The effect of ergosterol on dipalmitoylphosphatidylcholine bilayers: a deuterium NMR and calorimetric study.

We have studied the effect of ergosterol, an important component of fungal plasma membranes, on the physical properties of dipalmitoylphosphatidylcholine (DPPC) multibilayers using deuterium nuclear magnetic resonance ((2)H NMR) and differential scanning calorimetry (DSC). For the (2)H NMR experiments the sn-1 chain of DPPC was perdeuterated and NMR spectra were taken as a function of temperature and ergosterol concentration. The phase diagram, constructed from the NMR spectra and the DSC thermograms, exhibits both solid-ordered (so) + liquid-ordered (lo) and liquid-disordered (ld) + lo phase coexistence regions with a clear three-phase line. This is the first demonstration that lo domains exist in liquid crystalline membranes containing ergosterol. The domain sizes in the ld+lo phase coexistence region were estimated by analyzing the exchange of labeled DPPC between the two regions, and depend on ergosterol concentration. The DPPC-ergosterol phase diagram is similar to that of the DPPC-cholesterol multibilayer system except that the so+lo and ld+lo phase coexistence regions are considerably broader.

Binding of small alcohols to a lipid bilayer membrane: does the partitioning coefficient express the net affinity?

The total vapor pressures at 26 degreesC of binary (water-alcohol) and ternary (water-alcohol-vesicle) systems were measured for six short chain alcohols. The vesicles were unilamellar dipalmitoyl phosphatidylcholine (DMPC). The data was used to evaluate the effect of vesicles on the chemical potential of alcohols expressed as the preferential binding parameter of the alcohol-lipid interaction, gamma23. This quantity is a thermodynamic (model-free) measure of the net strength of membrane-alcohol interactions. For the smaller investigated alcohols (methanol, ethanol and 1-propanol) gamma23 was negative. This is indicative of so-called preferential hydration, a condition where the affinity of the membrane for water is higher than the affinity for the alcohol. For the longer alcohols (1-butanol, 1-pentanol, 1-hexanol) gamma23 was positive and increasing with increasing chain length. This demonstrates preferential binding, i.e. enrichment of alcohol in the membrane and a concomitant depletion of the solute in the aqueous bulk. The measured values of gamma23 were compared to the number of alcohol-membrane contacts specified by partitioning coefficients from the literature. It was found that for the small alcohols the number of alcohol-membrane contacts is much larger than the number of preferentially bound solutes. This discrepancy, which is theoretically expected in cases of very weak binding, becomes less pronounced with increasing alcohol chain length, and when the partitioning coefficient exceeds approximately 3 on the molal scale (10(2) in mole fraction units) it vanishes. Based on this, relationships between structural and thermodynamic interpretations of membrane partitioning are discussed.

A thermodynamic study of the effects of cholesterol on the interaction between liposomes and ethanol.

The association of ethanol with unilamellar dimyristoyl phosphatidylcholine (DMPC) liposomes of varying cholesterol content has been investigated by isothermal titration calorimetry over a wide temperature range (8-45 degrees C). The calorimetric data show that the interaction of ethanol with the lipid membranes is endothermic and strongly dependent on the phase behavior of the mixed lipid bilayer, specifically whether the lipid bilayer is in the solid ordered (so), liquid disordered (ld), or liquid ordered (lo) phase. In the low concentration regime (<10 mol%), cholesterol enhances the affinity of ethanol for the lipid bilayer compared to pure DMPC bilayers, whereas higher levels of cholesterol (>10 mol%) reduce affinity of ethanol for the lipid bilayer. Moreover, the experimental data reveal that the affinity of ethanol for the DMPC bilayers containing small amounts of cholesterol is enhanced in the region around the main phase transition. The results suggest the existence of a close relationship between the physical structure of the lipid bilayer and the association of ethanol with the bilayer. In particular, the existence of dynamically coexisting domains of gel and fluid lipids in the transition temperature region may play an important role for association of ethanol with the lipid bilayers. Finally, the relation between cholesterol content and the affinity of ethanol for the lipid bilayer provides some support for the in vivo observation that cholesterol acts as a natural antagonist against alcohol intoxication.

Slow relaxation of the sub-main transition in multilamellar phosphatidylcholine vesicles.

The influence of ionic strength and equilibration time on the appearance of the sub-main transition in fully hydrated multilamellar vesicles composed of phosphatidylcholines has been investigated by means of calorimetry and densitometry. The heat capacity measurements show that the transition enthalpy of the sub-main transition is affected by both salt concentration (KCl) and equilibration time. The small heat capacity peak appearing in vesicles made in pure water is significantly increased upon addition of salt. Furthermore, equilibration of the multilamellar vesicles at low temperatures for several weeks results in a pronounced enhancement of the transition enthalpy of the sub-main transition. Neither salt concentration nor equilibration time affected the transition temperature of the sub-main transition. In the densitometry measurements a small volume change is detectable for high salt concentrations. In order to gain further insight into the physical mechanisms involved in the sub-main transition, a Monte Carlo computer simulation study has been carried out using a microscopic model. The combined experimental and simulation results suggest that the sub-main transition involves an acyl chain disordering of phospholipids in lipid bilayer regions that are characterized by a locally decreased lateral pressure most likely caused by a curvature stress.

Thermodynamics of alcohol-lipid bilayer interactions: application of a binding model.

Several recent reports have provided evidence that interactions of small alcohols with lipid bilayer membranes are dominated by adsorption to the membrane-water interface. This mode of interaction is better modeled by binding models than solution theories. In the present study, alcohol-membrane interactions are examined by applying the 'solvent exchange model' [J.A. Schellmann, Biophys. Chem. 37 (1990) 121] to calorimetric measurements. Binding constants (in mole fraction units) for small alcohols to unilamellar liposomes of dimyristoyl phosphatidylcholine were found to be close to unity, and in contrast to partitioning coefficients they decrease through the sequence ethanol, 1-propanol, 1-butanol. Thus, the direct (intrinsic) affinity of the bilayer for these alcohols is lower the longer the acyl chain. A distinction between binding and partitioning is discussed, and it is demonstrated that a high concentration of solute in the bilayer (large partitioning coefficients) can be obtained even in cases of weak binding. Other results from the model suggest that the number of binding sites on the lipid bilayer interface is 1-3 times the number of lipid molecules and that the binding is endothermic with an enthalpy change of 10-15 kJ/mol. Close to the main phase transition of the lipid bilayer the results suggest the presence of two distinct classes of binding sites: 'normal' sites similar to those observed at higher temperatures, and a lower number of high-affinity sites with binding constants larger by one or two orders of magnitude. The occurrence of high-affinity sites is discussed with respect to fluctuating gel and fluid domains in bilayer membranes close to the main phase transition.

Association of ethanol with lipid membranes containing cholesterol, sphingomyelin and ganglioside: a titration calorimetry study.

The association of ethanol at physiologically relevant concentrations with lipid bilayers of different lipid composition has been investigated by use of isothermal titration calorimetry (ITC). The liposomes examined were composed of combinations of lipids commonly found in neural cell membranes: dimyristoyl phosphatidylcholine (DMPC), ganglioside (GM(1)), sphingomyelin and cholesterol. The calorimetric results show that the interaction of ethanol with fluid lipid bilayers is endothermic and strongly dependent on the lipid composition of the liposomes. The data have been used to estimate partitioning coefficients for ethanol into the fluid lipid bilayer phase and the results are discussed in terms of the thermodynamics of partitioning. The presence of 10 mol% sphingomyelin or ganglioside in DMPC liposomes enhances the partitioning coefficient by a factor of 3. Correspondingly, cholesterol (30 mol%) reduces the partitioning coefficient by a factor of 3. This connection between lipid composition and partitioning coefficient correlates with in vivo observations. Comparison of the data with the molecular structure of the lipid molecules suggests that ethanol partitioning is highly sensitive to changes in the lipid backbone (glycerol or ceramide) while it appears much less sensitive to the nature of the head group.