Differences in the physical properties of lipid monolayers and bilayers on a spherical solid support.

A monolayer of 1,2-dipalmitoyl-d62-glycero-3-phosphocholine (DPPC-d62) coated onto silanized silica beads (spherical supported monolayer: SSM) is studied by 2H-NMR and DSC. The results are compared with those obtained from a single bilayer on the same solid support (spherical supported vesicles: SSV) and from multilamellar vesicles (MLV). The phase transition temperature (Tm) of the SSMs is significantly higher than that of the bilayer systems and the extent of this difference depends on the lipid density in the monolayer that is determined during its preparation. 2H-NMR reveals a gel and fluid phase coexistence in the SSM transition region. A comparison of the 2H-NMR line shapes suggests the presence of highly curved structures for the fluid phase of the SSM samples. From a comparison of SSM and SSV transverse relaxation in the fluid phase we can conclude that the lateral diffusion coefficient D1 in supported monolayers is similar to that in bilayers.

The effect of increasing membrane curvature on the phase transition and mixing behavior of a dimyristoyl-sn-glycero-3-phosphatidylcholine/ distearoyl-sn-glycero-3-phosphatidylcholine lipid mixture as studied by Fourier transform infrared spectroscopy and differential scanning calorimetry.

The phase transition behavior of a lipid bilayer of dimyristoyl-sn-glycero-3-phosphatidylcholine/distearoyl-sn-glycero-3- phosphatidylcholine (DMPC-d54/DSPC) (1:1) on a solid support with varying curvatures was investigated with differential scanning calorimetry, infrared spectroscopy, and model calculations. With increasing curvature the temperatures of the liquidus and solidus points are shifted to lower values by up to 7 degrees C and 15 degrees C, and the mixing of the two lipid species in the two phase region is altered. With increasing curvature the DSPC dominates the gel phase, whereas the DMPC-d54 is expelled to the fluid phase. Whereas the planar system shows a nearly simultaneous phase transition of DSPC and DMPC-d54, the spherical system with the highest curvature exhibits an almost complete separation of the phase transitions of the two lipids. Model calculations suggest that the shift of the liquidus point can be understood as a reduction of the lateral pressure in the bilayer with increasing curvature. The shift of the solidus line is interpreted as a result of the increased demixing of the two components in the two-phase region with increasing curvature due to lowering of the lateral pressure.

Macroscopic orientation effects in broadline NMR-spectra of model membranes at high magnetic field strength: A method preventing such effects.

The partial orientation of multilamellar vesicles (MLV) in high magnetic fields has been studied and a method to prevent such effects is herewith proposed. The orientation effect was measured with (2)H-, (31)P-NMR and electron microscopy on MLVs of dipalmitoyl phosphatidylcholine with 30 mol% cholesterol. We present the first freeze-etch electron microscopy data obtained from MLV samples that were frozen directly in the NMR magnet at a field strength of 9.4 Tesla. These experiments clearly show that the MLVs adopt an ellipsoidal (but not a cylindrical) shape in the magnetic field. Best fit (31)P-NMR lineshape calculations assuming an ellipsoidal distribution of molecular director axes to the experimentally obtained spectra provide a quantitative measure of the average semiaxis ratio of the ellipsoidal MLVs and its change with temperature. The application of so-called spherical supported vesicles (SSV) is found to prevent any partial orientation effects so that undistorted NMR powder pattern of the bilayer can be measured independently of magnetic field strength and temperature.The usefulness of SSVs is further demonstrated by a direct comparison of spectral data such as (31)P-and (2)H-NMR lineshapes and relaxation times as well as (2)H-NMR dePaked spectra obtained for both model systems. These experiments show that spectral data obtained from partially oriented MLVs are not unambiguous to interpret, in particular, if an external parameter such as temperature is varied.

Changes of the physical properties of the liquid-ordered phase with temperature in binary mixtures of DPPC with cholesterol: A H-NMR, FT-IR, DSC, and neutron scattering study.

The structure of the so-called liquid-ordered (lo) phase of binary mixtures of DPPC-d(62) with cholesterol was studied between 20-50 mol% cholesterol using (2)H-NMR, FT-IR, DSC, and neutron specular reflection. Different model systems such as multilamellar vesicles, spherical supported vesicles, and oriented multilayers were used. We observed significant changes of the lo phase structure in the physiological relevant temperature region between 30-45 degrees C. (2)H-NMR in combination with lineshape simulations provides evidence for a drastic effect of cholesterol on the shape of multilamellar vesicles due to magnetic field orientation. Moreover, the data indicates a significant change of the extent of this partial orientation for DPPC-d(62) multilamellar vesicles containing 25 mol% cholesterol between 36-42 degrees C. The semiaxes ratio of the (due to magnetic field orientation) ellipsoidal multilamellar vesicles changes over this temperature range by approximately 25%. (2)H-NMR and FT-IR measurements indicate changes of the average orientational order at the bilayer center in the same temperature range and a significant increase of the number of end-gauche conformers while the majority of the methylene groups remain in an all-trans conformation. Additionally, specular reflection of neutrons shows a concomitant reduction of the bilayer thickness by 4 A. Based on a model of the arrangement of DPPC and cholesterol in the lo phase, a molecular mechanism is proposed in which increasing the temperature between 30 and 45 degrees C has the effect of driving cholesterol from the bilayer center towards the head group region.

Phase transition behavior of single phosphatidylcholine bilayers on a solid spherical support studied by DSC, NMR and FT-IR.

For the first time, the chain melting transition from the gel phase to the liquid crystalline phase of a single DPPC bilayer on a solid, spherical support (silica beads) is observed by differential scanning calorimetry (DSC). This transition is remarkably cooperative, exhibits a transition temperature T(m) which is 2 degrees C lower than usually found for DPPC multilamellar vesicles and its excess enthalpy is about 25% less than in DPPC multilayers. 31P- and 2H-NMR data as well as FT-IR data provide evidence that despite the highly asymmetric characteristic of the model system, the whole single bilayer undergoes the transition at T(m), i.e., there is no decoupling of the two monolayer leaflets at the main phase transition. Furthermore, our results show that the formation of the ripple (P(beta')) phase is inhibited in single bilayers on a solid support. This result confirms a conclusion which we reached previously on the basis of neutron scattering data obtained on planar supported bilayers. The most likely reason for this inhibition as well as for the above mentioned thermodynamic differences between multilamellar vesicles and supported membranes is a significantly higher lateral stress in the latter. Moreover, the exchange of lipids between two populations of spherical supported vesicles (DMPC and chain perdeuterated DMPC) is studied by DSC. It is shown that this exchange process is symmetric and its half-time is a factor of 3-4 higher than observed for small sonicated DMPC vesicles.