Phospholipid vesicles experimental

The NMR measurements were made on sonicated phospholipid vesicles to obtain relatively narrow 31P-NMR signals while the electrophoretic mobility measurements were made on unsonicated vesicles. Since there are differences between the two systems (e.g., area per molecule (13,14)), we do not attempt to quantitatively compare the 31P-NMR data with the electrophoretic data, but rather use the 31P-NMR data as an independent demonstration of the difference between the binding of calcium and magnesium. We note, however, that the linewidth ratio for the outer monolayer of the sonicated vesicles were identical within experimental error (see Table II). This implies that the Ca++/Mg++ selectivity of the two monolayers is identical. We had expected the selectivity to be greater for the inner monolayer because the polar head groups of the lipids in this monolayer occupy a smaller area (13). 

Figure 5 Schematic drawing of the experimental setup used in the permeation studies with the phospholipid vesicle-based barrier (reprinted from Reference 15).

A EXPERIMENTAL FIGURE 17-8 Vesicle buds can be visualized during in vitro budding reactions. When purified COPII coat components are incubated with isolated ER vesicles or artificial phospholipid vesicles (liposomes), polymerization of the coat proteins on the vesicle surface induces emergence of highly curved buds. In this electron micrograph of an in vitro budding reaction, note the distinct membrane coat, visible as a dark protein layer, present on the vesicle buds. [From K. Matsuoka etal., 1988, Ce//93(2) 263.[... 

Liposomes were used as simple experimental models of the lipid bUayer for these studies. The type used were monodisperse, unilamellar phospholipid vesicles (PLVs) of 100 nm diameter, prepared using a pressnre extrusion technique (see Sect. 6.2.3) [1], PLVs of varying compositions were prepared, comprising phosphatidylcholine (PC), cholesterol (Choi) and sphingomyelin (SM) (Fig. 3.2). 

ABSTRACT. Kinetics of proton transfer photoreactions in simple model systems is analyzed from the point of view of reaction kinetics in microphases. Protolytic photodissociation of some hydroxyaromatic compounds ArOH ( 1- and 2-na-phthol, chlorosubstituted naphthols ) was studied in micellar solutions and phospholipid vesicles by fluorescence spectra and kinetics. Experimental results give evidence of at least two localization sites of naphthols in the microphase of these systems. In lipid bilayer membranes of vesicles there are two comparable fractions of ArOH molecules, one of which undergo photodissociation, but another do not dissociate. In micelles only minor fraction ( few per cent ) of ArOH molecules do not take part in excited-state proton transfer reaction. These phenomena reflect heterogeneous structure and dynamic properties of lipid bilayer membranes and micelles. A correlation between proton transfer rate constants and equilibrium constants in microphases similar to that in homogeneous solutions is observed. Microphase approach give a possibility to discuss reactions in dynamical organized molecular systems in terms of classical chemical kinetics. 

A schematic representation of the experimental setup is shown in Fig. 12. A thin teflon film separates two aqueous compartments. One compartment contains phospholipid vesicles into which RCs were incorporated. The vesicles break up at the air-water interface to form a monolayer (30) which is adsorbed onto the teflon film. Illumination of the adsorbed layer evokes an electrical signal. From the time dependence of the signal, the particular transfer... 

The total curvature energy of a spherical vesicle is given by 4tt(2/cc + k). As all experimental data on phospholipids indicate that kc is not small, one is inclined to conclude that the vesicles are thermodynamically unstable the reduction of the number of vesicles, e.g. by vesicle fusion or by Ostwald ripening, will reduce the overall curvature energy. However, such lines of thought overlook the possibility that k is sufficiently negative to allow the overall curvature free energy of vesicles to remain small. 

This review describes experimental techniques, then gives some selected results of H, and NMR studies of pressure effects on the structure, dynamics and phase transitions of phospholipid bilayers. Other examples deal with 2D-NOESY experiments on lipid vesicles and pressure effects on the interaction of anaesthetics with phospholipid bilayers. Furthermore, we discuss... 

Figure 11.9. Contrast formation in cryo-TEM. (a) Schematic image of a vesicle formed with phospholipid molecules, (b) Schematic representation of a phospholipid molecule with polar headgroup and apolar tail. (c)(d) Projection of the polar head group, which is the strongest scattering center, (e) Calculated line scan considering the projection of the polar head groups, (d) Schematic image of a vesicle. (e)(f) Experimental images of vesicles where the double layer with a thickness of about 3.5 nm is clearly seen. Adapted from Sagalowicz et al. 2003.

Among the several drug delivery systems, liposomes - phospholipid nanosized vesicles with a bilayered membrane structure - have drawn a lot of interest as advanced and versatile pharmaceutical carriers for both low and high molecular weight pharmaceuticals. At present, liposomal formulations span multiple areas, from clinical application of the liposomal drugs to the development of various multifunctional liposomal systems to be used in therapy and diagnostics. This chapter provides a brief overview of various liposomal products currently under development at experimental and preclinical level. 

In this work, we will mainly focus on DMPC and DMPG lipids but some other phospholipids work as well. However, if the geometry of a phospholipid molecule (i.e., the contribution of the polar headgroup versus the apolar part - see ref (22, 23)) does not allow the formation of small unilamellar vesicles it will be difficult to generate stable MLs. For instance, in selected experimental conditions, pure phosphatidyletha-nolamine membranes are known to be destabilized (24). 

Experimentally, we observed that the stability of DMPC vesicles increases by adding a small amount (>1 mol% is enough) of negatively charged phospholipids such as DMPG. 

We note here that systematic studies of the melting transition of dry or nearly dry phospholipids bilayers (e.g., vesicles) have been scarce. While there is an abundant experimental and theoretical literature concerning the structure and properties of bilayers in water, less is known about their behavior when water is removed. We have therefore initiated a systematic experimental study of the gel-liquid crystal transition of pure DPPC and DPPC-cholesterol vesicles freeze-dried with and without disaccharides and oxyanion-disaccharide complexes. Some of our results to date are shown in Figure 9.3. 

This experimental approach has been used previously to study the spontaneous transfer of phospholipids between artificial membranes (Martin and MacDonald, 1976 Duckwitz-Peterlein et al., 1977). Xti et al. (1982) used the fluorescence anisotropy of diphenylhexatriene (DPH) in the phosphatidylcholine bilayer to measure the change in the physical state of DPPC and DMPC vesicles upon mixing in the presence of transfer protein. The fluorescent measurements were recorded at a temperature intermediate between the phase transition of the two initially pure vesicles. By using flow cytometry, it was possible to measure the fluorescence... 

Regarding the choice of phospholipids most receptors, ion channels, and transporters work in membranes made from very different phospholipids, albeit with different degrees of cheerfulness. Phospholipids from yolk, brain, or soy beans with addition of cholesterol make good prolis. The experimenter who would like to fuse the vesicles with other membranes afterward (e.g., for electrophysiological investigations), needs special phospholipids (Rehm et al. 1989). 

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