathogenic bacteria, interacting with the host organism. Working with membrane proteins in the lab can be more challenging than working with soluble proteins because of difficulties in their recombinant expression and purification. This protocol describes a standard method to express, solubilize, and purify bacterial integral membrane proteins. The recombinant protein of interest with a 6His affinity tag is expressed in E. coli. After harvesting the cultures and isolating cellular membranes, mild detergents are used to solubilize the membrane proteins. Protein-detergent complexes are then purified using IMAC column chromatography. Support protocols are included to help select a detergent for protein solubilization and for use of gel filtration chromatography for further purification.

  8. Factors determining rock phosphate solubilization by microorganisms isolated from soil.

    Nahas, E


    Forty two soil isolates (31 bacteria and 11 fungi) were studied for their ability to solubilize rock phosphate and calcium phosphate in culture medium. Eight bacteria and 8 fungi possessed solubilizing ability. Pseudomonas cepacia and Penicillium purpurogenum showed the highest activity. There was a correlation between final pH value and titratable acidity (r=-0.29 to -0.87) and between titratable acidity and soluble phosphate (r=0.22 to 0.99). Correlation values were functions of insoluble phosphate and of the group of microorganisms considered. A high correlation was observed between final pH and soluble phosphate only for the rock phosphates inoculated with the highest concentration of solubilizing bacteria (r=-0.73 to -0.98).

  9. Confocal Raman Microscopy of Hybrid-Supported Phospholipid Bilayers within Individual C18-Functionalized Chromatographic Particles.

    Kitt, Jay P; Harris, Joel M


    Measuring lipid-membrane partitioning of small molecules is critical to predicting bioavailability and investigating molecule-membrane interactions. A stable model membrane for such studies has been developed through assembly of a phospholipid monolayer on n-alkane-modified surfaces. These hybrid bilayers have recently been generated within n-alkyl-chain (C18)-modified porous silica and used in chromatographic retention studies of small molecules. Despite their successful application, determining the structure of hybrid bilayers within chromatographic silica is challenging because they reside at buried interfaces within the porous structure. In this work, we employ confocal Raman microscopy to investigate the formation and temperature-dependent structure of hybrid-phospholipid bilayers in C18-modified, porous-silica chromatographic particles. Porous silica provides sufficient surface area within a confocal probe volume centered in an individual particle to readily measure, with Raman microscopy, the formation of an ordered hybrid bilayer of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) with the surface C18 chains. The DMPC surface density was quantified from the relative Raman scattering intensities of C18 and phospholipid acyl chains and found to be ∼40% of a DMPC vesicle membrane. By monitoring Raman spectra acquired versus temperature, the bilayer main phase transition was observed to be broadened and shifted to higher temperature compared to a DMPC vesicle, in agreement with differential scanning calorimetry (DSC) results. Raman scattering of deuterated phospholipid was resolved from protonated C18 chain scattering, showing that the lipid acyl and C18 chains melt simultaneously in a single phase transition. The surface density of lipid in the hybrid bilayer, the ordering of both C18 and lipid acyl chains upon bilayer formation, and decoupling of C18 methylene C-H vibrations by deuterated lipid acyl chains all suggest an interdigitated acyl chain

  10. Autonomous movement of chemically powered vesicle

    Gupta, Shivam; Thakur, Snigdha


    A mechanism for self propulsion of deformable vesicle has been proposed, vesicle moves by sensing the self-generated chemical gradient. Like many molecular motors they suffer strong perturbations from the environment in which they move as a result of thermal fluctuations and do not rely on inertia for their propulsion. Motion of the vesicle is driven by an asymmetric distribution of reaction products. The propulsive velocity of the device is calculated as well as the scale of the velocity fluctuations. We present the simulation results on velocity of vesicle, reorientation of vesicle, and shape transformation of vesicles.

  11. Solubilization of bovine gelatin using power ultrasound: gelation without heating.

    Farahnaky, Asgar; Zendeboodi, Fatemeh; Azizi, Rezvan; Mesbahi, Gholamreza; Majzoobi, Mahsa


    The aim of this study was to investigate the efficacy of power ultrasound without using any heating stage in solubilizeing gelatin dispersions, and to characterize the mechanical and microstructural properties of the resulting gels using texture analysis and scanning electron microscopy, respectively. Usually to prepare a gel from gelatin, a primary heating stage of at about 40C or above is required to solubilize gelatin macromolecules. In this study solubilizing gelatin dispersions using power ultrasound without any heating was successfully performed. For solubilising gelatin, an ultrasound equipment with a frequency of 20 kHz, amplitude of 100% and power range of 50-150 W was used. Aqueous gelatin dispersions (4% w/v) were subjected to ultrasound for different times (40-240 s) at a constant temperature of 13C. Applying ultrasound to gelatin dispersions caused increases in water absorption and water solubility of the hydrocolloid. The textural parameters of the resulting gelatin gels, increased with increasing time and power of ultrasound. Moreover, a generalized Maxwell model with three elements was used for calculating relaxation times of the gels. The microstructural observations by SEM showed that the structural cohesiveness of the gels increased by increasing ultrasonication time. Ultrasound-assisted solubilization of gelatin can have emerging implications for industrial uses in pharmaceuticals, food and non-food systems. Usually to prepare a gel from gelatin, a primary heating stage of at about 40C or above is required to solubilize gelatin macromolecules. Therefore, the use of gelatin as a hydrocolloid in food processings or pharmaceutical formulations which lack a heating step has been a technological and practical challenge. In this study solubilizing gelatin dispersions using power ultrasound without any heating was successfully performed. Ultrasound-assisted solubilisation of gelatin can have emerging implications for industrial uses in pharmaceuticals

  12. Cystadenoma of the seminal vesicle

    Gil Antônio O.


    Full Text Available Primary tumors of the seminal vesicle are extremely rare. Among them, there is a spectrum of tumors derived from both epithelium and stroma and so classified as epithelial-stromal tumors. Herein, we report a case of a cystadenoma in a 49-year-old asymptomatic man, detected in a routine ultrasonography for liver disease follow-up. The digital rectal examination detected a large mass anterior to rectum and posterior to bladder. Computed tomography scan and magnetic resonance imaging showed a normal prostate and a 9.0 cm cystic tumor, replacing the left seminal vesicle. The gross appearance and microscopic aspect was compatible with cystadenoma of seminal vesicle. Patient's postoperative recovery was uneventful. He is currently alive, 3 years after the diagnosis, with no signs of recurrence.

  13. [Solubilization Specificities Interferon beta-1b from Inclusion Bodies].

    Zhuravko, A S; Kononova, N V; Bobruskin, A I


    A new solubilization method of recombinant interferon beta-1b (IFNβ-1b) from the inclusion bodies was developed. This method allows to extract the target protein selectively in the solutions of different alcohols, such as ethanol, propanol and isopropanol. It was shown that the more effective IFNβ-1b solubilization was achieved in the 55% propanol solution. This method allowed to extract the target protein from inclusion bodies around 85-90%, and significantly reduced Escherichia coli content in the solubilizate, in comparison with standard methods.

  14. Water-solubilization and functionalization of semiconductor quantum dots.

    Tyrakowski, Christina M; Isovic, Adela; Snee, Preston T


    Semiconductor quantum dots (QDs) are highly fluorescent nanocrystals that have abundant potential for uses in biological imaging and sensing. However, the best materials are synthesized in hydrophobic surfactants that prevent direct aqueous solubilization. While several methods have been developed to impart water-solubility, an aqueous QD dispersion has no inherent useful purpose and must be functionalized further. Due to the colloidal nature of QD dispersions, traditional methods of chemical conjugation in water either have low yields or cause irreversible precipitation of the sample. Here, we describe several methods to water-solubilize QDs and further functionalize the materials with chemical and/or biological vectors.

  15. Phospholipid vesicles encapsulated bacteriophage: A novel approach to enhance phage biodistribution.

    Singla, S; Harjai, K; Raza, K; Wadhwa, S; Katare, O P; Chhibber, S


    Phage therapy has been at the centre of attraction for combating multi-drug resistant strains. However, less stability and rapid clearance of phage by mononuclear phagocytic system (MPS) restricts its use in humans. In the present study, aim was to develop a liposomal delivery system for bacteriophage that can assure efficient phage delivery and retention at the site of infection. Different ratios of cholesterol, lipids and surfactant along with different charge inducers were employed to prepare liposomes. Phage was then entrapped in the liposomes and characterized on the basis of morphology, size, entrapment efficiency and stability. Further, in vivo biodistribution of free phage and liposome entrapped phage was compared in different organs of mice. A cationic liposomal formulation showed maximum encapsulation efficiency of 92%. Transmission electron microscopy (TEM) confirmed the entrapment of phages in liposomes. Liposome preparation was found to be most stable at 4°C during storage. Liposome entrapped bacteriophage was retained for longer duration in different organs i.e. upto day 4 in blood, day 6 in liver, lungs and kidney, 14days in spleen of mice as compared to free phage that became undetectable by 36th h in blood as well as lungs and by 48th h in all other organs. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. When to biopsy seminal vesicles.

    Panach-Navarrete, J; García-Morata, F; Hernández-Medina, J A; Martínez-Jabaloyas, J M


    The involvement of seminal vesicles in prostate cancer can affect the prognosis and determine the treatment. The objective of this study was to determine whether we could predict its infiltration at the time of the prostate biopsy to know when to indicate the biopsy of the seminal vesicles. observational retrospective study of 466 patients who underwent seminal vesicle biopsy. The indication for this biopsy was a prostate-specific antigen (PSA) level greater than 10 ng/ml or an asymmetric or obliterated prostatoseminal angle. The following variables were included in the analysis: PSA level, PSA density, prostate volume, number of cores biopsied, suspicious rectal examination, and preservation of the prostatoseminal angle, studying its relationship with the involvement of the seminal vesicles. Forty-one patients (8.8%) had infiltrated seminal vesicles and 425 (91.2%) had no involvement. In the univariate analysis, the cases with infiltration had a higher mean PSA level (P 19.60 ng/dL (P < .01) and 2.95 times higher if there is a suspicious rectal examination (P = .014). Furthermore, this probability increases by 1.04 times for each unit of prostate volume lower (P < .01). The ROC curves showed maximum sensitivity and specificity at 19.6 ng/mL for PSA and 0.39 for PSA density. In this series, greater involvement of seminal vesicles was associated with a PSA level ≥20 ng/ml, a suspicious rectal examination and a lack of prostatoseminal angle preservation. Copyright © 2014 AEU. Publicado por Elsevier España, S.L.U. All rights reserved.

  17. Health effects of dietary phospholipids

    Küllenberg Daniela


    Full Text Available Abstract Beneficial effects of dietary phospholipids (PLs have been mentioned since the early 1900's in relation to different illnesses and symptoms, e.g. coronary heart disease, inflammation or cancer. This article gives a summary of the most common therapeutic uses of dietary PLs to provide an overview of their approved and proposed benefits; and to identify further investigational needs. From the majority of the studies it became evident that dietary PLs have a positive impact in several diseases, apparently without severe side effects. Furthermore, they were shown to reduce side effects of some drugs. Both effects can partially be explained by the fact that PL are highly effective in delivering their fatty acid (FA residues for incorporation into the membranes of cells involved in different diseases, e.g. immune or cancer cells. The altered membrane composition is assumed to have effects on the activity of membrane proteins (e.g. receptors by affecting the microstructure of membranes and, therefore, the characteristics of the cellular membrane, e.g. of lipid rafts, or by influencing the biosynthesis of FA derived lipid second messengers. However, since the FAs originally bound to the applied PLs are increased in the cellular membrane after their consumption or supplementation, the FA composition of the PL and thus the type of PL is crucial for its effect. Here, we have reviewed the effects of PL from soy, egg yolk, milk and marine sources. Most studies have been performed in vitro or in animals and only limited evidence is available for the benefit of PL supplementation in humans. More research is needed to understand the impact of PL supplementation and confirm its health benefits.

  18. Nonenzymatic Reactions above Phospholipid Surfaces of Biological Membranes: Reactivity of Phospholipids and Their Oxidation Derivatives.

    Solís-Calero, Christian; Ortega-Castro, Joaquín; Frau, Juan; Muñoz, Francisco


    Phospholipids play multiple and essential roles in cells, as components of biological membranes. Although phospholipid bilayers provide the supporting matrix and surface for many enzymatic reactions, their inherent reactivity and possible catalytic role have not been highlighted. As other biomolecules, phospholipids are frequent targets of nonenzymatic modifications by reactive substances including oxidants and glycating agents which conduct to the formation of advanced lipoxidation end products (ALEs) and advanced glycation end products (AGEs). There are some theoretical studies about the mechanisms of reactions related to these processes on phosphatidylethanolamine surfaces, which hypothesize that cell membrane phospholipids surface environment could enhance some reactions through a catalyst effect. On the other hand, the phospholipid bilayers are susceptible to oxidative damage by oxidant agents as reactive oxygen species (ROS). Molecular dynamics simulations performed on phospholipid bilayers models, which include modified phospholipids by these reactions and subsequent reactions that conduct to formation of ALEs and AGEs, have revealed changes in the molecular interactions and biophysical properties of these bilayers as consequence of these reactions. Then, more studies are desirable which could correlate the biophysics of modified phospholipids with metabolism in processes such as aging and diseases such as diabetes, atherosclerosis, and Alzheimer's disease.

  19. The application of P-gp inhibiting phospholipids as novel oral bioavailability enhancers - An in vitro and in vivo comparison.

    Weinheimer, Manuel; Fricker, Gert; Burhenne, Jürgen; Mylius, Patricia; Schubert, Rolf


    The efflux transporter P-glycoprotein (P-gp) significantly modulates drug transport across the intestinal mucosa, strongly reducing the systemic absorption of various active pharmaceutical ingredients. P-gp inhibitors could serve as helpful tools to enhance the oral bioavailability of those substances. As a membrane-associated protein P-gp is surrounded and influenced by phospholipids. Some synthetic phospholipids have been found to strongly reduce P-gp's activity. In this study two representative phospholipids, 1,2-dioctanoyl-sn-glycero-3-phosphocholine (8:0 PC) and 1,2-didecanoyl-sn-glycero-3-phosphocholine (10:0 PC), were compared with Tween® 80 and Cremophor® EL, both commonly used surfactants with P-gp inhibitory properties. Their influence on the cellular transport of the P-gp substrate rhodamine 123 (RH123) was examined using Caco-2 cell layers. In addition, fluorescence anisotropy measurements were performed in order to investigate their effect on membrane fluidity. Finally, we compared the phospholipids with Tween® 80 and the competitive P-gp inhibitor verapamil in an in vivo study, testing their effects on the oral bioavailability of the P-gp substrate drug ritonavir. Both phospholipids not only led to the strongest absorption of RH123, but a permeability enhancing effect was detected in addition to the P-gp inhibition. Their effects on membrane fluidity were not consistent with their P-gp inhibiting effects, and therefore suggested a more complex mode of action. Both phospholipids significantly increased the area under the ritonavir plasma level curve (AUC) within 150min by more than tenfold, but were inferior to Tween® 80, which showed superior solubilizing effects. Finally, these phospholipids represent a novel substance class showing a high permeabilization potential for P-gp substrates. Because of their physiological structure and intestinal degradability, good tolerability without systemic absorption is expected. Formulating P-gp substrates with

  20. Quantification of phospholipids classes in human milk.

    Giuffrida, Francesca; Cruz-Hernandez, Cristina; Flück, Brigitte; Tavazzi, Isabelle; Thakkar, Sagar K; Destaillats, Frédéric; Braun, Marcel


    Phospholipids are integral constituents of the milk fat globule membranes and they play a central role in infants' immune and inflammatory responses. A methodology employing liquid chromatography coupled with evaporative light scattering detector has been optimized and validated to quantify the major phospholipids classes in human milk. Phospholipids were extracted using chloroform and methanol and separated on C18 column. Repeatability, intermediate reproducibility, and recovery values were calculated and a large sample set of human milk analyzed. In human milk, phospholipid classes were quantified at concentrations of 0.6 mg/100 g for phosphatidylinositol; 4.2 mg/100 g for phosphatidylethanolamine, 0.4 mg/100 g for phosphatidylserine, 2.8 mg/100 g for phosphatidylcholine, and 4.6 mg/100 g for sphingomyelin. Their relative standard deviation of repeatability and intermediate reproducibility values ranging between 0.8 and 13.4 % and between 2.4 and 25.7 %, respectively. The recovery values ranged between 67 and 112 %. Finally, the validated method was used to quantify phospholipid classes in human milk collected from 50 volunteers 4 weeks postpartum providing absolute content of these lipids in a relatively large cohort. The average content of total phospholipids was 23.8 mg/100 g that corresponds to an estimated mean intake of 140 mg phospholipids/day in a 4-week old infant when exclusively breast-fed.

  1. Phyto-liposomes as nanoshuttles for water-insoluble silybin-phospholipid complex.

    Angelico, Ruggero; Ceglie, Andrea; Sacco, Pasquale; Colafemmina, Giuseppe; Ripoli, Maria; Mangia, Alessandra


    Among various phospholipid-mediated drug delivery systems (DDS) suitable for topic and oral administration, phytosome technology represents an advanced innovation, widely used to incorporate standardized bioactive polyphenolic phytoconstituents into phospholipid molecular complexes. In order to extend their potential therapeutic efficiency also to other routes of administration, we proposed a novel phytosome carrier-mediated vesicular system (phyto-liposome) as DDS for the flavonolignan silybin (SIL), a natural compound with multiple biological activities related to its hepatoprotective, anticancer and antioxidant (radical scavenging) effects. We screened the optimum fraction of its phytosome, available in the market as Siliphos™, into liposomes prepared by extrusion, such that vesicle sizes and charges, monitored through dynamic light scattering and laser doppler velocimetry, satisfied several quality requirements. Special emphasis was placed on the study of host-guest interaction by performing UV-vis absorption, spectrofluorimetry and NMR experiments both in aqueous and non-polar solvents to probe the effect of the presence of phospholipids on the electronic properties of SIL and its propensity to engage H bonding with the lipid headpolar groups. Finally, fluorescence microscopy observations confirmed the ability of phyto-liposomes to be internalized in human hepatoma cells, which was promising for their potential application in the treatment of acute or chronic liver diseases.

  2. Inborn errors of metabolism in the biosynthesis and remodelling of phospholipids.

    Wortmann, Saskia B; Espeel, Marc; Almeida, Ligia; Reimer, Annette; Bosboom, Dennis; Roels, Frank; de Brouwer, Arjan P M; Wevers, Ron A


    Since the proposal to define a separate subgroup of inborn errors of metabolism involved in the biosynthesis and remodelling of phospholipids, sphingolipids and long chain fatty acids in 2013, this group is rapidly expanding. This review focuses on the disorders involved in the biosynthesis of phospholipids. Phospholipids are involved in uncountable cellular processes, e.g. as structural components of membranes, by taking part in vesicle and mitochondrial fusion and fission or signal transduction. Here we provide an overview on both pathophysiology and the extremely heterogeneous clinical presentations of the disorders reported so far (Sengers syndrome (due to mutations in AGK), MEGDEL syndrome (or SERAC defect, SERAC1), Barth syndrome (or TAZ defect, TAZ), congenital muscular dystrophy due to CHKB deficiency (CHKB). Boucher-Neuhäuser/Gordon Holmes syndrome (PNPLA6), PHARC syndrome (ABHD12), hereditary spastic paraplegia type 28, 54 and 56 (HSP28, DDHD1; HSP54, DDHD2; HSP56, CYP2U1), Lenz Majewski syndrome (PTDSS1), spondylometaphyseal dysplasia with cone-rod dystrophy (PCYT1A), atypical haemolytic-uremic syndrome due to DGKE deficiency (DGKE).

  3. Solubilization, purification and reconstitution of Ca(2+)-ATPase from bovine pulmonary artery smooth muscle microsomes by different detergents: preservation of native structure and function of the enzyme by DHPC.

    Mandal, Amritlal; Das, Sudip; Chakraborti, Tapati; Kar, Pulak; Ghosh, Biswarup; Chakraborti, Sajal


    The properties of Ca(2+)-ATPase purified and reconstituted from bovine pulmonary artery smooth muscle microsomes {enriched with endoplasmic reticulum (ER)} were studied using the detergents 1,2-diheptanoyl-sn-phosphatidylcholine (DHPC), poly(oxy-ethylene)8-lauryl ether (C(12)E(8)) and Triton X-100 as the solubilizing agents. Solubilization with DHPC consistently gave higher yields of purified Ca(2+)-ATPase with a greater specific activity than solubilization with C(12)E(8) or Triton X-100. DHPC was determined to be superior to C(12)E(8); while that the C(12)E(8) was determined to be better than Triton X-100 in active enzyme yields and specific activity. DHPC solubilized and purified Ca(2+)-ATPase retained the E1Ca-E1*Ca conformational transition as that observed for native microsomes; whereas the C(12)E(8) and Triton X-100 solubilized preparations did not fully retain this transition. The coupling of Ca(2+) transported to ATP hydrolyzed in the DHPC purified enzyme reconstituted in liposomes was similar to that of the native micosomes, whereas that the coupling was much lower for the C(12)E(8) and Triton X-100 purified enzyme reconstituted in liposomes. The specific activity of Ca(2+)-ATPase reconstituted into dioleoyl-phosphatidylcholine (DOPC) vesicles with DHPC was 2.5-fold and 3-fold greater than that achieved with C(12)E(8) and Triton X-100, respectively. Addition of the protonophore, FCCP caused a marked increase in Ca(2+) uptake in the reconstituted proteoliposomes compared with the untreated liposomes. Circular dichroism analysis of the three detergents solubilized and purified enzyme preparations showed that the increased negative ellipticity at 223 nm is well correlated with decreased specific activity. It, therefore, appears that the DHPC purified Ca(2+)-ATPase retained more organized and native secondary conformation compared to C(12)E(8) and Triton X-100 solubilized and purified preparations. The size distribution of the reconstituted liposomes measured

  4. Tryptophan probes reveal residue-specific phospholipid interactions of apolipoprotein C-III.

    Pfefferkorn, Candace M; Walker, Robert L; He, Yi; Gruschus, James M; Lee, Jennifer C


    Apolipoproteins are essential human proteins for lipid metabolism. Together with phospholipids, they constitute lipoproteins, nm to μm sized particles responsible for transporting cholesterol and triglycerides throughout the body. To investigate specific protein-lipid interactions, we produced and characterized three single-Trp containing apolipoprotein C-III (ApoCIII) variants (W42 (W54F/W65F), W54 (W42F/W65F), W65 (W42F/W54F)). Upon binding to phospholipid vesicles, wild-type ApoCIII adopts an α-helical conformation (50% helicity) as determined by circular dichroism spectroscopy with an approximate apparent partition constant of 3×10(4) M(-1). Steady-state and time-resolved fluorescence measurements reveal distinct residue-specific behaviors with W54 experiencing the most hydrophobic environment followed by W42 and W65. Interestingly, time-resolved anisotropy measurements show a converse trend for relative Trp mobility with position 54 being the least immobile. To determine the relative insertion depths of W42, W54, and W65 in the bilayer, fluorescence quenching experiments were performed using three different brominated lipids. W65 had a clear preference for residing near the headgroup while W54 and W42 sample the range of depths ~8-11 Å from the bilayer center. On average, W54 is slightly more embedded than W42. Based on Trp spectral differences between ApoCIII binding to phospholipid vesicles and sodium dodecyl sulfate micelles, we suggest that ApoCIII adopts an alternate helical conformation on the bilayer which could have functional implications.

  5. Extracellular vesicles: Exosomes, microvesicles, and friends

    Raposo, G.; Stoorvogel, W.|info:eu-repo/dai/nl/074352385


    Cells release into the extracellular environment diverse types of membrane vesicles of endosomal and plasma membrane origin called exosomes and microvesicles, respectively. These extracellular vesicles (EVs) represent an important mode of intercellular communication by serving as vehicles for

  6. Identification of the phospholipid lysobisphosphatidic acid in the protozoan Entamoeba histolytica: An active molecule in endocytosis

    Silvia Castellanos-Castro


    Full Text Available Phospholipids are essential for vesicle fusion and fission and both are fundamental events for Entamoeba histolytica phagocytosis. Our aim was to identify the lysobisphosphatidic acid (LBPA in trophozoites and investigate its cellular fate during endocytosis. LBPA was detected by TLC in a 0.5 Rf spot of total lipids, which co-migrated with the LBPA standard. The 6C4 antibody, against LBPA recognized phospholipids extracted from this spot. Reverse phase LC-ESI-MS and MS/MS mass spectrometry revealed six LBPA species of m/z 772.58–802.68. LBPA was associated to pinosomes and phagosomes. Intriguingly, during pinocytosis, whole cell fluorescence quantification showed that LBPA dropped 84% after 15 min incubation with FITC-Dextran, and after 60 min, it increased at levels close to steady state conditions. Similarly, during erythrophagocytosis, after 15 min, LBPA also dropped in 36% and increased after 60 and 90 min. EhRab7A protein appeared in some vesicles with LBPA in steady state conditions, but after phagocytosis co-localization of both molecules increased and in late phases of erythrophagocytosis they were found in huge phagosomes or multivesicular bodies with many intraluminal vacuoles, and surrounding ingested erythrocytes and phagosomes. The 6C4 and anti-EhADH (EhADH is an ALIX family protein antibodies and Lysotracker merged in about 50% of the vesicles in steady state conditions and throughout phagocytosis. LBPA and EhADH were also inside huge phagosomes. These results demonstrated that E. histolytica LBPA is associated to pinosomes and phagosomes during endocytosis and suggested differences of LBPA requirements during pinocytosis and phagocytosis.

  7. Effect of temperature, water content and free fatty acid on reverse micelle formation of phospholipids in vegetable oil.

    Lehtinen, Olli-Pekka; Nugroho, Robertus Wahyu N; Lehtimaa, Tuula; Vierros, Sampsa; Hiekkataipale, Panu; Ruokolainen, Janne; Sammalkorpi, Maria; Österberg, Monika


    The self-assembly of phospholipids in oil, specifically lecithin in rapeseed oil, was investigated by combining experimental and computational methods The influence of temperature, water, and free fatty acids on the onset of lecithin aggregation in the rapeseed oil was determined using the 7,7,8,8 -tetracyanoquinodimethane dye (TCNQ) solubilization method and the size and shape of the self-assembled lecithin structures were investigated by small-angle X-ray scattering and cryogenic transmission electron microscopy. In the absence of excess water in the system (0.03wt-% water in oil), stable cylindrical lecithin reverse micelles were observed above the critical micelle concentration (CMC). Comparing the aggregation response in room temperature and at 70°C revealed that CMC decreased with increasing temperature. Furthermore, already a modest amount of added water (0.3wt-% water in oil) was sufficient to induce the formation of lamellar lecithin structures, that phase separated from the oil. In low water content, oleic acid suppressed the formation of lecithin reverse micelles whereas in the presence of more water, the oleic acid stabilized the reverse micelles. Consequently, more water was needed to induce phase separation in the presence of oleic acid. Molecular dynamics simulations indicated that the stabilizing effect of oleic acid resulted from oleic acid enhancing phospholipid solubilization in the oil by forming a solvating shell around the phosphate head group. The findings showed that the response of the mixed surfactant system is a delicate interplay of the different components and variables. The significance of the observations is that multiple parameters need to be controlled for desired system response, for example towards vegetable oil purification or phospholipid based microemulsions. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Amphiphilic biopolymers (amphibiopols) as new surfactants for membrane protein solubilization

    Duval-Terrié, Caroline; Cosette, Pascal; Molle, Gérard; Muller, Guy; Dé, Emmanuelle


    The aim of this study was to develop new surfactants for membrane protein solubilization, from a natural, biodegradable polymer: the polysaccharide pullulan. A set of amphiphilic pullulans (HMCMPs), differing in hydrophobic modification ratio, charge ratio, and the nature of the hydrophobic chains introduced, were synthesized and tested in solubilization experiments with outer membranes of Pseudomonas fluorescens. The membrane proteins were precipitated, and then resolubilized with various HMCMPs. The decyl alkyl chain (C10) was the hydrophobic graft that gave the highest level of solubilization. Decyl alkyl chain-bearing HMCMPs were also able to extract integral membrane proteins from their lipid environment. The best results were obtained with an amphiphilic pullulan bearing 18% decyl groups (18C10). Circular dichroism spectroscopy and membrane reconstitution experiments were used to test the structural and functional integrity of 18C10-solubilized proteins (OmpF from Escherichia coli and bacteriorhodopsin from Halobacterium halobium). Whatever their structure type (α or β), 18C10 did not alter either the structure or the function of the proteins analyzed. Thus, HMCMPs appear to constitute a promising new class of polymeric surfactants for membrane protein studies. PMID:12649425

  9. Storage stability of marine phospholipids emulsions

    Lu, Henna Fung Sieng; Nielsen, Nina Skall; Baron, Caroline Pascale

    Marine phospholipids (MPL) are believed to provide more advantages than fish oil from the same source. They are considered to have a better bioavailability, a better resistance towards oxidation and a higher content of polyunsaturated fatty acids such as eicosapentaenoic (EPA) and docosahexaenoic...... of secondary volatile compounds by Solid Phase Microextraction at several time intervals at 2°C storage. Preliminary results showed that marine phospholipids emulsion has a good oxidative stability....... acids (DHA) than oily triglycerides (fish oil). Therefore, the objective of this study is to explore the feasibility of using marine phospholipids emulsions as delivery system through investigation of the physical, oxidative and hydrolytic stability of MPL emulsions with or without addition of fish oil...

  10. Bile salt/phospholipid mixed micelle precursor pellets prepared by fluid-bed coating

    Dong F


    Full Text Available Fuxia Dong,1,2 Yunchang Xie,1 Jianping Qi,1 Fuqiang Hu,3 Yi Lu,1 Sanming Li,2 Wei Wu1 1School of Pharmacy, Fudan University, Key Laboratory of Smart Drug Delivery of Ministry of Education and PLA, Shanghai, People’s Republic of China; 2School of Pharmacy, Shenyang Pharmaceutical University, Liaoning, People’s Republic of China; 3School of Pharmacy, Zhejiang University, Hangzhou, People’s Republic of China Abstract: Bile salt/phospholipid mixed micelles (MMs are potent carriers used for oral absorption of drugs that are poorly soluble in water; however, there are many limitations associated with liquid formulations. In the current study, the feasibility of preparing bile salt/phospholipid MM precursor (preMM pellets with high oral bioavailability, using fluid-bed coating technology, was examined. In this study, fenofibrate (FB and sodium deoxycholate (SDC were used as the model drug and the bile salt, respectively. To prepare the MMs and to serve as the micellular carrier, a weight ratio of 4:6 was selected for the sodium deoxycholate/phospholipids based on the ternary phase diagram. Polyethylene glycol (PEG 6000 was selected as the dispersion matrix for precipitation of the MMs onto pellets, since it can enhance the solubilizing ability of the MMs. Coating of the MMs onto the pellets using the fluid-bed coating technology was efficient and the pellets were spherical and intact. MMs could be easily reconstituted from preMM pellets in water. Although they existed in a crystalline state in the preMM pellets, FB could be encapsulated into the reconstituted MMs, and the MMs were redispersed better than solid dispersion pellets (FB:PEG = 1:3 and Lipanthyl®. The redispersibility of the preMM pellets increased with the increase of the FB/PEG/micellar carrier. PreMM pellets with a FB:PEG:micellar carrier ratio of 1:1.5:1.5 showed 284% and 145% bioavailability relative to Lipanthyl® and solid dispersion pellets (FB:PEG = 1:3, respectively. Fluid

  11. Partition of alkane by an extracellular vesicle derived from hexadecane-grown Acinetobacter

    Kappeli, O.; Finnerty, W.R.


    The enhanced solubility of hexadecane in the growth medium of hexadecane-grown Acinetobacter species has been related to the accumulation of an extra-cellular vesicular component. The partition of hexadecane was determined by measuring the amount of (/sup 3/H)hexadecane bound to the vesicular particle. The vesicle was characterized as a phospholipid-rich, lipopolysaccharide-rich particle with a polypeptide composition similar to the outer membrane of Acinetobacter. The accumulation of an extracellular vesicular component that binds hexadecane in the form of a microemulsion represents another example of molecules produced by microorganisms in response to paraffinic substrates.

  12. Zinc solubilizing Bacillus spp. potential candidates for biofortification in maize.

    Mumtaz, Muhammad Zahid; Ahmad, Maqshoof; Jamil, Moazzam; Hussain, Tanveer


    Bioaugmentation of Zn solubilizing rhizobacteria could be a sustainable intervention to increase bioavailability of Zn in soil which can be helpful in mitigation of yield loss and malnutrition of zinc. In present study, a number of pure rhizobacterial colonies were isolated from maize rhizosphere and screened for their ability to solubilize zinc oxide. These isolates were screened on the basis of zinc and phosphate solubilization, IAA production, protease production, catalase activity and starch hydrolysis. All the selected isolates were also positive for oxidase activity (except ZM22), HCN production (except ZM27) and utilization of citrate. More than 70% of isolates produces ammonia, hydrogen cyanide, siderophores, exopolysaccharides and cellulase. More than half of isolates also showed potential for urease activity and production of lipase. The ZM31 and S10 were the only isolates which showed the chitinase activity. All these isolates were evaluated in a jar trial for their ability to promote growth of maize under axenic conditions. Results revealed that inoculation of selected zinc solubilizing rhizobacterial isolates improved the growth of maize. In comparison, isolates ZM20, ZM31, ZM63 and S10 were best compared to other tested isolates in stimulating the growth attributes of maize like shoot length, root length, plant fresh and dry biomass. These strains were identified as Bacillus sp. (ZM20), Bacillus aryabhattai (ZM31 and S10) and Bacillus subtilis (ZM63) through 16S rRNA sequencing. This study indicated that inoculation of Zn solubilizing strains have potential to promote growth and can be the potential bio-inoculants for biofortification of maize to overcome the problems of malnutrition. Copyright © 2017 Elsevier GmbH. All rights reserved.

  13. Characterization of Phosphate Solubilizing and Potassium Decomposing Strains and Study on their Effects on Tomato Cultivation

    Tin Mar Lynn; Hlaing Swe Win; Ei Phyu Kyaw; Zaw Ko Latt; San San Yu


    Seven strains were collected for phosphate solubilizing and potassium decomposing activities from Microbiology Laboratory, Department of Biotechnology, Shweziwa Biofertilizer Plant. When phosphate solubilizing activity of selected strains was qualitatively determined, all strains except from B1 strain, gave clear zone formation on NBRIP media. But when quantitatively determined by spectrophotometric method, all strains solubilized insoluble tricalcium phosphate. Among seven strains, Ps strain...

  14. Self-assembly of gold nanorods coated with phospholipids: a coarse-grained molecular dynamics study

    Wan, Mingwei; Li, Xiaoxu; Gao, Lianghui; Fang, Weihai


    The self-assembly of phospholipid-coated gold nanorods (GNRs) was investigated by coarse-grained molecular dynamics simulations. We predict that in addition to the formation of deformed vesicles encapsulating GNRs with diverse orientations, the lipid-coated GNRs can form a semi-ring attached to an excess vesicle phase, a branch with excess vesicle phase, a ring phase, a branch phase, a stack phase, and a vortex phase. The morphologies of the lipid-GNR complexes depend on the lipid/GNR molar ratio and the interaction strength between the nanorod surface and the lipid head groups. At given lipid-nanorod interactions, removing the lipid induces a phase transition from an isolated ring or branch phase to an aggregated vortex or stack phase and vice versa. As the lipid-coated GNRs transit from an isolated phase to an aggregated phase, the structure of the lipid at the nanorod surface converts from a bilayer state to a non-bilayer state.

  15. Distinct behaviour of the homeodomain derived cell penetrating peptide penetratin in interaction with different phospholipids.

    Ofelia Maniti

    Full Text Available BACKGROUND: Penetratin is a protein transduction domain derived from the homeoprotein Antennapedia. Thereby it is currently used as a cell penetrating peptide to introduce diverse molecules into eukaryotic cells, and it could also be involved in the cellular export of transcription factors. Moreover, it has been shown that it is able to act as an antimicrobial agent. The mechanisms involved in all these processes are quite controversial. METHODOLOGY/PRINCIPAL FINDINGS: In this article, we report spectroscopic, calorimetric and biochemical data on the penetratin interaction with three different phospholipids: phosphatidylcholine (PC and phosphatidylethanolamine (PE to mimic respectively the outer and the inner leaflets of the eukaryotic plasma membrane and phosphatidylglycerol (PG to mimic the bacterial membrane. We demonstrate that with PC, penetratin is able to form vesicle aggregates with no major change in membrane fluidity and presents no well defined secondary structure organization. With PE, penetratin aggregates vesicles, increases membrane rigidity and acquires an α-helical structure. With PG membranes, penetratin does not aggregate vesicles but decreases membrane fluidity and acquires a structure with both α-helical and β-sheet contributions. CONCLUSIONS/SIGNIFICANCE: These data from membrane models suggest that the different penetratin actions in eukaryotic cells (membrane translocation during export and import and on prokaryotes may result from different peptide and lipid structural arrangements. The data suggest that, for eukaryotic cell penetration, penetratin does not acquire classical secondary structure but requires a different conformation compared to that in solution.

  16. Vesicle formation and stability in the surfactant sodium 4-(1'-heptylnonyl) benzenesulfonate

    Franses, E.I.; Talmon, Y.; Scriven, L.E.; Davis, H.T.; Miller, W.G.


    Surfactants composed of a hydrophilic moiety covalently attached to the end of a hydrocarbon chain (e.g., sodium dodecyl sulfate), spontaneously form micelles, equilibrium aggregates, in solution if the surfactant concentration exceeds a critical value called the CMC. Naturally occurring double-tail surfactants (e.g., phospholipids) are not known to form micelles. Over a considerable range in surfactant concentration, 2 phases coexist in equilibrium: a hydrated, multilamellar (smectic) surfactant phase and an aqueous phase saturated with surfactant. In this report the preparation of vesicles, their direct, unstained visualization by electron microscopy, and investigation of their stability and structure by turbidimetry, conductimetry, light microscopy, densitometry, scanning calorimetry, and nuclear magnetic resonance spectroscopy are discussed. Dispersed liquid crystal was studied by the same means. For comparison, parallel studies on bovine lecithin are presented. From the results, it is concluded that these vesicles may be stable for many months, but eventually revert to multilamellar liquid crystals.

  17. A Time-Resolved Study on Nanodisc-to-Vesicle Transformation

    Nieh, Mu-Ping; Mahabir, Suanne; Wan, Wan Kei; Kastaras, John


    Structural phase diagram of a phospholipid mixture composed of dimyristoyl phosphatidylcholine (DMPC), dihexanoyl phosphatidylcholine (DHPC) and dimyristoyl phosphatidylglycerol (DMPG) contains many rich morphologies, e.g., nanodiscs also known as ``bicelles'', bilayered ribbons, unilamellar vesicles (ULVs), multi-lamellar vesicles (MLVs) and perforated lamellae. In this report, we will present time-resolved small angle neutron scattering and dynamic light scattering measurements of the structural transformation from nanodiscs to ULVs as a function of temperature, lipid concentration and charge density. The result will reveal the growth rate of nanodiscs and all the intermediate structures along the transformation process. Through the understanding of the kinetic pathway, the size and polydispersity of the self-assembled nano-size ULVs can be well-controlled. These ULVs can be used as a carrier for therapeutics or imaging probes.

  18. Comparison between advected-field and level-set methods in the study of vesicle dynamics

    Maitre, E; Peyla, P; Raoult, A


    Phospholipidic membranes and vesicles constitute a basic element in real biological functions. Vesicles are viewed as a model system to mimic basic viscoelastic behaviors of some cells, like red blood cells. Phase field and level-set models are powerful tools to tackle dynamics of membranes and their coupling to the flow. These two methods are somewhat similar, but to date no bridge between them has been made. This is a first focus of this paper. Furthermore, a constitutive viscoelastic law is derived for the composite fluid: the ambient fluid and the membranes. We present two different approaches to deal with the membrane local incompressibility, and point out differences. Some numerical results following from the level-set approach are presented.

  19. Application of potential phosphate-solubilizing bacteria and organic acids on phosphate solubilization from phosphate rock in aerobic rice.

    Panhwar, Qurban Ali; Jusop, Shamshuddin; Naher, Umme Aminun; Othman, Radziah; Razi, Mohd Ismail


    A study was conducted at Universiti Putra Malaysia to determine the effect of phosphate-solubilizing bacteria (PSB) and organic acids (oxalic & malic) on phosphate (P) solubilization from phosphate rock (PR) and growth of aerobic rice. Four rates of each organic acid (0, 10, 20, and 30 mM), and PSB strain (Bacillus sp.) were applied to aerobic rice. Total bacterial populations, amount of P solubilization, P uptake, soil pH, and root morphology were determined. The results of the study showed significantly high P solubilization in PSB with organic acid treatments. Among the two organic acids, oxalic acid was found more effective compared to malic acid. Application of oxalic acid at 20 mM along with PSB16 significantly increased soluble soil P (28.39 mg kg(-1)), plant P uptake (0.78 P pot(-1)), and plant biomass (33.26 mg). Addition of organic acids with PSB and PR had no influence on soil pH during the planting period. A higher bacterial population was found in rhizosphere (8.78 log10 cfu g(-1)) compared to the nonrhizosphere and endosphere regions. The application of organic acids along with PSB enhanced soluble P in the soil solution, improved root growth, and increased plant biomass of aerobic rice seedlings without affecting soil pH.

  20. Application of Potential Phosphate-Solubilizing Bacteria and Organic Acids on Phosphate Solubilization from Phosphate Rock in Aerobic Rice

    Qurban Ali Panhwar


    Full Text Available A study was conducted at Universiti Putra Malaysia to determine the effect of phosphate-solubilizing bacteria (PSB and organic acids (oxalic & malic on phosphate (P solubilization from phosphate rock (PR and growth of aerobic rice. Four rates of each organic acid (0, 10, 20, and 30 mM, and PSB strain (Bacillus sp. were applied to aerobic rice. Total bacterial populations, amount of P solubilization, P uptake, soil pH, and root morphology were determined. The results of the study showed significantly high P solubilization in PSB with organic acid treatments. Among the two organic acids, oxalic acid was found more effective compared to malic acid. Application of oxalic acid at 20 mM along with PSB16 significantly increased soluble soil P (28.39 mg kg−1, plant P uptake (0.78 P pot−1, and plant biomass (33.26 mg. Addition of organic acids with PSB and PR had no influence on soil pH during the planting period. A higher bacterial population was found in rhizosphere (8.78 log10 cfu g−1 compared to the nonrhizosphere and endosphere regions. The application of organic acids along with PSB enhanced soluble P in the soil solution, improved root growth, and increased plant biomass of aerobic rice seedlings without affecting soil pH.

  1. Pellet-free isolation of human and bovine milk extracellular vesicles by size-exclusion chromatography

    Blans, Kristine; Hansen, Maria S.; Sørensen, Laila V.; Hvam, Michael L.; Howard, Kenneth A.; Möller, Arne; Wiking, Lars; Larsen, Lotte B.; Rasmussen, Jan T.


    ABSTRACT Studies have suggested that nanoscale extracellular vesicles (EV) in human and bovine milk carry immune modulatory properties which could provide beneficial health effects to infants. In order to assess the possible health effects of milk EV, it is essential to use isolates of high purity from other more abundant milk structures with well-documented bioactive properties. Furthermore, gentle isolation procedures are important for reducing the risk of generating vesicle artefacts, particularly when EV subpopulations are investigated. In this study, we present two isolation approaches accomplished in three steps based on size-exclusion chromatography (SEC) resulting in effective and reproducible EV isolation from raw milk. The approaches do not require any EV pelleting and can be applied to both human and bovine milk. We show that SEC effectively separates phospholipid membrane vesicles from the primary casein and whey protein components in two differently obtained casein reduced milk fractions, with one of the fractions obtained without the use of ultracentrifugation. Milk EV isolates were enriched in lactadherin, CD9, CD63 and CD81 compared to minimal levels of the EV-marker proteins in other relevant milk fractions such as milk fat globules. Nanoparticle tracking analysis and electron microscopy reveals the presence of heterogeneous sized vesicle structures in milk EV isolates. Lipid analysis by thin layer chromatography shows that EV isolates are devoid of triacylglycerides and presents a phospholipid profile differing from milk fat globules surrounded by epithelial cell plasma membrane. Moreover, the milk EV fractions are enriched in RNA with distinct and diverging profiles from milk fat globules. Collectively, our data supports that successful milk EV isolation can be accomplished in few steps without the use of ultracentrifugation, as the presented isolation approaches based on SEC effectively isolates EV in both human and bovine milk. PMID:28386391

  2. Formation of Oligovesicular Vesicles by Micromanipulation

    Yukihisa Okumura


    Full Text Available Cell-sized lipid bilayer membrane vesicles (giant vesicles, GVs or semi-vesicles were formed from egg yolk phosphatidylcholine on a platinum electrode under applied electric voltage by electroformation. Micromanipulation of the semi-vesicle by first pressing its membrane with a glass microneedle and then withdrawing the needle left a GV in the interior of the vesicle. During the process, an aqueous solution of Ficoll that filled the needle was introduced into the newly formed inner vesicle and remained encapsulated. Approximately 50% of attempted micromanipulation resulted in the formation of an inner daughter vesicle, “microvesiculation”. By repeating the microvesiculation process, multiple inner GVs could be formed in a single parent semi-vesicle. A semi-vesicle with inner GVs could be detached from the electrode by scraping with a microneedle, yielding an oligovesicular vesicle (OVV with desired inner aqueous contents. Microvesiculation of a GV held on the tip of a glass micropipette was also possible, and this also produced an OVV. Breaking the membrane of the parent semi-vesicle by micromanipulation with a glass needle after microvesiculation, released the inner GVs. This protocol may be used for controlled formation of GVs with desired contents.

  3. Rheology of a vesicle suspension with finite concentration: a numerical study.

    Thiébaud, Marine; Misbah, Chaouqi


    Vesicles, closed membranes made of a bilayer of phospholipids, are considered as a biomimetic system for the mechanics of red blood cells. The understanding of their dynamics under flow and their rheology is expected to help the understanding of the behavior of blood flow. We conduct numerical simulations of a suspension of vesicles in two dimensions at a finite concentration in a shear flow imposed by countertranslating rigid bounding walls by using an appropriate Green's function. We study the dynamics of vesicles, their spatial configurations, and their rheology, namely, the effective viscosity η(eff). A key parameter is the viscosity contrast λ (the ratio between the viscosity of the encapsulated fluid over that of the suspending fluid). For small enough λ, vesicles are known to exhibit tank treading (TT), while at higher λ they exhibit tumbling (TB). We find that η(eff) decreases in the TT regime, passes a minimum at a critical λ=λ(c), and increases in the TB regime. This result confirms previous theoretical and numerical works performed in the extremely dilute regime, pointing to the robustness of the picture even in the presence of hydrodynamic interactions. Our results agree also with very recent numerical simulations performed in three dimensions both in the dilute and more concentrated regime. This points to the fact that dimensionality does not alter the qualitative features of η(eff). However, they disagree with recent simulations in two dimensions. We provide arguments about the possible sources of this disagreement.

  4. Small-Angle Neutron Scattering and Spontaneous Formation of Unilamellar Vesicles: Potential Vehicles for Drug Delivery

    Katsaras, John


    Unilamellar vesicles (ULVs) are single-bilayer shells with radii commonly between 10 and 100 nm, and are widely used as model membranes, drug delivery systems, microreactors and substrates for a variety of enzymes and proteins. A common method of making ULVs is the extrusion of multilamellar vesicles (MLVs) through synthetic membranes of known pore size. These extruded ULVs are invariably unstable and in due time, revert back to MLVs. Over the years there have been reports of the spontaneous formation of stable ULVs in surfactant, lipid, and lipid/detergent mixtures. These ULVs have sometimes been shown to be monodisperse and their radii were found, almost without exception, to vary with concentration. We have carried-out small-angle neutron scattering (SANS) experiments on a biomimetic system composed of the phospholipids dimyristoyl and dihexanoyl phosphorylcholine (DMPC and DHPC, respectively). Doping DMPC/DHPC multilamellar vesicles with either the negatively charged lipid dimyristoyl phosphorylglycerol (DMPG, net charge -1) or the divalent cation, calcium (Ca2+) leads to the spontaneous formation of monodisperse unilamellar vesicles whose radii are concentration independent, in contrast to previous experimental observations.

  5. Separation and identification of phospholipid peroxidation products.

    Milne, G L; Porter, N A


    The molecular species in mixtures of phospholipid hydroperoxides are difficult to separate and identify by typical chromatographic and mass spectrometric techniques. As reported by Havrilla and coworkers, silver ion coordination ion-spray mass spectrometry (CIS-MS) has proven to be a powerful technique for the identification of mixtures of hydroperoxides. This ionization technique, which involves the formation of Ag+ adducts of the hydroperoxides, provides valuable, unambiguous structural information about the hydroperoxides. Herein, we report a method for the analysis and identification of phospholipid hydroperoxides using CIS-MS. We also report an improved method for the separation of phospholipid hydroperoxides by reversed-phase high-performance liquid chromatography (RP-HPLC), which, for the first time, separates some of the hydroperoxide isomers. CIS-MS can be coupled with this RP-HPLC method by the addition of AgBF4 to the mobile phase or to the HPLC effluent postcolumn, thus allowing powerful HPLC-MS techniques to be used to identify complex mixtures of phospholipid hydroperoxides.

  6. Pseudocritical Behavior and Unbinding of Phospholipid Bilayers

    Lemmich, Jesper; Mortensen, Kell; Ipsen, John Hjorth;


    The temperature dependence of the small-angle neutron scattering from fully hydrated multilamellar phospholipid bilayers near the main phase transition is analyzed by means of a simple geometric model which yields both the lamellar repeat distance as well as the hydrophobic thickness of the bilayer...

  7. Computer simulations of phospholipid - membrane thermodynamic fluctuations

    Pedersen, U.R.; Peters, Günther H.j.; Schröder, T.B.


    This paper reports all-atom computer simulations of five phospholipid membranes, DMPC, DPPC, DMPG, DMPS, and DMPSH, with a focus on the thermal equilibrium fluctuations of volume, energy, area, thickness, and order parameter. For the slow fluctuations at constant temperature and pressure (defined...

  8. Fundamental Studies of Assembly and Mechanical Properties of Lipid Bilayer Membranes and Unilamellar Vesicles

    Wang, Xi

    This dissertation work focuses on: (i) obtaining a phospholipid bilayer membrane (LBM)/conducting electrode system with low defect density and optimized rigidity; (ii) investigating vesicle stability and mechanical properties. LBM is a simplified yet representative cell membrane model. LBMs assembled on conductive surfaces can probe protein-LBM interactions activities electrochemically. Sterically stabilized vesicles could be used as cell models or for drug delivery. The main challenges for LBM assembly on gold are vesicles do not spontaneously rupture to form LBMs on gold and the roughness of the gold substrate has considerable influence on molecular film defect density. In this study, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) vesicles were functionalized with 1,2-distearoyl-sn-glycero-3-phosphoethanolamine- N-poly(ethylene glycol)-2000-N-[3-(2-pyridyldithio)propionate] (DSPE-PEG-PDP) to yield stable LBMs on gold without surface modification. A template-stripping method was used to obtain atomically flat and pristine gold surfaces. The critical force to initiate vesicle rupture decreases with increasing DSPE-PEG-PDP concentration, indicating that gold-thiolate bonding between DSPE-PEG-PDP and gold substrates promotes LBM formation. Mechanical properties of LBMs and vesicles were investigated as a function of DSPE-PEG-PDP concentration via Atomic Force Microscopy. The elastic moduli of LBMs were determined with DSPE-PEG-PDP concentration ranging from 0mol% to 24mol% and were found to depend on PEG chain conformation. Incorporating DSPE-PEG-PDP molecules with PEG in mushroom conformation results in a decrease of LBM rigidity, while incorporating PEG in brush conformation leads to LBM stiffening. Contrarily, mechanical properties of functionalized vesicles did not vary significantly by varying DSPE-PEG-PDP concentration. LBM with tunable rigidity by adjusting DSPE-PEG-PDP concentration provides a versatile cell membrane model for studying protein or

  9. Solubilization properties of polycyclic aromatic hydrocarbons by saponin, a plant-derived biosurfactant

    Zhou Wenjun, E-mail: [Department of Environmental Science, Zhejiang University, Hangzhou, Zhejiang 310028 (China); Zhejiang Provincial Key Laboratory of Organic Pollution Process and Control, Hangzhou, Zhejiang 310028 (China); Yang Juanjuan; Lou Linjie [Department of Environmental Science, Zhejiang University, Hangzhou, Zhejiang 310028 (China); Zhu Lizhong [Department of Environmental Science, Zhejiang University, Hangzhou, Zhejiang 310028 (China); Zhejiang Provincial Key Laboratory of Organic Pollution Process and Control, Hangzhou, Zhejiang 310028 (China)


    The enhanced solubilization of polycyclic aromatic hydrocarbons (PAHs) by saponin, a plant-derived non-ionic biosurfactant, was investigated. The results indicated that the solubilization capabilities of saponin for PAHs were greater than some representative synthetic non-ionic surfactants and showed strong dependence on solution pH and ionic strength. The molar solubilization ratio (MSR) of saponin for phenanthrene was about 3-6 times of those of the synthetic non-ionic surfactants, and decreased by about 70% with the increase of solution pH from 4.0 to 8.0, but increased by approximately 1 times with NaCl concentration increased from 0.01 to 1.0 M. Heavy metal ions can enhance saponin solubilization for phenanthrene and the corresponding MSR values increased by about 25% with the presence of 0.01 M of Cd{sup 2+} or Zn{sup 2+}. Saponin is more effective in enhancing PAHs solubilization than synthetic non-ionic surfactants and has potential application in removing organic pollutants from contaminated soils. - Highlights: > The enhanced solubilization of PAHs by saponin was investigated in this study. > Saponin showed great solubilization capability for PAHs. > Saponin is more effective in enhancing HOCs solubilization at lower solution pH. > Increasing ionic strength can enhance HOCs solubilization in saponin solution. > Heavy metal ions can also enhance phenanthrene solubilization in saponin solution. - Saponin showed different solubilization properties for PAHs from the synthetic non-ionic surfactants and anionic rhamnolipid biosurfactants.

  10. Blood clotting reactions on nanoscale phospholipid bilayers.

    Morrissey, James H; Pureza, Vincent; Davis-Harrison, Rebecca L; Sligar, Stephen G; Ohkubo, Y Zenmei; Tajkhorshid, Emad


    Blood clotting reactions, such as those catalyzed by the tissue factor:factor VIIa complex (TF:FVIIa), assemble on membrane surfaces containing anionic phospholipids such as phosphatidylserine (PS). In fact, membrane binding is critical for the function of most of the steps in the blood clotting cascade. In spite of this, our understanding of how the membrane contributes to catalysis, or even how these proteins interact with phospholipids, is incomplete. Making matters more complicated, membranes containing mixtures of PS and neutral phospholipids are known to spontaneously form PS-rich membrane microdomains in the presence of plasma concentrations of calcium ions, and it is likely that blood-clotting proteases such as TF:FVIIa partition into these PS-rich microdomains. Unfortunately, little is known about how membrane microdomain composition influences the activity of blood-clotting proteases, which is typically not under experimental control even in "simple" model membranes. Our laboratories have developed and applied new technologies for studying membrane proteins to gain insights into how blood-clotting protease-cofactor pairs assemble and function on membrane surfaces. This includes using a novel, nanoscale bilayer system (Nanodiscs) that permits assembling blood-clotting protease-cofactor pairs on stable bilayers containing from 65 to 250 phospholipid molecules per leaflet. We have used this system to investigate how local (nanometer-scale) changes in phospholipid bilayer composition modulate TF:FVIIa activity. We have also used detailed molecular-dynamics simulations of nanoscale bilayers to provide atomic-scale predictions of how the membrane-binding domain of factor VIIa interacts with PS in membranes.

  11. Solubilization of Australian lignites by fungi and other microorganisms

    Catcheside, D.E.A.; Mallett, K.J. (Flinders University, Bedford Park, SA (Australia). School of Biological Sciences)

    Lignites (brown coals) from the Latrobe Valley in Victoria are solubilized by {ital Coriolus versicolor}, {ital Phanerochaete chrysosporium}, and five other species known to be active on Leonardite and various acid-treated North America lignites. Run-of-mine coal from Morwell and Loy Yang is refractory but is soluble after pretreatment with acid. A weathered deposit at Loy Yang, like Leonardite, is susceptible to biosolubilization without pretreatment. The white rot fungi {ital Ganoderma applanatum}, {ital Perenniporia tephropora} ({ital Fomes lividus}), {ital Pleurotus ostreatus}, {ital Pycnoporus cinnabarinus}, {ital Rigidoporus ulmarius}, and {ital Xylaria hypoxylon} were found to be capable of solubilizing lignite. In contrast, brown rot fungi were weakly active or inactive under the same test conditions. Lignite-degrading fungi, actinomycetes, and other bacteria, including some active on untreated run-of-mine coal, were isolated from natural lignite exposures and mining sites. 15 refs., 5 tabs.

  12. Recovery Act - Large Scale SWNT Purification and Solubilization

    Michael Gemano; Dr. Linda B. McGown


    The goal of this Phase I project was to establish a quantitative foundation for development of binary G-gels for large-scale, commercial processing of SWNTs and to develop scientific insight into the underlying mechanisms of solubilization, selectivity and alignment. In order to accomplish this, we performed systematic studies to determine the effects of G-gel composition and experimental conditions that will enable us to achieve our goals that include (1) preparation of ultra-high purity SWNTs from low-quality, commercial SWNT starting materials, (2) separation of MWNTs from SWNTs, (3) bulk, non-destructive solubilization of individual SWNTs in aqueous solution at high concentrations (10-100 mg/mL) without sonication or centrifugation, (4) tunable enrichment of subpopulations of the SWNTs based on metallic vs. semiconductor properties, diameter, or chirality and (5) alignment of individual SWNTs.

  13. Spontaneous Vesicle Recycling in the Synaptic Bouton

    Sven eTruckenbrodt


    Full Text Available The trigger for synaptic vesicle exocytosis is Ca2+, which enters the synaptic bouton following action potential stimulation. However, spontaneous release of neurotransmitter also occurs in the absence of stimulation in virtually all synaptic boutons. It has long been thought that this represents exocytosis driven by fluctuations in local Ca2+ levels. The vesicles responding to these fluctuations are thought to be the same ones that release upon stimulation, albeit potentially triggered by different Ca2+ sensors. This view has been challenged by several recent works, which have suggested that spontaneous release is driven by a separate pool of synaptic vesicles. Numerous articles appeared during the last few years in support of each of these hypotheses, and it has been challenging to bring them into accord. We speculate here on the origins of this controversy, and propose a solution that is related to developmental effects. Constitutive membrane traffic, needed for the biogenesis of vesicles and synapses, is responsible for high levels of spontaneous membrane fusion in young neurons, probably independent of Ca2+. The vesicles releasing spontaneously in such neurons are not related to other synaptic vesicle pools and may represent constitutively releasing vesicles (CRVs rather than bona fide synaptic vesicles. In mature neurons, constitutive traffic is much dampened, and the few remaining spontaneous release events probably represent bona fide spontaneously releasing synaptic vesicles (SRSVs responding to Ca2+ fluctuations, along with a handful of CRVs that participate in synaptic vesicle turnover.

  14. Spontaneous vesicle recycling in the synaptic bouton.

    Truckenbrodt, Sven; Rizzoli, Silvio O


    The trigger for synaptic vesicle exocytosis is Ca(2+), which enters the synaptic bouton following action potential stimulation. However, spontaneous release of neurotransmitter also occurs in the absence of stimulation in virtually all synaptic boutons. It has long been thought that this represents exocytosis driven by fluctuations in local Ca(2+) levels. The vesicles responding to these fluctuations are thought to be the same ones that release upon stimulation, albeit potentially triggered by different Ca(2+) sensors. This view has been challenged by several recent works, which have suggested that spontaneous release is driven by a separate pool of synaptic vesicles. Numerous articles appeared during the last few years in support of each of these hypotheses, and it has been challenging to bring them into accord. We speculate here on the origins of this controversy, and propose a solution that is related to developmental effects. Constitutive membrane traffic, needed for the biogenesis of vesicles and synapses, is responsible for high levels of spontaneous membrane fusion in young neurons, probably independent of Ca(2+). The vesicles releasing spontaneously in such neurons are not related to other synaptic vesicle pools and may represent constitutively releasing vesicles (CRVs) rather than bona fide synaptic vesicles. In mature neurons, constitutive traffic is much dampened, and the few remaining spontaneous release events probably represent bona fide spontaneously releasing synaptic vesicles (SRSVs) responding to Ca(2+) fluctuations, along with a handful of CRVs that participate in synaptic vesicle turnover.

  15. Mobilization and micellar solubilization of NAPL contaminants in aquifer rocks

    Javanbakht, Gina; Goual, Lamia


    Surfactant-enhanced aquifer remediation is often performed to overcome the capillary forces that keep residual NAPL phases trapped within contaminated aquifers. The surfactant selection and displacement mechanism usually depend on the nature of NAPL constituents. For example, micellar solubilization is often used to cleanup DNAPLs from aquifers whereas mobilization is desirable in aquifers contaminated by LNAPLs. Although the majority of crude oils are LNAPLs, they often contain heavy organic macromolecules such as asphaltenes that are classified as DNAPLs. Asphaltenes contain surface-active components that tend to adsorb on rocks, altering their wettability. Previous studies revealed that surfactants that formed Winsor type III microemulsions could promote both mobilization and solubilization. However the extent by which these two mechanisms occur is still unclear, particularly in oil-contaminated aquifers. In this study we investigated the remediation of oil-contaminated aquifers using an environmentally friendly surfactant such as n-Dodecyl β-D-maltoside. Focus was given on asphaltenes to better understand the mechanisms of surfactant cleanup. Through phase behavior, spontaneous imbibition, dynamic interfacial tension and contact angle measurements, we showed that microemulsions formed by this surfactant are able to mobilize bulk NAPL (containing 9 wt.% asphaltenes) in the porous rock and solubilize DNAPL (i.e., 4-6 wt.% adsorbed asphaltenes) from the rock surface. Spontaneous imbibition tests, in particular, indicated that the ratio of mobilized to solubilized NAPL is about 6:1. Furthermore, aging the cores in NAPL beyond 3 days allowed for more NAPL to be trapped in the large pores of the rock but did not alter the amount of asphaltenes adsorbed on the mineral surface.

  16. Synthesis and function of phospholipids in Staphylococcus aureus.

    Kuhn, Sebastian; Slavetinsky, Christoph J; Peschel, Andreas


    Phospholipids are the major components of bacterial membranes, and changes in phospholipid composition affect important cellular processes such as metabolism, stress response, antimicrobial resistance, and virulence. The most prominent phospholipids in Staphylococcus aureus are phosphatidylglycerol, lysyl-phosphatidylglycerol, and cardiolipin, whose biosynthesis is mediated by a complex protein machinery. Phospholipid composition of the staphylococcal membrane has to be continuously adjusted to changing external conditions, which is achieved by a series of transcriptional and biochemical regulatory mechanisms. This mini-review outlines the current state of knowledge concerning synthesis, regulation, and function of the major staphylococcal phospholipids.

  17. Hydrotropic Solubilization by Urea Derivatives: A Molecular Dynamics Simulation Study

    Yong Cui


    Full Text Available Hydrotropy is a phenomenon where the presence of a large quantity of one solute enhances the solubility of another solute. The mechanism of this phenomenon remains a topic of debate. This study employed molecular dynamics simulation to investigate the hydrotropic mechanism of a series of urea derivatives, that is, urea (UR, methylurea (MU, ethylurea (EU, and butylurea (BU. A poorly water-soluble compound, nifedipine (NF, was used as the model solute that was solubilized. Structural, dynamic, and energetic changes upon equilibration were analyzed to supply insights to the solubilization mechanism. The study demonstrated that NF and urea derivatives underwent significant nonstoichiometric molecular aggregation in the aqueous solution, a result consistent with the self-aggregation of urea derivatives under the same conditions. The analysis of hydrogen bonding and energy changes revealed that the aggregation was driven by the partial restoration of normal water structure. The energetic data also suggested that the promoted solubilization of NF is favored in the presence of urea derivatives. While the solutes aggregated to a varying degree, the systems were still in single-phase liquid state as attested by their active dynamics.

  18. Extractant efficiency in the solubilization of alternative sources of potassium

    Daniele Nogueira dos Reis


    Full Text Available Knowledge of a rock’s composition allows for inferences regarding several properties, ranging from its physical characteristics to its solubility. This study aimed to evaluate the use of different extractants to solubilize the K present in rocks as a potential source of nutrients and the effects of extractant contact time and temperature on rock solubilization. Samples of two rocks and a mineral concentrated from a granitic rock were treated with ammonium dihydrogen phosphate (NH4(H2PO4, calcium dihydrogen phosphate (Ca(H2PO42, sodium hydroxide (NaOH and water (control. Sample-extractant treatments were performed using a water bath shaker at temperatures of 25 and 50°C for periods of 3, 7, 10, 20, and 30 days. The amounts of K extracted from rocks using the extractants were in the following order: NH4H2PO4>Ca(HPO42>NaOH>water. The sequence of K release (ppm based on the rocks studied was as follows: nepheline syenite>green banded argillite>concentrated biotite. Increasing the extractant contact time and temperature enhanced the solubilized K content.

  19. Solubilization of diabase and phonolite dust by filamentous fungus

    Juliana Andréia Vrba Brandão


    Full Text Available The objective of this study was to evaluate the effect of the fungus Aspergillus niger strain CCT4355 in the release of nutrients contained in two types of rock powder (diabase and phonolite by means of in vitro solubilization trials. The experimental design was completely randomized in a 5 x 4 factorial design with three replications. It was evaluated five treatments (phonolite dust + culture medium; phonolite dust + fungus + culture medium; diabase powder + culture medium; diabase powder + fungus + culture medium and fungus + culture medium and four sampling dates (0, 10, 20 and 30 days. Rock dust (0.4% w/v was added to 125 mL Erlenmeyer flasks containing 50 mL of liquid culture medium adapted to A. niger. The flasks were incubated at 30°C for 30 days, and analysis of pH (in water, titratable acidity, and concentrations of soluble potassium, calcium, magnesium, zinc, iron and manganese were made. The fungus A. niger was able to produce organic acids that solubilized ions. This result indicates its potential to alter minerals contained in rock dust, with the ability to interact in different ways with the nutrients. A significant increase in the amount of K was found in the treatment with phonolite dust in the presence of the fungus. The strain CCT4355 of A. niger can solubilize minerals contained in these rocks dust.

  20. Alignment of synaptic vesicle macromolecules with the macromolecules in active zone material that direct vesicle docking.

    Harlow, Mark L; Szule, Joseph A; Xu, Jing; Jung, Jae Hoon; Marshall, Robert M; McMahan, Uel J


    Synaptic vesicles dock at active zones on the presynaptic plasma membrane of a neuron's axon terminals as a precondition for fusing with the membrane and releasing their neurotransmitter to mediate synaptic impulse transmission. Typically, docked vesicles are next to aggregates of plasma membrane-bound macromolecules called active zone material (AZM). Electron tomography on tissue sections from fixed and stained axon terminals of active and resting frog neuromuscular junctions has led to the conclusion that undocked vesicles are directed to and held at the docking sites by the successive formation of stable connections between vesicle membrane proteins and proteins in different classes of AZM macromolecules. Using the same nanometer scale 3D imaging technology on appropriately stained frog neuromuscular junctions, we found that ∼10% of a vesicle's luminal volume is occupied by a radial assembly of elongate macromolecules attached by narrow projections, nubs, to the vesicle membrane at ∼25 sites. The assembly's chiral, bilateral shape is nearly the same vesicle to vesicle, and nubs, at their sites of connection to the vesicle membrane, are linked to macromolecules that span the membrane. For docked vesicles, the orientation of the assembly's shape relative to the AZM and the presynaptic membrane is the same vesicle to vesicle, whereas for undocked vesicles it is not. The connection sites of most nubs on the membrane of docked vesicles are paired with the connection sites of the different classes of AZM macromolecules that regulate docking, and the membrane spanning macromolecules linked to these nubs are also attached to the AZM macromolecules. We conclude that the luminal assembly of macromolecules anchors in a particular arrangement vesicle membrane macromolecules, which contain the proteins that connect the vesicles to AZM macromolecules during docking. Undocked vesicles must move in a way that aligns this arrangement with the AZM macromolecules for docking

  1. Phase Transition Induced Fission in Lipid Vesicles

    Leirer, C; Myles, V M; Schneider, M F


    In this work we demonstrate how the first order phase transition in giant unilamellar vesicles (GUVs) can function as a trigger for membrane fission. When driven through their gel-fluid phase transition GUVs exhibit budding or pearl formation. These buds remain connected to the mother vesicle presumably by a small neck. Cooling these vesicles from the fluid phase (T>Tm) through the phase transition into the gel state (Tvesicle remains intact. Pearling tubes which formed upon heating break-up and decay into multiple individual vesicles which then diffuse freely. Finally we demonstrate that mimicking the intracellular bulk viscosity by increasing the bulk viscosity to 40cP does not affect the overall fission process, but leads to a significant decrease in size of the released vesicles.

  2. Synaptic vesicle proteins and active zone plasticity

    Robert J Kittel


    Full Text Available Neurotransmitter is released from synaptic vesicles at the highly specialized presynaptic active zone. The complex molecular architecture of active zones mediates the speed, precision and plasticity of synaptic transmission. Importantly, structural and functional properties of active zones vary significantly, even for a given connection. Thus, there appear to be distinct active zone states, which fundamentally influence neuronal communication by controlling the positioning and release of synaptic vesicles. Vice versa, recent evidence has revealed that synaptic vesicle components also modulate organizational states of the active zone.The protein-rich cytomatrix at the active zone (CAZ provides a structural platform for molecular interactions guiding vesicle exocytosis. Studies in Drosophila have now demonstrated that the vesicle proteins Synaptotagmin-1 (Syt1 and Rab3 also regulate glutamate release by shaping differentiation of the CAZ ultrastructure. We review these unexpected findings and discuss mechanistic interpretations of the reciprocal relationship between synaptic vesicles and active zone states, which has heretofore received little attention.

  3. Synaptic Vesicle Proteins and Active Zone Plasticity.

    Kittel, Robert J; Heckmann, Manfred


    Neurotransmitter is released from synaptic vesicles at the highly specialized presynaptic active zone (AZ). The complex molecular architecture of AZs mediates the speed, precision and plasticity of synaptic transmission. Importantly, structural and functional properties of AZs vary significantly, even for a given connection. Thus, there appear to be distinct AZ states, which fundamentally influence neuronal communication by controlling the positioning and release of synaptic vesicles. Vice versa, recent evidence has revealed that synaptic vesicle components also modulate organizational states of the AZ. The protein-rich cytomatrix at the active zone (CAZ) provides a structural platform for molecular interactions guiding vesicle exocytosis. Studies in Drosophila have now demonstrated that the vesicle proteins Synaptotagmin-1 (Syt1) and Rab3 also regulate glutamate release by shaping differentiation of the CAZ ultrastructure. We review these unexpected findings and discuss mechanistic interpretations of the reciprocal relationship between synaptic vesicles and AZ states, which has heretofore received little attention.

  4. Stokesian jellyfish: Viscous locomotion of bilayer vesicles

    Evans, Arthur A; Lauga, Eric


    Motivated by recent advances in vesicle engineering, we consider theoretically the locomotion of shape-changing bilayer vesicles at low Reynolds number. By modulating their volume and membrane composition, the vesicles can be made to change shape quasi-statically in thermal equilibrium. When the control parameters are tuned appropriately to yield periodic shape changes which are not time-reversible, the result is a net swimming motion over one cycle of shape deformation. For two classical vesicle models (spontaneous curvature and bilayer coupling), we determine numerically the sequence of vesicle shapes through an enthalpy minimization, as well as the fluid-body interactions by solving a boundary integral formulation of the Stokes equations. For both models, net locomotion can be obtained either by continuously modulating fore-aft asymmetric vesicle shapes, or by crossing a continuous shape-transition region and alternating between fore-aft asymmetric and fore-aft symmetric shapes. The obtained hydrodynamic e...

  5. Biophysical studies of cholesterol in unsaturated phospholipid model membranes

    Williams, Justin Adam

    PUFAs can incorporate into lipid rafts, which are domains enriched in SM and chol in the plasma membrane, and potentially disrupt the activity of signaling proteins that reside therein. DHA, furthermore, may be the more potent component of fish oil. PUFA-chol interactions were also examined through affinity measurements. A novel method utilizing electron paramagnetic resonance (EPR) was developed, to monitor the partitioning of a spin-labeled analog of chol, 3beta-doxyl-5alpha-cholestane (chlstn), between large unilamellar vesicles (LUVs) and methyl-beta-cyclodextrin (mbetaCD). The EPR spectra for chlstn in the two environments are distinguishable due to the substantial differences in tumbling rates, allowing the population distribution ratio to be determined by spectral simulation. Advantages of this approach include speed of implementation and avoidance of potential artifacts associated with physical separation of LUV and mbetaCD. Additionally, in a check of the method, the relative partition coefficients between lipids measured for the spin label analog agree with values obtained for chol by isothermal titration calorimetry (ITC). Results from LUV with different composition confirmed a hierarchy of decreased sterol affinity for phospholipids with increasing acyl chain unsaturation, PDPC possessing half the affinity of the corresponding monounsaturated phospholipid. Taken together, the results of these studies on model membranes demonstrate the potential for PUFA-driven alteration of the architecture of biomembranes, a mechanism through which human health may be impacted.

  6. Constraint for the Existence of Ellipsoidal Vesicles

    XIE Yu-Zhang


    Under the spontaneous curvature model of lipid bilayers, the constraints for the existence of equilibrium axisym metric oblate and prolate ellipsoidal vesicles are obtained from the general shape equation. They degenerate either to the constraint for the existence of a spherical vesicle or to that of a circular cylindrical vesicle given by Ou-Yang and Helfrich [Phys. Rev. Lett. 59 (1987) 2486; 60(1988)120; Phys. Rev. A 39 (1989) 5280].

  7. Synthesis of magnetic Fe sub 3 O sub 4 particles covered with a modifiable phospholipid coat

    Cuyper, M D; Lueken, H; Hodenius, M


    This work reports the synthesis of iron oxide cores by coprecipitation of Fe sup 2 sup + and Fe sup 3 sup + ions with NaHCO sub 3 or NH sub 3. Depending on the experimental conditions, particles of two different sizes (13 or 130 nm diameter) were produced. X-ray diffractometry revealed Fe sub 3 O sub 4 (magnetite) to be the main constituent. The smaller particles, which, in contrast to the larger ones, are superparamagnetic, were stabilized with a phospholipid bilayer consisting of a 9:1 molar ratio of dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol, thereby creating so-called magnetoliposomes. In a subsequent step, poly(ethylene glycol)-(PEG-) derivatized dipalmitoylphosphatidylethanolamine was introduced into the lipid envelope by incubating the magnetoliposomes with pre-formed sonicated vesicles containing the PEGylated lipid. The mechanism by which lipid transfer occurred was determined from the kinetic profiles. The relevance of these observations to a wide range of biomedical applicat...

  8. Transcriptome of extracellular vesicles released by hepatocytes.

    Felix Royo

    Full Text Available The discovery that the cells communicate through emission of vesicles has opened new opportunities for better understanding of physiological and pathological mechanisms. This discovery also provides a novel source for non-invasive disease biomarker research. Our group has previously reported that hepatocytes release extracellular vesicles with protein content reflecting the cell-type of origin. Here, we show that the extracellular vesicles released by hepatocytes also carry RNA. We report the messenger RNA composition of extracellular vesicles released in two non-tumoral hepatic models: primary culture of rat hepatocytes and a progenitor cell line obtained from a mouse foetal liver. We describe different subpopulations of extracellular vesicles with different densities and protein and RNA content. We also show that the RNA cargo of extracellular vesicles released by primary hepatocytes can be transferred to rat liver stellate-like cells and promote their activation. Finally, we provide in vitro and in vivo evidence that liver-damaging drugs galactosamine, acetaminophen, and diclofenac modify the RNA content of these vesicles. To summarize, we show that the extracellular vesicles secreted by hepatocytes contain various RNAs. These vesicles, likely to be involved in the activation of stellate cells, might become a new source for non-invasive identification of the liver toxicity markers.

  9. Interaction of isopropylthioxanthone with phospholipid liposomes.

    Momo, Federico; Fabris, Sabrina; Stevanato, Roberto


    Isopropylthioxanthone (ITX) is a highly lipophilic molecule which can be released in foods and beverages from the packages, where it is present as photoinitiator of inks in printing processes. Recently it was found in babies milk, and its toxicity cannot be excluded. The structure of the molecule suggests a possible strong interaction with the lipid moiety of biological membranes, and this is the first study of its effects on phospholipid organization, using differential scanning calorimetry (DSC) and spin labelling techniques. The data obtained with multilamellar liposomes of saturated phospholipids of different length, with and without cholesterol, point out that the molecule changes the lipid structure; in particular, in the gel state, behaving like a disordering agent it increases the mobility of the bilayer, while, in the fluid state, tends to rigidify the membrane, in a cholesterol like way. This behavior supports the hypothesis that ITX experiences a relocation process when the lipid matrix passes from the gel to the fluid state.

  10. Planar bilayer membranes from photoactivable phospholipids.

    Borle, F; Sänger, M; Sigrist, H


    Planar bilayer membranes formed from photoactivable phospholipids have been characterized by low frequency voltametry. Cyclic voltametric measurements were applied for simultaneous registration of planar membrane conductivity and capacitance. The procedure has been utilized to characterize the formation and stability of planar bilayer membranes. Bilayer membranes were formed from N'-(1,2-dimyristoyl-sn-glycero-3-phosphoethyl)-N-((m-3- trifluoromethyldiazirine)phenyl)thiourea (C14-PED), a head-group photosensitive phospholipid. In situ photoactivation of C14-PED at wavelengths greater than or equal to 320 nm altered neither the mean conductivity nor the capacitance of the bilayer. Ionophore (valinomycin) and ion channel (gramicidin) activities were not impaired upon photoactivation. In contrast, bilayer membranes formed from 1,2-bis(hexadeca-2,4-dienoyl)-sn- glycero-3-phosphocholine (C16-DENPC) revealed short life times. In situ photopolymerization of the diene fatty acids significantly increased the membrane conductivity or led to membrane rupture.

  11. Vesicles

    ... poison ivy) Herpes simplex (cold sores, genital herpes ) Herpes zoster (shingles) Impetigo Fungal infections Burns Home Care It ... disease on the soles Herpes simplex - close-up Herpes zoster (shingles) - close-up of lesion Poison ivy on ...

  12. Molecular phospholipid films on solid supports

    Czolkos, Ilja; Jesorka, Aldo; Orwar, Owe


    Phospholipid membranes are versatile structures for mimicking biological surfaces. Bilayer and monolayer membranes can be formed on solid supports, leading to enhanced stability and accessibility of the biomimetic molecular film. This has facilitated functional studies of membrane proteins...... stable lipid membranes. In this review, the current state of the art of molecularly thin lipid layer fabrication is presented with an emphasis on support materials, film formation mechanisms, characterisation methods, and applications....

  13. Phospholipids as Biomarkers for Excessive Alcohol Use


    is designed to evaluate the utility of levels of two phospholipids in serum as a marker of past drinking behavior across month- level time an attempt to improve ability to measure alcohol quantity consumed and associated damage better than can be done with ethyl alcohol level measures...and other existing tests that only measure very recent exposure and poorly reflect quantity consumed . This will be achieved by correlating detailed

  14. Precise detection of pH inside large unilamellar vesicles using membrane-impermeable dendritic porphyrin-based nanoprobes.

    Leiding, Thom; Górecki, Kamil; Kjellman, Tomas; Vinogradov, Sergei A; Hägerhäll, Cecilia; Arsköld, Sindra Peterson


    Accurate real-time measurements of proton concentration gradients are pivotal to mechanistic studies of proton translocation by membrane-bound enzymes. Here we report a detailed characterization of the pH-sensitive fluorescent nanoprobe Glu(3), which is well suited for pH measurements in microcompartmentalized biological systems. The probe is a polyglutamic porphyrin dendrimer in which multiple carboxylate termini ensure its high water solubility and prevent its diffusion across phospholipid membranes. The probe's pK is in the physiological pH range, and its protonation can be followed ratiometrically by absorbance or fluorescence in the ultraviolet-visible spectral region. The usefulness of the probe was enhanced by using a semiautomatic titration system coupled to a charge-coupled device (CCD) spectrometer, enabling fast and accurate titrations and full spectral coverage of the system at millisecond time resolution. The probe's pK was measured in bulk solutions as well as inside large unilamellar vesicles in the presence of physiologically relevant ions. Glu(3) was found to be completely membrane impermeable, and its distinct spectroscopic features permit pH measurements inside closed membrane vesicles, enabling quantitative mechanistic studies of membrane-spanning proteins. Performance of the probe was demonstrated by monitoring the rate of proton leakage through the phospholipid bilayer in large vesicles with and without the uncoupler gramicidin present. Overall, as a probe for biological proton translocation measurements, Glu(3) was found to be superior to the commercially available pH indicators.

  15. Annexin-Phospholipid Interactions. Functional Implications

    Javier Turnay


    Full Text Available Annexins constitute an evolutionary conserved multigene protein superfamily characterized by their ability to interact with biological membranes in a calcium dependent manner. They are expressed by all living organisms with the exception of certain unicellular organisms. The vertebrate annexin core is composed of four (eight in annexin A6 homologous domains of around 70 amino acids, with the overall shape of a slightly bent ring surrounding a central hydrophilic pore. Calcium- and phospholipid-binding sites are located on the convex side while the N-terminus links domains I and IV on the concave side. The N-terminus region shows great variability in length and amino acid sequence and it greatly influences protein stability and specific functions of annexins. These proteins interact mainly with acidic phospholipids, such as phosphatidylserine, but differences are found regarding their affinity for lipids and calcium requirements for the interaction. Annexins are involved in a wide range of intra- and extracellular biological processes in vitro, most of them directly related with the conserved ability to bind to phospholipid bilayers: membrane trafficking, membrane-cytoskeleton anchorage, ion channel activity and regulation, as well as antiinflammatory and anticoagulant activities. However, the in vivo physiological functions of annexins are just beginning to be established.

  16. Hepatic Bel-7402 Cell Proliferation on Different Phospholipid Surfaces


    Phospholipids are believed to be important biomaterials.However, limited information is available on their cytocompatibilities.The objective of this study is to evaluate the effects of different phospholipids on the proliferation of hepatic Bel-7402 cells by comparing the adhesion, viability and proliferation of Bel-7402 cells cultured on different phospholipid surfaces.The cell adhesion, determined by counting the number of adhered cells to the surface, indicated that the cell adhesion was enhanced on charged phospolipid membranes.The cell viability evaluated by MTT[3 (4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium-bromide] showed that cells cultured on charged phospholipids have greater viability than those cultured on the control, while cells cultured on neutral phospholipids showed lower viability.The cell cycle analysis using flow cytometry demonstrated that S phase entry increased on charged phospholipids, while S phase entry decreased on neutral phospholipids.The results suggested that charged phospholipids, especially positively charged phospholipids, show better cytocompatibilities than neutral phospholipids to hepatic Bel-7402 cell.

  17. Specific Genomic Fingerprints of Phosphate Solubilizing Pseudomonas Strains Generated by Box Elements

    Javadi Nobandegani, Mohammad Bagher; Saud, Halimi Mohd; Yun, Wong Mui


    Primers corresponding to conserved bacterial repetitive of BOX elements were used to show that BOX-DNA sequences are widely distributed in phosphate solubilizing Pseudomonas strains. Phosphate solubilizing Pseudomonas was isolated from oil palm fields (tropical soil) in Malaysia. BOX elements were used to generate genomic fingerprints of a variety of Pseudomonas isolates to identify strains that were not distinguishable by other classification methods. BOX-PCR, that derived genomic fingerprints, was generated from whole purified genomic DNA by liquid culture of phosphate solubilizing Pseudomonas. BOX-PCR generated the phosphate solubilizing Pseudomonas specific fingerprints to identify the relationship between these strains. This suggests that distribution of BOX elements' sequences in phosphate solubilizing Pseudomonas strains is the mirror image of their genomic structure. Therefore, this method appears to be a rapid, simple, and reproducible method to identify and classify phosphate solubilizing Pseudomonas strains and it may be useful tool for fast identification of potential biofertilizer strains. PMID:25580434

  18. Hybrid phospholipid bilayer coatings for separations of cationic proteins in capillary zone electrophoresis.

    Gallagher, Elyssia S; Adem, Seid M; Bright, Leonard K; Calderon, Isen A C; Mansfield, Elisabeth; Aspinwall, Craig A


    Protein separations in CZE suffer from nonspecific adsorption of analytes to the capillary surface. Semipermanent phospholipid bilayers have been used to minimize adsorption, but must be regenerated regularly to ensure reproducibility. We investigated the formation, characterization, and use of hybrid phospholipid bilayers (HPBs) as more stable biosurfactant capillary coatings for CZE protein separations. HPBs are formed by covalently modifying a support with a hydrophobic monolayer onto which a self-assembled lipid monolayer is deposited. Monolayers prepared in capillaries using 3-cyanopropyldimethylchlorosilane (CPDCS) or n-octyldimethylchlorosilane (ODCS) yielded hydrophobic surfaces with lowered surface free energies of 6.0 ± 0.3 or 0.2 ± 0.1 mJ m(-2) , respectively, compared to 17 ± 1 mJ m(-2) for bare silica capillaries. HPBs were formed by subsequently fusing vesicles comprised of 1,2-dilauroyl-sn-glycero-3-phosphocholine or 1,2-dioleoyl-sn-glycero-3-phosphocholine to CPDCS- or ODCS-modified capillaries. The resultant HPB coatings shielded the capillary surface and yielded reduced electroosmotic mobility (1.3-1.9 × 10(-4) cm(2) V(-1) s(-1) ) compared to CPDCS- and ODCS-modified or bare capillaries (3.6 ± 0.2 × 10(-4) cm(2) V(-1) s(-1) , 4.8 ± 0.4 × 10(-4) cm(2) V(-1) s(-1) , and 6.0 ± 0.2 × 10(-4) cm(2) V(-1) s(-1) , respectively), with increased stability compared to phospholipid bilayer coatings. HPB-coated capillaries yielded reproducible protein migration times (RSD ≤ 3.6%, n ≥ 6) with separation efficiencies as high as 200 000 plates/m.

  19. Fluidized-bed bioreactor system for the microbial solubilization of coal

    Scott, C.D.; Strandberg, G.W.


    A fluidized-bed bioreactor system for the conversion of coal into microbially solubilized coal products. The fluidized-bed bioreactor continuously or periodically receives coal and bio-reactants and provides for the production of microbially solubilized coal products in an economical and efficient manner. An oxidation pretreatment process for rendering coal uniformly and more readily susceptible to microbial solubilization may be employed with the fluidized-bed bioreactor. 2 figs.

  20. Fixed-bed bioreactor system for the microbial solubilization of coal

    Scott, C.D.; Strandberg, G.W.


    A fixed-bed bioreactor system for the conversion of coal into microbially solubilized coal products. The fixed-bed bioreactor continuously or periodically receives coal and bio-reactants and provides for the large scale production of microbially solubilized coal products in an economical and efficient manner. An oxidation pretreatment process for rendering coal uniformly and more readily susceptible to microbial solubilization may be employed with the fixed-bed bioreactor. 1 fig., 1 tab.

  1. Biochemical solubilization of toxic salts from residual geothermal brines and waste waters

    Premuzic, Eugene T.; Lin, Mow S.


    A method of solubilizing metal salts such as metal sulfides in a geothermal sludge using mutant Thiobacilli selected for their ability to metabolize metal salts at high temperature is disclosed, The method includes the introduction of mutated Thiobacillus ferrooxidans and Thiobacillus thiooxidans to a geothermal sludge or brine. The microorganisms catalyze the solubilization of metal salts, For instance, in the case of metal sulfides, the microorganisms catalyze the solubilization to form soluble metal sulfates.

  2. Fatty acid solubilizer from the oral disk of the blowfly.

    Yuko Ishida

    Full Text Available BACKGROUND: Blowflies are economic pests of the wool industry and potential vectors for epidemics. The establishment of a pesticide-free, environmentally friendly blowfly control strategy is necessary. Blowflies must feed on meat in order to initiate the cascade of events that are involved in reproduction including juvenile hormone synthesis, vitellogenesis, and mating. During feeding blowflies regurgitate salivary lipase, which may play a role in releasing fatty acids from triglycerides that are found in food. However, long-chain fatty acids show low solubility in aqueous solutions. In order to solubilize and ingest the released hydrophobic fatty acids, the blowflies must use a solubilizer. METHODOLOGY: We applied native PAGE, Edman degradation, cDNA cloning, and RT-PCR to characterize a protein that accumulated in the oral disk of the black blowfly, Phormia regina. In situ hybridization was carried out to localize the expression at the cellular level. A fluorescence competitive binding assay was used to identify potential ligands of this protein. CONCLUSION: A protein newly identified from P. regina (PregOBP56a belonged to the classic odorant-binding protein (OBP family. This gene was expressed in a cluster of cells that was localized between pseudotracheae on the oral disk, which are not accessory cells of the taste peg chemosensory sensilla that normally synthesize OBPs. At pH 7 and pH 6, PregOBP56a bound palmitic, stearic, oleic, and linoleic acids, that are mainly found in chicken meat. The binding affinity of PregOBP56a decreased at pH 5. We propose that PregOBP56a is a protein that solubilizes fatty acids during feeding and subsequently helps to deliver the fatty acids to the midgut where it may help in the process of reproduction. As such, PregOBP56a is a potential molecular target for controlling the blowfly.

  3. Molecular underpinnings of synaptic vesicle pool heterogeneity.

    Crawford, Devon C; Kavalali, Ege T


    Neuronal communication relies on chemical synaptic transmission for information transfer and processing. Chemical neurotransmission is initiated by synaptic vesicle fusion with the presynaptic active zone resulting in release of neurotransmitters. Classical models have assumed that all synaptic vesicles within a synapse have the same potential to fuse under different functional contexts. In this model, functional differences among synaptic vesicle populations are ascribed to their spatial distribution in the synapse with respect to the active zone. Emerging evidence suggests, however, that synaptic vesicles are not a homogenous population of organelles, and they possess intrinsic molecular differences and differential interaction partners. Recent studies have reported a diverse array of synaptic molecules that selectively regulate synaptic vesicles' ability to fuse synchronously and asynchronously in response to action potentials or spontaneously irrespective of action potentials. Here we discuss these molecular mediators of vesicle pool heterogeneity that are found on the synaptic vesicle membrane, on the presynaptic plasma membrane, or within the cytosol and consider some of the functional consequences of this diversity. This emerging molecular framework presents novel avenues to probe synaptic function and uncover how synaptic vesicle pools impact neuronal signaling.

  4. Amyloglucosidase enzymatic reactivity inside lipid vesicles

    Kim Jin-Woo


    Full Text Available Abstract Efficient functioning of enzymes inside liposomes would open new avenues for applications in biocatalysis and bioanalytical tools. In this study, the entrapment of amyloglucosidase (AMG (EC from Aspergillus niger into dipalmitoylphosphatidylcholine (DPPC multilamellar vesicles (MLVs and large unilamellar vesicles (LUVs was investigated. Negative-stain, freeze-fracture, and cryo-transmission electron microscopy images verified vesicle formation in the presence of AMG. Vesicles with entrapped AMG were isolated from the solution by centrifugation, and vesicle lamellarity was identified using fluorescence laser confocal microscopy. The kinetics of starch hydrolysis by AMG was modeled for two different systems, free enzyme in aqueous solution and entrapped enzyme within vesicles in aqueous suspension. For the free enzyme system, intrinsic kinetics were described by a Michaelis-Menten kinetic model with product inhibition. The kinetic constants, Vmax and Km, were determined by initial velocity measurements, and Ki was obtained by fitting the model to experimental data of glucose concentration-time curves. Predicted concentration-time curves using these kinetic constants were in good agreement with experimental measurements. In the case of the vesicles, the time-dependence of product (glucose formation was experimentally determined and simulated by considering the kinetic behavior of the enzyme and the permeation of substrate into the vesicle. Experimental results demonstrated that entrapped enzymes were much more stable than free enyzme. The entrapped enzyme could be recycled with retention of 60% activity after 3 cycles. These methodologies can be useful in evaluating other liposomal catalysis operations.

  5. Leiomyoma of the seminal vesicles: laparoscopic excision.

    Casado Varela, Javier; Hermida Gutiérrez, Juan Francisco; Castillón Vela, Ignacio T; León Rueda, Maria Eugenia; Ortega Medina, Luis; Moreno Sierra, Jesús


    Leiomyoma of the seminal vesicles is an extremely rare type of benign tumor of the genitourinary system and can cause lower urinary tract symptoms. Despite their low incidence, these tumors can be identified with transrectal ultrasound of the seminal vesicles during prostate examination. The removal of these tumors is facilitated by a laparoscopic approach.

  6. Functional Advantages of Porphyromonas gingivalis Vesicles.

    Meng-Hsuan Ho

    Full Text Available Porphyromonas gingivalis is a keystone pathogen of periodontitis. Outer membrane vesicles (OMVs have been considered as both offense and defense components of this bacterium. Previous studies indicated that like their originating cells, P. gingivalis vesicles, are able to invade oral epithelial cells and gingival fibroblasts, in order to promote aggregation of some specific oral bacteria and to induce host immune responses. In the present study, we investigated the invasive efficiency of P. gingivalis OMVs and compared results with that of the originating cells. Results revealed that 70-90% of human primary oral epithelial cells, gingival fibroblasts, and human umbilical vein endothelial cells carried vesicles from P. gingivalis 33277 after being exposed to the vesicles for 1 h, while 20-50% of the host cells had internalized P. gingivalis cells. We also detected vesicle-associated DNA and RNA and a vesicle-mediated horizontal gene transfer in P. gingivalis strains, which represents a novel mechanism for gene transfer between P. gingivalis strains. Moreover, purified vesicles of P. gingivalis appear to have a negative impact on biofilm formation and the maintenance of Streptococcus gordonii. Our results suggest that vesicles are likely the best offence weapon of P. gingivalis for bacterial survival in the oral cavity and for induction of periodontitis.

  7. Bio-solubilization of Chinese lignite Ⅱ: protein adsorption onto the lignite surface

    YIN Su-dong; TAO Xiu-xiang; SHI Kai-yi


    Lignite bio-solubilization is a promising technology for converting solid lignite into oil. This study concerns the adsorption of lignite-solubilizing enzymes onto the lignite surface. Adsorption capacity, infrared spectral analysis and driving forces analysis are studied as a way to help understand the bio-solubilization mechanism. The results show that the amount of lignite bio-solubilization is proportional to the amount of adsorbed lignite-solubilizing enzymes. An increase in lignite-solubilizing enzyme adsorption of 10% leads to a 7% increase in lignite bio-solubilization. However, limited amounts of enzymes can be adsorbed by the lignite, thus resulting in low percentages of bio-solubilization. Infrared spectral analysis shows that side chains, such as hydroxyl and carbonyl, of the lignite structure are the main, and necessary, structures where lignite-solubilizing enzymes attachto the lignite. Furthermore, driving force analysis indicates that the electrostatic force between lignite and enzymes is the main adsorption mechanism. The forces are influenced by solution pH levels, the zeta potential of the lignite and the isoelectric points of the enzymes.

  8. A standardized method for quantitating the complement-mediated immune complex solubilizing capacity of human serum

    Baatrup, G; Peterson, I; Svehag, S E;


    A standardized radioassay for measuring the complement-mediated immune complex solubilizing capacity (CMSC) and the initial kinetics of the solubilization (IKS) reaction is described. The total complement (C)-mediated solubilizing capacity was determined after incubation of diluted serum and 125I......-BSA-anti-BSA. Percentage C-mediated solubilization (CMS) was measured after centrifugation by determining the distribution of radioactivity. The dependency of CMSC upon factors such as serum dilution and buffer system used, amount of IC added to serum, serum storage conditions and centrifugation conditions...

  9. Complement-mediated solubilization of immune complexes and their interaction with complement C3 receptors

    Petersen, Ivan; Baatrup, Gunnar; Jepsen, H H;


    Some of the molecular events in the complement (C)-mediated solubilization of immune complexes (IC) have been clarified in recent years. The solubilization is primarily mediated by alternative C pathway proteins whereas factors in the classical pathway accelerate the process. Components of the me......Some of the molecular events in the complement (C)-mediated solubilization of immune complexes (IC) have been clarified in recent years. The solubilization is primarily mediated by alternative C pathway proteins whereas factors in the classical pathway accelerate the process. Components...

  10. Effect of acute thioacetamide administration on rat brain phospholipid metabolism

    Osada, J.; Aylagas, H.; Miro-Obradors, M.J.; Arce, C.; Palacios-Alaiz, E.; Cascales, M. (Tufs Univ., Boston, MA (USA))


    Brain phospholipid composition and the ({sup 32}P)orthophosphate incorporation into brain phospholipids of control and rats treated for 3 days with thioacetamide were studied. Brain phospholipid content, phosphatidylcholine, phosphatidylethanolamine, lysolecithin and phosphatidic acid did not show any significant change by the effect of thioacetamide. In contrast, thioacetamide induced a significant decrease in the levels of phosphatidylserine, sphingomyelin, phosphatidylinositol and diphosphatidylglycerol. After 75 minutes of intraperitoneal label injection, specific radioactivity of all the above phospholipids with the exception of phosphatidylethanolamine and phosphatidylcholine significantly increased. After 13 hours of isotope administration the specific radioactivity of almost all studied phospholipid classes was elevated, except for phosphatidic acid, the specific radioactivity of which did not change and for diphosphatidylglycerol which showed a decrease in specific radioactivity. These results suggest that under thioacetamide treatment brain phospholipids undergo metabolic transformations that may contribute to the hepatic encephalopathy induced by thioacetamide.

  11. A Vesicle Superpool Spans Multiple Presynaptic Terminals in Hippocampal Neurons

    Staras, K.; Branco, T.; Burden, J. J.; Pozo, K.; Darcy, K.; Marra, V; Ratnayaka, A.; Goda, Y


    Synapse-specific vesicle pools have been widely characterized at central terminals. Here, we demonstrate a vesicle pool that is not confined to a synapse but spans multiple terminals. Using fluorescence imaging, correlative electron microscopy, and modeling of vesicle dynamics, we show that some recycling pool vesicles at synapses form part of a larger vesicle "superpool." The vesicles within this superpool are highly mobile and are rapidly exchanged between terminals (turnover: similar to 4%...

  12. [Transvesical Removal of Seminal Vesicle Cystadenoma].

    Takayasu, Kenta; Harada, Jiro; Kawa, Gen; Ota, Syuichi; Sakurai, Takanori


    Primary tumors of the seminal vesicles are extremely rare. There have been 25 reports of this tumor from overseas and most cases are cystadenoma. We report a case of seminal vesicle cystadenoma in a 70-year-old man who presented with lower abdominal pain and urinary frequency. A digital rectal examination detected a projecting and hard mass in the right side of the prostate. Magnetic resonance imaging (MRI) showed a 15 cm multiple cystic mass continuous with the right seminal vesicle. A transrectal needle biopsy revealed benign tissue. The tumor was resected using an open transvesical approach that enabled full exposure of the seminal vesicle without damaging the nerves and blood supply of the bladder. Pathology was consistent with a benign seminal vesicle cystadenoma. We describe the natural history, pathology,and surgical approach in this case.

  13. Alternative methods for characterization of extracellular vesicles

    Fatemeh eMomen-Heravi


    Full Text Available Extracellular vesicles are nano-sized vesicles released by all cells in vitro as well as in vivo. Their role has been implicated mainly in cell-cell communication, but also in disease biomarkers and more recently in gene delivery. They represent a snapshot of the cell status at the moment of release and carry bioreactive macromolecules such as nucleic acids, proteins and lipids. A major limitation in this emerging new field is the availability/awareness of techniques to isolate and properly characterize Extracellular vesicles. The lack of gold standards makes comparing different studies very difficult and may potentially hinder some Extracellular vesicles -specific evidence. Characterization of Extracellular vesicles has also recently seen many advances with the use of Nanoparticle Tracking Analysis (NTA, flow cytometry, cryo-EM instruments and proteomic technologies. In this review, we discuss the latest developments in translational technologies involving characterization methods including the facts in their support and the challenges they face.

  14. Transport vesicle formation in plant cells.

    Hwang, Inhwan; Robinson, David G


    In protein trafficking, transport vesicles bud from donor compartments and carry cargo proteins to target compartments with which they fuse. Thus, vesicle formation is an essential step in protein trafficking. As for mammals, plant cells contain the three major types of vesicles: COPI, COPII, and CCV and the major molecular players in vesicle-mediated protein transport are also present. However, plant cells generally contain more isoforms of the coat proteins, ARF GTPases and their regulatory proteins, as well as SNAREs. In addition, plants have established some unique subfamilies, which may reflect plant cell-specific conditions such as the absence of an ER-Golgi intermediate compartment and the combined activities of the TGN and early endosome. Thus, even though we are still at an early stage in understanding the physiological function of these proteins, it is already clear that vesicle-mediated protein transport in plant cells displays both similarities as well as differences in animal cells.

  15. Phospholipid Bicelles Improve the Conformational Stability of Rhodopsin Mutants Associated with Retinitis Pigmentosa.

    Dong, Xiaoyun; Ramon, Eva; Herrera-Hernández, María Guadalupe; Garriga, Pere


    Mutations in the visual photoreceptor rhodopsin are the cause of the retinal degenerative disease retinitis pigmentosa. Some naturally occurring mutations can lead to protein conformational instability. Two such mutations, N55K and G90V, in the first and second transmembrane helices of the receptor, have been associated with sector and classical retinitis pigmentosa, respectively, and showed enhanced thermal sensitivity. We have carefully analyzed the effect of phospholipid bicelles on the stability and ligand binding properties of these two mutants and compared it with those of the detergent-solubilized samples. We have used a phospholipid bilayer consisting of 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) and 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC). We find that DMPC/DHPC bicelles dramatically increase the thermal stability of the rhodopsin mutants G90V and N55K. The chromophore stability and regeneration of the mutants were also increased in bicelles when compared to their behavior in a dodecyl maltoside detergent solution. The retinal release process was slowed in bicelles, and chromophore entry, after illumination, was improved for the G90V mutant but not for N55K. Furthermore, fluorescence spectroscopy measurements showed that bicelles allowed more exogenous retinal binding to the photoactivated G90V mutant than in a detergent solution. In contrast, N55K could not reposition any chromophore either in the detergent or in bicelles. The results demonstrate that DMPC/DHPC bicelles can counteract the destabilizing effect of the disease-causing mutations and can modulate the structural changes in the ensuing receptor photoactivation in a distinct specific manner for different retinitis pigmentosa mutant phenotypes.

  16. Bending elasticity modulus of giant vesicles composed of aeropyrum pernix k1 archaeal lipid.

    Genova, Julia; Ulrih, Nataša Poklar; Kralj-Iglič, Veronika; Iglič, Aleš; Bivas, Isak


    Thermally induced shape fluctuations were used to study elastic properties of giant vesicles composed of archaeal lipids C25,25-archetidyl (glucosyl) inositol and C25,25-archetidylinositol isolated from lyophilised Aeropyrum pernix K1 cells. Giant vesicles were created by electroformation in pure water environment. Stroboscopic illumination using a xenon flash lamp was implemented to remove the blur effect due to the finite integration time of the camera and to obtain an instant picture of the fluctuating vesicle shape. The mean weighted value of the bending elasticity modulus kc of the archaeal membrane determined from the measurements meeting the entire set of qualification criteria was (1.89 ± 0.18) × 10-19 J, which is similar to the values obtained for a membrane composed of the eukaryotic phospholipids SOPC (1.88 ± 0.17) × 10-19 J and POPC (2.00 ± 0.21) ´ 10-19 J. We conclude that membranes composed of archaeal lipids isolated from Aeropyrum pernix K1 cells have similar elastic properties as membranes composed of eukaryotic lipids. This fact, together with the importance of the elastic properties for the normal circulation through blood system, provides further evidence in favor of expectations that archaeal lipids could be appropriate for the design of drug delivery systems.

  17. Mass-spectrometry-based molecular characterization of extracellular vesicles: lipidomics and proteomics.

    Kreimer, Simion; Belov, Arseniy M; Ghiran, Ionita; Murthy, Shashi K; Frank, David A; Ivanov, Alexander R


    This review discusses extracellular vesicles (EVs), which are submicron-scale, anuclear, phospholipid bilayer membrane enclosed vesicles that contain lipids, metabolites, proteins, and RNA (micro and messenger). They are shed from many, if not all, cell types and are present in biological fluids and conditioned cell culture media. The term EV, as coined by the International Society of Extracellular Vesicles (ISEV), encompasses exosomes (30-100 nm in diameter), microparticles (100-1000 nm), apoptotic blebs, and other EV subsets. EVs have been implicated in cell-cell communication, coagulation, inflammation, immune response modulation, and disease progression. Multiple studies report that EV secretion from disease-affected cells contributes to disease progression, e.g., tumor niche formation and cancer metastasis. EVs are attractive sources of biomarkers due to their biological relevance and relatively noninvasive accessibility from a range of physiological fluids. This review is focused on the molecular profiling of the protein and lipid constituents of EVs, with emphasis on mass-spectrometry-based "omic" analytical techniques. The challenges in the purification and molecular characterization of EVs, including contamination of isolates and limitations in sample quantities, are discussed along with possible solutions. Finally, the review discusses the limited but growing investigation of post-translational modifications of EV proteins and potential strategies for future in-depth molecular characterization of EVs.

  18. Huntingtin interactions with membrane phospholipids: strategic targets for therapeutic intervention?

    Kegel-Gleason, Kimberly B


    The Huntington's disease gene encodes the protein huntingtin (Htt), a soluble protein that largely distributes to the cytoplasm where about half the protein is found in association with membranes. Early studies on Huntington's disease patients suggested changes in membrane phospholipids. Furthermore, changes in phospholipid biosynthetic enzymes have been found in HD cell models using genetic methods. Recent investigations prove that Htt associates with membranes by direct interactions with phospholipids in membranes. Htt contains at least two membrane binding domains, which may work in concert with each other, to target to the appropriate intracellular membranes for diverse functions. Htt has a particular affinity for a specific class of phospholipids called phosphatidylinositol phosphates; individual species of these phospholipids propagate signals promoting cell survival and regulating changes in morphology. Mutant Htt fragments can disrupt synthetic phospholipid bilayers and full-length mutant Htt shows increased binding to numerous phospholipids, supporting the idea that mutant Htt can introduce pathology at the level of phospholipid interactions. There is a great potential to develop therapeutic agents since numerous enzymes regulate the both the biosynthesis/metabolism of lipids and the post-translational modifications of Htt that direct membrane interactions. Understanding the relationship of Htt with membrane phospholipids, and the impact of mutant Htt on membrane-related functions and lipid metabolism, may help identify new modes of therapeutic intervention for Huntington's disease.

  19. Lung protection by inhalation of exogenous solubilized extracellular matrix

    Wu, Jinglei; Ravikumar, Priya; Nguyen, Kytai T.; Hsia, Connie C. W.


    Decellularized extracellular matrix (ECM) contains complex tissue-specific components that work in concert to promote tissue repair and constructive remodeling and has been used experimentally and clinically to accelerate epithelial wound repair, leading us to hypothesize that lung-derived ECM could mitigate acute lung injury. To explore the therapeutic potential of ECM for noninvasive delivery to the lung, we decellularized and solubilized porcine lung ECM, then characterized the composition, concentration, particle size and stability of the preparation. The ECM preparation at 3.2 mg/mL with average particle size <3 μm was tested in vitro on human A549 lung epithelial cells exposed to 95% O2 for 24 hours, and in vivo by tracheal instillation or nebulization into the lungs of rats exposed intermittently or continuously to 90% O2 for a cumulative 72 hours. Our results showed that the preparation was enriched in collagen, reduced in glycosaminoglycans, and contained various bioactive molecules. Particle size was concentration-dependent. Compared to the respective controls treated with cell culture medium in vitro or saline in vivo, ECM inhalation normalized cell survival and alveolar morphology, and reduced hyperoxia-induced apoptosis and oxidative damage. This proof-of-concept study established the methodology, feasibility and therapeutic potential of exogenous solubilized ECM for pulmonary cytoprotection, possibly as an adjunct or potentiator of conventional therapy. PMID:28151947

  20. Studies on the solubilization of German coal by fungi

    Reiss, J. (Grahamhaus Stadt kg, Bad Kreuznach (Germany))


    The capability of seven basidiomycetes (Trametes versicolor, Poria placenta, Pleurotus florida, P. ostreatus, P. sajor-caju, P. eryngii, Stropharia sp.), one ascomycete (Chaetomium globosum) and five hyphomycetes and moulds (Humicola grisea, Trichoderma viride, Aspergillus terreus, Paecilomyces varioti, Papulaspora immersa) to solubilize medium and high volatile bituminous coals (types A and B) as well as four types of lignite B from Germany was tested in surface cultures. The intensity of bioliquefaction was determined by estimating the rate of droplet formation and by measuring the loss of weight of the coal granules gravimetrically. The bituminous coals with a relative high degree of coalification were only moderately converted by Trametes versicolor, Pleurotus florida, P. ostreatus and P. sajor-caju. The three species of Pleurotus caused the greatest rate of biosolubilization of lignite, yielding a loss of weight of the coal granules of more than 5.8% with a maximum of 7.6% with P. florida. The non-basidiomycetes proved to be less active with a liquefaction rate of up to 3.5% with Trichoderma viride. In general, the geologically younger lignite coals were more effectively solubilized than the older hard coals. The volatile matter and the oxygen content proved to be the principal factors influencing the intensity of bioconversion.

  1. Solubilization and electrophoretic characterization of select edible nut seed proteins.

    Sathe, Shridhar K; Venkatachalam, Mahesh; Sharma, Girdhari M; Kshirsagar, Harshal H; Teuber, Suzanne S; Roux, Kenneth H


    The solubility of almond, Brazil nut, cashew nut, hazelnut, macadamia, pecan, pine nut, pistachio, walnut, and peanut proteins in several aqueous solvents was qualitatively and quantitatively assessed. In addition, the effects of extraction time and ionic strength on protein solubility were also investigated. Electrophoresis and protein determination (Lowry, Bradford, and micro-Kjeldahl) methods were used for qualitative and quantitative assessment of proteins, respectively. Depending on the seed, buffer type and ionic strength significantly affected protein solubility. The results suggest that buffered sodium borate (BSB; 0.1 M H(3)BO(3), 0.025 M Na(2)B(4)O(7), 0.075 M NaCl, pH 8.45) optimally solubilizes nut seed proteins. Qualitative differences in seed protein electrophoretic profiles were revealed. For a specific seed type, these differences were dependent on the solvent(s) used to solubilize the seed proteins. SDS-PAGE results suggest the polypeptide molecular mass range for the tree nut seed proteins to be 3-100 kDa. The results of native IEF suggested that the proteins were mainly acidic, with a pI range from >4.5 to <7.0. Western immunoblotting experiments indicated that rabbit polyclonal antibodies recognized substantially the same polypeptides as those recognized by the corresponding pooled patient sera IgE.

  2. Maximizing mutagenesis with solubilized CRISPR-Cas9 ribonucleoprotein complexes.

    Burger, Alexa; Lindsay, Helen; Felker, Anastasia; Hess, Christopher; Anders, Carolin; Chiavacci, Elena; Zaugg, Jonas; Weber, Lukas M; Catena, Raul; Jinek, Martin; Robinson, Mark D; Mosimann, Christian


    CRISPR-Cas9 enables efficient sequence-specific mutagenesis for creating somatic or germline mutants of model organisms. Key constraints in vivo remain the expression and delivery of active Cas9-sgRNA ribonucleoprotein complexes (RNPs) with minimal toxicity, variable mutagenesis efficiencies depending on targeting sequence, and high mutation mosaicism. Here, we apply in vitro assembled, fluorescent Cas9-sgRNA RNPs in solubilizing salt solution to achieve maximal mutagenesis efficiency in zebrafish embryos. MiSeq-based sequence analysis of targeted loci in individual embryos using CrispRVariants, a customized software tool for mutagenesis quantification and visualization, reveals efficient bi-allelic mutagenesis that reaches saturation at several tested gene loci. Such virtually complete mutagenesis exposes loss-of-function phenotypes for candidate genes in somatic mutant embryos for subsequent generation of stable germline mutants. We further show that targeting of non-coding elements in gene regulatory regions using saturating mutagenesis uncovers functional control elements in transgenic reporters and endogenous genes in injected embryos. Our results establish that optimally solubilized, in vitro assembled fluorescent Cas9-sgRNA RNPs provide a reproducible reagent for direct and scalable loss-of-function studies and applications beyond zebrafish experiments that require maximal DNA cutting efficiency in vivo.

  3. Contribution of matrix vesicles and alkaline phosphatase to ectopic bone formation

    Ciancaglini P.


    Full Text Available Endochondral calcification involves the participation of matrix vesicles (MVs, but it remains unclear whether calcification ectopically induced by implants of demineralized bone matrix also proceeds via MVs. Ectopic bone formation was induced by implanting rat demineralized diaphyseal bone matrix into the dorsal subcutaneous tissue of Wistar rats and was examined histologically and biochemically. Budding of MVs from chondrocytes was observed to serve as nucleation sites for mineralization during induced ectopic osteogenesis, presenting a diameter with Gaussian distribution with a median of 306 ± 103 nm. While the role of tissue-nonspecific alkaline phosphatase (TNAP during mineralization involves hydrolysis of inorganic pyrophosphate (PPi, it is unclear how the microenvironment of MV may affect the ability of TNAP to hydrolyze the variety of substrates present at sites of mineralization. We show that the implants contain high levels of TNAP capable of hydrolyzing p-nitrophenylphosphate (pNPP, ATP and PPi. The catalytic properties of glycosyl phosphatidylinositol-anchored, polidocanol-solubilized and phosphatidylinositol-specific phospholipase C-released TNAP were compared using pNPP, ATP and PPi as substrates. While the enzymatic efficiency (k cat/Km remained comparable between polidocanol-solubilized and membrane-bound TNAP for all three substrates, the k cat/Km for the phosphatidylinositol-specific phospholipase C-solubilized enzyme increased approximately 108-, 56-, and 556-fold for pNPP, ATP and PPi, respectively, compared to the membrane-bound enzyme. Our data are consistent with the involvement of MVs during ectopic calcification and also suggest that the location of TNAP on the membrane of MVs may play a role in determining substrate selectivity in this micro-compartment.

  4. The oxidized phospholipid PazePC promotes permeabilization of mitochondrial membranes by Bax.

    Lidman, Martin; Pokorná, Šárka; Dingeldein, Artur P G; Sparrman, Tobias; Wallgren, Marcus; Šachl, Radek; Hof, Martin; Gröbner, Gerhard


    Mitochondria play a crucial role in programmed cell death via the intrinsic apoptotic pathway, which is tightly regulated by the B-cell CLL/lymphoma-2 (Bcl-2) protein family. Intracellular oxidative stress causes the translocation of Bax, a pro-apoptotic family member, to the mitochondrial outer membrane (MOM) where it induces membrane permeabilization. Oxidized phospholipids (OxPls) generated in the MOM during oxidative stress directly affect the onset and progression of mitochondria-mediated apoptosis. Here we use MOM-mimicking lipid vesicles doped with varying concentrations of 1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine (PazePC), an OxPl species known to significantly enhance Bax-membrane association, to investigate three key aspects of Bax's action at the MOM: 1) induction of Bax pores in membranes without additional mediator proteins, 2) existence of a threshold OxPl concentration required for Bax-membrane action and 3) mechanism by which PazePC disturbs membrane organization to facilitate Bax penetration. Fluorescence leakage studies revealed that Bax-induced leakage, especially its rate, increased with the vesicles' PazePC content without any detectable threshold neither for OxPl nor Bax. Moreover, the leakage rate correlated with the Bax to lipid ratio and the PazePC content. Solid state NMR studies and calorimetric experiments on the lipid vesicles confirmed that OxPl incorporation disrupted the membrane's organization, enabling Bax to penetrate into the membrane. In addition, 15N cross polarization (CP) and insensitive nuclei enhanced by polarization transfer (INEPT) MAS NMR experiments using uniformly (15)N-labeled Bax revealed dynamically restricted helical segments of Bax embedded in the membrane, while highly flexible protein segments were located outside or at the membrane surface. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Protein-phospholipid interactions in blood clotting.

    Morrissey, James H; Davis-Harrison, Rebecca L; Tavoosi, Narjes; Ke, Ke; Pureza, Vincent; Boettcher, John M; Clay, Mary C; Rienstra, Chad M; Ohkubo, Y Zenmei; Pogorelov, Taras V; Tajkhorshid, Emad


    Most steps of the blood clotting cascade require the assembly of a serine protease with its specific regulatory protein on a suitable phospholipid bilayer. Unfortunately, the molecular details of how blood clotting proteins bind to membrane surfaces remain poorly understood, owing to a dearth of techniques for studying protein-membrane interactions at high resolution. Our laboratories are tackling this question using a combination of approaches, including nanoscale membrane bilayers, solid-state NMR, and large-scale molecular dynamics simulations. These studies are now providing structural insights at atomic resolution into clotting protein-membrane interactions. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  6. Drug induced `softening' in phospholipid monolayers

    Basak, Uttam Kumar; Datta, Alokmay; Bhattacharya, Dhananjay


    Compressibility measurements on Langmuir monolayers of the phospholipid Dimystoryl Phospatidylcholine (DMPC) in pristine form and in the presence of the Non-steroidal Anti-inflammatory Drug (NSAID) Piroxicam at 0.025 drug/lipid (D/L) molecular ratio at different temperatures, show that the monolayer exhibits large increase (and subsequent decrease) in compressibility due to the drug in the vicinity of the Liquid Expanded - Liquid Condensed (LE-LC) phase transition. Molecular dynamics simulations of the lipid monolayer in presence of drug molecules show a disordering of the tail tilt, which is consistent with the above result.

  7. Motional Coherence in Fluid Phospholipid Membranes

    Rheinstadter, Maikel C; Flenner, Elijah J; Bruening, Beate; Seydel, Tilo; Kosztin, Ioan


    We report a high energy-resolution neutron backscattering study, combined with in-situ diffraction, to investigate slow molecular motions on nanosecond time scales in the fluid phase of phospholipid bilayers of 1,2-dimyristoyl-sn-glycero-3-phoshatidylcholine (DMPC) and DMPC/40% cholesterol (wt/wt). A cooperative structural relaxation process was observed. From the in-plane scattering vector dependence of the relaxation rates in hydrogenated and deuterated samples, combined with results from a 0.1 microsecond long all atom molecular dynamics simulation, it is concluded that correlated dynamics in lipid membranes occurs over several lipid distances, spanning a time interval from pico- to nanoseconds.

  8. Hyaluronan and phospholipid association in biolubrication

    Wang, Min; Liu, Chao; Thormann, Esben


    load bearing capacity. With DPPC as the last adsorbed component, a friction coefficient of 0.01 was found up to pressures significantly above what is encountered in healthy synovial joints. Hyaluronan as the last added component increases the friction coefficient to 0.03 and decreases the load bearing...... capacity somewhat (but still above what is needed in the synovial joint). Our data demonstrate that self-assembly structures formed by hyaluronan and phospholipids at interfaces are efficient aqueous lubricants, and it seems plausible that such self-assembly structures contribute to the exceptional...

  9. Nutritional influences on the solubilization of metal phosphate by ericoid mycorrhizal fungi.

    Gibson, Brian R; Mitchell, Derek T


    Four ericoid mycobionts (two isolates of Hymenoscyphus ericae, and two dark, sterile ericoid mycobionts isolated from metal-contaminated mine sites) were grown on solid agar plates supplemented with zinc phosphate (0.25 %) containing different forms of nitrogen (nitrate, ammonium or alanine) and different concentrations of carbon (glucose) and phosphorus (K2HPO4). The influence of nutrient variation on solubilizing ability of the fungi was assessed by measuring the zones of solubilization appearing beneath the growing colonies. All four mycobionts were capable of zinc phosphate solubilization in the presence of all three nitrogen sources and in media containing no nitrogen. No solubilization was observed at 0 mM glucose-C but was observed with increasing glucose concentration from 300 to 600 mM C. Increasing phosphorus concentration (0-5 mM P) had no effect on the solubilizing ability of the isolates. All but one of the mycobionts were capable of solubilizing calcium phosphate (CaHPO4), while no solubilization was observed in media containing aluminium phosphate (AlPO4), iron phosphate (FePO4 x 4H2O) or copper phosphate (Cu3O8P2 x 2H2O) under conditions which were found to be optimal for zinc phosphate solubilization. Under conditions of glucose at 300 mM C and alanine as the N source in the zinc phosphate-amended agar medium, one of the mycobionts produced new crystals, which were morphologically distinct from the original zinc phosphate crystals. It is concluded that medium composition influences the metal-phosphate solubilizing ability of ericoid mycobionts. The results are discussed in relation to the possible mechanisms involved in solubilization and the potential benefits of metal-phosphate solubilization to ericoid mycobionts and their host plants.

  10. On the Computing Potential of Intracellular Vesicles.

    Mayne, Richard; Adamatzky, Andrew


    Collision-based computing (CBC) is a form of unconventional computing in which travelling localisations represent data and conditional routing of signals determines the output state; collisions between localisations represent logical operations. We investigated patterns of Ca2+-containing vesicle distribution within a live organism, slime mould Physarum polycephalum, with confocal microscopy and observed them colliding regularly. Vesicles travel down cytoskeletal 'circuitry' and their collisions may result in reflection, fusion or annihilation. We demonstrate through experimental observations that naturally-occurring vesicle dynamics may be characterised as a computationally-universal set of Boolean logical operations and present a 'vesicle modification' of the archetypal CBC 'billiard ball model' of computation. We proceed to discuss the viability of intracellular vesicles as an unconventional computing substrate in which we delineate practical considerations for reliable vesicle 'programming' in both in vivo and in vitro vesicle computing architectures and present optimised designs for both single logical gates and combinatorial logic circuits based on cytoskeletal network conformations. The results presented here demonstrate the first characterisation of intracelluar phenomena as collision-based computing and hence the viability of biological substrates for computing.

  11. Vesicles Are Persistent Features of Different Plastids.

    Lindquist, Emelie; Solymosi, Katalin; Aronsson, Henrik


    Peripheral vesicles in plastids have been observed repeatedly, primarily in proplastids and developing chloroplasts, in which they are suggested to function in thylakoid biogenesis. Previous observations of vesicles in mature chloroplasts have mainly concerned low temperature pretreated plants occasionally treated with inhibitors blocking vesicle fusion. Here, we show that such vesicle-like structures occur not only in chloroplasts and proplastids, but also in etioplasts, etio-chloroplasts, leucoplasts, chromoplasts and even transforming desiccoplasts without any specific pretreatment. Observations are made both in C3 and C4 species, in different cell types (meristematic, epidermis, mesophyll, bundle sheath and secretory cells) and different organs (roots, stems, leaves, floral parts and fruits). Until recently not much focus has been given to the idea that vesicle transport in chloroplasts could be mediated by proteins, but recent data suggest that the vesicle system of chloroplasts has similarities with the cytosolic coat protein complex II system. All current data taken together support the idea of an ongoing, active and protein-mediated vesicle transport not only in chloroplasts but also in other plastids, obviously occurring regardless of chemical modifications, temperature and plastid developmental stage.

  12. On the Computing Potential of Intracellular Vesicles.

    Richard Mayne

    Full Text Available Collision-based computing (CBC is a form of unconventional computing in which travelling localisations represent data and conditional routing of signals determines the output state; collisions between localisations represent logical operations. We investigated patterns of Ca2+-containing vesicle distribution within a live organism, slime mould Physarum polycephalum, with confocal microscopy and observed them colliding regularly. Vesicles travel down cytoskeletal 'circuitry' and their collisions may result in reflection, fusion or annihilation. We demonstrate through experimental observations that naturally-occurring vesicle dynamics may be characterised as a computationally-universal set of Boolean logical operations and present a 'vesicle modification' of the archetypal CBC 'billiard ball model' of computation. We proceed to discuss the viability of intracellular vesicles as an unconventional computing substrate in which we delineate practical considerations for reliable vesicle 'programming' in both in vivo and in vitro vesicle computing architectures and present optimised designs for both single logical gates and combinatorial logic circuits based on cytoskeletal network conformations. The results presented here demonstrate the first characterisation of intracelluar phenomena as collision-based computing and hence the viability of biological substrates for computing.

  13. Behavior of inorganic elements during sludge ozonation and their effects on sludge solubilization.

    Sui, Pengzhe; Nishimura, Fumitake; Nagare, Hideaki; Hidaka, Taira; Nakagawa, Yuko; Tsuno, Hiroshi


    The behavior of inorganic elements (including phosphorus, nitrogen, and metals) during sludge ozonation was investigated using batch tests and the effects of metals on sludge solubilization were elucidated. A decrease of ∼ 50% in the ratio of sludge solubilization was found to relate to a high iron content 80-120 mgFe/gSS than that of 4.7-7.4 mgFe/gSS. During sludge ozonation, the pH decreased from 7 to 5, which resulted in the dissolution of chemically precipitated metals and phosphorus. Based on experimental results and thermodynamic calculation, phosphate precipitated by iron and aluminum was more difficult to release while that by calcium released with decrease in pH. The release of barium, manganese, and chrome did not exceed 10% and was much lower than COD solubilization; however, that of nickel, copper, and zinc was similar to COD solubilization. The ratio of nitrogen solubilization was 1.2 times higher than that of COD solubilization (R(2)=0.85). Of the total nitrogen solubilized, 80% was organic nitrogen. Because of their high accumulation potential and negative effect on sludge solubilization, high levels of iron and aluminum in both sewage and sludge should be considered carefully for the application of the advanced sewage treatment process with sludge ozonation and phosphorus crystallization. Copyright © 2010 Elsevier Ltd. All rights reserved.

  14. Improving the aqueous solubility of triclosan by solubilization, complexation, and in situ salt formation.

    Grove, Christine; Liebenberg, Wilna; du Preez, Jan L; Yang, Wenzhan; de Villiers, Melgardt M


    Triclosan, an antimicrobial, although widely incorporated into many skin care products, toothpastes, and liquid soaps, presents formulation difficulties because it is practically insoluble in water. The objective of this study was to improve the aqueous solubility of triclosan through solubilization, complexation, and salt formation. The solubility of triclosan in distilled water and in phosphate buffers (pH 7.4) was determined at 30 degrees C. The order of solubilizing performance of the solubilizers was: N-methylglucamine> or =L-arginine>sodium lauryl sulfate>beta-cyclodextrin> or =hydroxypropyl-beta-cyclodextrin>ethanolamine>sodium benzoate>sodium methyl 4-hydroxybenzoate>triethanolamine> or =diethanolamine. These solubilizers increased the solubility of triclosan from 80- to 6000-fold. Micellar solubilization and the formation of either salts or complexes are postulated as possible mechanisms for the increase in the solubility of triclosan by the surfactant sodium lauryl sulphate, the cyclic sugar derivatives beta-cyclodextrin and 2-hydropropyl-beta-cyclodextrin, the amino acid L-arginine, and the amino sugar alcohol N-methylglucamine. Furthermore, although the bacteriostatic efficacy of triclosan was significantly increased when solubilized with N-methylglucamine, L-arginine, and ethanolamine, increased solubilization did not increase the effectiveness of triclosan for all solubilizers tested.

  15. Effect of an organophosphate pesticide, monocrotophos, on phosphate-solubilizing efficiency of soil fungal isolates.

    Jain, Rachna; Garg, Veena; Saxena, Jyoti


    Soil is a sink of pesticide residues as well as microorganisms. Fungi are well known for solubilization of inorganic phosphates, and this activity of fungal isolates may be affected by the presence of pesticide residues in the soil. In the present study, five generically different fungal isolates, viz. Aspergillus niger JQ660373, Aspergillus flavus, Penicillium aculeatum JQ660374, Fusarium pallidoroseum and Macrophomina sp., were tested and compared for their phosphate-solubilizing ability in the absence and presence of monocrotophos (500 mg L(-1)). After 168 h of incubation, four times high amount of tricalcium phosphate was solubilized by isolates in the growth medium containing monocrotophos in comparison to control (without monocrotophos). Concurrently, 78 % of the applied monocrotophos was degraded by these fungal isolates. Kinetics of phosphate solubilization shifted from logarithmic to power model in the presence of monocrotophos. Similarly, the phosphatase activity was also found significantly high in the presence of monocrotophos. The combined order of phosphate solubilization as well as monocrotophos degradation was found to be A. niger JQ660373 > P. aculeatum JQ660374 > A. flavus > F. pallidoroseum > Macrophomina sp. On the contrary, phosphate solubilization negatively correlated with the pH of the growth medium. Hence, it could be concluded that these fungal species efficiently solubilize inorganic phosphates and monocrotophos poses a positive effect on their ability and in turn degraded by them. To the best of our knowledge, this is the first report on P solubilization by Macrophomina sp. and F. pallidoroseum.

  16. Carbon and Nitrogen Sources Influence Tricalcium Phosphate Solubilization and Extracellular Phosphatase Activity by Talaromyces flavus.

    Stefanoni Rubio, P J; Godoy, M S; Della Mónica, I F; Pettinari, M J; Godeas, A M; Scervino, J M


    The aim of this work was to study phosphate (P) solubilization (and the processes involved in this event) by Talaromyces flavus (BAFC 3125) as a function of carbon and/or nitrogen sources. P solubilization was evaluated in NBRIP media supplemented with different carbon (glucose, sorbitol, sucrose, and fructose) and nitrogen (L-asparagine, urea, ammonium sulfate (AS), and ammonium nitrate (AN) combinations. The highest P solubilization was related to the highest organic acid production (especially gluconic acid) and pH drop for those treatments where glucose was present. Also P solubilization was higher when an inorganic nitrogen source was supplemented to the media when compared to an organic one. Although not being present an organic P source, phosphatase activity was observed. This shows that P mineralization and P solubilization can occur simultaneously, and that P mineralization is not induced by the enzyme substrate. The combination that showed highest P solubilization was for AN-glucose. The highest acid phosphatase activity was for AS-fructose, while for alkaline phosphatase were for AS-fructose and AN-fructose. Acid phosphatase activity was higher than alkaline. P solubilization and phosphatase activity (acid and alkaline) were influenced by the different carbon-nitrogen combinations. A better understanding of phosphate-solubilizing fungi could bring a better use of soil P.

  17. Biochar enhances Aspergillus niger rock phosphate solubilization by increasing organic acid production and alleviating fluoride toxicity.

    Mendes, Gilberto de Oliveira; Zafra, David Lopez; Vassilev, Nikolay Bojkov; Silva, Ivo Ribeiro; Ribeiro, José Ivo; Costa, Maurício Dutra


    During fungal rock phosphate (RP) solubilization, a significant quantity of fluoride (F(-)) is released together with phosphorus (P), strongly inhibiting the process. In the present study, the effect of two F(-) adsorbents [activated alumina (Al2O3) and biochar] on RP solubilization by Aspergillus niger was examined. Al2O3 adsorbed part of the F(-) released but also adsorbed soluble P, which makes it inappropriate for microbial RP solubilization systems. In contrast, biochar adsorbed only F(-) while enhancing phosphate solubilization 3-fold, leading to the accumulation of up to 160 mg of P per liter. By comparing the values of F(-) measured in solution at the end of incubation and those from a predictive model, it was estimated that up to 19 mg of F(-) per liter can be removed from solution by biochar when added at 3 g liter(-1) to the culture medium. Thus, biochar acted as an F(-) sink during RP solubilization and led to an F(-) concentration in solution that was less inhibitory to the process. In the presence of biochar, A. niger produced larger amounts of citric, gluconic, and oxalic acids, whether RP was present or not. Our results show that biochar enhances RP solubilization through two interrelated processes: partial removal of the released F(-) and increased organic acid production. Given the importance of organic acids for P solubilization and that most of the RPs contain high concentrations of F(-), the proposed solubilization system offers an important technological improvement for the microbial production of soluble P fertilizers from RP.

  18. Isolation of phosphate solubilizing bacteria and their potential for lead immobilization in soil.

    Park, Jin Hee; Bolan, Nanthi; Megharaj, Mallavarapu; Naidu, Ravi


    Lead (Pb), a highly toxic heavy metal forms stable compounds with phosphate (P). The potential of phosphate solubilizing bacteria (PSB) to immobilize Pb by enhancing solubilization of insoluble P compounds was tested in this research. Eighteen different PSB strains isolated from P amended and Pb contaminated soils were screened for their efficiency in P solubilization. The PSB isolated from P amended soils solubilized 217-479 mg/L of P while the PSB from Pb contaminated soil solubilized 31-293 mg/L of P. Stepwise multiple regression analysis and P solubility kinetics indicated that the major mechanism of P solubilization by PSB is the pH reduction through the release of organic acids. From the isolated bacteria, two PSB were chosen for Pb immobilization and these bacteria were identified as Pantoea sp. and Enterobacter sp., respectively. The PSB significantly increased P solubilization by 25.0% and 49.9% in the case of Pantoea sp., and 63.3% and 88.6% in the case of Enterobacter sp. for 200 and 800 mg/kg of rock phosphate (RP) addition, respectively, thereby enhancing the immobilization of Pb by 8.25-13.7% in the case of Pantoea sp. and 14.7-26.4% in the case of Enterobacter sp. The ability of PSB to solubilize P, promote plant growth, and immobilize Pb can be used for phytostabilization of Pb contaminated soils.

  19. Isolation and functional characterization of hydrocarbon emulsifying and solubilizing factors produced by a Pseudomonas species

    Reddy, P.G.; Banuah, J.N.; Bhagat, S.D.; Pathak, M.G.; Singh, H.D.


    Pseudomonas PG-1 cultivated on pristane produced in good amount a heat-stable polymeric substance which showed strong hydrocarbon emulsifying and solubilizing properties. The substance was isolated in crude form and was found to contain 34% protein, 16% carbohydrate, and 40% lipid. The hydrocarbon solubilizing activity of the isolate was strongly inhibited by EDTA but the chelating agent had no effect on the hydrocarbon emulsifying activity. Both activities of the isolate were strongly inhibited by chymotrypsin treatment indicating the importance of the protein moiety for its activity. Hydrocarbon solubilization by the isolate showed a certain degree of specificity to pristane in modest agitation generally used in microbial cultivation, but this specificity was lost by vigorous agitation in a Waring blender. It was proposed that in the first case, solubilization was effected by a solubilizing factor specific to pristane, whereas in the latter ease, nonspecific solubilization occurred due to the action of the emulsifying factor. The rate of pristane solubilization by heat-treated culture broth under the conditions of agitation used in cultivation (rotary shaker, 120 rpm) was found to be ca. 750 mg L/sup -1/h/sup -1/ which was much larger than the maximal pristane uptake rate of 170 mg L/sup -1/h/sup -1/ observed during microbial growth on the substrate. It was concluded that hydrocarbon solubilization could satisfactorily account for the substrate uptake and growth.

  20. Chelating agents improve enzymatic solubilization of pectinaceous co-processing streams

    Ravn, Helle Christine; Meyer, Anne S.


    /substrate] at 60 °C, pH 6.0 for 1 min. Characterization of the released fractions demonstrated a significantly improved effect of chelating agents for polysaccharide solubilization from FiberBind 400, PUF, and citrus peel, whereas only low amounts of polysaccharides were solubilized from the sugar beet pulp...

  1. Phospholipid imprinted polymers as selective endotoxin scavengers

    Sulc, Robert; Szekely, Gyorgy; Shinde, Sudhirkumar; Wierzbicka, Celina; Vilela, Filipe; Bauer, David; Sellergren, Börje


    Herein we explore phospholipid imprinting as a means to design receptors for complex glycolipids comprising the toxic lipopolysaccharide endotoxin. A series of polymerizable bis-imidazolium and urea hosts were evaluated as cationic and neutral hosts for phosphates and phosphonates, the latter used as mimics of the phospholipid head groups. The bis-imidazolium hosts interacted with the guests in a cooperative manner leading to the presence of tight and well defined 1:2 ternary complexes. Optimized monomer combinations were subsequently used for imprinting of phosphatidic acid as an endotoxin dummy template. Presence of the aforementioned ternary complexes during polymerization resulted in imprinting of lipid dimers - the latter believed to crudely mimic the endotoxin Lipid A motif. The polymers were characterized with respect to template rebinding, binding affinity, capacity and common structural properties, leading to the identification of polymers which were thereafter subjected to an industrially validated endotoxin removal test. Two of the polymers were capable of removing endotoxin down to levels well below the accepted threshold (0.005 EU/mg API) in pharmaceutical production.

  2. Low phospholipid-associated cholestasis and cholelithiasis.

    Erlinger, Serge


    Low phospholipid-associated cholestasis and cholelithiasis (LPAC) is a genetic disorder characterized by cholesterol gallbladder and intrahepatic stones. It is caused by a mutation of the gene ABCB4, which encodes the canalicular protein ABCB4/MDR3, a flippase that plays an essential role in the secretion of phosphatidylcholine into bile. Failure of this protein leads to secretion of bile that is poor in phospholipids and, hence, highly lithogenic, with potent detergent properties. This, in turn, leads to cholangiocyte luminal membrane injury and biliary lesions causing cholestasis. The diagnosis should be suspected when at least two of the following criteria are present: onset of symptoms before the age of 40 years; recurrence of biliary symptoms (biliary colic, jaundice, cholangitis, acute pancreatitis) after cholecystectomy; presence of echogenic foci within the liver indicative of intrahepatic stones or biliary sludge; previous episode(s) of intrahepatic cholestasis of pregnancy; and family history of gallstones in first-degree relatives. Intrahepatic stones can be demonstrated by ultrasonography with color Doppler examination, computed tomography and magnetic resonance imaging with magnetic resonance cholangiography, and the diagnosis confirmed by ABCB4 genotyping. Therapy with ursodeoxycholic acid offers prompt relief of symptoms and usually prevents complications. In some cases, however, surgery may be necessary. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  3. Phospholipid imprinted polymers as selective endotoxin scavengers

    Sulc, Robert; Szekely, Gyorgy; Shinde, Sudhirkumar; Wierzbicka, Celina; Vilela, Filipe; Bauer, David; Sellergren, Börje


    Herein we explore phospholipid imprinting as a means to design receptors for complex glycolipids comprising the toxic lipopolysaccharide endotoxin. A series of polymerizable bis-imidazolium and urea hosts were evaluated as cationic and neutral hosts for phosphates and phosphonates, the latter used as mimics of the phospholipid head groups. The bis-imidazolium hosts interacted with the guests in a cooperative manner leading to the presence of tight and well defined 1:2 ternary complexes. Optimized monomer combinations were subsequently used for imprinting of phosphatidic acid as an endotoxin dummy template. Presence of the aforementioned ternary complexes during polymerization resulted in imprinting of lipid dimers – the latter believed to crudely mimic the endotoxin Lipid A motif. The polymers were characterized with respect to template rebinding, binding affinity, capacity and common structural properties, leading to the identification of polymers which were thereafter subjected to an industrially validated endotoxin removal test. Two of the polymers were capable of removing endotoxin down to levels well below the accepted threshold (0.005 EU/mg API) in pharmaceutical production. PMID:28303896

  4. Phospholipids as inhibitors of amyloid fibril formation

    K. O. Vus


    Full Text Available Amyloid fibrils are the protein aggregates, whose formation is involved in the pathogenesis of Alzheimer’s disease, systemic amyloidosis, etc. Since there is no effective ways to treat these diseases, developing the new anti-amyloid drugs is of great importance. In this study a series of phospholipids have been tested for their ability to inhibit lysozyme and insulin amyloid fibril formation at acidic or neutral pH and elevated temperature.  The lag time, elongation rate and fibrillization extent were estimated using Thioflavin T fluorescence assay. It is found that the oxidized and charged phospholipids, included into the liposomes, were the most effective inhibitors of the protein fibrillization. By comparing the magnitude and direction of the lipid effect in different lipid-protein systems it was concluded that the reduction of the amyloid fibril formation is governed by hydrophobic and specific liposome-protein interactions. It is hypothesized that the presence of the surface formed by the lipid polar heads is critical for reducing the protein fibrillization extent.

  5. Shape fluctuations of large unilamellar lipid vesicles observed by laser light scattering: influence of the small-scale structure.

    Brocca, Paola; Cantù, Laura; Corti, Mario; Del Favero, Elena; Motta, Simona


    In the present paper, we apply the dynamic laser light scattering technique to investigate the dependence of the characteristic times of thermally induced shape fluctuation of large unilamellar vesicles (LUVs) on bilayer composition. After addressing single-component LUVs made of two common phospholipids, dimyristoylphosphatidylcholine (DMPC) and dipalmitoylphosphatidylcholine (DPPC), we investigate the changes in vesicle shape fluctuation times due to the presence of cholesterol and gangliosides (GM1), added in small amounts. The experimental results show that the addition of a second component, even in small amount, to DMPC vesicles induces a change in membrane fluctuation times. Moreover, in the case of ganglioside addition, also the disposition of GM1 within the bilayer is of importance. Quite unexpectedly, the symmetric or asymmetric disposition of GM1 has opposite effects on bilayer dynamics, the first resulting in a "hardening" and the second in a "softening" of the membrane. Those results support that the small-scale structure of the bilayer is important in determining the overall dynamics of the vesicle. They also suggest that the physiological disposition of GM1 in the outer leaflet of real cells has a significative result in mechanical terms, positively affecting the dynamics of the membrane.

  6. Solubilization of zinc compounds by the diazotrophic, plant growth promoting bacterium Gluconacetobacter diazotrophicus.

    Saravanan, V S; Madhaiyan, M; Thangaraju, M


    Gluconacetobacter diazotrophicus an endophytic diazotroph also encountered as rhizosphere bacterium is reported to possess different plant growth promoting characteristics. In this study, we assessed the zinc solubilizing potential of G. diazotrophicus under in vitro conditions with different Zn compounds using glucose or sucrose as carbon sources. G. diazotrophicus showed variations in their solubilization potential with the strains used and the Zn compounds tested. G. diazotrophicus PAl5 efficiently solubilized the Zn compounds tested and ZnO was effectively solubilized than ZnCO(3) or Zn(3)(PO(4))(2). The soluble Zn concentration was determined in the culture supernatant through Atomic Absorption Spectrophotometer. Gas chromatography coupled Mass Spectrometry analysis revealed 5-ketogluconic acid, a derivative of gluconic acid as the major organic acid produced by G. diazotrophicus PAl5 cultured with glucose as carbon source. This organic anion may be an important agent that helped in the solubilization of insoluble Zn compounds.

  7. Zinc metal solubilization by Gluconacetobacter diazotrophicus and induction of pleomorphic cells.

    Saravanan, Venkatakrishnan Sivaraj; Osborne, Jabez; Madhaiyan, Munusamy; Mathew, Lazar; Chung, Jongbae; Ahn, Kisup; Sa, Tongmin


    Gluconacetobacter diazotrophicus strain PAl5 exhibited a minimum inhibitory concentration value of 11 mM in an LGI medium amended with ZnCl2. When an LGI medium was amended with Zn metal, solubilization halos were observed in a plate assay, and further solubilization was confirmed in a broth assay. The maximum solubilization was recorded after 120 h with a 0.1% Zn metal amendment. During solubilization, the culture growth and pH of the broth were indirectly correlated. Using a Fourier Transform Infrared Spectroscopy analysis, one of the agents solubilizing the Zn metal was identified as gluconic acid. When the Zn-amended broth was observed under a bright field microscope, long involution cells were observed, and further analysis with Atomic Force Microscopy revealed highly deformed, pleomorphic, aggregate-like cells.

  8. Visualization of surfactant enhanced NAPL mobilization and solubilization in a two-dimensional micromodel



    Surfactant-enhanced aquifer remediation is an emerging technology for aquifers contaminated with nonaqueous phase liquids (NAPLs). A two-dimensional micromodel and image capture system were applied to observe NAPL mobilization and solubilization phenomena. In each experiment, a common residual NAPL field was established, followed by a series of mobilization and solubilization experiments. Mobilization floods included pure water floods with variable flow rates and surfactant floods with variations in surfactant formulations. At relatively low capillary numbers (N{sub ca}<10{sup {minus}3}), the surfactant mobilization floods resulted in higher NAPL saturations than for the pure water flood, for similar N{sub ca}.These differences in macroscopic saturations are explained by differences in micro-scale mobilization processes. Solubilization of the residual NAPL remaining after the mobilization stage was dominated by the formation of dissolution fingers, which produced nonequilibrium NAPL solubilization. A macroemulsion phase also as observed to form spontaneously and persist during the solubilization stage of the experiments.

  9. Co-assembly of chitosan and phospholipids into hybrid hydrogels

    Mendes, Ana Carina Loureiro; Shekarforoush, Elhamalsadat; Engwer, Christoph


    Novel hybrid hydrogels were formed by adding chitosan (Ch) to phospholipids (P) self-assembled particles in lactic acid. The effect of the phospholipid concentration on the hydrogel properties was investigated and was observed to affect the rate of hydrogel formation and viscoelastic properties...

  10. Depletion of acidic phospholipids influences chromosomal replication in Escherichia coli.

    Fingland, Nicholas; Flåtten, Ingvild; Downey, Christopher D; Fossum-Raunehaug, Solveig; Skarstad, Kirsten; Crooke, Elliott


    In Escherichia coli, coordinated activation and deactivation of DnaA allows for proper timing of the initiation of chromosomal synthesis at the origin of replication (oriC) and assures initiation occurs once per cell cycle. In vitro, acidic phospholipids reactivate DnaA, and in vivo depletion of acidic phospholipids, results in growth arrest. Growth can be restored by the expression of a mutant form of DnaA, DnaA(L366K), or by oriC-independent DNA synthesis, suggesting acidic phospholipids are required for DnaA- and oriC-dependent replication. We observe here that when acidic phospholipids were depleted, replication was inhibited with a concomitant reduction of chromosomal content and cell mass prior to growth arrest. This global shutdown of biosynthetic activity was independent of the stringent response. Restoration of acidic phospholipid synthesis resulted in a resumption of DNA replication prior to restored growth, indicating a possible cell-cycle-specific growth arrest had occurred with the earlier loss of acidic phospholipids. Flow cytometry, thymidine uptake, and quantitative polymerase chain reaction data suggest that a deficiency in acidic phospholipids prolonged the time required to replicate the chromosome. We also observed that regardless of the cellular content of acidic phospholipids, expression of mutant DnaA(L366K) altered the DNA content-to-cell mass ratio.

  11. Different oxidized phospholipid molecules unequally affect bilayer packing.

    Megli, Francesco M; Russo, Luciana


    The aim of this study was to gain more detailed knowledge about the effect of the presence of defined oxidized phospholipid molecules in phospholipid bilayers. After chromatographic and mass spectrometry analysis, the previously used product of the Fenton reaction with unsaturated lecithins proved to consist of a plethora of oxidatively modified lecithins, useless either for the detailed study of the effects brought about in the bilayer or as the source of defined oxidized phospholipid molecules. The latter, particularly 2-(omega-carboxyacyl)- and 2-(n-hydroperoxyacyl)-lecithins, can be more conveniently prepared by chemical or enzymatic synthesis rather than by chemical or physical oxidation. The effect of those molecules and of commercially available 12-hydroxy-stearic and dodecanedioic acid was studied in planar supported phospholipid bilayers (SPBs) by use of EPR spectrometry. The SPBs also contained 2-(5-doxylstearoyl)-lecithin as the spin probe, and the EPR spectral anisotropy loss, indicative of bilayer disordering, was measured as a function of the molar percentage of oxidized lipid. Most oxidized lipid molecules examined in this study were able to induce bilayer disordering, while hydroperoxyl group-bearing acyl chains appeared to be much less effective. It is concluded that the effects of different oxidized phospholipids on phospholipid bilayer structure cannot be generalized, as happens with batch-oxidized phospholipids, and that the use of defined oxidized phospholipid molecular species for membrane oxidative stress guarantees a more reliable and detailed response.

  12. Marine actinobacteria showing phosphate-solubilizing efficiency in Chorao Island, Goa, India.

    Dastager, Syed G; Damare, Samir


    The occurrence and distribution of an actinobacteria group of bacteria capable of dissolving insoluble phosphates were investigated in this study in marine environments, especially in sediments of Chorao Island, Goa Province, India. A total of 200 bacterial isolates of actinobacteria was isolated. All isolates were screened for phosphate-solubilizing activity on Pikovskaya's agar. Thirteen different isolates exhibiting maximum formation of halos (zone of solubilization) around the bacterial colonies were selected for quantitative estimations of P-solubilization. Quantitative estimations for P-solubilization were analyzed for up to 10 days at intervals of 24 h. Maximum solubilization from 89.3 ± 3.1 to 164.1 ± 4.1 μg ml(-1) was observed after 6 days of incubation in six of all isolates, while the isolate NII-1020 showed maximum P-solubilization. The increase in solubilization coincided with the drop in pH. Many of these species showed wide range of tolerance to temperature, pH, and salt concentrations. Further, 16S rRNA gene sequence analyses were carried to identify the bacterial groups which are actively solubilized phosphate in vitro. Gene sequencing results reveal that all isolates were clustered into six different actinobacterial genera: Streptomyces, Microbacterium, Angustibacter, Kocuria, Isoptericola, and Agromyces. The presence of phosphate-solubilizing microorganisms and their ability to solubilize phosphate were indicative of the important role played by bacteria in the biogeochemical cycle of phosphorus and the plant growth in coastal ecosystems.

  13. Solubilization of glycoproteins of envelope viruses by detergents

    Berezin, V.E.; Zaides, V.M.; Artamsnov, A.F.; Isaeva, E.S.; Zhdanov, V.M.


    The action of a number of known ionic and nonionic detergents, as well as the new nonionic detergent MESK, on envelope viruses was investigated. It was shown that the nonionic detergents MESK, Triton X-100, and octyl-..beta..-D-glucopyranoside selectively solubilize the outer glycoproteins of the virus particles. The nonionic detergent MESK has the mildest action. Using MESK, purified glycoproteins of influenza, parainfluenza, Venezuelan equine encephalomyelitis, vesicular stomatitis, rabies, and herpes viruses were obtained. The procedure for obtaining glycoproteins includes incubation of the virus suspension with the detergent MESK, removal of subvirus structures by centrifuging, and purification of glycoproteins from detergents by dialysis. Isolated glycoproteins retain a native structure and biological activity and possess high immunogenicity. The detergent MESK is promising for laboratory tests and with respect to the production of subunit vaccines.

  14. Phosphate solubilizing bacteria: Comparison between coastal and deep sea sediments

    Biche, S.; Pandey, S.; Gonsalves, M.J.B.D.; Das, A.; Mascarenhas-Pereira, M.B.L.; LokaBharathi, P.A.

    Phosphorus (P) is an essential nutrient for sustaining marine life (Karl, 2000). It is an important macromolecule and it plays vital role in the primary production in marine environment. In some organisms it can also be present as intracellular... organic compounds (Hoppe and Ullrich, 1999). Fig:3a Scatter plot of phosphate solubilizing activity v/s Pore water phosphate in the coast R² = 0.1423 0 0.01 0.02 0.03 0.04 0.05 0.06 0.00 0.50 1.00 1.50 Ph os ph at e uM PSA µg P ml-1 h-1 PSA v/s P...

  15. Kinetic regulation of coated vesicle secretion

    Foret, Lionel


    The secretion of vesicles for intracellular transport often rely on the aggregation of specialized membrane-bound proteins into a coat able to curve cell membranes. The nucleation and growth of a protein coat is a kinetic process that competes with the energy-consuming turnover of coat components between the membrane and the cytosol. We propose a generic kinetic description of coat assembly and the formation of coated vesicles, and discuss its implication to the dynamics of COP vesicles that traffic within the Golgi and with the Endoplasmic Reticulum. We show that stationary coats of fixed area emerge from the competition between coat growth and the recycling of coat components, in a fashion resembling the treadmilling of cytoskeletal filaments. We further show that the turnover of coat components allows for a highly sensitive switching mechanism between a quiescent and a vesicle producing membrane, upon a slowing down of the exchange kinetics. We claim that the existence of this switching behaviour, also tri...

  16. Physiopathologic dynamics of vesicle traffic in astrocytes.

    Potokar, Maja; Stenovec, Matjaž; Kreft, Marko; Gabrijel, Mateja; Zorec, Robert


    The view of how astrocytes, a type of glial cells, contribute to the functioning of the central nervous system (CNS) has changed greatly in the last decade. Although glial cells outnumber neurons in the mammalian brain, it was considered for over a century that they played a subservient role to neurons. This view changed. Functions thought to be exclusively present in neurons, i.e. excitability mediated release of chemical messengers, has also been demonstrated in astrocytes. In this process, following an increase in cytosolic calcium activity, membrane bound vesicles, storing chemical messengers (gliotransmitters), fuse with the plasma membrane, a process known as exocytosis, permitting the exit of vesicle cargo into the extracellular space. Vesicles are delivered to and are removed from the site of exocytosis by an amazingly complex set of processes that we have only started to learn about recently. In this paper we review vesicle traffic, which is subject to physiological regulation and may be changed under pathological conditions.

  17. Stability of Spherical Vesicles in Electric Fields


    The stability of spherical vesicles in alternating (ac) electric fields is studied theoretically for asymmetric conductivity conditions across their membranes. The vesicle deformation is obtained from a balance between the curvature elastic energies and the work done by the Maxwell stresses. The present theory describes and clarifies the mechanisms for the four types of morphological transitions observed experimentally on vesicles exposed to ac fields in the frequency range from 500 to 2 × 107 Hz. The displacement currents across the membranes redirect the electric fields toward the membrane normal to accumulate electric charges by the Maxwell−Wagner mechanism. These accumulated electric charges provide the underlying molecular mechanism for the morphological transitions of vesicles as observed on the micrometer scale. PMID:20575588

  18. Proteomics of Aggregatibacter actinomycetemcomitans Outer Membrane Vesicles

    Kieselbach, Thomas; Zijnge, Vincent; Granstrom, Elisabeth; Oscarsson, Jan


    Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen associated with aggressive forms of periodontitis and with endocarditis. Outer membrane vesicles (OMVs) released by this species have been demonstrated to deliver effector proteins such as cytolethal distending toxin (CDT) and

  19. Combination of surfactant solubilization with permanganate oxidation for DNAPL remediation.

    Li, Zhaohui; Hanlie, Hong


    A combination of surfactant solubilization with permanganate oxidation of trichloroethylene (TCE) was studied in batch, flow-through column, and three-dimensional (3-D) tank tests. Batch results showed that chloride production, an indication of TCE degradation, followed a pseudo-first-order reaction kinetics with respect to KMnO4 in the presence of free-phase TCE. A higher chloride production rate was achieved when anionic surfactants were present. The observed pseudo-first-order reaction rate constant increased as the concentrations of anionic surfactants Ninate 411 and Calfax increased from 0% to 0.1%, 0.3%, and 1.0%. Column experiments on TCE reduction by permanganate in the presence and absence of surfactants were carried out using well-sorted coarse Ottawa sand. The peak effluent TCE concentration reached 1700 mg/L due to enhanced solubilization when both sodium dodecyl sulfate (SDS) and permanganate were used, in contrast to less than 300 mg/L when only permanganate solution was used. In addition, the effluent TCE concentration decreased much faster when SDS was present in the permanganate solution, compared with the case when SDS was absent. With an initial 1 mL of TCE emplaced in the columns, the effluent TCE concentration dropped to breakthrough occurred after 21-25 h of injection when SDS was present compared with 45-70 h later when SDS was absent. A slightly higher chloride concentration was observed in the earlier stage of the column experiment and the chloride concentration decreased quickly once KMnO4 was seen in the effluent. The 3-D tank test showed that the MnO2 precipitation front formed more quickly when 1% SDS was present, which further confirmed the observation from the column study.

  20. Improved solubilization of curcumin with a microemulsification formulation


    Full Text Available Due to the large number of bioactive substances, with low and very low solubility in water, new and improved investigation methods were developed. Researches in this area have shown that lipid systems in lipophilic substances formulation increase their bioavailability and prevent or reduce the toxicological risk because most of the components involved in the formulation are of natural origin, with a structure compatible with biological membranes components. Among the lipid systems used in the leaching, transport and release of lipophilic substances there are: liposomes, solid lipid nanoparticles, double and single emulsions, autoemulsionante and auto-microemulsionante lipid systems. The last are the subject of the present research and meet specialists in concern for the harmonization of cost-benefit-risk in order to improve population health. Curcumin [(1E, 6E-1,7-bis(4-hydroxy-3-methoxyphenyl-1,6-heptadiene-3,5-dione] is a yellow pigment derived from the rhizome of the plant Curcuma Longa with phenol groups and conjugated double bounds which is unstable at light and basic pH, degrading within 30 minutes. The aim of this study is curcumin solubilization used as alimentary dye in automicroemulsionante systems. Dye/oil/surfactant/cosurfactant mixing ratio was made, based on quaternary phase diagrams. Mesofazice structures were revealed by conductivity and viscosimetric analysis. A curcumin solubilization system in aqueous medium was obtained. On the other hand, this paper studies the colour evolution of these automicroemulsionante systems comparing with hexane dye solution. The use of the chromatic attributes L*, a* and b* and L*, C* and hab, suggested by the Commission Internationale de l’Eclairage (CIE (i.e., the CIELAB system, obtained from direct transmitance measurements, which made it possible to follow the evolution of colour.

  1. Geochemical hosts of solubilized radionuclides in uranium mill tailings

    Landa, E.R.; Bush, C.A.


    The solubilization and subsequent resorption of radionuclides by ore components or by reaction products during the milling of uranium ores may have both economic and environmental consequences. Particle-size redistribution of radium during milling has been demonstrated by previous investigators; however, the identification of sorbing components in the tailings has received little experimental attention. In this study, uranium-bearing sandstone ore was milled, on a laboratory scale, with sulfuric acid. At regular intervals, filtrate from this suspension was placed in contact with mixtures of quartz sand and various potential sorbents which occur as gangue in uranium ores; the potential sorbents included clay minerals, iron and aluminum oxides, feldspar, fluorspar, barite, jarosite, coal, and volcanic glass. After equilibration, the quartz sand-sorbent mixtures were separated from the filtrate and radioassayed by gamma-spectrometry to determine the quantities of 238U, 230Th, 226Ra, and 210Pb sorbed, and the radon emanation coefficients. Sorption of 238U was low in all cases, with maximal sorptions of 1-2% by the bentonite- and coal-bearing samples. 230Th sorption also was generally less than 1%; maximal sorption here was observed in the fluorspar-bearing sample and appears to be associated with the formation of gypsum during milling. 226Ra and 210 Pb generally showed higher sorption than the other nuclides - more than 60% of the 26Ra solubilized from the ore was sorbed on the barite-bearing sample. The mechanism (s) for this sorption by a wide variety of substrates is not yet understood. Radon emanation coefficients of the samples ranged from about 5 to 30%, with the coal-bearing samples clearly demonstrating an emanating power higher than any of the other materials. ?? 1990.

  2. Tetraspanins in Extracellular Vesicle Formation and Function

    Andreu, Zoraida; Yáñez-Mó, María


    Extracellular vesicles (EVs) represent a novel mechanism of intercellular communication as vehicles for intercellular transfer of functional membrane and cytosolic proteins, lipids, and RNAs. Microvesicles, ectosomes, shedding vesicles, microparticles, and exosomes are the most common terms to refer to the different kinds of EVs based on their origin, composition, size, and density. Exosomes have an endosomal origin and are released by many different cell types, participating in different phy...

  3. Tetraspanins in Extracellular Vesicle formation and function

    Zoraida Andreu Martínez; María eYáñez-Mó


    Extracellular vesicles (EVs) represent a novel mechanism of intercellular communication as vehicles for intercellular transfer of functional membrane and cytosolic proteins, lipids, and RNAs. Microvesicles, ectosomes, shedding vesicles, microparticles and exosomes are the most common terms to refer to the different kinds of EVs based on their origin, composition, size and density. Exosomes have an endosomal origin and are released by many different cell types, participating in different physi...

  4. Oriented reconstitution of a membrane protein in a giant unilamellar vesicle: experimental verification with the potassium channel KcsA.

    Yanagisawa, Miho; Iwamoto, Masayuki; Kato, Ayako; Yoshikawa, Kenichi; Oiki, Shigetoshi


    We report a method for the successful reconstitution of the KcsA potassium channel with either an outside-out or inside-out orientation in giant unilamellar vesicles, using the droplet-transfer technique. The procedure is rather simple. First, we prepared water-in-oil droplets lined with a lipid monolayer. When solubilized KcsA was encapsulated in the droplet, it accumulated at monolayers of phosphatidylglycerol (PG) and phosphoethanolamine (PE) but not at a monolayer of phosphatidylcholine (PC). The droplet was then transferred through an oil/water interface having a preformed monolayer. The interface monolayer covered the droplet so as to generate a bilayer vesicle. By creating chemically different lipid monolayers at the droplet and oil/water interface, we obtained vesicles with asymmetric lipid compositions in the outer and inner leaflets. KcsA was spontaneously inserted into vesicles from the inside or outside, and this was accelerated in vesicles that contained PE or PG. Integrated insertion into the vesicle membrane and the KcsA orientation were examined by functional assay, exploiting the pH sensitivity of the opening of the KcsA when the pH-sensitive cytoplasmic domain (CPD) faces toward acidic media. KcsA loaded from the inside of the PG-containing vesicles becomes permeable only when the intravesicular pH is acidic, and the KcsA loaded from the outside becomes permeable when the extravesicular pH is acidic. Therefore, the internal or external insertion of KcsA leads to an outside-out or inside-out configuration so as to retain its hydrophilic CPD in the added aqueous side. The CPD-truncated KcsA exhibited a random orientation, supporting the idea that the CPD determines the orientation. Further application of the droplet-transfer method is promising for the reconstitution of other types of membrane proteins with a desired orientation into cell-sized vesicles with a targeted lipid composition of the outer and inner leaflets.

  5. Hierarchical unilamellar vesicles of controlled compositional heterogeneity.

    Maik Hadorn

    Full Text Available Eukaryotic life contains hierarchical vesicular architectures (i.e. organelles that are crucial for material production and trafficking, information storage and access, as well as energy production. In order to perform specific tasks, these compartments differ among each other in their membrane composition and their internal cargo and also differ from the cell membrane and the cytosol. Man-made structures that reproduce this nested architecture not only offer a deeper understanding of the functionalities and evolution of organelle-bearing eukaryotic life but also allow the engineering of novel biomimetic technologies. Here, we show the newly developed vesicle-in-water-in-oil emulsion transfer preparation technique to result in giant unilamellar vesicles internally compartmentalized by unilamellar vesicles of different membrane composition and internal cargo, i.e. hierarchical unilamellar vesicles of controlled compositional heterogeneity. The compartmentalized giant unilamellar vesicles were subsequently isolated by a separation step exploiting the heterogeneity of the membrane composition and the encapsulated cargo. Due to the controlled, efficient, and technically straightforward character of the new preparation technique, this study allows the hierarchical fabrication of compartmentalized giant unilamellar vesicles of controlled compositional heterogeneity and will ease the development of eukaryotic cell mimics that resemble their natural templates as well as the fabrication of novel multi-agent drug delivery systems for combination therapies and complex artificial microreactors.

  6. Sucrose induces vesicle accumulation and autophagy.

    Higuchi, Takahiro; Nishikawa, Jun; Inoue, Hiroko


    It has been shown that the treatment of mammalian cells with sucrose leads to vacuole accumulation associated with lysosomes and upregulation of lysosomal enzyme expression and activity. Autophagy is an evolutionarily conserved homeostatic process by which cells deliver cytoplasmic material for degradation into lysosomes, thus it is probable that sucrose affects the autophagic activity. The role of sucrose in autophagy is unknown; however, another disaccharide, trehalose has been shown to induce autophagy. In the current study, we used mouse embryonic fibroblasts to investigate whether sucrose induces autophagy and whether vesicle formation is associated with autophagy. The results showed that sucrose induces autophagy while being accumulated within the endosomes/lysosomes. These vesicles were swollen and packed within the cytoplasm. Furthermore, trehalose and the trisaccharide raffinose, which are not hydrolyzed in mammalian cells, increased the rate of vesicles accumulation and LC3-II level (a protein marker of autophagy). However, fructose and maltose did not show the same effects. The correlation between the two processes, vesicle accumulation and autophagy induction, was confirmed by treatment of cells with sucrose plus invertase, or maltose plus acarbose-the α-glucosidase inhibitor-and by sucrose deprivation. Results also showed that vesicle accumulation was not affected by autophagy inhibition. Therefore, the data suggest that sucrose-induced autophagy through accumulation of sucrose-containing vesicles is caused by the absence of hydrolysis enzymes.

  7. Investigation of nano lipid vesicles of methotrexate for anti-rheumatoid activity

    Prabhu P


    Full Text Available Prabhakara Prabhu1, Rakshith Shetty1, Marina Koland1, K Vijayanarayana3, KK Vijayalakshmi2, M Harish Nairy1, GS Nisha11Department of Pharmaceutics, Nitte University, NGSM Institute of Pharmaceutical Sciences, Paneer, Deralakatte, Mangalore, Karnataka, India; 2Department of Applied Zoology, Mangalore University, Konaje, Mangalore, Karnataka, India; 3Department of Pharmacy Practice, Manipal University, Manipal College of Pharmaceutical Sciences, Manipal, Karnataka, IndiaBackground: The purpose of this study was to formulate and evaluate nano lipid vesicles of methotrexate (MTX for its anti-rheumatoid activity.Methods: In this study the principle of both active as well as passive targeting using MTX-loaded stealth liposomes as per the magic gun approach was followed. Stealth liposomes of MTX were prepared by thin-film hydration method using a PEGylated phospholipid-like DSPE-MPEG 2000. Similarly, conventional liposomes were prepared using phospholipids like DPPC and DSPC. Conventional liposomes were coated with a hydrophilic biocompatible polymer like chitosan. They were investigated for their physical properties and in vitro release profile. Further, in vivo screening of the formulations for their anti-rheumatoid efficacy was carried out in rats. Rheumatoid arthritis was induced in male Wistar-Lewis rats using complete Freund’s adjuvant (1 mg/mL Mycobacterium tuberculosis, heat killed in mineral oil.Results: It was found that chitosan coating of the conventional liposomes increased the physical stability of the liposomal suspension as well as its entrapment efficiency. The size of the unsonicated lipid vesicles was found to be in the range of 8–10 µm, and the sonicated lipid vesicles in the range of 210–260 nm, with good polydispersity index. Further, chitosan-coated conventional liposomes and the PEGylated liposomes released the drug for a prolonged period of time, compared to the uncoated conventional liposomes. It was found that there

  8. The impact of phospholipids and phospholipid removal on bioanalytical method performance.

    Carmical, Jennifer; Brown, Stacy


    Phospholipids (PLs) are a component of cellmembranes, biological fluids and tissues. These compounds are problematic for the bioanalytical chemist, especially when PLs are not the analytes of interest. PL interference with bioanalysis highly impacts reverse-phase chromatographic methods coupled with mass spectrometric detection. Phospholipids are strongly retained on hydrophobic columns, and can cause significant ionization suppression in the mass spectrometer, as they outcompete analyte molecules for ionization. Strategies for improving analyte detection in the presence of PLs are reviewed, including in-analysis modifications and sample preparation strategies. Removal of interfering PLs prior to analysis seems to be most effective atmoderating thematrix effects fromthese endogenous cellular components, and has the potential to simplify chromatography and improve column lifetime. Products targeted at PL removal for sample pre-treatment, as well as products that combine multiplemodes of sample preparation (i.e. Hybrid SPE), show significant promise inmediating the effect on PL interference in bioanalysis.

  9. Packing of ganglioside-phospholipid monolayers

    Majewski, J.; Kuhl, T.L.; Kjær, K.


    DPPE monolayer and does not distort the hexagonal in-plane unit cell or out-of-plane two-dimensional (2-D) packing compared with a pure DPPE monolayer. The oligosaccharide headgroups were found to extend normally from the monolayer surface, and the incorporation of these glycolipids into DPPE...... monolayers did not affect hydrocarbon tail packing (fluidization or condensation of the hydrocarbon region). This is in contrast to previous investigations of lipopolymer-lipid mixtures, where the packing structure of phospholipid monolayers was greatly altered by the inclusion of lipids bearing hydrophilic...... polymer groups. Indeed, the lack of packing disruptions by the oligosaccharide groups indicates that protein-GM, interactions, including binding, insertion, chain fluidization, and domain formation (lipid rafts), can be studied in 2-D monolayers using scattering techniques....

  10. Lessons from the "Euro-Phospholipid" project.

    Cervera, Ricard


    The "Euro-Phospholipid" project started in 1999 with a multicentre, consecutive and prospective design. A total cohort of 1000 patients with antiphospholipid syndrome (APS), derived from 13 countries (Belgium, Bulgaria, Denmark, France, Germany, Greece, Hungary, Israel, Italy, the Netherlands, Portugal, Spain and United Kingdom), has been followed since then by a European consortium that was created as part of the network promoted by the "European Forum on Antiphospholipid Antibodies", a study group devoted to the development of multicentre projects with large populations of APS patients. This project allowed the identification of the prevalence and characteristics of the main clinical and immunological manifestations at the onset and during the evolution of APS and demonstrated that it is possible to recognize more homogeneous subsets of clinical significance.

  11. Food enrichment with marine phospholipid emulsions

    Lu, Henna Fung Sieng; Nielsen, Nina Skall; Baron, Caroline P.

    Many studies have shown that marine phospholipids (PL) provide more advantages than fish oil. They seem to have better bioavailability, better resistance towards oxidation and higher content of eicosapentaenoic acids and docosahexaenoic acids than fish oil, which essentially contains triglycerides...... marine PL emulsions with and without addition of fish oil. The oxidative stability of marine PL emulsions was significantly influenced by the chemical composition of marine PL used for emulsions preparation. For instance, emulsions with good oxidative stability could be obtained when using raw materials...... with high purity, low fish oil content and high PL, cholesterol and α-tocopherol content. In addition, non-enzymatic browning reactions may also affect the oxidative stability of the marine PL emulsion. These reactions included Strecker degradation and pyrrolization, and their occurrence were due...

  12. Structure and homogeneity of pseudo-physiological phospholipid bilayers and their deposition characteristics on carboxylic acid terminated self-assembled monolayers.

    Mechler, Adam; Praporski, Slavica; Piantavigna, Stefania; Heaton, Steven M; Hall, Kristopher N; Aguilar, Marie-Isabel; Martin, Lisandra L


    Supported phospholipid bilayers are frequently used to establish a pseudo-physiological environment required for the study of protein function or the design of enzyme-based biosensors and biocatalytic reactors. These membranes are deposited from bilayer vesicles (liposomes) that rupture and fuse into a planar membrane upon adhesion to a surface. However, the morphology and homogeneity of the resulting layer is affected by the characteristics of the precursor liposome suspension and the substrate. Here we show that two distinct liposome populations contribute to membrane formation--equilibrium liposomes and small unilamellar vesicles. Liposome deposition onto carboxylic acid terminated self-assembled monolayers resulted in planar mono- and multilayer, vesicular and composite membranes, as a function of liposome size and composition. Quartz crystal microbalance data provided estimates for layer thicknesses and sheer moduli and were used for classification of the final structure. Finally, atomic force microscopy data illustrated the inherently inhomogeneous and dynamic nature of these membranes.

  13. Conformation and Orientation of Phospholipid Molecule in Pure Phospholipid Monolayer During Compressing

    XUE Weilan; WANG Dan; ZENG Zuoxiang; GAO Xuechao


    On the basis of energy conservation law and surface pressure isotherm,the conformation energy changes of dipalmitoylphosphatidylcholine(DPPC)and dipalmitoylphosphatidylglycerol(DPPG)in pure phospholipid monolayer at the air/water interface during compression are derived.The optimized conformations of phospholipids at absolute freedom state are simulated by Gaussian 98 software.Based on following assumptions:(1)the conformation energy change is mainly caused by the rotation of one special bond;(2)the atoms of glycerol near the water surface are active;(3)the rotation is motivated by hydrogen-bond action;(4)the rotation of bond is inertial,one simplified track of conformational change is suggested and the conformations of DPPC and DPPG at different states are determined by the plots of conformation energy change vs.dihedral angle.The thickness of the simulated phospholipid monolayer is consistent with published experimental result.According to molecular areas at different states,the molecular orientations in the compressing process are also developed.

  14. Integrated Microfluidic System for Size-Based Selection and Trapping of Giant Vesicles.

    Kazayama, Yuki; Teshima, Tetsuhiko; Osaki, Toshihisa; Takeuchi, Shoji; Toyota, Taro


    Vesicles composed of phospholipids (liposomes) have attracted interest as artificial cell models and have been widely studied to explore lipid-lipid and lipid-protein interactions. However, the size dispersity of liposomes prepared by conventional methods was a major problem that inhibited their use in high-throughput analyses based on monodisperse liposomes. In this study, we developed an integrative microfluidic device that enables both the size-based selection and trapping of liposomes. This device consists of hydrodynamic selection and trapping channels in series, which made it possible to successfully produce an array of more than 60 monodisperse liposomes from a polydisperse liposome suspension with a narrow size distribution (the coefficient of variation was less than 12%). We successfully observed a size-dependent response of the liposomes to sequential osmotic stimuli, which had not clarified so far, by using this device. Our device will be a powerful tool to facilitate the statistical analysis of liposome dynamics.

  15. Preparation of Giant Vesicles Encapsulating Microspheres by Centrifugation of a Water-in-oil Emulsion.

    Natsume, Yuno; Wen, Hsin-I; Zhu, Tong; Itoh, Kazumi; Sheng, Li; Kurihara, Kensuke


    The constructive biology and the synthetic biology approach to creating artificial life involve the bottom-up assembly of biological or nonbiological materials. Such approaches have received considerable attention in research on the boundary between living and nonliving matter and have been used to construct artificial cells over the past two decades. In particular, Giant Vesicles (GVs) have often been used as artificial cell membranes. In this paper, we describe the preparation of GVs encapsulating highly packed microspheres as a model of cells containing highly condensed biomolecules. The GVs were prepared by means of a simple water-in-oil emulsion centrifugation method. Specifically, a homogenizer was used to emulsify an aqueous solution containing the materials to be encapsulated and an oil containing dissolved phospholipids, and the resulting emulsion was layered carefully on the surface of another aqueous solution. The layered system was then centrifuged to generate the GVs. This powerful method was used to encapsulate materials ranging from small molecules to microspheres.

  16. Solubilization and degradation of perchloroethylene (PCE) in cationic and nonionic surfactant solutions

    Sivaram Harendra; Cumaraswamy Vipulanandan


    Solubilization of perchloroethylene (PCE) in a nonionic (Triton X-100) and a cationic (cetyltrimethylammonium bromide (CTAB)) surfactant solutions and the degradation of surfactant solubilized PCE using fine to nanosize Fe and bi-metallic Fe-Ni particles were investigated.Micelle partition coefficients (Km) and molar solubility ratio (MSR) for PCE in 10 g/L of surfactant solutions have been quantified and the solubility of PCE (100 mg/L in water) in the surfactant solutions increased by about ten fold.Of the two surfactants studied,Triton X-100 solubilized the higher amount of PCE per gram of surfactant.To degrade solubilized PCE,both iron and bimetallic Fe-Ni particles were used in continuously stirred batch reactors.The iron and bi-metallic particles were synthesized using the solution method and the particles were characterized using the SEM,EDS,TEM and XRD.The PCE solubilized up to 500 mg/L in both surfactant solutions were totally degraded at various rates by 200 g/L of bi-metallic Fe-Ni particles in less than 20 hr,which is the highest concentration of PCE degraded in the shortest time compared to data in the literature.The degradations of PCE solubilized in surfactant solutions were represented by nonlinear kinetic relationships which depended on the type of surfactant used for solubilizing the PCE.

  17. Phosphate Solubilization Potential of Rhizosphere Fungi Isolated from Plants in Jimma Zone, Southwest Ethiopia

    Elias, Firew


    Phosphorus (P) is one of the major bioelements limiting agricultural production. Phosphate solubilizing fungi play a noteworthy role in increasing the bioavailability of soil phosphates for plants. The present study was aimed at isolating and characterizing phosphate solubilizing fungi from different rhizospheres using both solid and liquid Pikovskaya (PVK) medium. A total of 359 fungal isolates were obtained from 150 rhizosphere soil samples of haricot bean, faba bean, cabbage, tomato, and sugarcane. Among the isolates, 167 (46.52%) solubilized inorganic phosphate. The isolated phosphate solubilizing fungi belonged to genera of Aspergillus (55.69%), Penicillium spp. (23.35%), and Fusarium (9.58%). Solubilization index (SI) ranged from 1.10 to 3.05. Isolates designated as JUHbF95 (Aspergillus sp.) and JUFbF59 (Penicillium sp.) solubilized maximum amount of P 728.77 μg·mL−1 and 514.44 μg mL−1, respectively, from TCP (tricalcium phosphate) after 15 days of incubation. The highest (363 μg mL−1) soluble-P was released from RP with the inoculation of JUHbF95 in the PVK broth after 10 days of incubation. The present study indicated the presence of diverse plant associated P-solubilizing fungi that may serve as potential biofertilizers. PMID:27688771

  18. Rock inhabiting potassium solubilizing bacteria from Kerala, India: characterization and possibility in chemical K fertilizer substitution.

    Anjanadevi, Indira Parameswaran; John, Neetha Soma; John, Kuzhivilayil Susan; Jeeva, Muthulekshmi Lajapathy; Misra, Raj Shekhar


    The role of rock inhabiting bacteria in potassium (K) solubilization from feldspar and their application in crop nutrition through substitution of fertilizer K was explored through the isolation of 36 different bacteria from rocks of a major hill station at Ponmudi in Thiruvananthapuram, Kerala, India. A comprehensive characterization of K solubilization from feldspar was achieved with these isolates which indicated that the K solubilizing efficiency increases with decrease in pH and increase in viscosity and viable cell count. Based on the level of K solubilization, two potent isolates were selected and identified as Bacillus subtilis ANctcri3 and Bacillus megaterium ANctcri7. Exopolysaccharide production, scanning electron microscopic and fourier transform infrared spectroscopic studies with these efficient strains conclusively depicted the role of low pH, increase in viscosity, and bacterial attachment in K solubilization. They were also found to be efficient in phosphorus (P) solubilization, indole acetic acid production as well as tolerant to wide range of physiological conditions. Moreover, the applicability of K containing rock powder as a carrier for K solubilizing bacteria was demonstrated. A field level evaluation on the yield of a high K demanding tuberous vegetable crop, elephant foot yam (Amorphophallus paeoniifolius (dennst.) nicolson) established the possibility of substituting chemical K fertilizer with these biofertilizer candidates successfully.

  19. Solubilization of octane in cationic surfactant-anionic polymer complexes: effect of polymer concentration and temperature.

    Zhang, Hui; Deng, Lingli; Zeeb, Benjamin; Weiss, Jochen


    Polymers may alter the ability of oppositely charged surfactant micelles to solubilize hydrophobic molecules depending on surfactant-polymer interactions. This study was conducted to investigate the effects of polymer concentration and temperature on the solubilization thermodynamics of an octane oil-in-water emulsion in mixtures of an anionic polymer (carboxymethyl cellulose) and cationic cetyltrimethylammonium bromide (CTAB) surfactant micelles using isothermal titration calorimetry (ITC). Results showed that the CTAB binding capacity of carboxymethyl cellulose increased with increasing temperature from 301 to 323 K, and correspondingly the thermodynamic behavior of octane solubilization in CTAB micelles, either in the absence or presence of polymer, was found to depend on temperature. The addition of carboxymethyl cellulose caused the solubilization in CTAB micelles to be less endothermic, and increased the solubilization capacity. Based on the phase separation model, the solubilization was suggested to be mainly driven by enthalpy gains. Results suggest that increasing concentrations of the anionic polymer gave rise to a larger Gibbs energy decrease and a larger unfavorable entropy increase for octane solubilization in cationic surfactant micelles.

  20. Empigen BB: a useful detergent for solubilization and biochemical analysis of keratins.

    Lowthert, L A; Ku, N O; Liao, J; Coulombe, P A; Omary, M B


    Intermediate filament (IF) proteins make up some of the most insoluble proteins known, and within the IF protein family, keratins are the least soluble. We compared the efficiency of nonionic, cationic, mixed nonionic and anionic, and zwitterionic detergents in solubilizing keratins from insect cells that express recombinant human keratins and from human colonic cell lines and normal keratinocytes. The cationic detergent cetyltrimethylammonium bromide was similar to the zwitterionic detergent Empigen BB in its ability to efficiently solubilize keratins, but the latter detergent was superior in that it maintained antibody reactivity and allowed for immunoprecipitation of the keratins. Although Nonidet-P40 partially solubilizes keratins, Empigen BB solubilizes a significant amount of keratins not solubilized by Nonidet-P40. In the case of vimentin, differences in solubilization efficiency among the detergents was not as dramatic as with keratins. Our results show that Empigen BB solubilizes a significant amount of epidermal and glandular keratins while preserving antigenicity. This detergent should prove useful for carrying out biochemical and molecular studies on these proteins and may be similarly beneficial for other IF proteins.

  1. Effect of carbon substrates on rock phosphate solubilization by bacteria from composts and macrofauna.

    Hameeda, B; Reddy, Y Harish Kumar; Rupela, O P; Kumar, G N; Reddy, Gopal


    Five of the 207 isolates from different composts, farm waste compost (FWC), rice straw compost (RSC), Gliricidia vermicompost (GVC), and macrofauna, showed rock phosphate (RP) solubilization in buffered medium in plate culture. When tested in RP broth medium, all five strains, Enterobacter cloacae EB 27, Serratia marcescens EB 67, Serratia sp. EB 75, Pseudomonas sp. CDB 35, and Pseudomonas sp. BWB 21, showed gluconic acid production and solubilized RP. Based on cellulose-degrading and P-solubilizing ability, two strains were selected for further studies. In the presence of different carbon sources, both strains showed a drop in pH and solubilized RP. P released was maximum with glucose (1212 and 522 micromol) and minimum with cellobiose (455 and 306 micromol) by S. marcescens EB 67 and Pseudomonas sp. CDB 35, respectively. Glucose dehydrogenase (GDH) activity was 63 and 77% with galactose and 35 and 46% with cellobiose when compared to glucose (100%) by EB 67 and CDB 35, respectively. Both strains solubilized RP in the presence of different crop residues. EB 67 and CDB 35 showed maximum cellulase activity (0.027 units) in the presence of rice straw and a mixture of rice straw and root. P solubilized from RP in the presence of pigeonpea root was 134 and 140 micromol with EB 67 and CDB 35. Significantly, these bacteria isolated from composts and macrofauna solubilized rock phosphate in the presence of various pure carbon substrates and crop residues and their importance in soil/rhizosphere conditions is discussed.

  2. Physical characterization and antioxidant activity of thymol solubilized Tween 80 micelles

    Deng, Ling-Li; Taxipalati, Maierhaba; Que, Fei; Zhang, Hui


    Attempts were made to solubilize thymol in Tween 80 micelle to study the solubilization mechanism of thymol and the effect of solubilization on its antioxidant activity. The maximum solubilized concentration of thymol in a 2.0% (w/v) Tween 80 micelle solution is 0.2 wt%. There was no significant difference in Z-average diameter between the empty micelles and thymol solubilized micelles. 1H NMR spectra indicated that 3-H and 4-H on the benzene ring of thymol interacted with the ester group between the hydrophilic head group and the hydrophobic tail group of Tween 80 by Van der Waals’ force. Ferric reducing antioxidant potential (FRAP) and cupric ion reducing antioxidant capacity (CUPRAC) assays showed that the reducing antioxidant activity of free thymol did not change after solubilized in Tween 80 micelles. Compared to free thymol, the solubilized thymol showed higher activities to scavenge DPPH (2,2-diphenyl-1-picrylhydrazyl) and hydroxyl radicals. The present study suggested a possible preparation of thymol-carrying micelles with enhanced antioxidant activities that could be applied in food beverages.

  3. Mechanism of enhancement of prochymosin renaturation by solubilization of inclusion bodies at alkaline pH

    张治洲; 张渝英; 杨开宇


    The renaturation efficiency of recombinant prochymosin depends on not only the renaturation condi-tions but also the solubilization (denaturation) conditions. Compared with pH 8, solubilization of prochymosin-contain-ing inclusion bodies at pH 11 (8 mol/L urea) results in onefold increase of renaturation efficiency ( ~ 40% vs. ~ 20 % ). Alkaline pH facilitates the solubilization of inclusion bodies via the breakage of intermolecular disulfide bonds. Moreover, alkaline pH renders prochymosin molecules to be in a more reduced and more unfolded state which undergoes refolding readily.

  4. EDTA-type Polymer Based on Diazacrown Ether as the Solubilizer of Barium Sulfate to Water

    Bo ZHOU; Jian Zhang LI; Chun Hong HE; Sheng Ying QIN


    The EDTA dianhydride reacted with diazacrown ethers to obtain the water-soluble EDTA-diazacrown ether polymers 1~3. The effects of crown ether ring in the polymer chains including its cavity size on the solubilization of barium sulfate to water were investigated by comparison with the crown ring-free analogue 4. The result shows that the polymer 2 is the efficient solubilizer of BaSO4 and the highest solubilization efficiency of the BaSO4 to water is up to 72.5%.

  5. A two phase field model for tracking vesicle-vesicle adhesion.

    Gu, Rui; Wang, Xiaoqiang; Gunzburger, Max


    A multi-phase-field model for simulating the adhesion between two vesicles is constructed. Two phase field functions are introduced to simulate each of the two vesicles. An energy model is defined which accounts for the elastic bending energy of each vesicle and the contact potential energy between the two vesicles; the vesicle volume and surface area constraints are imposed using a penalty method. Numerical results are provided to verify the efficacy of our model and to provide visual illustrations of the different types of contact. The method can be adjusted to solve endocytosis problems by modifying the bending rigidity coefficients of the two elastic bending energies. The method can also be extended to simulate multi-cell adhesions, one example of which is erythrocyte rouleaux. A comparison with laboratory observations demonstrates the effectiveness of the multi-phase field approach.

  6. Hybrid electrospun chitosan-phospholipids nanofibers for transdermal drug delivery

    Mendes, Ana Carina Loureiro; Gorzelanny, Christian; Halter, Natalia


    Chitosan (Ch) polysaccharide was mixed with phospholipids (P) to generate electrospun hybrid nanofibers intended to be used as platforms for transdermal drug delivery. Ch/P nanofibers exibithed average diameters ranging from 248 +/- 94 nm to 600 +/- 201 nm, depending on the amount of phospholipids...... used. Fourier Transformed Infra-Red (FTIR) spectroscopy and Dynamic Light Scattering (DLS) data suggested the occurrence of electrostatic interactions between amine groups of chitosan with the phospholipid counterparts. The nanofibers were shown to be stable for at least 7 days in Phosphate Buffer...

  7. Nanoporous Silicified Phospholipids and Application to Controlled Glycolic Acid Release

    Kang SangHwa


    Full Text Available Abstract This work demonstrates the synthesis and characterization of novel nanoporous silicified phospholipid bilayers assembled inorganic powders. The materials are obtained by silicification process with silica precursor at the hydrophilic region of phospholipid bilayers. This process involves the co-assembly of a chemically active phospholipids bilayer within the ordered porosity of a silica matrix and holds promise as a novel application for controlled drug release or drug containers with a high level of specificity and throughput. The controlled release application of the synthesized materials was achieved to glycolic acid, and obtained a zero-order release pattern due to the nanoporosity.

  8. Phospholipid remodeling and eicosanoid signaling in colon cancer cells.

    Das, Siddhartha; Martinez, Leobarda Robles; Ray, Suparna


    Phospholipid remodeling and eicosanoid synthesis are central to lipid-based inflammatory reactions. Studies have revealed that membrane phospholipid remodeling by fatty acids through deacylation/reacylation reactions increases the risk of colorectal cancers (CRC) by allowing the cells to produce excess inflammatory eicosanoids, such as prostaglandins, thromboxanes and leukotrienes. Over the years, efforts have been made to understand the lipid remodeling pathways and to design anti-cancer drugs targeting the enzymes of eicosanoid biosynthesis. Here, we discuss the recent progress in phospholipid remodeling and eicosanoid biosynthesis in CRC.

  9. Astrocytic Vesicle Mobility in Health and Disease

    Robert Zorec


    Full Text Available Astrocytes are no longer considered subservient to neurons, and are, instead, now understood to play an active role in brain signaling. The intercellular communication of astrocytes with neurons and other non-neuronal cells involves the exchange of molecules by exocytotic and endocytotic processes through the trafficking of intracellular vesicles. Recent studies of single vesicle mobility in astrocytes have prompted new views of how astrocytes contribute to information processing in nervous tissue. Here, we review the trafficking of several types of membrane-bound vesicles that are specifically involved in the processes of (i intercellular communication by gliotransmitters (glutamate, adenosine 5'-triphosphate, atrial natriuretic peptide, (ii plasma membrane exchange of transporters and receptors (EAAT2, MHC-II, and (iii the involvement of vesicle mobility carrying aquaporins (AQP4 in water homeostasis. The properties of vesicle traffic in astrocytes are discussed in respect to networking with neighboring cells in physiologic and pathologic conditions, such as amyotrophic lateral sclerosis, multiple sclerosis, and states in which astrocytes contribute to neuroinflammatory conditions.

  10. Getting to know the extracellular vesicle glycome.

    Gerlach, Jared Q; Griffin, Matthew D


    Extracellular vesicles (EVs) are a diverse population of complex biological particles with diameters ranging from approximately 20 to 1000 nm. Tremendous interest in EVs has been generated following a number of recent, high-profile reports describing their potential utility in diagnostic, prognostic, drug delivery, and therapeutic roles. Subpopulations, such as exosomes, are now known to directly participate in cell-cell communication and direct material transfer. Glycomics, the 'omic' portion of the glycobiology field, has only begun to catalog the surface oligosaccharide and polysaccharide structures and also the carbohydrate-binding proteins found on and inside EVs. The EV glycome undoubtedly contains vital clues essential to better understanding the function, biogenesis, release and transfer of vesicles, however getting at this information is technically challenging and made even more so because of the small physical size of the vesicles and the typically minute yield from physiological-scale biological samples. Vesicle micro-heterogeneity which may be related to specific vesicle origins and functions presents a further challenge. A number of primary studies carried out over the past decade have turned up specific and valuable clues regarding the composition and roles of glycan structures and also glycan binding proteins involved EV biogenesis and transfer. This review explores some of the major EV glycobiological research carried out to date and discusses the potential implications of these findings across the life sciences.

  11. Astrocytic vesicle mobility in health and disease.

    Potokar, Maja; Vardjan, Nina; Stenovec, Matjaž; Gabrijel, Mateja; Trkov, Saša; Jorgačevski, Jernej; Kreft, Marko; Zorec, Robert


    Astrocytes are no longer considered subservient to neurons, and are, instead, now understood to play an active role in brain signaling. The intercellular communication of astrocytes with neurons and other non-neuronal cells involves the exchange of molecules by exocytotic and endocytotic processes through the trafficking of intracellular vesicles. Recent studies of single vesicle mobility in astrocytes have prompted new views of how astrocytes contribute to information processing in nervous tissue. Here, we review the trafficking of several types of membrane-bound vesicles that are specifically involved in the processes of (i) intercellular communication by gliotransmitters (glutamate, adenosine 5'-triphosphate, atrial natriuretic peptide), (ii) plasma membrane exchange of transporters and receptors (EAAT2, MHC-II), and (iii) the involvement of vesicle mobility carrying aquaporins (AQP4) in water homeostasis. The properties of vesicle traffic in astrocytes are discussed in respect to networking with neighboring cells in physiologic and pathologic conditions, such as amyotrophic lateral sclerosis, multiple sclerosis, and states in which astrocytes contribute to neuroinflammatory conditions.


    A. V. Oberemko


    Full Text Available This review presents a generalized definition of vesicles as bilayer extracellular organelles of all celular forms of life: not only eu-, but also prokaryotic. The structure and composition of extracellular vesicles, history of research, nomenclature, their impact on life processes in health and disease are discussed. Moreover, vesicles may be useful as clinical instruments for biomarkers, and they are promising as biotechnological drug. However, many questions in this area are still unresolved and need to be addressed in the future. The most interesting from the point of view of practical health care represents a direction to study the effect of exosomes and microvesicles in the development and progression of a particular disease, the possibility of adjusting the pathological process by means of extracellular vesicles of a particular type, acting as an active ingredient. Relevant is the further elucidation of the role and importance of exosomes to the surrounding cells, tissues and organs at the molecular level, the prospects for the use of non-cellular vesicles as biomarkers of disease.

  13. Mechanics of post-fusion exocytotic vesicle

    Stephens, Thomas; Wu, Zhanghan; Liu, Jian


    Exocytosis is an important cellular process controlled by metabolic signaling. It involves vesicle fusion to the plasma membrane, followed by the opening of a fusion pore, and the subsequent release of the vesicular lumen content into the extracellular space. While most modeling efforts focus on the events leading to membrane fusion, how the vesicular membrane remodels after fusing to plasma membrane remains unclear. This latter event dictates the nature and the efficiency of exocytotic vesicular secretions, and is thus critical for exocytotic function. We provide a generic membrane mechanical model to systematically study the fate of post-fusion vesicles. We show that while membrane stiffness favors full-collapse vesicle fusion into the plasma membrane, the intravesicular pressure swells the vesicle and causes the fusion pore to shrink. Dimensions of the vesicle and its associated fusion pore further modulate this mechanical antagonism. We systematically define the mechanical conditions that account for the full spectrum of the observed vesicular secretion modes. Our model therefore can serve as a unified theoretical framework that sheds light on the elaborate control mechanism of exocytosis.

  14. Haloarchaea and the Formation of Gas Vesicles

    Felicitas Pfeifer


    Full Text Available Halophilic Archaea (Haloarchaea thrive in salterns containing sodium chloride concentrations up to saturation. Many Haloarchaea possess genes encoding gas vesicles, but only a few species, such as Halobacterium salinarum and Haloferax mediterranei, produce these gas-filled, proteinaceous nanocompartments. Gas vesicles increase the buoyancy of cells and enable them to migrate vertically in the water body to regions with optimal conditions. Their synthesis depends on environmental factors, such as light, oxygen supply, temperature and salt concentration. Fourteen gas vesicle protein (gvp genes are involved in their formation, and regulation of gvp gene expression occurs at the level of transcription, including the two regulatory proteins, GvpD and GvpE, but also at the level of translation. The gas vesicle wall is solely formed of proteins with the two major components, GvpA and GvpC, and seven additional accessory proteins are also involved. Except for GvpI and GvpH, all of these are required to form the gas permeable wall. The applications of gas vesicles include their use as an antigen presenter for viral or pathogen proteins, but also as a stable ultrasonic reporter for biomedical purposes.

  15. Sterols associated with small unilamellar vesicles (SUVs): intrinsic mobility role for 1H NMR detection.

    Mouret, Liza; Da Costa, Grégory; Bondon, Arnaud


    Small unilamellar vesicles (SUVs) of phospholipids are often used as a membrane model system for studying the interaction of molecules. When using NMR under the standard liquid-state conditions, SUV phospholipid proton spectra can be recorded, exhibiting sharp signals. This is not only because of the fast vesicular tumbling but also because of the combination of this tumbling with the individual motion of the lipids inside the bilayer. This appears evident because addition of cholesterol is responsible of broader resonances because of the slowing down of the lipid motion. On the other hand, no (1)H signal is detected for cholesterol in the bilayer. This lack of detection of the inserted molecules explains why generally SUVs are not considered as a good model for NMR studies under the standard liquid-state conditions. Here, we use two other sterols in order to demonstrate that an increase of the molecular mobility inside the bilayer could allow the detection of their proton resonances. For desmosterol and lanosterol, which show higher mobility inside the bilayer, with increasing lateral diffusion rates, (1)H sterol signals are detected in contrast to cholesterol. For the fast diffusing lanosterol, no significant improvement in detection is observed using deuterated lipids, demonstrating that homonuclear dipolar coupling is fully averaged out. Furthermore, in the case of low mobility such as for cholesterol, the use of a fast magic angle spinning probe is shown to be efficient to recover the full proton spectrum. Copyright © 2014 John Wiley & Sons, Ltd.

  16. Production of humic substances through coal-solubilizing bacteria

    Nelson Valero


    Full Text Available In this paper, the production of humic substances (HS through the bacterial solubilization of low rank coal (LRC was evaluated. The evaluation was carried out by 19 bacterial strains isolated in microenvironments with high contents of coal wastes. The biotransformed LRC and the HS produced were quantified in vitro in a liquid growth medium. The humic acids (HA obtained from the most active bacterial strain were characterized via elemental composition (C, H, N, O, IR analyses, and the E4/E6 ratio; they were then compared with the HA extracted chemically using NaOH. There was LRC biotransformation ranged from 25 to 37%, and HS production ranged from 127 to 3100 mg.L-1. More activity was detected in the isolated strains of Bacillus mycoides, Microbacterium sp, Acinetobacter sp, and Enterobacter aerogenes. The HA produced by B. mycoides had an IR spectrum and an E4/E6 ratio similar to those of the HA extracted with NAOH, but their elemental composition and their degree of aromatic condensation was different. Results suggest that these bacteria can be used to exploit the LRC resulting from coal mining activities and thus produce HS in order to improve the content of humified organic matter in soils.

  17. [Phosphate solubilization of Aureobasidium pullulan F4 and its mechanism].

    Wang, Dan; Zhan, Jing; Sun, Qing-Ye


    The Aureobasidium pullulans F4 was isolated from the rhizosphere of Hippochaete ramosissimum in Tongguanshan mine wasteland in Tongling City, Anhui Province. Liquid culture was conducted with four kinds of phosphorus sources, calcium phosphate, aluminum phosphate, ferric phosphate and rock phosphate to determine the pH, dissolved phosphorus, phosphorus in the bacteria and organic acid in the solution. The results showed that the phosphate solubilization by A. pullulans F4 varied with phosphorus sources, which decreased in order of aluminum phosphate > ferric phosphate, calcium phosphate > rock phosphate. The amounts of dissolved phosphorus in the different treatments were all higher than 200 mg x L(-1). The pH of the medium dropped immediately in 48 h, and the aluminum phosphate and ferric phosphate treatments showed a greater decrease in pH than the calcium phosphate and rock phosphate treatments. The organic acid synthesized by A. pullulans F4 included oxalic acid, citric acid and tartaric acid, and oxalic acid, among which oxalic acid was the dominated component. The phosphate dissolving capacity of A. pullulans F4 showed no significant correlation with organic acid, but significantly correlated with the pH. The available phosphorus was significantly improved with the combined application of A. pullulans F4 and glucose, suggesting A. pullulans F4 was a potent candidate for remediation of copper mine wastelands.

  18. Production of humic substances through coal-solubilizing bacteria

    Valero, Nelson; Gómez, Liliana; Pantoja, Manuel; Ramírez, Ramiro


    In this paper, the production of humic substances (HS) through the bacterial solubilization of low rank coal (LRC) was evaluated. The evaluation was carried out by 19 bacterial strains isolated in microenvironments with high contents of coal wastes. The biotransformed LRC and the HS produced were quantified in vitro in a liquid growth medium. The humic acids (HA) obtained from the most active bacterial strain were characterized via elemental composition (C, H, N, O), IR analyses, and the E4/E6 ratio; they were then compared with the HA extracted chemically using NaOH. There was LRC biotransformation ranged from 25 to 37%, and HS production ranged from 127 to 3100 mg.L−1. More activity was detected in the isolated strains of Bacillus mycoides, Microbacterium sp, Acinetobacter sp, and Enterobacter aerogenes. The HA produced by B. mycoides had an IR spectrum and an E4/E6 ratio similar to those of the HA extracted with NAOH, but their elemental composition and their degree of aromatic condensation was different. Results suggest that these bacteria can be used to exploit the LRC resulting from coal mining activities and thus produce HS in order to improve the content of humified organic matter in soils. PMID:25477925

  19. Interactions between solubilized polymer molecules and blood components.

    Liu, Zonghua; Jiao, Yanpeng; Wang, Tong; Zhang, Yuanming; Xue, Wei


    In biomedical field, a variety of natural or synthetic polymers are being exponentially developed and used in vivo to improve human health. In practical applications, these biopolymers would enter blood circulation directly or indirectly, positively or passively, rapidly or slowly. Blood is a special tissue of human body, which fulfills many important missions to sustain normal metabolism. The contact with blood of the biopolymers, which are seen as foreign substances by the body, would be harmful or even instantaneously lethal, depending on the nature and the dose of the biopolymers administered. Therefore, it is critical to clearly understand the potential influences of the foreign polymers on blood before the polymers are applied from the lab to bedside. In this review, we discuss the recent studies on the interactions of foreign, solubilized polymer molecules (excluding formed polymer materials) with blood constituents (red blood cells, white blood cells, platelets, plasma proteins, etc.), to gain insight into the potential in vivo applications of the biopolymers in the biomedical field. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. New competition assay for the solubilized hepatic galactosyl receptor

    Ray, D.A.; Weigel, P.H.


    The present method of quantitating soluble asialoglycoprotein (galactosyl) receptor activity relies on the selective precipitation of receptor-ligand complexes to allow separation from free ligand. To provide an alternative to selective precipitation procedures, a simple and rapid method to assay for detergent-solubilized galactosyl receptor activity has been developed which uses permeabilized, fixed cells as a source of immobilized solid-phase receptors. Isolated rat hepatocytes were treated with digitonin to make available the internal as well as the external receptors. The permeable cells were also treated with glutaraldehyde to prevent further protein loss during subsequent exposure to detergents such as Triton X-100. The permeable/fixed cells, which retained about 70% of their total /sup 125/I-asialo-orosomucoid (/sup 125/I-ASOR)-binding activity, with 89% specific binding, were insoluble even in 0.5% Triton X-100 and were easily pelleted. The permeable/fixed cells can be prepared in advance and stored frozen for months. A detergent extract of receptor is mixed with a constant amount of both /sup 125/I-ASOR and permeable/fixed cells. Soluble active receptors compete with immobilized receptors on the treated cell for binding of the /sup 125/I-ASOR. The assay is reproducible, linear over a broad range of soluble receptor concentration, and can quantitate receptor activity from as few as 10(5) hepatocytes. A modified purification procedure for the rat hepatic galactosyl receptor using this competition assay is also described.

  1. Looking Beyond Structure: Membrane Phospholipids of Skeletal Muscle Mitochondria.

    Heden, Timothy D; Neufer, P Darrell; Funai, Katsuhiko


    Skeletal muscle mitochondria are highly dynamic and are capable of tremendous expansion to meet cellular energetic demands. Such proliferation in mitochondrial mass requires a synchronized supply of enzymes and structural phospholipids. While transcriptional regulation of mitochondrial enzymes has been extensively studied, there is limited information on how mitochondrial membrane lipids are generated in skeletal muscle. Herein we describe how each class of phospholipids that constitute mitochondrial membranes are synthesized and/or imported, and summarize genetic evidence indicating that membrane phospholipid composition represents a significant modulator of skeletal muscle mitochondrial respiratory function. We also discuss how skeletal muscle mitochondrial phospholipids may mediate the effect of diet and exercise on oxidative metabolism. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Structure and mechanism of ATP-dependent phospholipid transporters

    Lopez Marques, Rosa Laura; Poulsen, Lisbeth Rosager; Bailly, Aurélien


    Background ATP-binding cassette (ABC) transporters and P4-ATPases are two large and seemingly unrelated families of primary active pumps involved in moving phospholipids from one leaflet of a biological membrane to the other. Scope of review This review aims to identify common mechanistic features...... in the way phospholipid flipping is carried out by two evolutionarily unrelated families of transporters. Major conclusions Both protein families hydrolyze ATP, although they employ different mechanisms to use it, and have a comparable size with twelve transmembrane segments in the functional unit. Further......, despite differences in overall architecture, both appear to operate by an alternating access mechanism and during transport they might allow access of phospholipids to the internal part of the transmembrane domain. The latter feature is obvious for ABC transporters, but phospholipids and other hydrophobic...

  3. Herpes simplex virus 1 induces de novo phospholipid synthesis

    Sutter, Esther [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Oliveira, Anna Paula de; Tobler, Kurt [Electron microscopy, Institute of Virology, University of Zuerich (Switzerland); Schraner, Elisabeth M. [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland); Sonda, Sabrina [Institute of Parasitology, University of Zuerich (Switzerland); Kaech, Andres [Center for Microscopy and Image Analysis, University of Zuerich (Switzerland); Lucas, Miriam S. [Electron Microscopy ETH Zuerich (EMEZ), Swiss Federal Institute of Technology, Zuerich (Switzerland); Ackermann, Mathias [Electron microscopy, Institute of Virology, University of Zuerich (Switzerland); Wild, Peter, E-mail: [Electron Microscopy, Institute of Veterinary Anatomy, University of Zuerich (Switzerland)


    Herpes simplex virus type 1 capsids bud at nuclear membranes and Golgi membranes acquiring an envelope composed of phospholipids. Hence, we measured incorporation of phospholipid precursors into these membranes, and quantified changes in size of cellular compartments by morphometric analysis. Incorporation of [{sup 3}H]-choline into both nuclear and cytoplasmic membranes was significantly enhanced upon infection. [{sup 3}H]-choline was also part of isolated virions even grown in the presence of brefeldin A. Nuclei expanded early in infection. The Golgi complex and vacuoles increased substantially whereas the endoplasmic reticulum enlarged only temporarily. The data suggest that HSV-1 stimulates phospholipid synthesis, and that de novo synthesized phospholipids are inserted into nuclear and cytoplasmic membranes to i) maintain membrane integrity in the course of nuclear and cellular expansion, ii) to supply membrane constituents for envelopment of capsids by budding at nuclear membranes and Golgi membranes, and iii) to provide membranes for formation of transport vacuoles.

  4. Directed vesicle transport by diffusio-osmosis

    Michler, D.; Shahidzadeh, N.; Sprik, R.; Bonn, D.


    We present a study on surfactant vesicles that spontaneously move towards an oil droplet that is deposited on a glass substrate. Tracer particles in the surfactant solution show that the motion is not self-propelled: the vesicles are entrained by a macroscopic hydrodynamic flow. Measurements of the flow velocity suggest that the flow is of diffusio-osmotic nature. The surfactant is observed to move into the oil phase which creates a gradient in ion concentration in the vicinity of the droplet. As the diffusion coefficients of the surfactant's co- and counter-ions differ, a charge separation takes place and an electric field arises. This electric field then generates a hydrodynamic flow along the charged glass substrate in which the vesicles are entrained.

  5. Functionalization of Block Copolymer Vesicle Surfaces

    Wolfgang Meier


    Full Text Available In dilute aqueous solutions certain amphiphilic block copolymers self-assemble into vesicles that enclose a small pool of water with a membrane. Such polymersomes have promising applications ranging from targeted drug-delivery devices, to biosensors, and nanoreactors. Interactions between block copolymer membranes and their surroundings are important factors that determine their potential biomedical applications. Such interactions are influenced predominantly by the membrane surface. We review methods to functionalize block copolymer vesicle surfaces by chemical means with ligands such as antibodies, adhesion moieties, enzymes, carbohydrates and fluorophores. Furthermore, surface-functionalization can be achieved by self-assembly of polymers that carry ligands at their chain ends or in their hydrophilic blocks. While this review focuses on the strategies to functionalize vesicle surfaces, the applications realized by, and envisioned for, such functional polymersomes are also highlighted.

  6. Asymmetric Vesicle Instability in Extensional Flow

    Spann, Andrew; Zhao, Hong; Shaqfeh, Eric


    Previous researchers have chronicled the breakup of drops in an extensional flow as they stretch into a dumbbell shape with a long thin neck. Motivated by recent experimental observations, we study an apparently similar problem with vesicles, which are deformable but incompressible membranes that conserve area and volume. First, we simulate vesicles in an unbounded uniaxial extensional flow which are given general radial perturbations from an initially stable symmetric equilibrium state. For sufficiently low reduced volume (outer viscosity) there exists a capillary number at which an asymmetric perturbation mode will grow, resulting in the formation of an asymmetric dumbbell shape with a thin connecting cylindrical bridge analogous to the shapes associated with drop breakup. Our simulations help elucidate a mechanism for this instability based on a competition between internal pressure differentials in the vesicle resulting from the membrane bending force and ambient flow. We compare and contrast this transition to the ``standard'' drop breakup transition. Funded by NSF GRFP and Stanford Graduate Fellowship.

  7. Microbial Efficacy of Phosphate Solubilization in Agro-Saline Soils of Various Areas of Sindh Region

    Agha Asad Noor


    Full Text Available Microorganisms are the most prominent entities for solubilization of phosphate in various soils of different areas of Sindh Province including Tando Muhammad Khan, Tando Allah Yar, Nawabshah, Rato Dero-Larkana, Shikarpur and Umer Kot. These soils, having varying concentrations of chemicals, different climatic conditions, pH and varying numbers of microorganisms for PSA (Phosphate Solubilization Activity. This presentation shows the isolation of different fungi and bacteria capable Psa including fungi (Fusarium sp. Aspergillus sp. Penicillium sp. and Rhizopus sp. and bacteria (Bacillus sp. Pseudomonas sp. and Arthrobacter sp.. From the observations, it was revealed that fungi Aspergillus sp. and Bacillus sp. showed greater phosphate solubilization activity as compared to other fungi and bacteria showing 60 and 53.33% Psa (Phosphate Solubilizing Activity respectively

  8. Isolation and characterization of phosphate-solubilizing bacteria from seagrass rhizosphere soil

    Ghosh, Upasana; Subhashini, Ponnambalam; Dilipan, Elangovan; Raja, Subramanian; Thangaradjou, Thirunavukarassu; Kannan, Lakshmanan


    Phosphate-solubilizing bacterial strains (6 Nos.) were isolated from the rhizosphere soils of two seagrasses ( Halophila ovalis (R. Br.) Hook and Halodule pinifolia (Miki) Hartog) in the Vellar estuary. Experimental studies found that the strain PSSG6 was effective in phosphate solubilization with Phosphate Solubilization efficiency index E = 375 ± 8.54, followed by the strain PSSG5 with Phosphate Solubilization efficiency index E = 275 ± 27.3. Of the 6 strains isolated, the strains PSSG4 and PSSG5 belonged to the genus Bacillus, and PSSG1, PSSG2 and PSSG3 were identified as Citrobacter sp., Shigella sp., and Klebsiella sp., respectively, by conventional method, and PSSG6 was identified as Bacillus circulans using conventional and molecular methods.

  9. Identification and characterization of Gluconacetobacter diazotrophicus mutants defective in the solubilization of phosphorus and zinc.

    Intorne, Aline C; de Oliveira, Marcos Vinicius V; Lima, Mariana L; da Silva, Juliana F; Olivares, Fábio L; de Souza Filho, Gonçalo Apolinário


    Gluconacetobacter diazotrophicus is a plant-growth-promoting bacterium, which is able to colonize sugarcane and other plant species of economic importance. The potentially beneficial effects promoted by this bacterium on plants are nitrogen-fixation, production of phythormones, action against pathogens and mineral nutrient solubilization. In this study, the molecular mechanisms associated with phosphorus and zinc solubilization were analyzed. A transposon mutant library was constructed and screened to select for mutants defective for phosphorous [Ca(5)(PO(4))(3)OH] and zinc (ZnO) solubilization. A total of five mutants were identified in each screen. Both screenings, performed independently, allowed to select the same mutants. The interrupted gene in each mutant was identified by sequencing and the results demonstrate that the production of gluconic acid is a required pathway for solubilization of such nutrients in G. diazotrophicus.

  10. Isolation and Characterization of Phosphate-Solubilizing Bacteria from Seagrass Rhizosphere Soil

    Upasana Ghosh; Ponnambalam Subhashini; Elangovan Dilipan; Subramanian Raja; Thirunavukarassu Thangaradjou; Lakshmanan Kannan


    Phosphate-solubilizing bacterial strains (6 Nos.) were isolated from the rhizosphere soils of two seagrasses (Halophila ovalis (R.Br.) Hook and Halodule pinifolia (Miki) Hartog) in the Vellar estuary.Experimental studies found that the strain PSSG6was effective in phosphate solubilization with Phosphate Solubilization efficiency index E=375 ± 8.54,followed by the strain PSSG5with Phosphate Solubilization efficiency index E=275 ± 27.3.Of the 6 strains isolated,the strains PSSG4 and PSSG5 belonged to the genus Bacillus,and PSSG1,PSSG2 and PSSG3 were identified as Citrobacter sp.,Shigella sp.,and Klebsiella sp.,respectively,by conventional method,and PSSG6 was identified as Bacillus circulans using conventional and molecular methods.


    Although surfactants have received considerable attention as a potential means for enhancing the recovery of organic compounds from the subsurface, only limited information is available regarding the micellar solubilization of common groundwater contaminants by nonionic surfactan...

  12. Mechanisms of coal solubilization by the deuteromycetes Trichoderma atroviride and Fusarium oxysporum

    Hoelker, U.; Ludwig, S.; Scheel, T.; Hoefer, M. [Bonn Univ. (Germany). Botanisches Inst. und Botanischer Garten


    Three different mechanisms can be envisaged that are used by fungi to solubilize coal: the production of alkaline substances, the extrusion of chelators and, of special interest in the scope of biotechnology, the action of enzymes. Whether these mechanisms are operating separately or in various combinations has not yet been finally assessed. The two deuteromycetes Fusarium oxysporum and Trichoderma atroviride solubilize coal by synergistic effects of various different mechanisms depending on the cell metabolism. F. oxysporum seems to solubilize coal by increasing the pH of the mycelial surroundings and by the action of chelators induced during growth in glutamate-containing media (without involvement of enzymes). T. atroviride, on the other hand, appears to use, in addition to an alkaline pH and a high chelator activity, at least two classes of enzyme activity to attack coal: hydrolytic activity for coal solubilization and ligninolytic activity for degradation of humic acids. (orig.)

  13. Regional distribution of phospholipids in porcine vitreous humor.

    Schnepf, Abigail; Yappert, Marta Cecilia; Borchman, Douglas


    This project explores the regional phospholipid distribution in porcine vitreous humor, retina, and lens. Matrix-assisted laser desorption mass spectrometry has been used previously to image lipids, proteins, and other metabolites in retinas and lenses. However, the regional composition of phospholipids in vitreous humors is not known. To address this issue, we have applied this mass spectral method to explore the regional phospholipid distribution in porcine vitreous humor both ex-situ and in-vitro. To establish the possible source(s) of phospholipids in the vitreous humor, compositional studies of the lens and retina were also pursued. Due to the overall low levels of phospholipids in vitreous humor, it was necessary to optimize the experimental approaches for ex-situ and in-vitro studies. The sensitivity observed in the spectra of methanol extracts from the lens and retina was higher than that for methanol:chloroform extracts, but the compositional trends were the same. A fourfold improvement in sensitivity was observed in the analysis of vitreous humor extracts obtained with the Bligh and Dyer protocol relative to the other two extraction methods. For ex-situ studies, the 'stamp method' with para-nitroaniline as the matrix was chosen. Throughout the vitreous humor, phosphatidylcholines were the most abundant phospholipids. In-vitro results showed higher relative levels of phospholipids compared to the 'stamp' method. However, more details in the regional phospholipid distribution were provided by the ex-situ approach. Both in-vitro and ex-situ results indicated higher levels of phospholipids in the posterior vitreous region, followed by the anterior and central regions. The posterior region contained more unsaturated species whereas more saturated phospholipids were detected in the anterior region. The observed trends suggest that the phospholipids detected in the posterior vitreous humor migrate from the retina and associated vasculature while those present in

  14. Screening for the drug-phospholipid interaction: correlation to phospholipidosis

    Alakoskela, Juha-Matti; Vitovic, Pavol; Kinnunen, Paavo K J


    Phospholipid bilayers represent a complex, anisotropic environment fundamentally different from bulk oil or octanol, for instance. Even "simple" drug association to phospholipid bilayers can only be fully understood if the slab-of-hydrocarbon approach is abandoned and the complex, anisotropic pro...... of these interactions in PLD in particular. We also focus on a potential causal connection between drug-induced PLD and steatohepatitis, which is induced by some cationic amphiphilic drugs....

  15. Phospholipid synthesis rates in the eastern subtropical South Pacific Ocean

    B. A. S. Van Mooy


    Full Text Available Membrane lipid molecules are a major component of planktonic organisms and this is particularly true of the microbial picoplankton that dominate the open ocean; with their high surface-area to volume ratios, the synthesis of membrane lipids places a major demand on their overall cell metabolism. Specifically, the synthesis of cell membrane phospholipids creates a demand for the nutrient phosphorus, and we sought to refine our understanding of the role of phospholipids in the upper ocean phosphorus cycle. We measured the rates of phospholipid synthesis in a transect of the eastern subtropical South Pacific from Easter Island to Concepcion, Chile as part of the BIOSOPE program. Our approach combined standard phosphorus radiotracer incubations and lipid extraction methods. We found that phospholipid synthesis rates varied from less than 1 to greater than 200 pmol P L−1 h−1, and that phospholipid synthesis contributed between less than 5% to greater than 22% of the total PO43− incorporation rate. Changes in the percentage that phospholipid synthesis contributed to total PO43− uptake were strongly correlated with the ratio of primary production to bacterial production, which supported our hypothesis that heterotrophic bacteria were the primary agents of phospholipid synthesis. The spatial variation in phospholipid synthesis rates underscored the importance of heterotrophic bacteria in the phosphorus cycle of the eastern subtropical South Pacific, particularly the hyperoligotrophic South Pacific subtropical gyre.

  16. Phosphate-solubilizing microbes and their occurrence in the rhizospheres of Piper betel in Karnataka, India



    Low phosphate solubility is one of the most important factors limiting the plant growth in Indian soils. Many microorganisms can enhance phosphate solubility, but little is known about the magnitude of their phosphorus-solubilizing ability. The native populations of phosphate-solubilizing bacteria and fungi were studied in different rhizospheric soil samples obtained from betel vine plants (Piper betel L.) in order to compare the results. The present study focuses on the phosphate-solubilizin...

  17. Diversity and phosphate solubilization by bacteria isolated from Laki Island coastal ecosystem



    Widawati S (2011) Diversity and phosphate solubilization by bacteria isolated from Laki Island coastal ecosystem. Biodiversitas 12: 17-21. Soil, water, sand, and plant rhizosphere samples collected from coastal ecosystem of Laki Island-Jakarta were screened for phosphate solubilizing bacteria (PSB). While the population was dependent on the cultivation media and the sample type, the highest bacterial population was observed in the rhizosphere of Ipomea aquatica. The PSB strains isolated from ...

  18. Performance of pineapple slips inoculated with diazotrophic phosphate-solubilizing bacteria and rock phosphate

    Lílian Estrela Borges Baldotto; Marihus Altoé Baldotto; Fábio Lopes Olivares; Adriane Nunes de Souza


    Besides fixing N2, some diazotrophic bacteria or diazotrophs, also synthesize organic acids and are able to solubilize rock phosphates, increasing the availability of P for plants. The application of these bacteria to pineapple leaf axils in combination with rock phosphate could increase N and P availability for the crop, due to the bacterial activity of biological nitrogen fixation and phosphate solubilization. The objectives of this study were: (i) to select and characterize diazotrophs abl...

  19. Selection and evaluation of phosphate-solubilizing bacteria from grapevine rhizospheres for use as biofertilizers

    Min Liu; Xu Liu; Bao-Sen Cheng; Xue-Lei Ma; Xiao-Tong Lyu; Xian-Fang Zhao; Yan-Lun Ju; Zhuo Min; Yu-Lin Fang


    Phosphate-solubilizing bacteria (PSB) have the ability to solubilize insoluble phosphorus (P) and release soluble P. Extensive research has been performed with respect to PSB isolation from the rhizospheres of various plants, but little is known about the prevalence of PSB in the grapevine rhizosphere. In this study, we aimed to isolate and identify PSB from the grapevine rhizosphere in five vineyards of Northwest China, to characterize their plant-growth-promoting (PGP) traits, evaluate the ...

  20. Phosphate-Solubilizing and -Mineralizing Abilities of Bacteria Isolated from Soils

    TAO Guang-Can; TIAN Shu-Jun; CAI Miao-Ying; XIE Guang-Hui


    Microorganisms capable of solubilizing and mineralizing phosphorus (P) pools in soils are considered vital in promoting P bioavailability. The study was conducted to screen and isolate inorganic P-solubilizing bacteria (IPSB) and organic P-mineralizing bacteria (OPMB) in soils taken from subtropical flooded and temperate non-flooded soils, and to compare inorganic P-solubilizing and organic P-solubilizing abilities between IPSB and OPMB. Ten OPMB strains were isolated and identified as Bacillus cereus and Bacillus megaterium, and five IPSB strains as B. megaterium, Burkholderia caryophylli,Pseudomonas ciehorii, and Pseudomonas syringae. P-solubilizing and -mineralizing abilities of the strains were measured using the methods taking cellular P into account. The IPSB strains exhibited inorganic P-sohibilizing abilities ranging between 25.4-41.7 μg P mL-1 and organic P-mineralizing abilities between 8.2-17.8 μg P mL-1. Each of the OPMB strains also exhibited both solubilizing and mineralizing abilities varying from 4.4 to 26.5 μg P mL-1 and from 13.8 to 62.8 μg P mL-1, respectively. For both IPSB and OPMB strains, most of the P mineralized from the organic P source was incorporated into the bacterial cells as cellular P. A significantly negative linear correlation (P < 0.05) was found between culture pH and P solubilized from inorganic P by OPMB strains. The results suggested that P solubilization and mineralization could coexist in the same bacterial strain.

  1. The mechanism of detergent solubilization of liposomes and protein-containing membranes.

    Kragh-Hansen, U; le Maire, M; Møller, J V


    The present study explores intermediate stages in detergent solubilization of liposomes and Ca2+-ATPase membranes by sodium dodecyl sulfate (SDS) and medium-sized ( approximately C12) nonionic detergents. In all cases detergent partitioning in the membranes precedes cooperative binding and solubilization, which is facilitated by exposure to detergent micelles. Nonionic detergents predominantly interact with the lipid component of Ca2+-ATPase membranes below the CMC (critical micellar concentr...

  2. Toroidal membrane vesicles in spherical confinement

    Bouzar, Lila; Müller, Martin Michael


    We investigate the morphology of a toroidal fluid membrane vesicle confined inside a spherical container. The equilibrium shapes are assembled in a geometrical phase diagram as a function of scaled area and reduced volume of the membrane. For small area the vesicle can adopt its free form. When increasing the area, the membrane cannot avoid contact and touches the confining sphere along a circular contact line, which extends to a zone of contact for higher area. The elastic energies of the equilibrium shapes are compared to those of their confined counterparts of spherical topology to predict under which conditions a topology change is favored energetically.

  3. Toroidal membrane vesicles in spherical confinement

    Bouzar, Lila; Menas, Ferhat; Müller, Martin Michael


    We investigate the morphology of a toroidal fluid membrane vesicle confined inside a spherical container. The equilibrium shapes are assembled in a geometrical phase diagram as a function of scaled area and reduced volume of the membrane. For small area the vesicle can adopt its free form. When increasing the area, the membrane cannot avoid contact and touches the confining sphere along a circular contact line, which extends to a zone of contact for higher area. The elastic energies of the equilibrium shapes are compared to those of their confined counterparts of spherical topology to predict under which conditions a topology change is favored energetically.

  4. Molecular Insights into Phospholipid -- NSAID Interactions

    Babu Boggara, Mohan; Krishnamoorti, Ramanan


    Non steroidal anti inflammatory drugs (NSAIDs) e.g. Aspirin and Ibuprofen, with chronic usage cause gastro intestinal (GI) toxicity. It has been shown experimentally that NSAIDs pre-associated with phospholipids reduce the GI toxicity and also increase the therapeutic activity of these drugs compared to the unmodified ones. Using all atomistic simulations and two different methodologies, we studied the partitioning behavior of two model NSAIDs (Aspirin and Ibuprofen) as a function of pH and drug loading. The results from two methodologies are consistent in describing the equilibrium drug distribution in the bilayers. Additionally, the heterogeneity in density and polarity of the bilayer in the normal direction along with the fact that NSAIDs are amphiphilic (all of them have a carboxylic acid group and a non-polar part consisting of aromatic moieties), indicate that the diffusion mechanism in the bilayer is far different compared to the same in a bulk medium. This study summarizes the various effects of NSAIDs and their behavior inside the lipid bilayer both as a function of pH and drug concentration.

  5. Enhancement by cytidine of membrane phospholipid synthesis

    G-Coviella, I. L.; Wurtman, R. J.


    Cytidine, as cytidine 5'-diphosphate choline, is a major precursor in the synthesis of phosphatidylcholine in cell membranes. In the present study, we examined the relationships between extracellular levels of cytidine, the conversion of [14C]choline to [14C]phosphatidylcholine, and the net syntheses of phosphatidylcholine and phosphatidylethanolamine by PC12 cells. The rate at which cytidine (as [3H]cytidine) was incorporated into the PC12 cells followed normal Michaelis-Menten kinetics (Km = 5 microM; Vmax = 12 x 10(-3) mmol/mg of protein/min) when the cytidine concentrations in the medium were below 50 microM; at higher concentrations, intracellular [3H]cytidine nucleotide levels increased linearly. Once inside the cell, cytidine was converted mainly into cytidine triphosphate. In pulse-chase experiments, addition of cytidine to the medium caused a time- and dose-dependent increase (by up to 30%) in the incorporation of [14C]choline into membrane [14C]-phosphatidylcholine. When the PC12 cells were supplemented with both cytidine and choline for 14 h, small but significant elevations (p less than 0.05) were observed in their absolute contents of membrane phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine, all increasing by 10-15% relative to their levels in cells incubated with choline alone. Exogenous cytidine, acting via cytidine triphosphate, can thus affect the synthesis and levels of cell membrane phospholipids.

  6. Electrohydrodynamics of a compound vesicle under an AC electric field

    Priti Sinha, Kumari; Thaokar, Rochish M.


    Compound vesicles are relevant as simplified models for biological cells as well as in technological applications such as drug delivery. Characterization of these compound vesicles, especially the inner vesicle, remains a challenge. Similarly their response to electric field assumes importance in light of biomedical applications such as electroporation. Fields lower than that required for electroporation cause electrodeformation in vesicles and can be used to characterize their mechanical and electrical properties. A theoretical analysis of the electrohydrodynamics of a compound vesicle with outer vesicle of radius R o and an inner vesicle of radius λ {{R}o} , is presented. A phase diagram for the compound vesicle is presented and elucidated using detailed plots of electric fields, free charges and electric stresses. The electrohydrodynamics of the outer vesicle in a compound vesicle shows a prolate-sphere and prolate-oblate-sphere shape transitions when the conductivity of the annular fluid is greater than the outer fluid, and vice-versa respectively, akin to single vesicle electrohydrodynamics reported in the literature. The inner vesicle in contrast shows sphere-prolate-sphere and sphere-prolate-oblate-sphere transitions when the inner fluid conductivity is greater and smaller than the annular fluid, respectively. Equations and methodology are provided to determine the bending modulus and capacitance of the outer as well as the inner membrane, thereby providing an easy way to characterize compound vesicles and possibly biological cells.

  7. Biomimetic surface modification of polyurethane with phospholipids grafted carbon nanotubes.

    Tan, Dongsheng; Liu, Liuxu; Li, Zhen; Fu, Qiang


    To improve blood compatibility of polyurethane (PU), phospholipids grafted carbon nanotubes (CNTs) were prepared through zwitterion-mediated cycloaddition reaction and amide condensation, and then were added to the PU as fillers via solution mixing to form biomimetic surface. The properties of phospholipids grafted CNTs (CNT-PC) were investigated by thermal gravimetric analysis (TGA), X-ray photoelectron spectroscopy (XPS) and proton nuclear magnetic resonance ((1) H NMR). The results indicated that the phospholipids were grafted onto CNTs in high efficiency, and the hydrophilicity and dispersibility of the modified CNTs were improved effectively. The structures and properties of composites containing CNT-PC were investigated by optical microscope, XPS, and water contact angles. The results indicated that phospholipids were enriched on the surface with addition of 0.1 wt % of CNT-PC, which significantly reduced protein adsorption and platelet adhesion. The method of carrying phospholipids on the nanofiller to modify polymers has provided a promising way of constructing biomimetic phospholipid membrane on the surface to improve blood compatibility.

  8. Phospholipid Synthesis in Sindbis Virus-Infected Cells

    Waite, Marilynn R. F.; Pfefferkorn, E. R.


    We investigated the metabolic requirements for the decrease in phospholipid synthesis previously observed by Pfefferkorn and Hunter in primary cultures of chick embryo fibroblasts infected with Sindbis virus. The incorporation of 32PO4 into all classes of phospholipids was found to decline at the same rate and to the same extent; thus, incorporation of 14C-choline into acid-precipitable form provided a convenient measure of phospholipid synthesis that was used in subsequent experiments. Experiments with temperature-sensitive mutants suggested that some viral ribonucleic acid (RNA) synthesis was essential for the inhibition of choline incorporation, but that functional viral structural proteins were not required. The reduction in phospholipid synthesis was probably a secondary effect of infection resulting from viral inhibition of the cellular RNA and protein synthesis. All three inhibitory effects required about the same amount of viral RNA synthesis; the inhibition of host RNA and protein synthesis began sooner than the decline in phospholipid synthesis; and both actinomycin D and cycloheximide inhibited 14C-choline incorporation in uninfected cells. In contrast, incorporation of 14C-choline into BHK-21 cells was not decreased by 10 hr of exposure to actinomycin D and declined only slowly after cycloheximide treatment. Growth of Sindbis virus in BHK cells did not cause the marked stimulation of phospholipid synthesis seen in picornavirus infections of other mammalian cells; however, inhibition was seen only late in infection. PMID:5530011

  9. Regulation of phospholipid synthesis in Saccharomyces cerevisiae by zinc.

    Iwanyshyn, Wendy M; Han, Gil-Soo; Carman, George M


    Zinc is an essential nutrient required for the growth and metabolism of eukaryotic cells. In this work, we examined the effects of zinc depletion on the regulation of phospholipid synthesis in the yeast Saccharomyces cerevisiae. Zinc depletion resulted in a decrease in the activity levels of the CDP-diacylglycerol pathway enzymes phosphatidylserine synthase, phosphatidylserine decarboxylase, phosphatidylethanolamine methyltransferase, and phospholipid methyltransferase. In contrast, the activity of phosphatidylinositol synthase was elevated in response to zinc depletion. The level of Aut7p, a marker for the induction of autophagy, was also elevated in zinc-depleted cells. For the CHO1-encoded phosphatidylserine synthase, the reduction in activity in response to zinc depletion was controlled at the level of transcription. This regulation was mediated through the UAS(INO) element and by the transcription factors Ino2p, Ino4p, and Opi1p that are responsible for the inositol-mediated regulation of UAS(INO)-containing genes involved in phospholipid synthesis. Analysis of the cellular composition of the major membrane phospholipids showed that zinc depletion resulted in a 66% decrease in phosphatidylethanolamine and a 29% increase in phosphatidylinositol. A zrt1Delta zrt2Delta mutant (defective in the plasma membrane zinc transporters Zrt1p and Zrt2p) grown in the presence of zinc exhibited a phospholipid composition similar to that of wild type cells depleted for zinc. These results indicated that a decrease in the cytoplasmic levels of zinc was responsible for the alterations in phospholipid composition.

  10. Diversity and phosphate solubilization by bacteria isolated from Laki Island coastal ecosystem



    Full Text Available Widawati S (2011 Diversity and phosphate solubilization by bacteria isolated from Laki Island coastal ecosystem. Biodiversitas 12: 17-21. Soil, water, sand, and plant rhizosphere samples collected from coastal ecosystem of Laki Island-Jakarta were screened for phosphate solubilizing bacteria (PSB. While the population was dependent on the cultivation media and the sample type, the highest bacterial population was observed in the rhizosphere of Ipomea aquatica. The PSB strains isolated from the sample registered 18.59 g-1L-1, 18.31 g-1L-1, and 5.68 g-1L-1 of calcium phosphate (Ca-P, Al-P and rock phosphate solubilization after 7-days. Phosphate solubilizing capacity was the highest in the Ca-P medium. Two strains, 13 and 14, registered highest Phosphomonoesterase activities (2.01 µgNP.g-1.h-1 and 1.85NP µg.g-1.h-1 were identified as Serattia marcescens, and Pseudomonas fluorescense, respectively. Both strains were isolated from the crops of Amaranthus hybridus and I. aquatica, respectively, which are commonly observed in coastal ecosystems. The presence of phosphate solubilizing microorganisms and their ability to solubilize various types of phosphate species are indicative of the important role of both species of bacteria in the biogeochemical cycle of phosphorus and the plant growth in coastal ecosystems.

  11. Biochemical analysis and solubilization of central alpha 1-adrenoceptors in the rat

    Guicheney, P.; Rappaport, A.; Meyer, P.


    To study the structure and the molecular mechanisms of action of brain alpha-adrenoceptors, their solubilization was undertaken. alpha 1-Adrenoceptors were first successfully solubilized from fresh rat brain membranes by treatment with 0.3 percent deoxycholate, after prelabeling of the binding site by the highly specific tritiated antagonist /sup 3/H-prazosin. The complex thus solubilized underwent a rapid loss of activity at 25 degrees C. Direct solubilization of brain alpha 1-adrenoceptors was obtained by treatment with a new zwitterionic derivative of cholic acid (CHAPS) at a concentration of 5 to 10 mM. The soluble complex was detected by precipitation by polyethylene glycol 6,000 with gamma globulin as a carrier. Binding of /sup 3/H-prazosin at 25 degrees C was rapid; at 4 degrees C the steady state was obtained within two hours and remained unchanged for at least six hours. The affinity of the soluble binding site, determined by Scatchard analysis (congruent to 0.6 nM), varied with the concentration of detergent. Specificity of the membrane-bound receptor was preserved as demonstrated by incubation in the presence of alpha 1- and alpha 2-antagonists at various concentrations (by order of potency: prazosin greater than phentolamine greater than yohimbine). Stereoselectivity was also retained in the solubilized binding protein. The solubilization of an active brain alpha 1-adrenoceptor will allow further investigation at the molecular level.

  12. Ethyl methanesulfonate mutagenesis-enhanced mineral phosphate solubilization by groundnut-associated Serratia marcescens GPS-5.

    Tripura, Chaturvedula; Sashidhar, Burla; Podile, Appa Rao


    Twenty-three bacterial isolates were screened for their mineral phosphate-solubilizing (MPS) ability on Pikovskaya and National Botanical Research Institute's phosphate (NBRIP) agar. The majority of the isolates exhibited a strong ability to solubilize hydroxyapatite in both solid and liquid media. The solubilization in liquid medium corresponded with a decrease in the pH of the medium. Serratia marcescens GPS-5, known for its biocontrol of late leaf spot in groundnut, emerged as the best solubilizer. S. marcescens GPS-5 was subjected to ethyl methanesulfonate (EMS) mutagenesis, and a total of 1700 mutants, resulting after 45 minutes of exposure, were screened on buffered NBRIP medium for alterations in MPS ability compared with that of the wild type. Seven mutants with increased (increased-MPS mutants) and 6 mutants with decreased (decreased-MPS mutants) MPS ability were isolated. All seven increased-MPS mutants were efficient at solubilizing phosphate in both solid and liquid NBRIP medium. Among the increased-MPS mutants, EMS XVIII Sm-35 showed the maximum (40%) increase in the amount of phosphate released in liquid medium compared with wild-type S. marcescens GPS-5, therefore, it would be a useful microbial inoculant in groundnut cultivation. EMS III Sm W, a nonpigmented mutant, showed the lowest solubilization of phosphate among the 6 decreased-MPS mutants.

  13. Solubilization of Waste Activated Sludge and Nitrogenous Compounds Transformation During Solubilization by Thermophilic Enzyme (S-TE) Process.

    Yang, Qi; Luo, Kun; Li, Xiao-ming; Zhong, Yu; Chen, Hong-bo; Yang, Guo-jing; Shi, Yan-wei; Zeng, Guang-ming


    A representative thermophilic bacterial strain (AT06-1) capable of secreting protease was isolated from thermophilic aerobic digestion reactor, and 16S rRNA gene analysis indicated that it was Bacillus sp. The isolated strain was inoculated in waste activated sludge (WAS) to evaluate the performance of solubilization by thermophilic enzyme (S-TE) process under aerobic or microaerobic conditions at different temperatures (55-70 °C). Results showed that the inoculation of specific thermophilic strain significantly affected the volatile suspended solids (VSS) removal. At the optimal temperature of 65 °C, the maximum VSS removal of 43.6 % and highest SCOD of 4475 mg/L was achieved during microaerobic S-TE process. Compared to the noninoculation, more soluble protein was released during S-TE process due to the higher protease activity associated with the protein hydrolysis originated from cell lysis. The protease activity at aerobic and microaerobic S-TE process was respectively 1.73 and 1.88 times that of the noninoculation. Ammonia was the end nitrogenous compound of protein hydrolysis during S-TE process, which was stripped from the digestion system through continuous aeration.

  14. Exploring novel techniques for the single molecule toolkit: Vesicle encapsulation and immobilization

    Okumus, Burak

    Tracking asynchronous time evolution of single biological molecules provides unique insights into detailed reaction kinetics and pathways. Such measurements are frequently made on macromolecules that are tethered on a glass surface. However, there have been reports of variability of surface environment, and suspicion that observed heterogeneity of dynamic properties in single molecules might be an artifact of the local surface. A striking example is the hairpin ribozyme which was shown---in our lab---to exhibit two orders of magnitude variation in folding/unfolding kinetics between molecules. Moreover, a DNA with a sequence of human telomeric repeat exhibited extreme conformational diversity among six interconverting conformations. In order to find out the true nature of the observed heterogeneities, we encapsulated the ribozyme and the human telomeric DNA inside liposomes (i.e. artificially formed phospholipid vesicles) which were then tethered on the surface. Our data revealed similar behavior for encapsulated and the conventionally attached nucleic acid molecules. Although vesicle encapsulation offers a biologically relevant environment for many soluble proteins and nucleic acids, impermeability towards ions and other small molecules such as ATP hinders more general applications. We therefore developed methods to induce pores into vesicles which open up the possibility of using them as ultra-small, bio-mimetic, porous containers. Porous vesicles were then utilized to perform unique measurements for observing RecA filament formation, hairpin ribozyme cleavage and Rep helicase translocation within confined volumes. Novel features were revealed by such experiments unveiling new biological findings. We also discuss ideas to introduce pores that can be opened up via ultraviolet radiation for future applications. Aside from the encapsulation studies, we developed a new assay to detect the SNARE mediated membrane fusion by using surface attached proteliposomes. Such an

  15. A Novel C2-Domain Phospholipid-Binding Protein,OsPBP1.Is Required for Pollen Fertility in Rice

    Wen-Qiang Yang; Ying Lai; Mei-Na Li; Wen-Ying Xu; Yong-Biao Xue


    Pollen fertility is a crucial factor for successful pollination and essential for seed formation.Recent studies have suggested that a diverse range of internal and external factors,signaling components and their related pathways are likely involved in pollen fertility.Here,we reporta single C2-domain containing protein.OsPBPl.initially identified through cDNA microarray analysis.OsP8P1 is a single copy gene and preferentially expressed in pistil and pollen but downregulated by pollination.OsPBP1 had a calcium concentration-dependent phospholipid-binding activity and was localized mainly in cytoplasm and nucleus,but translocated onto the plasma membrane in response to an intracellular Ca2+increase.Pollen grains of antisense OsPBP1 transgenic Iines were largely nonviable.germinated poorly in vitro and of low fertility,OsPBP1 protein was localized in a region peripheral to pollen wall and vesicles of elongating pollen tube.and its repressed expression reduced substantially this association and led to alteration of microfilament polymerization during pollen germination.Taken together,these results indicate that OsPBP1 is a novel functional C2-domain phosphoIipids-binding protein that is required for pollen fertility likely by regulating Ca2+ and phospholipid signaling pathways.

  16. Characterisation of adipocyte-derived extracellular vesicles released pre- and post-adipogenesis

    Katherine D. Connolly


    Full Text Available Extracellular vesicles (EVs are submicron vesicles released from many cell types, including adipocytes. EVs are implicated in the pathogenesis of obesity-driven cardiovascular disease, although the characteristics of adipocyte-derived EVs are not well described. We sought to define the characteristics of adipocyte-derived EVs before and after adipogenesis, hypothesising that adipogenesis would affect EV structure, molecular composition and function. Using 3T3-L1 cells, EVs were harvested at day 0 and day 15 of differentiation. EV and cell preparations were visualised by electron microscopy and EVs quantified by nanoparticle tracking analysis (NTA. EVs were then assessed for annexin V positivity using flow cytometry; lipid and phospholipid composition using 2D thin layer chromatography and gas chromatography; and vesicular protein content by an immuno-phenotyping assay. Pre-adipogenic cells are connected via a network of protrusions and EVs at both time points display classic EV morphology. EV concentration is elevated prior to adipogenesis, particularly in exosomes and small microvesicles. Parent cells contain higher proportions of phosphatidylserine (PS and show higher annexin V binding. Both cells and EVs contain an increased proportion of arachidonic acid at day 0. PREF-1 was increased at day 0 whilst adiponectin was higher at day 15 indicating EV protein content reflects the stage of adipogenesis of the cell. Our data suggest that EV production is higher in cells before adipogenesis, particularly in vesicles <300 nm. Cells at this time point possess a greater proportion of PS (required for EV generation whilst corresponding EVs are enriched in signalling fatty acids, such as arachidonic acid, and markers of adipogenesis, such as PREF-1 and PPARγ.

  17. Study of Viability of Phosphate Solubilizing Bacteria in Phosphate granules

    hajar rajabi


    Full Text Available Introduction: sustainable development and the environment are interconnected. Sustainable agriculture is continuous utilization of a farm with respect to various aspects of environmental conditions by using fewer inputs (other than Bio-fertilizers. Phosphorus is one of the essential elements for the plants. Management of soil is possible by using biological fertilizers pillar of sustainable agriculture and providing some of the phosphorus needed by plants via bio-fertilizers. Phosphorus deficiency is extremely effective on the plant growth and productivity. The application of phosphorus fertilizers is expensive and dangerous. In addition, phosphorus in the soilmay become insoluble and will be unavailable to the plants. Studies showed that phosphate solubilizing bacteria in the soil rhizosphere are active and by root exudates solve insoluble phosphates such as tricalcium phosphate, and form absorbable P for plant. Consequently, the use of microbial fertilizers could reduce excessive use of chemical fertilizers and lead to decrease their harmful effects and protect the environment and conservation of available resources. The biological phosphate fertilizer industry uses sugar beet molasses as a binder and drying granules at high temperatures. Therefore, it is important to evaluate the durability of the bacteria in molasses at high temperature. Materials and Methods: This study was designed as completely randomized design in a factorial arrangement.10 isolates were selected and the ratios of 50%, 25%, 15% and 10% of the apatite, organic matter, sulfur and soluble granule (ratio 1: 1 and 2: 1 bacteria and molasses, respectively, for each isolate was prepared. The final product was dried at 28 and 40 °C and remained for 4 months and population counted at first day and 10, 20, 30, 60, 90 and 120 days after the preparing. The population was counted by the serial dilution technique and cultured at Sperber media. Results and Discussion:Comparing the

  18. Preparation of vesicles entrapped lycopene extract.

    Luxsuwong, Dhitaree; Indranupakorn, Ratana; Wongtrakul, Paveena


    Lycopene, a lipophilic carotenoid, has been known as an effective antioxidant in supporting the cutaneous defensive system. However, it is unstable when exposed to light and water. In this study, lycopene was isolated from tomatoes and a vesicular delivery system was developed to entrap and stabilize the lycopene in the aqueous system. A simple process, maceration in ethyl acetate, was used to extract lycopene from the tomatoes. The extract was then chromatographed on the Sephadex LH20 column using acetone as a solvent system to yield 995 μg of lycopene per gram of dried tomato weight. The vesicular delivery system was prepared from a combination of ascorbic acid-6-palmitate (AP), cholesterol and dicetyl phosphate using a thin film hydration method. The formulation was composed of AP, cholesterol and dicetyl phosphate at a 44:44:12 molar ratio and with 2.12 μmol/ml of the isolated lycopene. Both blank vesicles and lycopene loaded vesicles were kept for a period of 3 months at 4±2°C and at the room temperature (28±2°C) to evaluate the effect of the encapsulation on the characteristic of the vesicles and on the antioxidant activity of the encapsulated lycopene. The result implied that lycopene could be stabilized in the vesicles and its scavenging activity against DPPH free radicals was superior to that of the free lycopene solution.

  19. Extracellular vesicles: fundamentals and clinical relevance

    Wael Nassar


    Full Text Available All types of cells of eukaryotic organisms produce and release small nanovesicles into their extracellular environment. Early studies have described these vesicles as ′garbage bags′ only to remove obsolete cellular molecules. Valadi and colleagues, in 2007, were the first to discover the capability of circulating extracellular vesicles (EVs to horizontally transfer functioning gene information between cells. These extracellular vesicles express components responsible for angiogenesis promotion, stromal remodeling, chemoresistance, genetic exchange, and signaling pathway activation through growth factor/receptor transfer. EVs represent an important mode of intercellular communication by serving as vehicles for transfer between cells of membrane and cytosolic proteins, lipids, signaling proteins, and RNAs. They contribute to physiology and pathology, and they have a myriad of potential clinical applications in health and disease. Moreover, vesicles can pass the blood-brain barrier and may perhaps even be considered as naturally occurring liposomes. These cell-derived EVs not only represent a central mediator of the disease microenvironment, but their presence in the peripheral circulation may serve as a surrogate for disease biopsies, enabling real-time diagnosis and disease monitoring. In this review, we′ll be addressing the characteristics of different types of extracellular EVs, as well as their clinical relevance and potential as diagnostic markers, and also define therapeutic options.

  20. Vesicle dynamics in shear and capillary flows

    Noguchi, Hiroshi; Gompper, Gerhard


    The deformation of vesicles in flow is studied by a mesoscopic simulation technique, which combines multi-particle collision dynamics for the solvent with a dynamically triangulated surface model for the membrane. Shape transitions are investigated both in simple shear flows and in cylindrical capillary flows. We focus on reduced volumes, where the discocyte shape of fluid vesicles is stable, and the prolate shape is metastable. In simple shear flow at low membrane viscosity, the shear induces a transformation from discocyte to prolate with increasing shear rate, while at high membrane viscosity, the shear induces a transformation from prolate to discocyte, or tumbling motion accompanied by oscillations between these two morphologies. In capillary flow, at small flow velocities the symmetry axis of the discocyte is found not to be oriented perpendicular to the cylinder axis. With increasing flow velocity, a transition to a prolate shape occurs for fluid vesicles, while vesicles with shear-elastic membranes (like red blood cells) transform into a coaxial parachute-like shape.

  1. The Bretherton Problem for a Vesicle

    Barakat, Joseph; Spann, Andrew; Shaqfeh, Eric


    The motion of a lipid bilayer vesicle through a circular tube is investigated by singular perturbation theory in the limit of vanishing clearance. The vesicle is treated as a sac of fluid enclosed by a thin, elastic sheet that admits a bending stiffness. It is assumed that the vesicle is axisymmetric and swollen to a near-critical volume such that the clearance "e" between the membrane and the tube wall is very small. In this limit, bending resistance is of negligible importance compared to the isotropic tension, allowing the vesicle to be treated as a "no-slip bubble." The effective membrane tension is found to scale inversely with "e" raised to the 3/2 power with a comparatively weak Marangoni gradient. The extra pressure drop is found to have a leading contribution due to the cylindrical midsection, which scales inversely with "e," as well as a correction due to the end caps, which scales inversely with the square root of "e." The apparent viscosity is predicted as a unique function of the geometry. The theory exhibits excellent agreement with a simplified, "quasi-parallel" theory and with direct numerical simulations using the boundary element method. The results of this work are compared to those for bubbles, rigid particles, and red blood cells in confined flows.

  2. Vesicle Pools: Lessons from Adrenal Chromaffin Cells

    David R Stevens


    Full Text Available The adrenal chromaffin cell serves as a model system to study fast Ca2+-dependent exocytosis. Membrane capacitance measurements in combination with Ca2+ uncaging offers a temporal resolution in the millisecond range and reveals that catecholamine release occurs in three distinct phases. Release of a readily releasable (RRP and a slowly releasable (SRP pool are followed by sustained release, due to maturation and release of vesicles which were not release-ready at the start of the stimulus. Trains of depolarizations, a more physiological stimulus, induce release from a small immediately releasable pool of vesicles residing adjacent to calcium channels, as well as from the RRP. The SRP is poorly activated by depolarization. A sequential model, in which non-releasable docked vesicles are primed to a slowly releasable state, and then further mature to the readily releasable state, has been proposed. The docked state, dependent on membrane proximity, requires SNAP-25, synaptotagmin and syntaxin. The ablation or modification of SNAP-25 and syntaxin, components of the SNARE complex, as well as of synaptotagmin, the calcium sensor, and modulators such complexins and Snapin alter the properties and/or magnitudes of different phases of release, and in particular can ablate the RRP. These results indicate that the composition of the SNARE complex and its interaction with modulatory molecules drives priming and provides a molecular basis for different pools of releasable vesicles.

  3. Impact of Micellar Surfactant on Supersaturation and Insight into Solubilization Mechanisms in Supersaturated Solutions of Atazanavir.

    Indulkar, Anura S; Mo, Huaping; Gao, Yi; Raina, Shweta A; Zhang, Geoff G Z; Taylor, Lynne S


    The goals of this study were to determine: 1) the impact of surfactants on the "amorphous solubility"; 2) the thermodynamic supersaturation in the presence of surfactant micelles; 3) the mechanism of solute solubilization by surfactant micelles in supersaturated solutions. The crystalline and amorphous solubility of atazanavir was determined in the presence of varying concentrations of micellar sodium dodecyl sulfate (SDS). Flux measurements, using a side-by-side diffusion cell, were employed to determine the free and micellar-bound drug concentrations. The solubilization mechanism as a function of atazanavir concentration was probed using fluorescence spectroscopy. Pulsed gradient spin-echo proton nuclear magnetic resonance (PGSE-NMR) spectroscopy was used to determine the change in micelle size with a change in drug concentration. Changes in the micelle/water partition coefficient, K m/w , as a function of atazanavir concentration led to erroneous estimates of the supersaturation when using concentration ratios. In contrast, determining the free drug concentration using flux measurements enabled improved determination of the thermodynamic supersaturation in the presence of micelles. Fluorescence spectroscopic studies suggested that K m/w changed based on the location of atazanavir solubilization which in turn changed with concentration. Thus, at a concentration equivalent to the crystalline solubility, atazanavir is solubilized by adsorption at the micelle corona, whereas in highly supersaturated solutions it is also solubilized in the micellar core. This difference in solubilization mechanism can lead to a breakdown in the prediction of amorphous solubility in the presence of SDS as well as challenges with determining supersaturation. PGSE-NMR suggested that the size of the SDS micelle is not impacted at the crystalline solubility of the drug but increases when the drug concentration reaches the amorphous solubility, in agreement with the proposed changes in

  4. Formation of Giant Protein Vesicles by a Lipid Cosolvent Method

    Hansen, Jesper S.; Vararattanavech, Ardcharaporn; Vissing, Thomas;


    This paper describes a method to create giant protein vesicles (GPVs) of ≥10 μm by solvent‐driven fusion of large vesicles (0.1–0.2 μm) with reconstituted membrane proteins. We found that formation of GPVs proceeded from rotational mixing of protein‐reconstituted large unilamellar vesicles (LUVs...

  5. Genetically Controlled Fusion, Exocytosis and Fission of Artificial Vesicles

    Bönzli, Eva; Hadorn, Maik; De Lucrezia, Davide

    Artificial vesicles represent ideal candidates as a model for artificial cells. It was shown that artificial genetic programs and the required cellular machinery (cell-free expression systems) can be incorporated into vesicles and allow the synthesis of proteins. Vesicles were shown to fuse...

  6. Functional Characterization of ABCB4 Mutations Found in Low Phospholipid-Associated Cholelithiasis (LPAC).

    Kim, Tae Hee; Park, Hyo Jin; Choi, Ji Ha


    Multidrug resistance 3 (MDR3) is expressed on the canalicular membrane of the hepatocytes and plays an important role in protecting the liver from bile acids. Altered ABCB4 gene expression can lead to a rare hepatic disease, low phospholipid-associated cholelithiasis (LPAC). In this study, we characterized 3 ABCB4 mutations in LPAC patients using various in vitro assay systems. We first measured the ability of each mutant to transport paclitaxel and then the mechanisms by which these mutations might change MDR3 transport activity were determined using immunoblotting, cell surface protein biotinylation, and immunofluorescence. Through a membrane vesicular transport assay, we observed that the uptake of paclitaxel was significantly reduced in membrane vesicles expressing 2 ABCB4 mutations, F165I and S320F. Both mutants showed significantly decreased total and cell surface MDR3 expression. These data suggest two missense mutations of ABCB4 may alter function of MDR3 and ultimately can be determined as LPAC-causing mutations.

  7. PEG-Phospholipids Coated Quantum Rods as Amplifiers of the Photosensitization Process by FRET.

    Timor, Reut; Weitman, Hana; Waiskopf, Nir; Banin, Uri; Ehrenberg, Benjamin


    Singlet oxygen ((1)O2) generated upon photostimulation of photosensitizer molecules is a highly reactive specie which is utilized in photodynamic therapy. Recent studies have shown that semiconductor nanoparticles can be used as donors in fluorescence resonance energy transfer (FRET) process to excite attached photosensitizer molecules. In these studies, their unique properties, such as low nanoscale size, long-term photostability, wide broad absorbance band, large absorption cross section, and narrow and tunable emission bands were used to provide advantages over the traditional methods to produce singlet oxygen. Previous studies that achieved this goal, however, showed some limitations, such as low FRET efficiency, poor colloidal stability, nonspecific interactions, and/or complex preparation procedure. In this work, we developed and characterized a novel system of semiconductor quantum rods (QRs) and the photosensitizer meso-tetra(hydroxyphenyl) chlorin (mTHPC), as a model system that produces singlet oxygen without these limitations. A simple two-step preparation method is shown; Hydrophobic CdSe/CdS QRs are solubilized in aqueous solutions by encapsulation with lecithin and PEGylated phospholipid (PEG-PL) of two lipid lengths: PEG350 or PEG2000. Then, the hydrophobic photosensitizer mTHPC, was intercalated into the new amphiphilic PEG-PL coating of the QR, providing a strong attachment to the nanoparticle without covalent linkage. These PEGylated QR (eQR)-mTHPC nanocomposites show efficient FRET processes upon light stimulation of the QR component which results in efficient production of singlet oxygen. The results demonstrate the potential for future use of this concept in photodynamic therapy schemes.

  8. Potentiality of Acidithiobacillus thiooxidans in Microbial Solubilization of Phosphate Mine Tailings

    S Dhakar


    Full Text Available This paper deals with the solubilization behavior of the tailings produced by the floatation of a complex low grade phosphate ore. The composition of the tailings was essentially dolomite (52.04% with minor amounts of phosphate, iron and aluminium oxides (10.4 and 0.5% respectively. The presence of these products created uncontrolled land pollution and severely affected groundwater. An initiative has been taken up for utilization of this waste to generate an eco-friendly product. First step towards this panorama is incorporation of suitable microorganisms for the biodegradation of this effluent. Sulphur oxidizing bacteria Acidithiobacillus thiooxidans produces sulphuric acid which neutralizes the dolomitic tailings and convert it into plant available forms. The solubilization activity was tested in sulphur medium with 5, 10, 15 and 20% concentration of tailings. The solubilization is graded on the basis of pH, Electrical conductivity (EC, soluble calcium and magnesium and soluble phosphate. The results from ex-situ experiments showed that the treatment with 15% tailings ended with highest solubilization. The values of pH, EC, soluble calcium and magnesium and soluble phosphate for this treatment were 4.92, 31.6 dS/m, 10.8 mL EDTA and 17.24 µg/mL respectively. Also, the results proved that sulphur oxidizing bacteria Acidithiobacillus thiooxidans is capable of solubilizing dolomitic tailings from the Jhamarkotra mines. Finally, an important factor taken into account was solubilization of residual phosphate along with dolomite in the tailings. This combined action affects the solubilization behaviour of the residue, which was also showed successfully with the assayed laboratory studies.

  9. Mineral phosphate solubilization by wild type and radiation induced mutants of pantoea dispersa and pantoea terrae

    Murugesan, Senthilkumar; Lee, Young Keun; Kim, Jung Hun [Korea Atomic Energy Research Institte, Jeongeup (Korea, Republic of)


    Three mineral phosphate solubilizing (MPS) bacteria where isolated from rhizosphere soil samples of common bean and weed plants. 16S rDNA analysis indicated that the isolate P2 and P3 are closely related to Pantoea dispersa while isolate P4 is closely related to Pantoea terrae. Islates P2 and P3 recorded 381.60 {mu}g ml{sup -1} of tricalcium phosphate (TCP) solubilization respectively on 3 days incubation. Isolate P4 recorded the TCP solubilization of 215.85 {mu}g ml{sup -1} and the pH was dropped to 4.44 on 24 h incubation. Further incubation of P4 sharply decreased the available phosphorous to 28.94 {mu}g ml{sup -1} and pH level was raised to 6.32. Gamma radiation induced mutagenesis was carried out at LD{sub 99} dose of the wild type strains. The total of 14 mutant clones with enhanced MPS activity and 4 clones with decreased activity were selected based on solubilization index (SI) and phosphate solubilization assay. Mutant P2-M1 recorded the highest P-solubilizing potential among any other wild or mutnat clones by releasing 504.21 {mu}g ml{sup -1} of phosphorous i.e. 35% higher than its wild type by the end of day 5. A comparative evaluation of TCP solubilization by wild type isolates of Pantoea and their mutants, led to select three MPS mutant clones such as P2-M1, P3-M2 and P3-M4 with a potential to release >471.67 {mu}g ml{sup {sub {sup 1}}} of phosphorous from TCP. These over expressing mutant clones are considered as suitable candidates for biofertilization.

  10. Assessment of genetic and functional diversity of phosphate solubilizing fluorescent pseudomonads isolated from rhizospheric soil

    Narayanan Kannan


    Full Text Available Abstract Background Phosphorus is an essential macronutrient for the growth of plants. However, in most soils a large portion of phosphorus becomes insoluble and therefore, unavailable to plants. Knowledge on biodiversity of phosphate-solubilizing fluorescent pseudomonads is essential to understand their ecological role and their utilization in sustainable agriculture. Results Of 443 fluorescent pseudomonad strains tested, 80 strains (18% showed positive for the solubilization of tri-calcium phosphate (Ca3(PO42 by the formation of visible dissolution halos on Pikovskaya's agar. These phosphate solubilizing strains showed high variability in utilizing various carbon sources. Numerical taxonomy of the phosphate solubilizing strains based on their carbon source utilization profiles resulted into three major phenons at a 0.76 similarity coefficient level. Genotypic analyses of strains by BOX (bacterial repetitive BOX element-polymerase chain reaction (PCR resulted into three distinct genomic clusters and 26 distinct BOX profiles at a 80% similarity level. On the basis of phenotypic characterization and 16S rRNA gene phylogenetic analyses strains were identified as Pseudomonas aeruginosa, P. mosselii, P. monteilii, P. plecoglossicida, P. putida, P. fulva and P. fluorescens. These phosphate solubilizing strains also showed the production of plant growth promoting enzymes, hormones and exhibited antagonism against phytopathogenic fungi that attack on various crops. Gene specific primers have identified the putative antibiotic producing strains. These putative strains were grown in fermentation media and production of antibiotics was confirmed by thin layer chromatography (TLC and high performance liquid chromatography (HPLC. Conclusion Present study revealed a high degree of functional and genetic diversity among the phosphate solubilizing fluorescent pseudomonad bacteria. Due to their innate potential of producing an array of plant growth promoting

  11. Solubilization of Polycyclic Aromatic Hydrocarbons by Single and Binary Mixed Rhamnolipid-Sophorolipid Biosurfactants.

    Song, Dandan; Liang, Shengkang; Yan, Lele; Shang, Yujun; Wang, Xiuli


    Biosurfactants are promising additives for surfactant enhanced remediation (SER) technologies due to their low toxicity and high biodegradability. To develop green and efficient additives for SER, the aqueous solubility enhancements of polycyclic aromatic hydrocarbons (PAHs; naphthalene, phenanthrene, and pyrene) by rhamnolipid (RL) and sophorolipid (SL) biosurfactants were investigated in single and binary mixed systems. The solubilization capacities were quantified in terms of the solubility enhancement factor, molar solubilization ratio (MSR), and micelle-water partition coefficient (). Rughbin's model was applied to evaluate the interaction parameters (β) in the mixed RL-SL micelles. The solubility of the PAHs increased linearly with the glycolipid concentration above the critical micelle concentration (CMC) in both single and mixed systems. Binary RL-SL mixtures exhibited greater solubilization than individual glycolipids. At a SL molar fraction of 0.7 to 0.8, the solubilization capacity was the greatest, and the MSR and reached their maximum values, and β values became positive. These results suggest that the two biosurfactants act synergistically to increase the solubility of the PAHs. The solubilization capacity of the RL-SL mixtures increased with increasing temperature and decreased with increasing salinity. The aqueous solubility of phenanthrene reached a maximum value at pH of 5.5. Moreover, the mixed RL-SL systems exhibited a strong ability to solubilize PAHs, even in the presence of heavy metal ions. These mixed biosurfactant systems have the potential to improve the performance of SER technologies using biosurfactants to solubilize hydrophobic organic contaminants by decreasing the applied biosurfactant concentration, which reduces the costs of remediation.

  12. Optimized lysis buffer reagents for solubilization and preservation of proteins from cells and tissues.

    Hwang, Byeong Hee; Tsai, Kenneth Y; Mitragotri, Samir


    Reagents that facilitate solubilization of cells and tissues while preserving the biological activity of their constituents play a major role in various applications including drug delivery. Such reagents are necessary for the accurate determination of cellular and tissue concentrations of proteins, peptides, and nucleic acids, and to measure therapeutic efficacy of drug delivery technologies. Surfactant-based reagents are commonly used for this purpose; however, their utility is marred either by limited ability to solubilize or tendency to denature the proteins during solubilization. Here, we report on the screening and identification of combinations of nonionic and zwitterionic surfactants that possess excellent ability to solubilize mechanically strong and elastic tissues such as skin, while preserving its protein constituents. The leading combination, comprising an equi-mass mixture of 3-(N,N-dimethyl myristyl ammonio) propanesulfonate (TPS, CAS number:14933-09-6) and polyoxyethylene(10) cetyl ether (Brij® C10, CAS number: 9004-95-9) with a total surfactant concentration 0.5 % w/v, solubilized keratinocytes and preserved the activity of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) enzyme in its extracts at room temperature for 7 days. The ability of this mixture to preserve GAPDH activity far exceeded that of a commonly used reagent, Triton-X100. The same mixture also helped solubilize mouse skin to extract proteins and maintain detectable activity of GAPDH in the extract for 1 day. Several other mixtures of nonionic and zwitterionic surfactants were studied. These mixtures provide new reagents for solubilization of cells and tissues for research as well as technological applications.

  13. Decrease in membrane phospholipid unsaturation induces unfolded protein response.

    Ariyama, Hiroyuki; Kono, Nozomu; Matsuda, Shinji; Inoue, Takao; Arai, Hiroyuki


    Various kinds of fatty acids are distributed in membrane phospholipids in mammalian cells and tissues. The degree of fatty acid unsaturation in membrane phospholipids affects many membrane-associated functions and can be influenced by diet and by altered activities of lipid-metabolizing enzymes such as fatty acid desaturases. However, little is known about how mammalian cells respond to changes in phospholipid fatty acid composition. In this study we showed that stearoyl-CoA desaturase 1 (SCD1) knockdown increased the amount of saturated fatty acids and decreased that of monounsaturated fatty acids in phospholipids without affecting the amount or the composition of free fatty acid and induced unfolded protein response (UPR), evidenced by increased expression of C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78) mRNAs and splicing of Xbox-binding protein 1 (XBP1) mRNA. SCD1 knockdown-induced UPR was rescued by various unsaturated fatty acids and was enhanced by saturated fatty acid. Lysophosphatidylcholine acyltransferase 3 (LPCAT3), which incorporates preferentially polyunsaturated fatty acids into phosphatidylcholine, was up-regulated in SCD1 knockdown cells. Knockdown of LPCAT3 synergistically enhanced UPR with SCD1 knockdown. Finally we showed that palmitic acid-induced UPR was significantly enhanced by LPCAT3 knockdown as well as SCD1 knockdown. These results suggest that a decrease in membrane phospholipid unsaturation induces UPR.

  14. Phospholipid dynamics in graphene of different topologies: predictive modeling

    Glukhova, O. E.; Slepchenkov, M. M.


    The subject of our scientific interest is the dynamics of the phospholipid molecules into a corrugated graphene sheet. According to our assumption by changing the topology of graphene properly it is possible to find the ways for management of the selective localization of phospholipid molecules to form the desired configuration of these structures. We considered DPPC (dipalmitoylphosphatidylcholine) phospholipids, which are the part of cell membranes and lipoproteins. We investigated the behavior of the phospholipids on the graphene sheet consisting of 1710 atoms with the size of 6.9 nm along the zigzag edge and 6.25 nm along the armchair edge. The numerical experiment was carried out using the original AMBER/AIREBO hybrid method with Lennard-Jones potential to describe the interaction between unbound atoms of different structures. The temperature was maintained at 300 K during the numerical experiment. All numerical experiments were performed using KVAZAR software system. We considered several cases of corrugated graphene with different width and dept of the corrugation. Special attention in our work was paid to the orientation of the phospholipids in the plane of graphene sheet.

  15. Erythrocyte phospholipid and polyunsaturated fatty acid composition in diabetic retinopathy.

    Philippe Koehrer

    Full Text Available Long chain polyunsaturated fatty acids (LCPUFAs including docosahexaenoic acid and arachidonic acid are suspected to play a key role in the pathogenesis of diabetes. LCPUFAs are known to be preferentially concentrated in specific phospholipids termed as plasmalogens. This study was aimed to highlight potential changes in the metabolism of phospholipids, and particularly plasmalogens, and LCPUFAs at various stages of diabetic retinopathy in humans.We performed lipidomic analyses on red blood cell membranes from controls and mainly type 2 diabetes mellitus patients with or without retinopathy. The fatty acid composition of erythrocytes was determined by gas chromatography and the phospholipid structure was determined by liquid chromatography equipped with an electrospray ionisation source and coupled with a tandem mass spectrometer (LC-ESI-MS/MS. A significant decrease in levels of docosahexaenoic acid and arachidonic acid in erythrocytes of diabetic patients with or without retinopathy was observed. The origin of this decrease was a loss of phosphatidyl-ethanolamine phospholipids esterified with these LCPUFAs. In diabetic patients without retinopathy, this change was balanced by an increase in the levels of several phosphatidyl-choline species. No influence of diabetes nor of diabetic retinopathy was observed on the concentrations of plasmalogen-type phospholipids.Diabetes and diabetic retinopathy were associated with a reduction of erythrocyte LCPUFAs in phosphatidyl-ethanolamines. The increase of the amounts of phosphatidyl-choline species in erythrocytes of diabetic patients without diabetic retinopathy might be a compensatory mechanism for the loss of LC-PUFA-rich phosphatidyl-ethanolamines.

  16. Isolation and phosphate-solubilizing ability of a fungus, Penicillium sp. from soil of an alum mine.

    Chai, Bo; Wu, Yan; Liu, Pengming; Liu, Biao; Gao, Meiying


    The use of microorganisms to solubilize elemental phosphorus from insoluble rock phosphate is a promising method to greatly reduce not only environmental pollution but also production costs. Phosphate-solubilizing microorganisms were isolated from soils in China, and a fungus strain (PSM11-5) from a soil sample from an alum mine, with the highest phosphate solubilization potential, was selected and identified as a Penicillium sp. Strain PSM11-5 could grow in buffered medium with pH values between 3.0 and 8.0 and showed phosphate solubilizing activity at pH values from 5.0 to 8.0. It also exhibited a degree of tolerance to the heavy metal ions, Cd(2+), Co(2+), and Cr(6+). PSM11-5 could rapidly solubilize tricalcium phosphate, and a high phosphate-solubilizing efficiency of 98% was achieved in an optimized medium. The strain could solubilize rock phosphate and aluminum phosphate with a solubilizing efficiency of approximately 74.5%, but did not solubilize iron phosphate. Solubilization of phosphate depended on acidification. Analysis of PSM11-5 culture supernatants by capillary electrophoresis showed that tricalcium phosphate was solubilized to PO(4) (3-) and Ca(2+) , and that the organic acid produced by the fungus was mainly gluconic acid at approximately ca. 13 g l(-1). In addition, PSM11-5 produced ca. 830 mg l(-1) of citric acid when it was used to solubilize rock phosphate. These excellent properties of strain PSM11-5 suggest that the fungus has potential for agricultural and industrial utilization.

  17. Phosphate solubilization and multiple plant growth promoting properties of rhizobacteria isolated from chickpea (Cicer aeritinum L.) producing areas of Ethiopia

    Midekssa, Mulissa J.; Löscher, Carolin; Ruth A Schmitz; Assefa, Fassil


    Chickpea is one of the major legume crops widely grown in Ethiopia. The low availability of phosphorus in soil is among the stresses that constrain the production of this crop in the country. However, there are rhizobacteria capable of solubilizing insoluble forms of phosphorus in soil and make it available to the plant. Thus, this study was aimed at isolation and characterization of phosphate solubilizing bacteria from chickpea rhizosphere. Fifty phosphate solubilizing bacterial strains were...

  18. Salt-free vesicle-phases and their template effect


    Researches on the construction, structure, and formation of vesicles formed from surfactants have attracted great attention from colloid and interface chemists. The vesicles formed from salt-free cationic-anionic surfactant systems are very different from those with excess salts, having many particular properties. In this paper, we introduce the properties of vesicles prepared from salt-free surfactant systems, according to our own results, especially the vesicles formed from surfactants with divalent metal ions as counterions in aqueous solutions and room temperature ionic liquids. Moreover, the primary results on template effect of the metal-ligand vesicles have also been summarized.

  19. Interaction of insulin with SDS/CTAB catanionic Vesicles

    Tah, Bidisha; Pal, Prabir; Talapatra, G.B., E-mail:


    In the present study, a novel method was used for entrapping the protein, insulin into the catanionic SDS/CTAB vesicle membrane. The anionic SDS and cationic CTAB formed catanionic vesicles at particular concentration (35:65 by volume). In this study, vesicle membrane can be considered as model membrane. The vesicle formation and entrapment efficiency depend on the pH of the aqueous solution. The insulin molecules have attached with the vesicular membrane at pH 7.0. However, at acidic pH, the vesicles were ruptured and the insulin did not entrap into the vesicle membrane, whereas at alkaline pH insulin became fibriller. The scanning electron microscope (SEM), Dynamic light scattering (DLS), and Zeta potential studies established the self-assembled structure formation of insulin and catanionic vesicles. To know the protein confirmations, Circular dichroism (CD) was also employed. The temperature dependent steady state and time resolved emission spectroscopy show that at room temperature (25 °C), apart from the 305 nm tyrosine fluorescence, a new emission peak at 450 nm was observed only in case of insulin-vesicle system, and was assigned as the tyrosine phosphorescence. This phosphorescence peak is the signature of the entrapment of insulin into the vesicle membrane. Highlights: • SDS-CTAB based catanionic vesicle has been fabricated. • Insulin has been successfully immobilized on these vesicles. • Immobilized insulin shows room temperature phosphorescence.

  20. Bio-liquefaction/solubilization of low-rank Turkish lignites and characterization of the products

    Yesim Basaran; Adil Denizli; Billur Sakintuna; Alpay Taralp; Yuda Yurum [Hacettepe University, Ankara (Turkey). Department of Environmental Sciences


    The effect of some white-rot fungi on the bio-liquefaction/solubilization of two low-rank Turkish coals and the chemical composition of the liquid products and the microbial mechanisms of coal conversion were investigated. Turkish Elbistan and Beypazari lignites were used in this study. The white-rot fungi received from various laboratories used in the bio-liquefaction/solubilization of the lignites were Pleurotus sajor-caju, Pleurotus sapidus, Pleurotus florida, Pleurotus ostreatus, Phanerochaete chrysosporium, and Coriolus versicolor. FT-IR spectra of raw and treated coal samples were measured, and bio-liquefied/solubilized coal samples were investigated by FT-IR and LC-MS techniques. The Coriolus versicolor fungus was determined to be most effective in bio-liquefying/solubilizing nitric acid-treated Elbistan lignite. In contrast, raw and nitric acid-treated Beypazari lignite seemed to be unaffected by the action of any kind of white-rot fungi. The liquid chromatogram of the water-soluble bio-liquefied/solubilized product contained four major peaks. Corresponding mass spectra of each peak indicated the presence of very complicated structures. 17 refs., 9 figs., 2 tabs.

  1. Phosphate solubilizing ability of Emericella nidulans strain V1 isolated from vermicompost.

    Bhattacharya, Satya Sunder; Barman, Soma; Ghosh, Ranjan; Duary, Raj Kumar; Goswami, Linee; Mandal, Narayan C


    Phosphorus is one of the key factors that regulate soil fertility. Its deficiencies in soil are largely replenished by chemical fertilizers. The present study was aimed to isolate efficient phosphate solubilizing fungal strains from Eisenia fetida vermicompost. Out of total 30 fungal strains the most efficient phosphate solubilizing one was Emericella (Aspergillus) nidulans V1 (MTCC 11044), identified by custom sequencing of beta-tubulin gene and BLAST analysis. This strain solubilized 13 to 36% phosphate from four different rock phosphates. After three days of incubation of isolated culture with black Mussorie phosphate rock, the highest percentage of phosphate solubilization was 35.5 +/- 1.01 with a pH drop of 4.2 +/- 0.09. Kinetics of solubilization and acid production showed a linear relationship until day five of incubation. Interestingly, from zero to tenth day of incubation, solubility of soil phosphate increased gradually from 4.31 +/- 1.57 to 13.65 +/- 1.82 (mg kg(-1)) recording a maximum of 21.23 +/- 0.54 on day 45 in respect of the V1 isolate. Further, enhanced phosphorus uptake by Phaseolus plants with significant pod yield due to soil inoculation of Emericella nidulans V1 (MTCC 11044), demonstrated its prospect as an effective biofertilizer for plant growth.

  2. Solubilization of thiamine triphosphatase from the electric organ of Electrophorus electricus.

    Bettendorff, L; Longrée, I; Wins, P; Schoffeniels, E


    The membrane-associated, anion-regulated thiamine triphosphatase from Electrophorus electricus electric organ can be solubilized by various neutral detergents. Polyoxyethylene ethers are the most effective. Anionic detergents readily inactivate the enzyme. A 6.4-fold increase in specific activity is obtained by successive treatment of crude membranes with octanoyl-N-methylglucamide, which solubilized other proteins, and Lubrol-PX with releases 60% of the thiamine triphosphatase (TTPase) activity. Solubilization by Lubrol-PX rapidly modifies kinetic parameters. The Km, Vmax and pH optimum are decreased. However, the solubilized TTPase may be kept at 0 degrees C for many hours without further change in specific activity. At 35 degrees C, the half-life is still 83 min at pH 5.0, but denaturation becomes rapid at pH greater than or equal to 7. Solubilization modifies anion effects on TTPase activity. The activating effect of nitrate is nearly lost, while inhibition by sulfate is no longer time-dependent.

  3. Solubilization of Rock Phosphate in Liquid Culture by Fungal Isolates from Rhizosphere Soil

    WANG Guang-Hua; ZHOU De-Rui; YANG Qian; JIN Jian; LIU Xiao-Bing


    Rock phosphate (RP) is a low efficiency P fertilizer that is directly applied to the soil and can be solubilized by phosphate-solubilizing microorganisms (PSMs) in fermentation or soil conditions. This study investigated dynamic solubilization of 2 concentrations of rock phosphate in a liquid culture with different dosages of glucose by two fungal isolates,Aspergillus niger P39 and Penicillium oxalicum P66, from soybean and wheat rhizosphere soil. Although during the 20day culture period A. niger P39 had a stronger ability to acidify the culture media than P. oxalicum P66, soluble P concentrations at glucose dosages of 30 and 50 gL-1 with RP of 15 g L-1 in the culture solution were much higher by P. oxalicum P66. The greater effectiveness of P. oxalicum P66 compared to A. niger P39 in the solubilization of RP was strongly associated with the production of organic acids. This study suggested that for RP solubilization the type rather than the concentration of PSM-produced organic acids was more important.

  4. Solubilization properties of polycyclic aromatic hydrocarbons by saponin, a plant-derived biosurfactant.

    Zhou, Wenjun; Yang, Juanjuan; Lou, Linjie; Zhu, Lizhong


    The enhanced solubilization of polycyclic aromatic hydrocarbons (PAHs) by saponin, a plant-derived non-ionic biosurfactant, was investigated. The results indicated that the solubilization capabilities of saponin for PAHs were greater than some representative synthetic non-ionic surfactants and showed strong dependence on solution pH and ionic strength. The molar solubilization ratio (MSR) of saponin for phenanthrene was about 3-6 times of those of the synthetic non-ionic surfactants, and decreased by about 70% with the increase of solution pH from 4.0 to 8.0, but increased by approximately 1 times with NaCl concentration increased from 0.01 to 1.0 M. Heavy metal ions can enhance saponin solubilization for phenanthrene and the corresponding MSR values increased by about 25% with the presence of 0.01 M of Cd2+ or Zn2+. Saponin is more effective in enhancing PAHs solubilization than synthetic non-ionic surfactants and has potential application in removing organic pollutants from contaminated soils.

  5. Enhancing uranium solubilization in soils by citrate, EDTA, and EDDS chelating amendments.

    Lozano, J C; Blanco Rodríguez, P; Tomé, F Vera; Calvo, C Prieto


    A systematic study was made of the effects of three soil amendments on the solubilization of uranium from a granitic soil. The aim was to optimize solubilization so as to enhance bioavailability for the purposes of remediation. The three amendments tested were with citrate, EDTA, and EDDS as chelating agents. The effects of pH, chelator concentration, and leaching time were studied. The most important factor in uranium solubilization was found to be the pH. In the absence of chelating agents, the greatest solubilization was obtained for alkaline conditions, with values representing about 15% of the total uranium activity in the bulk soil. There were major differences in uranium solubilization between the different amendments. The citrate treatment was the most efficient at acidic pH, particularly with the greatest concentration of citrate tested (50 mmol kg(-1)) after 6 days of treatment. Under these conditions, the uranium concentration in solution was greater by a factor of 356 than in the control suspension, and represented some 63% of the uranium concentration in the bulk soil. Under alkaline conditions, the EDTA and EDDS treatments gave the greatest uranium activity concentrations in solution, but these concentrations were much lower than those with the citrate amendment, and were not very different from the control results. The uranium extraction yield with EDDS amendment was greater than with EDTA.

  6. Microbial Phosphorus Solubilization and Its Potential for Use in Sustainable Agriculture

    Alori, Elizabeth T.; Glick, Bernard R.; Babalola, Olubukola O.


    The use of excess conventional Phosphorus (P) fertilizers to improve agricultural productivity, in order to meet constantly increasing global food demand, potentially causes surface and ground water pollution, waterway eutrophication, soil fertility depletion, and accumulation of toxic elements such as high concentration of selenium (Se), arsenic (As) in the soil. Quite a number of soil microorganisms are capable of solubilizing/mineralizing insoluble soil phosphate to release soluble P and making it available to plants. These microorganisms improve the growth and yield of a wide variety of crops. Thus, inoculating seeds/crops/soil with Phosphate Solubilizing Microorganisms (PSM) is a promising strategy to improve world food production without causing any environmental hazard. Despite their great significance in soil fertility improvement, phosphorus-solubilizing microorganisms have yet to replace conventional chemical fertilizers in commercial agriculture. A better understanding of recent developments in PSM functional diversity, colonizing ability, mode of actions and judicious application should facilitate their use as reliable components of sustainable agricultural systems. In this review, we discussed various soil microorganisms that have the ability to solubilize phosphorus and hence have the potential to be used as bio fertilizers. The mechanisms of inorganic phosphate solubilization by PSM and the mechanisms of organic phosphorus mineralization are highlighted together with some factors that determine the success of this technology. Finally we provide some indications that the use of PSM will promote sustainable agriculture and conclude that this technology is ready for commercial exploitation in various regions worldwide. PMID:28626450

  7. Phosphorus solubilization and plant growth enhancement by arsenic-resistant bacteria.

    Ghosh, Piyasa; Rathinasabapathi, Bala; Ma, Lena Q


    Phosphorus is an essential nutrient, which is limited in most soils. The P solubilization and growth enhancement ability of seven arsenic-resistant bacteria (ARB), which were isolated from arsenic hyperaccumulator Pteris vittata, was investigated. Siderophore-producing ARB (PG4, 5, 6, 9, 10, 12 and 16) were effective in solubilizing P from inorganic minerals FePO4 and phosphate rock, and organic phytate. To reduce bacterial P uptake we used filter-sterilized Hoagland medium containing siderophores or phytase produced by PG12 or PG6 to grow tomato plants supplied with FePO4 or phytate. To confirm that siderophores were responsible for P release, we compared the mutants of siderophore-producing bacterium Pseudomonas fluorescens Pf5 (PchA) impaired in siderophore production with the wild type and test strains. After 7d of growth, mutant PchA solubilized 10-times less P than strain PG12, which increased tomato root biomass by 1.7 times. For phytate solubilization by PG6, tomato shoot biomass increased by 44% than control bacterium Pseudomonas chlororaphis. P solubilization by ARB from P. vittata may be useful in enhancing plant growth and nutrition in other crop plants.

  8. Effect of acid and alkaline solubilization on the properties of surimi based film

    Thummanoon Prodpran


    Full Text Available The effect of acid and alkaline solubilizing processes on the properties of the protein based film from threadfin bream surimi was investigated. Surimi films prepared from both processes had the similar light transmission, tensile strength (TS and elongation at break (EAB (P<0.05. However, film with alkaline process had slightly lower water vapor permeability (WVP, compared to that prepared by acid solubilizing process. The protein concentration in the film-forming solution directly affected the properties of the film. Increase in protein concentration resulted in an increase in TS, EAB as well as WVP. The film prepared by acid solubilizing process had an increase in yellowish color as evidenced by the continuous increase in b* and E* values during the storage at r oom temperature. The acid and alkali solubilizing processes caused the degradation of muscle protein in surimi, especially with increasing exposure time. Therefore, solubilizing process had the influence on the properties of the protein film from threadfin bream surimi.

  9. Microbial Phosphorus Solubilization and Its Potential for Use in Sustainable Agriculture

    Elizabeth T. Alori


    Full Text Available The use of excess conventional Phosphorus (P fertilizers to improve agricultural productivity, in order to meet constantly increasing global food demand, potentially causes surface and ground water pollution, waterway eutrophication, soil fertility depletion, and accumulation of toxic elements such as high concentration of selenium (Se, arsenic (As in the soil. Quite a number of soil microorganisms are capable of solubilizing/mineralizing insoluble soil phosphate to release soluble P and making it available to plants. These microorganisms improve the growth and yield of a wide variety of crops. Thus, inoculating seeds/crops/soil with Phosphate Solubilizing Microorganisms (PSM is a promising strategy to improve world food production without causing any environmental hazard. Despite their great significance in soil fertility improvement, phosphorus-solubilizing microorganisms have yet to replace conventional chemical fertilizers in commercial agriculture. A better understanding of recent developments in PSM functional diversity, colonizing ability, mode of actions and judicious application should facilitate their use as reliable components of sustainable agricultural systems. In this review, we discussed various soil microorganisms that have the ability to solubilize phosphorus and hence have the potential to be used as bio fertilizers. The mechanisms of inorganic phosphate solubilization by PSM and the mechanisms of organic phosphorus mineralization are highlighted together with some factors that determine the success of this technology. Finally we provide some indications that the use of PSM will promote sustainable agriculture and conclude that this technology is ready for commercial exploitation in various regions worldwide.

  10. Solubilization of octane in electrostatically-formed surfactant-polymer complexes.

    Zhang, Hui; Zeeb, Benjamin; Salminen, Hanna; Feng, Fengqin; Weiss, Jochen


    Polymers can be used to modulate the stability and functionality of surfactant micelles. The purpose of this study was to investigate the solubilization of an octane oil-in-water emulsion in mixtures of an anionic polymer (carboxymethyl cellulose) and anionic sodium dodecylsulphate (SDS), nonionic polyoxyethylene sorbitan monooleate (Tween 80) and cationic cetyltrimethylammonium bromide (CTAB) surfactant micelles using dynamic light scattering, microelectrophoresis and turbidity measurements. The results showed that the addition of anionic carboxymethyl cellulose accelerated octane solubilization in cationic CTAB and CTAB-Tween 80 micelles, but did not affect the solubilization behaviors of micelles that were nonionic and anionic. The surfactant-polymer interactions were also studied using isothermal titration calorimetry (ITC) to characterize different physiochemical interaction regions depending on surfactant concentration in surfactant-polymer systems. Upon octane solubilization in CTAB-carboxymethyl cellulose mixtures, shape transitions of polymer-micelle complexes may have taken place that altered light scattering behavior. Based on these results, we suggest a mechanism for oil solubilization in electrostatically-formed surfactant-polymer complexes.

  11. Thermo-tolerant phosphate-solubilizing microbes for multi-functional biofertilizer preparation.

    Chang, Cheng-Hsiung; Yang, Shang-Shyng


    In order to prepare the multi-functional biofertilizer, thermo-tolerant phosphate-solubilizing microbes including bacteria, actinomycetes, and fungi were isolated from different compost plants and biofertilizers. Except Streptomycesthermophilus J57 which lacked pectinase, all isolates possessed amylase, CMCase, chitinase, pectinase, protease, lipase, and nitrogenase activities. All isolates could solubilize calcium phosphate and Israel rock phosphate; various isolates could solubilize aluminum phosphate, iron phosphate, and hydroxyapatite. During composting, biofertilizers inoculated with the tested microbes had a significantly higher temperature, ash content, pH, total nitrogen, soluble phosphorus content, and germination rate than non-inoculated biofertilizer; total organic carbon and carbon-to-nitrogen ratio showed the opposite pattern. Adding these microbes can shorten the period of maturity, improve the quality, increase the soluble phosphorus content, and enhance the populations of phosphate-solubilizing and proteolytic microbes in biofertilizers. Therefore, inoculating thermo-tolerant phosphate-solubilizing microbes into agricultural and animal wastes represents a practical strategy for preparing multi-functional biofertilizer.

  12. Ultrastructural and functional fate of recycled vesicles in hippocampal synapses.

    Rey, Stephanie A; Smith, Catherine A; Fowler, Milena W; Crawford, Freya; Burden, Jemima J; Staras, Kevin


    Efficient recycling of synaptic vesicles is thought to be critical for sustained information transfer at central terminals. However, the specific contribution that retrieved vesicles make to future transmission events remains unclear. Here we exploit fluorescence and time-stamped electron microscopy to track the functional and positional fate of vesicles endocytosed after readily releasable pool (RRP) stimulation in rat hippocampal synapses. We show that most vesicles are recovered near the active zone but subsequently take up random positions in the cluster, without preferential bias for future use. These vesicles non-selectively queue, advancing towards the release site with further stimulation in an actin-dependent manner. Nonetheless, the small subset of vesicles retrieved recently in the stimulus train persist nearer the active zone and exhibit more privileged use in the next RRP. Our findings reveal heterogeneity in vesicle fate based on nanoscale position and timing rules, providing new insights into the origins of future pool constitution.

  13. [Tracheal phospholipid composition and respiratory distress syndrome of the newborn].

    Obladen, M


    Tracheal or pharyngeal aspirates were collected in 50 newborn infants with and without respiratory distress syndrome (RDS). After lipid extraction the phospholipids were analyzed with 2-dimensional thin layer chromatography. Surface-active are lecithin (PC), phosphatidylglycerol (PG), and phosphatidylinositol (PI). Newborn infants with RDS always have a complete lack of PG, which makes up to 11% of phospholipid-phosphors in mature newborns. In all infants with and without RDS, a sharp increase of PC occurs in the lung effluent after birth. The recovery from RDS is characterized by marked changes of PI: this phospholipid rises up to twice its initial value if the infants survive. The PI-increase parallels the clinical improvement and reaches its maximum usually on the 5th day of life. At the time of the PI-peak, the infants' surfactant function is sufficient to maintain alveolar stability with spontaneous breathing. In infants dying from RDS the PI-increase was not observed.

  14. Possible mechanism of adhesion in a mica supported phospholipid bilayer

    Pertsin, Alexander, E-mail: [Angewandte Physikalische Chemie, Universität Heidelberg, Im Neuenheimer Feld 253, 69120 Heidelberg (Germany); Institute of Organo-Element Compounds, Russian Academy of Sciences, Vavilov Str. 28, 117991 Moscow (Russian Federation); Grunze, Michael [Angewandte Physikalische Chemie, Universität Heidelberg, Im Neuenheimer Feld 253, 69120 Heidelberg (Germany); Institute for Functional Interfaces, Karlsruhe Institute of Technology, Hermann-von- Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany)


    Phospholipid bilayers supported on hydrophilic solids like silica and mica play a substantial role in fundamental studies and technological applications of phospholipid membranes. In both cases the molecular mechanism of adhesion between the bilayer and the support is of primary interest. Since the possibilities of experimental methods in this specific area are rather limited, the methods of computer simulation acquire great importance. In this paper we use the grand canonical Monte Carlo technique and an atomistic force field to simulate the behavior of a mica supported phospholipid bilayer in pure water as a function of the distance between the bilayer and the support. The simulation reveals a possible adhesion mechanism, where the adhesion is due to individual lipid molecules that protrude from the bilayer and form widely spaced links with the support. Simultaneously, the bilayer remains separated from the bilayer by a thin water interlayer which maintains the bilayer fluidity.

  15. Neutron diffraction studies of amphipathic helices in phospholipid bilayers

    Bradshaw, J.P.; Gilchrist, P.J. [Univ. of Edinburgh (United Kingdom); Duff, K.C. [Univ. of Edinburgh Medical School (United Kingdom); Saxena, A.M. [Brookhaven National Laboratory, Upton, NY (United States)


    The structural feature which is thought to facilitate the interaction of many peptides with phospholipid bilayers is the ability to fold into an amphipathic helix. In most cases the exact location and orientation of this helix with respect to the membrane is not known, and may vary with factors such as pH and phospholipid content of the bilayer. The growing interest in this area is stimulated by indications that similar interactions can contribute to the binding of certain hormones to their cell-surface receptors. We have been using the techniques of neutron diffraction from stacked phospholipid bilayers in an attempt to investigate this phenomenon with a number of membrane-active peptides. Here we report some of our findings with three of these: the bee venom melittin; the hormone calcitonin; and a synthetic peptide representing the ion channel fragment of influenza A M2 protein.

  16. Docking of Secretory Vesicles Is Syntaxin Dependent

    de Wit, Heidi; Cornelisse, L. Niels; Toonen, Ruud F.G.; Verhage, Matthijs


    Secretory vesicles dock at the plasma membrane before they undergo fusion. Molecular docking mechanisms are poorly defined but believed to be independent of SNARE proteins. Here, we challenged this hypothesis by acute deletion of the target SNARE, syntaxin, in vertebrate neurons and neuroendocrine cells. Deletion resulted in fusion arrest in both systems. No docking defects were observed in synapses, in line with previous observations. However, a drastic reduction in morphologically docked secretory vesicles was observed in chromaffin cells. Syntaxin-deficient chromaffin cells showed a small reduction in total and plasma membrane staining for the docking factor Munc18-1, which appears insufficient to explain the drastic reduction in docking. The sub-membrane cortical actin network was unaffected by syntaxin deletion. These observations expose a docking role for syntaxin in the neuroendocrine system. Additional layers of regulation may have evolved to make syntaxin redundant for docking in highly specialized systems like synaptic active zones. PMID:17205130

  17. Pulsatile lipid vesicles under osmotic stress

    Chabanon, Morgan; Liedberg, Bo; Parikh, Atul N; Rangamani, Padmini


    The response of lipid bilayers to osmotic stress is an important part of cellular function. Previously, in [Oglecka et al. 2014], we reported that cell-sized giant unilamellar vesicles (GUVs) exposed to hypotonic media, respond to the osmotic assault by undergoing a cyclical sequence of swelling and bursting events, coupled to the membrane's compositional degrees of freedom. Here, we seek to deepen our quantitative understanding of the essential pulsatile behavior of GUVs under hypotonic conditions, by advancing a comprehensive theoretical model for vesicle dynamics. The model quantitatively captures our experimentally measured swell-burst parameters for single-component GUVs, and reveals that thermal fluctuations enable rate dependent pore nucleation, driving the dynamics of the swell-burst cycles. We further identify new scaling relationships between the pulsatile dynamics and GUV properties. Our findings provide a fundamental framework that has the potential to guide future investigations on the non-equili...

  18. Docking of secretory vesicles is syntaxin dependent.

    Heidi de Wit

    Full Text Available Secretory vesicles dock at the plasma membrane before they undergo fusion. Molecular docking mechanisms are poorly defined but believed to be independent of SNARE proteins. Here, we challenged this hypothesis by acute deletion of the target SNARE, syntaxin, in vertebrate neurons and neuroendocrine cells. Deletion resulted in fusion arrest in both systems. No docking defects were observed in synapses, in line with previous observations. However, a drastic reduction in morphologically docked secretory vesicles was observed in chromaffin cells. Syntaxin-deficient chromaffin cells showed a small reduction in total and plasma membrane staining for the docking factor Munc18-1, which appears insufficient to explain the drastic reduction in docking. The sub-membrane cortical actin network was unaffected by syntaxin deletion. These observations expose a docking role for syntaxin in the neuroendocrine system. Additional layers of regulation may have evolved to make syntaxin redundant for docking in highly specialized systems like synaptic active zones.

  19. NMR analyses of deuterated phospholipids isolated from Pichia angusta

    Massou, S.; Augé, S.; Tropis, M.; Lindley, N. D.; Milon, A.


    The phospholipid composition of methylotrophic yeasts grown on deuterated and hydrogenated media has been determined by proton and phosphorus NMR. By using a line narrowing solvent, we could obtain linewidth lower than 2 Hz, and all the resonances could be resolved. Phospholipids were identified on the basis of their chemical shift and by 31P - H correlations (HMQC - HOHAHA gradient enhanced experiments). We have thus analysed qualitatively and quantitatively lipids mixtures directly after chloroform-methanol extraction. The lipid composition is deeply modified after growth in deuterated medium were phosphatidyl Inositol (PI) becomes the major lipid, instead of a PC, PS, PI mixture in hydrogenated conditions. La composition en phospholipides de levures méthylotrophes ayant poussé sur des milieux de cultures hydrogénés et deutériés a été déterminée par RMN du proton et du phosphore31. L'utilisation d'un solvant d'affinement a permis d'obtenir des largeurs de raies inférieures à 2Hz et de résoudre toutes les classes de phospholipides. Ils sont ensuite identifiés par leur déplacement chimique et par des corrélations phosphore - proton spécifiques (expériences HMQC-HOHAHA gradients). Cette approche a permis une analyse qualitative et quantitative de mélanges de phospholipides directement après extraction au chloroforme-méthanol. La composition en phospholipides est profondément modifiée lors de la croissance en milieu perdeutérié où l'on observe un lipide majoritaire, le phosphatidyl Inositol (PI), au lieu d'un mélange PC, PS PI en milieu hydrogéné.

  20. Fat lowers fat: purified phospholipids as emerging therapies for dyslipidemia.

    Sahebkar, Amirhossein


    Dyslipidemia is a major coronary heart disease (CHD) risk factor. In spite of the proven efficacy of statin drugs in reducing CHD burden, there is still much room for the discovery of novel therapeutic agents to address the considerable residual cardiovascular risk that remains after treatment with currently available medications. In particular, there is an urgent demand for drugs capable of boosting the concentration and/or function of high-density lipoprotein (HDL) and apolipoprotein A-I (apo A-I), thereby promoting reverse cholesterol transport. Phospholipids are naturally occurring fats that play indispensible role in human health via their structural, energy storage, signal transduction and metabolic functions. Supplementation with either purified or mixed preparations of bioactive phospholipids has been reported to ameliorate a range of nutritional and cardiovascular disorders. Moreover, several lines of evidence have supported the efficacy of dietary phospholipids in reducing serum and hepatic contents of cholesterol and triglycerides, while increasing HDL-C and apo A-I levels. These beneficial effects of phospholipids could be attributed to their ability in reducing intestinal cholesterol absorption, enhancing biliary cholesterol excretion and modulating the expression and activity of transcriptional factors and enzymes that are involved in lipoprotein metabolism. Given their extreme safety and biocompatibility, dietary supplementation with phospholipid preparations, in particular phosphatidylinositol, appears as a novel and effective strategy that could be used as an alternative or adjunctive therapy to the current medications. The present review outlines the in-vitro, in-vivo and clinical findings on the anti-dyslipidemic effects of three most abundant phospholipids in the human body and diet namely phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol.