Pure Dipalmitoylphosphatidylcholine Monolayer Research Articles

The Interaction between Anesthetic Isoflurane and Model-Biomembrane Monolayer Using Simultaneous Quartz Crystal Microbalance (QCM) and Quartz Crystal Impedance (QCI) Methods.

The interaction between anesthetic Isoflurane (Iso) and model-biomembrane on the water surface has been investigated using quartz crystal microbalance (QCM) and quartz crystal impedance (QCI) methods. The model-biomembranes used were dipalmitoyl phosphatidyl choline (DPPC), DPPC-palmitic acid (PA) mixture (DPPC:PA = 8:2), DPPC-Alamethicin (Al) mixture (DPPC:Al = 39:1), and DPPC-β-Lactoglobulin (βLG) mixture (DPPC:βLG = 139:1) monolayers, respectively. The quartz crystal oscillator (QCO) was attached horizontally to each monolayer, and QCM and QCI measurements were performed simultaneously. It was found that Iso hydrate physisorbed on each monolayer/water interface from QCM and changed those interfacial viscosities from QCI. With an increase in Iso concentration, pure DPPC, DPPC-PA mixed, and DPPC-Al mixed monolayers showed a two-step process of Iso hydrate on both physisorption and viscosity, whereas it was a one-step for the DPPC-βLG mixed monolayer. The viscosity change in the DPPC-βLG mixed monolayer with the physisorption of Iso hydrate was much larger than that of other monolayers, in spite of the one-step process. From these results, the action mechanism of anesthetics and their relevance to the expression of anesthesia were discussed, based on the "release of interfacial hydrated water" hypothesis on the membrane/water interface.

Novel colloidal associations of soyasaponins and lipid components (DPPC, cholesterol) as potential adjuvants for vaccines

Soyasaponins from soybean (Glycine max) represent promising new potent adjuvants for vaccine research because of their immunostimulating properties and weak hemolytic activity. In the present study, saponin microstructures of soyasaponins (soyasaponin Bb, soyasaponin Ab) with lipid components (cholesterol, DPPC (dipalmitoylphosphatidylcholine)) were designed by the lipid film method. In interaction studies between soyasaponins (soyasaponin Ab/Bb) and Langmuir monolayers (model membranes), composed of cholesterol and DPPC, marked interactions between soyasaponins and a pure cholesterol monolayer were observed. No interaction was detected for soyasaponins with a pure DPPC monolayer. The intercalation of soyasaponins in a mixed DPPC/cholesterol (3:1, w/w) monolayer was only observed for the monodesmosidic soyasaponin Bb whereas the second sugar chain of the bidesmosidic soyasaponin Ab impaired the access to the monolayer. Transmission electron microscopy was used for visualizing particle formation of soyasaponins and lipid components. Pseudo-binary systems (soyasaponin Ab/Bb, cholesterol) formed colloidal associations built up from ring-like subunits in the nanometer size range. In pseudo-ternary systems (soyasaponin, cholesterol, DPPC) soyasaponin Bb attacked the liposomal membrane by forming colloidal associations. Colloidal associations in pseudo-ternary systems with soyasaponin Ab, cholesterol and a phospholipid were only observed in the presence of PE (phosphatidylethanolamine) instead of DPPC. In an MTT assay with a HaCaT cell line (keratinocyte cell line) the cell viability was neither affected by the soyasaponins nor by the corresponding formulations. Both the pure soyasaponin solution and the saponin formulations may be promising adjuvant systems for the intradermal vaccine application. Furthermore, interaction studies between the model antigen ovalbumin and colloidal associations of saponins and cholesterol using MST (Microscale Thermophoresis) gave first indications of an antigen binding to colloidal associations. Ex vivo T-cell proliferation in the presence of soyasaponin Ab was confirmed.

The bidesmosidic triterpene saponins hederacoside C and ginsenoside Rb1 exhibit low affinity to cholesterol in liposomal membranes

The formation of nanoparticulate structures in the presence of liposomal components has recently been demonstrated for a few saponins, with cholesterol being the essential component.As a follow-up from previous studies, we have studied the interaction of two bidesmosidic triterpene saponins (ginsenoside Rb1, hederacoside C) with liposomal components (cholesterol, DPPC (dipalmitoylphosphatidylcholine)) using the Langmuir film balance technique. In addition, we examined the influence of saponins in aqueous pseudo-binary (saponin:cholesterol/DPPC) and pseudo-ternary systems (saponin:cholesterol:DPPC) regarding particle formation utilizing transmission electron microscopy (TEM). Saponin formulations with liposomal components were prepared by the lipid film method.Langmuir monolayer studies showed a weak affinity of both saponins to cholesterol. Saponin molecules from the subphase migrated into the cholesterol monolayer only at low surface pressure. Further compression of the monolayer led to an exclusion of the saponin molecules back into the subphase. In mixed monolayers (cholesterol, DPPC) as well as in a pure DPPC monolayer, the saponins remained completely in the subphase independent of the pressure applied. TEM images of aqueous pseudo-ternary systems showed no colloidal associations along with the weak affinity for cholesterol. These results emphasize the correlation between the affinity for cholesterol and the formation of nanoparticular structures with liposomal components.

Bicelle-to-Vesicle Transition of a Binary Phospholipid Mixture Guided by Controlled Local Lipid Compositions: A Molecular Dynamics Simulation Study.

An essential step of nanoliposome formation in an aqueous lipid solution is the transition from discoidal lipid aggregate (bicelle) to vesicle. We investigate here the bicelle-to-vesicle transition of a binary lipid mixture of saturated and unsaturated phosphatidylcholine by performing nonequilibrium molecular dynamics simulations with the coarse-grained representation of di-palmitoyl-phosphatidyl-choline (DPPC) and di-linoleoyl-phosphatidyl-choline (DLiPC). When the DPPC molecules of a stable DPPC bicelle are randomly replaced with the DLiPC molecules, the transition occurs for higher apparent DLiPC concentrations. On the other hand, when the DPPC molecules only in the core region of the bicelle are replaced, the transition occurs even for lower apparent DLiPC concentrations. For the bicelle where the head and tail layers are pure DPPC and DLiPC monolayers, respectively, the side of the DLiPC monolayer becomes the concave surface of bending bicelle. Controlling the local lipid compositions in a binary lipid bicelle has the potential to determine the success of vesicle formation and the direction of bicelle bending. Our findings help explain nanoliposome formation with sonication and give useful information for controlling encapsulation efficiencies of nanoliposomes.

Properties of β-carotene and retinoic acid in mixed monolayers with dipalmitoylphosphatidylcholine (DPPC) and Solutol

The liposoluble vitamins and carotenoids are sensitive molecules and their antioxidant properties can be dramatically affected when included in membrane environment or in encapsulated carriers as liposomes or lipid nanocapsules (LNCs). The formation and state of mixed monolayers at air–water interface of β-carotene (βC) or retinoic acid (RA) with dipalmitoylphosphatidylcholine (DPPC) as simplest membrane model or with Solutol (Sol) which forms the soft layer covering the LNCs were studied by means of surface pressure (π) and surface potential (ΔV) measurements. The mutual miscibility and interfacial organization of components in studied mixed monolayers was found to strongly depend on their composition. It was suggested a better inclusion of βC small quantities in pure DPPC monolayer at low surface pressure than for films with more βC at higher surface pressure. The measurements of surface properties were complemented by Atomic Force Microscopy (AFM) as a suitable technique for direct visualization at nanoscale level of the monolayer microheterogeneity and the presence of interfacial structures. It was also found that the addition of bigger amounts of βC led to the appearance of stripe like phase which is more deeply formed at higher surface pressure.

Effect of cholesterol nanodomains on monolayer morphology and dynamics.

At low mole fractions, cholesterol segregates into 10- to 100-nm-diameter nanodomains dispersed throughout primarily dipalmitoylphosphatidylcholine (DPPC) domains in mixed DPPC:cholesterol monolayers. The nanodomains consist of 6:1 DPPC:cholesterol "complexes" that decorate and lengthen DPPC domain boundaries, consistent with a reduced line tension, λ. The surface viscosity of the monolayer, ηs, decreases exponentially with the area fraction of the nanodomains at fixed surface pressure over the 0.1- to 10-Hz range of frequencies common to respiration. At fixed cholesterol fraction, the surface viscosity increases exponentially with surface pressure in similar ways for all cholesterol fractions. This increase can be explained with a free-area model that relates ηs to the pure DPPC monolayer compressibility and collapse pressure. The elastic modulus, G', initially decreases with cholesterol fraction, consistent with the decrease in λ expected from the line-active nanodomains, in analogy to 3D emulsions. However, increasing cholesterol further causes a sharp increase in G' between 4 and 5 mol% cholesterol owing to an evolution in the domain morphology, so that the monolayer is elastic rather than viscous over 0.1-10 Hz. Understanding the effects of small mole fractions of cholesterol should help resolve the controversial role cholesterol plays in human lung surfactants and may give clues as to how cholesterol influences raft formation in cell membranes.

Phospholipid packing and hydration in pulmonary surfactant membranes and films as sensed by LAURDAN

The efficiency of pulmonary surfactant to stabilize the respiratory surface depends critically on the ability of surfactant to form highly packed films at the air–liquid interface. In the present study we have compared the packing and hydration properties of lipids in native pulmonary surfactant and in several surfactant models by analyzing the pressure and temperature dependence of the fluorescence emission of the LAURDAN (1-[6-(dimethylamino)-2-naphthyl]dodecan-1-one) probe incorporated into surfactant interfacial films or free-standing membranes. In interfacial films, compression-driven changes in the fluorescence of LAURDAN, evaluated from the generalized polarization function (GPF), correlated with changes in packing monitored by surface pressure. Compression isotherms and GPF profiles of films formed by native surfactant or its organic extract were compared at 25 or 37°C to those of films made of dipalmitoylphosphatidylcholine (DPPC), palmitoyloleoylphosphatidylcholine (POPC), DPPC/phosphatidylglycerol (PG) (7:3, w/w), or the mixture DPPC/POPC/palmitoyloleoylphosphatidylglycerol (POPG)/cholesterol (Chol) (50:25:15.10), which simulates the lipid composition of surfactant. In general terms, compression of surfactant films at 25°C leads to LAURDAN GPF values close to those obtained from pure DPPC monolayers, suggesting that compressed surfactant films reach a dehydrated state of the lipid surface, which is similar to that achieved in DPPC monolayers. However, at 37°C, the highest GPF values were achieved in films made of full surfactant organic extract or the mixture DPPC/POPC/POPG/Chol, suggesting a potentially important role of cholesterol to ensure maximal packing/dehydration under physiological constraints. Native surfactant films reached high pressures at 37°C while maintaining relatively low GPF, suggesting that the complex three-dimensional structures formed by whole surfactant might withstand the highest pressures without necessarily achieving full dehydration of the lipid environments sensed by LAURDAN. Finally, comparison of the thermotropic profiles of LAURDAN GPF in surfactant model bilayers and monolayers of analogous composition shows that the fluorophore probes an environment that is in average intrinsically more hydrated at the interface than inserted into free-standing bilayers, particularly at 37°C. This effect suggests that the dependence of membrane and surfactant events on the balance of polar/non-polar interactions could differ in bilayer and monolayer models, and might be affected differently by the access of water molecules to confined or free-standing lipid structures.

Measuring the Interfacial Rheological Properties of Lung Surfactant Monolayers

Lung surfactant (LS) is essential to respiration because it reduces surface tension at the interface between water and air in the alveoli. In addition, LS stabilizes the alveoli against collapse during exhalation. LS is composed of lipids, mainly dipalmitoylphosphatidylcholine (DPPC), and proteins. We are interested in the rheological properties of a pure DPPC film as a first step towards understanding complex LS rheology. During exhalation, we hypothesize that a large surface tension gradient occurs between the alveolar sacs and the bronchioles which should cause LS to flow away from the alveoli. Because LS does not readily leave the alveoli through the trachea, there must be the drag effect with an opposing direction to the flow induced form the surface tension gradient. Previous work has shown that the surface shear viscosity of LS increases exponentially at low surface tensions, which may be sufficient to prevent the flow of surfactant out of the lungs. Additionally, a yield stress in the interfacial LS film would also help to prevent the flow. Here, we define the yield stress as a minimum stress below which no flow occurs. As a result, we have shown the existence of the yield stress in pure DPPC monolayer, and how to increase or decrease the yield stress based on the addition of other lipid molecules.

Effects of Fullerenes on Phospholipid Membranes: A Langmuir Monolayer Study

We investigate two-component Langmuir monolayers of dipalmitoylphosphatidylcholine (DPPC)/C(60) by recording surface pressure/area (pi/A) and surface potential/area (DeltaV/A) isotherms and by direct Brewster angle microscopy (BAM) imaging. Atomic force microscopy (AFM) is employed to study morphologies of the mixed monolayers transferred to a solid substrate by the Langmuir-Blodgett technique. C(60) is shown to have little influence on isotherms of the DPPC/C(60) monolayers even at a molar fraction as high as X(C60)=0.3. The elastic modulus (Cs(-1) versus pi curves of the DPPC/C(60) monolayers almost overlay each other, as well as that of pure DPPC, that is, the elasticities of pure DPPC monolayers and DPPC/C(60) monolayers are remarkably similar. AFM studies reveal that fullerene flocs form at low surface pressures (pi<or=15 mN m(-1)), are gradually disaggregated and dispersed in the DPPC monolayer with increasing surface pressure up to 35 mN m(-1), and are then progressively squeezed out to form protruded islands as the surface pressure increases up to 65 mN m(-1). Our work provides experimental support to the computational result that C(60) can dissolve in lipid bilayers without significantly compromising their mechanical properties, a finding which has important implications for the toxicity and development of drug vehicles from fullerene materials.

On the characteristics of mixed Langmuir monolayer templates containing dipalmitoyl phosphatidylcholine for gold nanoparticle formation

Mixed Langmuir monolayers containing dipalmitoyl phosphatidylcholine (DPPC) were applied as two-dimensional templates to incorporate with gold precursor AuCl 4 − in the subphases. The organic monolayer templates were then transferred onto solid substrates to form ultra-thin films by the Langmuir–Blodgett (LB) deposition technique. With an UV irradiation approach, gold nanoparticles were thus fabricated in the LB films of monolayer templates. Characteristics of the monolayer templates were studied by the surface pressure–area isotherm measurements and Brewster angle microscopy (BAM) observation. The factors affecting the formation of gold nanoparticle structures in the LB films of organic monolayer templates were elucidated by the atomic force microscopy (AFM). The monolayer isotherms and BAM images suggested that by changing the gold precursor concentration in the subphase, one could control the adsorption behavior of the gold precursor onto the monolayer templates. It was found that the association of the gold precursor with a pure DPPC monolayer template resulted in an unstable Langmuir monolayer, which was inappropriate for the following LB deposition. With the presence of n-hexadecanol in a DPPC monolayer, the monolayer template stability and corresponding LB deposition quality could be tremendously improved. Moreover, the distribution of DPPC molecules in the monolayer templates was possible to be regulated by the addition of n-hexadecanol, and the association behavior of the gold precursor with the monolayer templates was thus controlled. The AFM analysis then indicated that the number and size of gold nanoparticles fabricated in the LB films of the mixed DPPC/ n-hexadecanol monolayer templates by a photoreduction reaction could be manipulated by the mole fraction of n-hexadecanol and UV irradiation time.

Phase Transitions in Phospholipid Monolayers Studied by Atomic Force Microscopy and Langmuir-Blodgett Technique

The phase behaviour and surface structure of dipalmitoyl phosphatidyl choline (DPPC) monolayers at the air/water interface, in the absence and the presence of procaine, have been investigated by Langmuir-Blodgett (LB) technique and atomic force microscopy. The LB films were transferred on mica, at a controlled surface pressure, characteristic for the expanded liquid to condensed liquid phase transition of pure DPPC monolayers. The results indicate that procaine penetrates into and specifically interacts with phospholipid monolayers stabilizing the lipid membrane interface.

Phospholipase D Activity Is Regulated by Product Segregation and the Structure Formation of Phosphatidic Acid within Model Membranes

Phospholipase D from Streptomyces chromofuscus (scPLD) hydrolyzes phosphatidylcholines (PC) to produce choline and phosphatidic acid (PA), a lipid messenger molecule within biological membranes. To scrutinize the influence of membrane structure on scPLD activity, three different substrate-containing monolayers are used as model systems: pure dipalmitoylphosphatidylcholine (DPPC) as well as equimolar mixtures of DPPC/ n-hexadecanol (C 16OH) and DPPC/dipalmitoylglycerol (DPG). The activity of scPLD toward these monolayers is tested by infrared reflection-absorption spectroscopy and exhibits different dependencies on surface pressure. For pure DPPC, the catalytic turnover drastically drops above 20 mN/m. On addition of C 16OH, this strong decrease starts at 5 mN/m. For the DPPC/DPG system, the reaction yield linearly decreases between 5 and 25 mN/m. The difference in scPLD activity is correlated to the phase state of the monolayers as examined by x-ray diffraction, Brewster angle microscopy, and atomic force microscopy. Because the additives C 16OH and DPG mediate the miscibility of PC and PA, only a basal activity of scPLD is observed toward the mixed systems at higher surface pressures. At pure DPPC monolayers, scPLD is activated after the segregation of initially formed PA. Furthermore, scPLD is inhibited when the lipids in the PA-rich domains adopt an upright orientation. This phenomenon offers a self-regulating mechanism for the concentration of the second messenger PA within biological membranes.

Interaction of Dipalmitoyl Phosphatidylcholine with n‐Hexadecanol in Monolayer and Liposome

AbstractThe monolayer collapse behavior of n‐hexadecanol/dipalmitoyl phosphatidylcholine (DPPC) was investigated in this study at the air/water interface at 37 °C. Surface pressure variations with time for the mixed monolayers of DPPC with 20 mol% and 50 mol% n‐hexadecanol at corresponding collapse points were recorded by a Langmuir trough system. In addition, the interaction of n‐hexadecanol with a pure DPPC monolayer was identified by fluorescence microscopy (FM). The results demonstrated distinct differences between these systems; according to our observation, the higher the ratio of n‐hexadecanol to DPPC, the more nucleation domains can be induced. The FM images demonstrated that pronounced domain formation was associated with a longer relaxation time of the collapsed DPPC and DPPC/n‐hexadecanol monolayers, and the presence of n‐hexadecanol appeared to enhance the relaxation processes. The liposome was prepared by the thin‐film hydration method. The average diameter of DPPC and DPPC/n‐hexadecanol liposomes was investigated by dynamic light scattering. It is shown that the diameter of DPPC liposome with n‐hexadecanol is smaller than pure DPPC liposome at the initial state. After 24 hours, DPPC/n‐hexadecanol liposome became larger than pure DPPC liposome and lasted for the next four days. The effects of a greater ratio of n‐hexadecanol did not play an important role in DPPC liposome formation based on our dynamic light scattering analysis. Our result demonstrated that n‐hexadecanol might affect the DPPC liposome stability. The increased ratio of n‐hexadecanol in DPPC liposomes could only a play a minor role in DPPC liposome fusion.

Modifying dipalmitoylphosphatidylcholine monolayers by n-hexadecanol and dipalmitoylglycerol

The monolayer structure of pure dipalmitoylphosphatidylcholine (DPPC) and equimolar mixtures of DPPC/ n-hexadecanol (C 16OH) and DPPC/dipalmitoylglycerol (DPG) are studied by the film balance technique and grazing incidence X-ray diffraction measurements. At 20 °C, the binary systems exhibit complete miscibility. In contrast to pure DPPC monolayers, a condensing effect is observed in the presence of both non-phospholipid additives; but the phase transition behavior differs. The tilt angle of the hydrocarbon chains in the DPPC/C 16OH mixture is significantly smaller than in pure DPPC monolayers. The tilt of the chains is even further reduced in the mixed monolayer of DPPC/DPG. A comparison of the three systems reveals distinct structural features such as phase state, chain tilt, and molecular area over a wide range of surface pressures. Therefore, these monolayers provide a highly suitable model to investigate the influence of structural parameters on biological processes occurring at the membrane surface, e.g. enzymatic reactions and adsorption events.

Atomic force microscopy studies of lateral phase separation in mixed monolayers of dipalmitoylphosphatidylcholine and dilauroylphosphatidylcholine

Atomic force microscopy imaging of dipalmitoylphosphatidylcholine (DPPC)/dilauroylphosphatidylcholine (DLPC) monolayers deposited onto alkanethiol modified-gold surfaces by the Langmuir–Schaefer technique was used to investigate domain formation in a binary system where phase separation arises from a difference in the alkyl chain lengths of the lipids. We have established how the condensed domain structure (shape and size) in DPPC/DLPC monolayers depends on the surface pressure and lipid composition. The mixed monolayers exhibit a positive deviation from an ideal mixing behavior at surface pressures of ⩽32 mN/m. Lateral compression to pressures greater than the liquid-expanded-to-liquid-condensed (LE-to-LC) phase transition pressure of the mixed monolayer (∼8–16 mN/m) induces extensive separation into condensed DPPC-rich domains and a fluid DLPC matrix. The condensed structures observed at a few milliNeutons per meter above the LE-to-LC transition pressure resemble those reported for pure DPPC monolayers in the LE/LC co-existence region. At a bilayer equivalence pressure of 32 mN/m and 20 °C, condensed domains exist between x DPPC ∼0.25 and ∼0.80, analogous to aqueous DPPC/DLPC dispersions. Compression from 32 to 40 mN/m results in either a striking distortion of the DPPC domain shape or a break-up of the microscopic DPPC domains into a network of nanoscopic islands (at higher DPPC mol fractions), possibly reflecting a critical mixing behavior. The results of this study provide a fundamental framework for understanding and controlling the formation of lateral domain structures in mixed phospholipid monolayers.

Epifluorescence microscopic studies of monolayers containing mixtures of dioleoyl- and dipalmitoylphosphatidylcholines

Monolayers of dipalmitoylphosphatidylcholine (DPPC), dioleoylphosphatidylcholine (DOPC), and some mixtures of these lipids were investigated using an epifluorescence microscopic surface balance. Monolayers were visualized at 23 +/- 1 degree C through the fluorescence of 1 mol% of two different fluorescent probes, 1-palmitoyl-2-(12-[(7-nitro-2-1,3-benzoxadizole-4- yl)amino]dodecanoyl)phosphatidylcholine (NBD-PC), which partitions into the liquid expanded (LE) or disordered lipid phase and 3,3'-dioctadecyloxacarbocyanine perchlorate (DiO-C18), which preferentially associates with the liquid condensed (LC) phase or lipid with ordered chains. LC domains were observed in pure DPPC monolayers at relatively low surface pressures (pi), and these domains grew with increasing surface pressure. Only liquid expanded phase was observed in pure DOPC monolayers up to the point of monolayer collapse. In monolayers containing 29:70:1, 49:50:1, and 69:30:1 (mol/mol/mol) of DPPC:DOPC:probe the domains of LC phase were smaller than those seen in DPPC monolayers at equivalent surface pressures. Quantitative analysis of the visual fields shown by the mixed monolayers showed a distribution of sizes of condensed domains at any given pi. At pi = 30 mN m-1, liquid-expanded, or fluid, regions occupied more than 70% of the total monolayer area in all three mixtures studied, whereas DPPC monolayers were more than 75% condensed or solid at that pressure. For monolayers of DPPC:DOPC:NBD-PC 49:50:1 and 69:30:1 the average domain size and the percentage of the total area covered with LC, or rigid, areas increased to a maximum at pi around 35 mN m-1 followed by a decrease at higher pi. Repetitive compression and expansion of the monolayers containing DPPC:DOPC:NBD-PC 49:50:1 at an initial rate of 3.2 A2 molecule-1 s-1 produced monolayers with visual properties consistent with there being a preferential exclusion of the unsaturated lipid from the monolayer.

The interaction of glycophorin with dipalmitoylphosphatidylcholine at the air—water interface

The interaction between glycophorin, the major sialoglycoprotein of the human erythrocyte membrane, and dipalmitoylphosphatidylcholine (DPPC) has been investigated in mixed monolayers at the air—water interface. Surface pressure—area isotherms have been measured for monolayers covering a range of glycophorin—DPPC molar ratios from 1:1765 to 1:44 at 20°C. The phase transition from the liquid-expanded to liquid-condensed monolayer states characteristic of pure DPPC monolayers is no longer observable in mixed monolayers at glycophorin to DPPC molar ratios in the range 1:355 to 1:44 and the isotherms at low areas per lipid molecule have a lower surface pressure than that calculated for an ideal mixture of the pure components. Analysis of the results indicates that in the range of surface pressure from 15–30 mNm −1, glycophorin withdraws between approximately 1300 to 200 DPPC molecules from participation in chain-melting in the monolayer.