On the microstructures of ethyl acetate/monoolein/water

A phase diagram, describing the behavior of the polar lipid monoolein (MO), water, and ethyl acetate (EtAc), is here presented as well as results from small angle X-ray scattering. MO is found to have a solubility of 60 wt.% in EtAc at 20 °C. No macroscopic aggregation of MO can, initially, be detected in the binary MO/EtAc solution even though MO forms solid crystals in concentrated samples when times goes by. In case of the ternary system small amounts of water, mainly bound to the lipid head groups, can be incorporated in the liquid EtAc/MO phase as water has a limited solubility in EtAc. For EtAc/water mass ratios below 2/3 EtAc is present into the reversed bicontinuous cubic and lamellar phases present in the binary MO/water system. To conclude, EtAc is mainly partitioned to the lipid membranes, with minor effects on spontaneous curvature. Hence, simple EtAc-addition has an effect similar to dehydration. For EtAc/water ratios above 2/3 the liquid crystalline phases dissolve. The phase behavior is here discussed in view of related phase behaviors for water-miscible solvent/MO/water systems. For instance, an interpretation of the swelling behavior of the sponge phase (L₃), present in the water-miscible solvent(s)/MO/water systems, shows that solvents partitioned to the polar domains strongly increases the spontaneous curvature of the MO-films. The reason is probably weaker hydrophobic interactions in interfacial regions. As expected, in case of water-miscible solvents, the ternary phase behaviors can be understood by consider water and water-miscible solvent as one “mixed solvent”.     

On the microstructures of ethyl acetate/monoolein/water

A phase diagram, describing the behavior of the polar lipid monoolein (MO), water, and ethyl acetate (EtAc), is here presented as well as results from small angle X-ray scattering. MO is found to have a solubility of 60 wt.% in EtAc at 20 °C. No macroscopic aggregation of MO can, initially, be detected in the binary MO/EtAc solution even though MO forms solid crystals in concentrated samples when times goes by. In case of the ternary system small amounts of water, mainly bound to the lipid head groups, can be incorporated in the liquid EtAc/MO phase as water has a limited solubility in EtAc. For EtAc/water mass ratios below 2/3 EtAc is present into the reversed bicontinuous cubic and lamellar phases present in the binary MO/water system. To conclude, EtAc is mainly partitioned to the lipid membranes, with minor effects on spontaneous curvature. Hence, simple EtAc-addition has an effect similar to dehydration. For EtAc/water ratios above 2/3 the liquid crystalline phases dissolve. The phase behavior is here discussed in view of related phase behaviors for water-miscible solvent/MO/water systems. For instance, an interpretation of the swelling behavior of the sponge phase (L₃), present in the water-miscible solvent(s)/MO/water systems, shows that solvents partitioned to the polar domains strongly increases the spontaneous curvature of the MO-films. The reason is probably weaker hydrophobic interactions in interfacial regions. As expected, in case of water-miscible solvents, the ternary phase behaviors can be understood by consider water and water-miscible solvent as one “mixed solvent”.     

Interlamellar forces and the thermodynamic characterization of lamellar phospholipid systems

In this review, we summarize a series of experimental studies of the swelling of zwitterionic lamellar phospholipid and phospholipid-cholesterol systems using a novel double twin calorimeter. With this method, one can obtain simultaneous measures of the partial molar free energy and the partial molar enthalpy, and the experimental studies thus provide a complete thermodynamic characterization of the isothermal swelling process. A major finding is that the swelling of lamellar zwitterionic phospholipid systems at higher water contents (> 4 water molecules per lipid) is endothermic. The enthalpy has the opposite sign relative to the free energy, thus demonstrating that the swelling process is entropy driven. The water uptake also triggers a transition from a gel to a liquid crystalline state showing that, at given water content, the swelling pressure is much higher in the liquid crystal than in the gel. When cholesterol is added to the system the liquid ordered phase is formed at all available water contents. In this phase the swelling pressure varies smoothly and takes relatively low values at water contents below two per phospholipid, while it is substantially higher than in the gel phase at higher water contents. Together, these data demonstrate that the swelling pressure is sensitive to the phase state of the lipids. We also describe a series of studies that demonstrate that the addition of a second polar solute to the phospholipid–water system has a remarkably small effect on the swelling behavior when analyzed with respect to solvent volume. The reviewed experimental studies provide a thermodynamic characterization of the swelling of lamellar zwitterionic phospholipid systems that should be encompassed in the mechanistic molecular interpretation of the “hydration force.”     

In Situ Separation and Analysis of Lipids by Paper Spray Ionization Mass Spectrometry

Paper spray ionization (PSI) is an extractive ambient ionization technique for mass spectrometry (MS), whereby a triangular paper tip serves as the sampling base and the electrospray tip. During PSI, analytes are extracted and transported to the edge of the paper tip by the applied spraying solvent. Analytes can be purified from a sample matrix and separated from each other by this transportation process. In this study, we investigated and utilized the analyte transportation process of PSI for the in situ separation and analysis of lipid mixtures. We found that differential transport of phosphatidylcholine (PC) and triacylglycerol (TAG), the two most abundant lipid classes in animals, occurred during PSI. We also found that the order in which these lipids moved strongly depended on how the spraying solvent was applied to the paper base. The more polar PC moved faster than the less polar TAG during PSI, when a polar solvent was slowly fed into a paper tip, whereas TAG was transported faster than PC when excess solvent was applied to the tip at once. In addition, we achieved a complete separation and detection of PC and TAG by slowly supplying a nonpolar solvent to a PSI tip.     

Dilution enthalpies of alkanols in concentrated aqueous solutions of urea at 25°C

Enthalpies of dilution of some aliphatic alcohols were determined at 25°C in aqueous 7M urea solutions by flow microcalorimetry. The excess enthalpies were expressed as power expansion series in molalities referred to 1 kg of constant composition urea-water mixture. This urea-water mixture was utilized throughout as a mixed solvent. The values of the second enthalpic virial coefficients were all found to be positive and generally lower than the corresponding values in water. Large differences were encountered, as in water, by comparing normal and branched isomeric propanols and butanols. For one system it was possible to measure the third coefficients, which were also positive. The second enthalpic coefficients were found to increase with the molecular weight of the alkanols. These facts suggest that in the presence of a large concentration of urea, the excess enthalpies are mainly determined by apolar interactions. This is surprising and potentially rich in consequences for a better understanding of the interactions among amino acid residues distantly situated in the primary sequences but topologically near in the loops of globular proteins. An analysis, carried out using the Savage-Wood additivity group method, shows that the enthalpic contributions (that appear to play a crucial role in water in making the polar interaction to be favorable) become essentially unfavorable in urea-water solvent. The hypothesis that the peptide-peptide interactions are prevented by the preferential solvation of urea is also discussed.     

Infrared spectroscopic properties of sodium bromide aerosols

The infrared extinction spectra of aqueous NaBr aerosols at ambient temperature have been measured as a function of relative humidity. Submicron-sized aerosol particles atomized from aqueous NaBr solutions at various concentrations are dried and/or mixed with nitrogen at different humidities and spectroscopically monitored as they flow through an infrared absorption cell. Estimated dry particle median diameters range from 0.24 to 0.15 microm, as calculated from Mie extinction theory. Measured deliquescence and efflorescence relative humidities (35-40% and 25-30%, respectively) are in accordance with previously reported ones. Our results show that NaBr particles take up water only moderately over the deliquescence point, with a significant increase at relative humidities above 70%. The effect of particle size onto water uptake properties has been studied, indicating that smaller particles take up lower amounts of water, and only increase their size significantly at relative humidities near saturation. Particle composition and diameter growth factors have been calculated from spectral data and are shown to be consistent with those predicted from thermodynamic data and Kohler theory. Band centers of liquid water in NaBr aerosols relative to pure water are blue-shifted up to 50 cm-1 at low humidities. Particle structure and phase, together with atmospheric implications, are also discussed.     

Local composition in the vicinity of a protein molecule in an aqueous mixed solvent

This paper is focused on the composition of a cosolvent in the vicinity of a protein surface (local composition) and its dependence on various factors. First, the Kirkwood-Buff theory of solution is used to obtain analytical expressions that connect the excess or deficit number of cosolvent and water molecules in the vicinity of a protein surface with experimentally measurable quantities such as the bulk concentration of the mixed solvent, the preferential binding parameter, and the molar volumes of water and cosolvent. Using these expressions, relations between the preferential binding parameter (at a molal concentration scale) and the above excesses (or deficits) are established. In addition, the obtained expressions are used to examine the effect of the nonideality of the water + cosolvent mixtures and of the molar volume of the cosolvent on the excess (or deficit) number of cosolvent molecules in the vicinity of the protein surface. It is shown that at least for the mixed solvents considered (water + urea and water + glucose) the nonideality of the mixed solvent is not an important factor in the local compositions around a protein molecule and that the main contribution is provided by the nonidealities of the protein-water and protein-cosolvent mixtures. Special attention is paid to urea as cosolvent, because urea is one of only a few compounds with a concentration at the protein surface larger than its concentration in the bulk. The composition dependence of the excess of urea around a protein molecule is calculated for the water + lysozyme + urea mixture at pH = 7.0 and 2.0. At pH = 7.0, the excess of urea becomes almost composition independent at high urea concentrations. Such independence could be explained by assuming that urea totally replaces water in some areas of the protein surface, whereas on the remaining areas of the protein surface both water and urea are present with concentration comparable to those in the bulk. The Schellman exchange model was used to relate the preferential binding parameter in water + lysozyme + urea mixtures to the urea concentration.     

Spectrophotometric determination of urea in dermatologic formulations and cosmetics

A rapid, relatively sensitive, and low-cost method for the determination of water-soluble urea content in dermatological therapy products and cosmetics is proposed using a new spectrophotometric assay with water as the only extraction solvent. Spectrophotometric methods involve addition of a known excess of bromate to urea in an acid medium, followed by the determination of residual bromine and chlorine reacting with methyl orange and measurement of absorbance at 505 nm. The absorbance increases linearly with urea concentration (r = 0.9998). The systems obey Beer's law for 6 - 90 microg ml(-1). The calculated apparent molar absorbance values are found to be 4.537 x 10(3) dm(3) mol(-1) cm(-1) and the Sandell's sensitivity is 0.013 microg cm(-2). The variables affecting the rate of the reaction were investigated. The relative standard deviation for five-replication determination of 60 microg ml(-1) urea was 2.1% and the detection limit of the method is 0.34 ng ml(-1).     

Osmotic coefficient data and an excess Gibbs energy function for single-phase complex system of glucose + alcohol + water

Thermodynamics of single-phase complex systems of glucose + alcohol + water is significantly important in food and pharmaceutical industries. The water activity is a practical parameter in thermodynamic characterization in food and pharmaceutical and other biological industries, which its value has an important role in growing microorganisms and rate of reactions. In this work, the NRTL-NRF excess Gibbs function has extended for ternary system of water, different alcohols (as second molecular solvent) and a special molecular solute like a sugar. The extended NRTL-NRF model with three parameters was used for correlation of the osmotic coefficient of the aqueous systems of glucose and alcohol at different concentrations. The data of the osmotic coefficient of water for these systems are obtained by an isopiestic method. The adjustable parameters are calculated by optimization of the experimental data of the osmotic coefficient using the Nedler–Mead algorithm. It is shown that the results of the modified NRTL-NRF model demonstrate low deviation from the experimental data.     

Thermodynamic studies of aqueous and CCl4 solutions of 15-crown-5 at 298.15 K: an application of McMillan-Mayer and Kirkwood-Buff theories of solutions

The density and osmotic coefficient data for solutions of 15-crown-5 (15C5) in water and in CCl4 solvent systems at 298.15 K have been reported using techniques of densitometry and vapor pressure osmometry in the concentration range of 0.01-2 mol kg-1. The data are used to obtain apparent molar and partial molar volumes, activity coefficients of the components as a function of 15C5 concentration. Using the literature heat of dilution data for aqueous system, it has become possible to calculate entropy of mixing (DeltaS(mix)), excess entropy of solution (DeltaS(E)), and partial molar entropies of the components at different concentrations. The results of all these are compared to those obtained for aqueous 18-crown-6 solutions reported earlier. It has been observed that the partial molar volume of 15C5 goes through a minimum and that of water goes through a maximum at approximately 1.2 mol kg(-1) in aqueous solutions whereas the opposite is true in CCl4 medium but at approximately 0.5 mol kg(-1). The osmotic and activity coefficients of 15C5 and excess free energy change for solution exhibit distinct differences in the two solvent systems studied. These results have been explained in terms of hydrophobic hydration and interactions in aqueous solution while weak solvophobic association of 15C5 molecules in CCl4 solutions is proposed. The data are further subjected to analysis by applying McMillan-Mayer and Kirkwood-Buff theories of solutions. The analysis shows that osmotic second virial coefficient value for 15C5 is marginally less than that of 18C6 indicating that reduction in ring flexibility does not affect the energetics of the interactions much in aqueous solution while the same gets influenced much in nonpolar solvent CCl4.     

Lyotropic stress in lipid-water model systems. Structural changes induced by freezing of the solvent

In a differential scanning calorimetry and X-ray study 12 different lipid-water model systems have been investigated in the temperature region from 250 to 290 K. Structural changes of the lipid system are observed in most cases in the course of freezing or melting of excess water, although the structures of the phases existing around 273 K were quite different. According to thermodynamic considerations a concentration jump (lyotropic stress) is predicted in all cases during the freezing process of excess water. The given lipid-water system reacts to the lyotropic stress in a specific way according to its individual phase diagram.     

Lyotropic stress in lipid-water model systems. Structural changes induced by freezing of the solvent

In a differential scanning calorimetry and X-ray study 12 different lipid-water model systems have been investigated in the temperature region from 250 to 290 K. Structural changes of the lipid system are observed in most cases in the course of freezing or melting of excess water, although the structures of the phases existing around 273 K were quite different. According to thermodynamic considerations a concentration jump (lyotropic stress) is predicted in all cases during the freezing process of excess water. The given lipid-water system reacts to the lyotropic stress in a specific way according to its individual phase diagram.     

Thermodynamic studies of binary methanol and ethanol solutions of 1-dodecanol and 1-tetradecanol at 25°C

The osmotic coefficients of binary methanol and ethanol solutions of 1-dodecanol and 1-tetradecanol wer measured at 25°C up to 8 mol-kg^–1 in methanol and 5.5 mol-kg^–1 in ethanol. The activity coefficients of the solute were calculated from Bjerrum's relation. From the osmotic and activity coeficients the excess Gibbs energies of solution as well as the respective partial molar functions of solute and solvent and the virial pair interaction coefficients for the excess Gibbs energies were calculated. In addition, the difference in the Gibbs energy of solvation for the solvent in solution relative to the pure solvent was calculated, as well as the partial molar volumes and excess partial molar volumes of solutes at infinite dilution, and the coefficients of pairwise contributions to the excess volume were determined. The thermodynamic parameters obtained are discussed on the basis of solute-solvent and solute-solute interactions.     

Water transport in aluminium oxide-poly(ethylene-co-butyl acrylate) nanocomposites

Polymer composites with metal oxide nanoparticles are emerging materials for use as insulation in electrical applications. However, the extensive interfacial surfaces and the presence of polar groups on the particle surfaces make these composites susceptible to water sorption. Water sorption kinetics were studied at 23 °C and different relative humidities (18–90%) for composites based on poly(ethylene-co-butyl acrylate) and aluminium oxide (?12 wt.%); the latter being in three different forms: uncoated and coated with either octyltriethoxysilane or aminopropyltriethoxysilane. The equilibrium water uptake increased linearly with increasing overall concentration of polar groups on the nanoparticle surfaces. Composites with well-dispersed nanoparticles showed Fickian diffusion (constant diffusivity and invariant boundary conditions) with a diffusivity that decreased with increasing filler content; the maximum factorial decrease in diffusivity was 300 with reference to that of the pristine polymer. This effect was most pronounced for composites with accessible polar groups on the particle surfaces, suggesting that water saturation of the composites is retarded by dual water sorption. Composites that contained a sizeable fraction of large nanoparticle agglomerates showed a two-stage sorption process: a rapid process associated with the saturation of the matrix phase and a slow diffusion process due to water sorption by the large nanoparticle agglomerates.     

A protein molecule in an aqueous mixed solvent: fluctuation theory outlook

In the present paper a procedure to calculate the properties of proteins in aqueous mixed solvents, particularly the excesses of the constituents of the mixed solvent near the protein molecule and the preferential binding parameters, is suggested. Expressions for the Kirkwood-Buff integrals in ternary mixtures and for the preferential binding parameter were derived and used to calculate various properties of infinitely dilute proteins in aqueous mixed solvents. The derived expressions and experimental information regarding the partial molar volumes and the preferential binding parameters were used to calculate the excesses (deficits) of water and cosolvent (in comparison with the bulk concentrations of protein-free mixed solvent) in the vicinity of ribonuclease A, ribonuclease T1, and lysozyme molecules. The calculations showed that water was in excess in the vicinity of ribonuclease A for water/glycerol and water/trehalose mixtures, and the cosolvent urea was in excess in the vicinity of ribonuclease T1 and lysozyme. The derivative of the activity coefficient of the protein with respect to the mole fraction of water was also calculated. This derivative was negative for the water/glycerol and water/trehalose mixed solvents and positive for the water/urea mixture. The mixture of lysozyme in the water/urea solvent is of particular interest, because the lysozyme at pH 7.0 is in its native state up to 9.3M urea, while at pH 2.0 it is denaturated between 2.5 and 5M and higher concentrations of urea. Our results demonstrated a striking similarity in the hydration of lysozyme at both pHs. It is worthwhile to note that the excesses of urea were only weakly composition dependent on both cases.     

The use of water structure breakers, urea and guanidium chloride, new mobile phase modifiers in reversed-phase partition high-performance liquid chromatography.

Organic solvent-free mobile-phase systems in ion-pair reversed-phase partition high-performance liquid chromatography (IPRP-HPLC) are demonstrated; using urea at 3.0-7.0 molal (mol kg-1) as a modifier in a mobile phase on an octadecylsilanized silica column, four nitrophenolates and metal 4-(2-pyridilazo)resorcinol (PAR) chelates (in PAR chelates system an aqueous mobile phase with 15 wt% methanol was used) were separated rapidly within 6 min at no sacrifice to the separation efficiency. On the addition of urea in the mobile phase, reduced retention times of nitrophenolates and naphthalenesulfonates and also diminution of the height equivalent to a theoretical plate were observed. The addition of urea and guanidium chloride (GuCl) in the mobile phase gave rise to a decrease in the mobile phase volume; in turn, this meant an increased volume of the stationary phase. As the concentration of urea and GuCl in the mobile phase increased, the volume of the mobile phase in the column decreased within about 70% and 40% at 7.0 molal of urea and GuCl, respectively. A decrease in the mobile phase volume suggests an increase in the extent of solvation of the bonded hydrocarbon chain of the stationary phase. The possible explanations for the LC behavior with the urea and GuCl are turned into reduction of hydrophobic interaction in LC processes, solute partitioning and entangling of alkyl chain brushes, with the addition of urea. The water structure breakers, urea and GuCl, most likely affect the solvation states of both solute molecules and the hydrocarboneous stationary phase by changing the nature of the water solvent, which provides a new technique for fine tuning of the LC resolution of the analytes.     

Stationary phase retention and preliminary application of a spiral disk assembly designed for high-speed counter-current chromatography

A spiral disk assembly composed of five single-channel units was designed for high-speed counter-current chromatography (HSCCC). The retention of different solvent systems ranging from moderately polar to polar organic-aqueous systems to aqueous two-phase systems (ATPs) was investigated under different elution modes. The results indicated that the spiral disk assembly can produce excellent retention of stationary phase for moderately polar organic-aqueous solvent systems, such as chloroform-methanol-water (4:3:2) and hexane-ethyl acetate-methanol-water (1:1:1:1) by pumping lower mobile phase from head (H) to tail (T), and upper mobile phase from tail (T) to head (H) even at a high flow-rate (8 mL/min, Sf>70%), regardless of whether the inlet is at the inner or outer terminal of the channel. This makes it possible for fast analysis of some small molecular compounds. This has been proved in the separation of mixtures of three flavones, including isorhamnetin, kaempferol, and quercetin. The spiral disk assembly can also provide satisfactory retention for polar to ATPS such as 1-butanol-acetic acid-water (4:1:5) (<3 mL/min, Sf>70%), 12.5% poly(ethylene glycol) (PEG) 1000-12.5% K2HPO4-75% water (< or =1 mL/min, Sf>70%) and 4% PEG 8000-5% Dextran T500-91% water (< or =0.5 mL/min, Sf>50%) by pumping lower mobile phase from inner terminal (I) to outer terminal (O), and upper mobile phase from outer terminal (O) to inner terminal (I) at a low flow-rate, while this is not possible with the multilayer coil column. Acceptable resolutions were achieved when it was used for the separation of peptides such as Leu-Tyr and Val-Tyr, and proteins including cytochrome c and myoglobin, lysozyme and myoglobin, and fresh chicken egg-white proteins.     

Organic high ionic strength aqueous two-phase solvent system series for separation of ultra-polar compounds by spiral high-speed counter-current chromatography

Existing two-phase solvent systems for high-speed countercurrent chromatography cover the separation of hydrophobic to moderately polar compounds, but often fail to provide suitable partition coefficient values for highly polar compounds, such as sulfonic acids, catecholamines and zwitter ions. The present paper introduces a new solvent series which can be applied for the separation of these polar compounds. It is composed of 1-butanol, ethanol, saturated ammonium sulfate and water at various volume ratios and consists of a series of 10 steps which are arranged according to the polarity of the solvent system so that the two-phase solvent system with suitable K values for the target compound(s) can be found in a few steps. Each solvent system gives proper volume ratio and high density difference between the two phases to provide a satisfactory level of retention of the stationary phase in the spiral column assembly. The method is validated by partition coefficient measurement of four typical polar compounds including methyl green (basic dye), tartrazine (sulfonic acid), tyrosine (zwitter ion) and epinephrine (a catecholamine), all of which show low partition coefficient values in the polar 1-butanol-water system. The capability of the method is demonstrated by separation of three catecholamines.     

Brewster angle microscopy and PMIRRAS study of DNA interactions with BGTC, a cationic lipid used for gene transfer

The lipid bis(guanidinium)-tris(2-aminoethyl)amine-cholesterol (BGTC) is a cationic cholesterol derivative bearing guanidinium polar headgroups which displays high transfection efficiency in vitro and in vivo when used alone or formulated as liposomes with the neutral colipid 1,2-di-[ cis-9-octadecenoyl]- sn-glycero-3-phosphoethanolamine (DOPE). Since transfection may be related to the structural and physicochemical properties of the self-assembled supramolecular lipid-DNA complexes, we used the Langmuir monolayer technique coupled with Brewster angle microscopy (BAM) and polarization modulation infrared reflection absorption spectroscopy (PMIRRAS) to investigate DNA-BGTC and DNA-BGTC/DOPE interactions at the air/water interface. We herein show that BGTC forms stable monolayers at the air/water interface. When DNA is injected into the subphase, it adsorbs to BGTC at 20 mN/m. Whatever the (+/-) charge ratio of the complexes used, defined as the ratio of positive charges of BGTC in the monolayer versus negative charges of DNA injected in the subphase, the DNA interacts with the cationic lipid and forms either an incomplete (no constituent in excess) or a complete (DNA in excess) monolayer of oriented double strands parallel to the lipid monolayer plan. We also show that, under a homogeneous BGTC/DOPE (3/2) monolayer at 20 mN/m, DNA adsorbs homogeneously to form an organized but incomplete layer whatever the charge ratio used (DNA in default or in excess). Compression beyond the collapse of these mixed DNA-BGTC/DOPE systems leads to the formation of dense DNA monolayers under an asymmetric lipid bilayer with a bottom layer of BGTC in contact with DNA and a top layer mainly constituted of DOPE. These results allow a better understanding of the mechanisms underlying the formation of the supramolecular BGTC-DNA complexes efficient for gene transfection.