Phase Transition Of Phospholipid Bilayers Research Articles

How Do Membranes Respond to Pressure?

Bilayers formed by phospholipids are fundamental structures of biological membranes. The mechanical perturbation brought about by pressure significantly affects the membrane states of phospholipid bilayers. In this chapter, we focus our attention on the pressure responsivity for bilayers of some major phospholipids contained in biological membranes. At first, the membrane states and phase transitions of phospholipid bilayers depending on water content, temperature and pressure are explained by using the bilayer phase diagrams of dipalmitoylphosphatidylcholine (DPPC), which is the most familiar phospholipid in model membrane studies. Subsequently, the thermotropic and barotropic bilayer phase behavior of various kinds of phospholipids with different molecular structures is discussed from the comparison of their temperature--pressure phase diagrams to that of the DPPC bilayer. It turns out that a slight change in the molecular structure of the phospholipids produces a significant difference in the bilayer phase behavior. The systematic pressure studies on the phase behavior of the phospholipid bilayers reveal not only the pressure responsivity for the bilayers but also the role and meaning of several important phospholipids existing in real biological membranes.

Intramitochondrial crystalline inclusions in nonalcoholic steatohepatitis

Mitochondrial dysfunction is an important element in the pathogenesis of nonalcoholic steatohepatitis (NASH). Intramitochondrial crystals (IMCs) are a well-documented morphological abnormality seen on transmission electron microscopy in this disease. It has been suggested that IMCs consist of phospholipids, but their exact composition remain uncertain many years after their discovery. Micellar phase transitions of phospholipid bilayers is a well-known but little-studied phenomenon in living systems. Its presence in the mitochondria of NASH would offer significant insight into the disease with possible therapeutic implications. We postulated that intramitochondrial disturbances in NASH are sufficient to produce such transitions and that their detection in fresh biopsies would therefore be a dynamic process. To test this, we performed a blinded, prospective analysis of fresh liver biopsy samples immediately fixed under different conditions. Quantitative transmission electron microscopy morphometry, performed by systematically counting total mitochondria and IMCs within areas of uniform dimension, showed a stepwise decline in IMCs with cooler fixation temperature in each subject studied. Randomization testing (Monte Carlo resampling) confirmed that the detection of IMCs was strongly dependent on fixation temperature (P < 0.0001). These results indicate that the intramitochondrial crystals characteristic of NASH are highly dynamic and unstable structures. The findings offer the strongest support yet for their origin in micellar phase transitions. We speculate that such transitions result from microenvironmental changes within the mitochondria and carry therapeutic implications, especially in regard to dietary manipulations of mitochondrial lipid composition.

Cooperativity and Kinetics of Phase Transitions in Nanopore-Confined Bilayers Studied by Differential Scanning Calorimetry

The first-order nature of the gel-to-liquid crystal phase transition of phospholipid bilayers requires very slow temperature rates in differential scanning calorimetry (DSC) experiments to minimize any rate-dependent distortions. Proportionality of the DSC signal to the rate poses a problem for studies of substrate-supported bilayers that contain very small volumes of the lipid phase. Recently, we described lipid bilayers self-assembled inside nanoporous substrates. The high density of the nanochannels in these structures provides at least a 500-fold increase in the bilayer surface area for the same size of the planar substrate chips. The increased surface area enables the DSC studies. The rate-dependent DSC curves were modeled as a convolution of a conventional van’t Hoff shape and a first-order decay curve of the lipid rearrangement. This analysis shows that although confinement of bilayers to the nanopores of ∼177nm in diameter results in a more than threefold longer characteristic time of the lipid rearrangement and a decrease in the cooperative unit number, the phase transition temperature is unaffected.

Effects of ethanol on lipid bilayers with and without cholesterol: the distearoylphosphatidylcholine system

Differential scanning calorimetry (DSC) and fluorescence spectroscopy are useful techniques for investigating the phase transitions of phospholipid bilayers. In this study, these methods have been extended to determine the effects of ethanol on DSPC and DSPC/2 mol.% cholesterol bilayers. The biphasic effect of the main transition was observed on the DSC heating scans above 0.60 M ethanol. In addition, the concentration at which the biphasic effect occurs is not significantly changed in the presence of 2 mol.% cholesterol. For the fluorescence studies, 1,6-diphenyl-1,3,5-hexatriene (DPH) has been incorporated into the bilayer to monitor the phase transitions through the displacement of DPH. This fluorescent probe is used to directly determine the onset of interdigitation in the bilayer systems as indicated by a large decrease in the DPH fluorescence intensity. The addition of cholesterol lowered and broadened the transition temperatures of the phosphatidylcholine (PC) system. However, 2 mol.% cholesterol did not have a significant effect on the induction of the interdigitated phase in DSPC as observed from the small difference in ethanol threshold concentration for the two systems. This suggests that DSPC forms a more stable interdigitated gel phase than other PCs with shorter acyl chains.

Differential Scanning Calorimetric Study of Bilayer Membrane Phase Transitions. A Biophysical Chemistry Experiment

In a series of experiments designed for a biologically-oriented physical chemistry lab, the gel to liquid-crystalline phase transition of phospholipid bilayers is examined by differential scanning calorimetry. The four interrelated experiments examine the effect of altering lipid bilayer composition on the thermodynamic properties of the phase transition. Bilayer composition is modified by (i) varying the length of the phospholipid acyl chains, (ii) varying the identity of the phosphate headgroup, (iii) adding cholesterol, or (iv) mixing phospholipids of two different length acyl chains. Results relating bilayer structure to phase transitions for all experiments are presented, and the relationship of these experiments to the study of naturally occurring biomembranes is examined.

23] Investigation of phase transitions in bilayer membranes

This article described three techniques used to study phase transitions in phospholipid bilayers. The complementarity of the three techniques in characterizing the thermotropic and structural properties of phospholipid bilayers has been demonstrated by describing their use to characterize a series of mixed-chain-length PCs. It has been shown that an understanding of the energetics that govern the packing of phospholipid chains in the gel phase can be used to construct a model to interpret thermodynamic data of the PCs. This model, in turn, provided a framework for designing and interpreting the Raman spectroscopic and X-ray diffraction experiments on this series of phospholipids. The result was a complete description of the phase transitions and gel phase packing properties of the mixed-chain-length PCs. The phase diagram of Fig. 5B has been expanded to include the mixed-chain-length PC series C18C18PC through C18C0PC. Furthermore, the phase diagram and the chain inequivalence parameter have been shown to describe the behavior of any mixed-chain-length PC, irrespective of the lengths of the hydrocarbon chains or the position of the chains on the glycerol backbone. This is demonstrated by the additional mixed-chain-length PCs plotted in Fig. 5B. With minor modifications, the phase diagram also accurately describes the behavior of mixed-chain-length phosphatidylethanolamines, sphingomyelins, and unsaturated PCs. Finally, it has been demonstrated that a correlation exists between the thermodynamic and the Raman spectroscopic parameters determined for the phase transition of phospholipid bilayers. This correlation is based on the common chain energetics being measured by these two techniques.

A Phase Diagram Including the Intermediate Rippled Phase of Phospholipid Bilayers

A Landau free energy proposed for the phase transitions in phospholipid bilayers has been reexamined, and the phase diagram in the adopted parameter space is presented. The temperature dependences of specific heat and the elastic stiffness constant associated with the order parameter derived from the given free energy function are shown.

Phenylbutazone Action on Dimiristoyl Phosphatidylcholine Liposome Phase Transition and 8-Anilino-1-naphthalene Sulfonate Binding

Phenylbutazone (PhB), a powerful anti-inflammatory drug, is able to modify the phase transition of phospholipid bilayers without changing its calorimetric enthalpy (ΔHcal), as can be shown by differential scanning calorimetry (DSC) experiments. Under PhB interaction, dimiristoyl phosphatidylcholine (DMPC) multilamellar liposomes (MLV) undergo lateral phase separation as a result of immiscibility in the bilayer plane. On the other hand, the binding of the anionic fluorescent probe 8-anilino-1-naphthalene sulfonate (ANS) to the surface of DMPC liposomes is altered by PhB. Even though the quantum efficacy of the probe fluorescence emission remains unaffected, the negative cooperativity of the binding process disappears, with the intrinsic dissociation constant showing only a minor variation. From these results it is concluded that PhB would be most likely located close to the lipid:water interface.

Phase transitions in phospholipid bilayers: Lateral phase separations play vital roles in biomembranes

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Analysis of the chainlength dependence of lipid phase transition temperatures: Main and pretransitions of phosphatidylcholines; main and non-lamellar transitions of phosphatidylethanolamines

The dependence on chainlength, n, of the temperatures at which the various thermotropic phase transitions in phospholipid bilayers take place can be fitted to high accuracy with an expression of the form: T t = T t ∞ (1 − A/( n − d)) where T t ∞, A and d are the constants to be fitted. This expression works well with literature values for the main and pretransitions of saturated straight-chain phosphatidylcholines, for the main transition of isobranched- and anteiso-branched-chain phosphatidylcholines, and for the main and non-lamellar phase transitions of saturated straight-chain phosphatidylethanolamines, and also for diacylglycerol glycolipids. The parameters in the fit can be related thermodynamically to the calorimetric properties of the phase transition. The constants A and d are related to the end effects in the chainlength dependence of the transition enthalpy and transition entropy, respectively, and the transition temperature extrapolated to infinite chainlength, T t ∞, is determined by the ratio of the incremental transition enthalpy and transition entropy per methylene group. A reasonable correspondence is found between the chainlength dependence of the transition temperature and of the transition enthalpy and transition entropy for saturated straight-chain diacylphosphatidylcholines. It is likely, however, that the expression for the chainlength dependence of the transition temperature may be of more general applicability, because, on the one hand of the greater inherent precision of the transition temperature measurements, and on the other hand because the expression for the transition temperature is of more general validity, provided that the chainlength dependence of the incremental transition enthalpy and entropy are the same.

A solid-solution theory of anesthetic interaction with lipid membranes: temperature span of the main phase transition

Anesthetics (or any other small additives) depress the temperature of the main phase transition of phospholipid bilayers. Certain anesthetics widen the temperature span of the transition, whereas others do not. The widening in a first-order phase transition is intriguing. In this report, the effects of additive molecules on the temperature and its span were explained by the solid-solution theory. By assuming coexistence of the liquid-crystal and solid-gel phases of lipid membranes at phase transition, the phase boundary is determined from the distribution of anesthetic molecules between the liquid-crystal membrane versus water and between the solid-gel membrane versus water. The theory shows that when the lipid concentration is large or when the lipid solubility of the drug is large, the width of the transition temperature increases, and vice versa. Highly lipid-soluble molecules, such as long-chain alkanols and volatile anesthetics, increase the width of the transition temperature when the lipid: water ratio is large, whereas highly water-soluble molecules, such as methanol and ethanol, do not. The aqueous phase serves as the reservoir for anesthetics. Depletion of the additive molecules from the aqueous phase is the cause of the widening. When the reservoir capacity is large, the temperature width does not increase. The theory also predicts asymmetry of the specific heat profile at the transition.

Effect of surface ionization of dimyristolyphosphatidic acid vesicle membranes on the main phase-transition enthalpy and temperature

The main phase transition of phospholipid bilayers is a property expressed by the order-disorder conformational change of the lipid tails. Nevertheless, with ionizable phospholipids, changes in the surface charge have large effects on the membrane properties. The free energy of a charged phospholipid membrane depends on the degree of ionization, area per phospholipid molecule, and the temperature. Here, the effect of surface electrostatic charges on the temperature and the enthalpy of the main phase transition of dimyristoylphosphatidic acid vesicle membranes is analyzed. A simple equation is presented that describes the relationship among the surface charge density, the phase-transition temperature, the surface area ratio between solid and liquid membranes, and the excess enthalpy. The theory indicated that the pH-induced shift in the excess enthalpy is attributable to the change in the surface area ratio between the solid and liquid membranes.

Detection of slow-motional changes in the phosphatidylcholine bilayers by 1H- 31P cross-polarization dynamics

Cross-polarization efficiency was shown to reflect the phase transitions of phospholipid bilayers. A quantitative analysis of the cross-polarization dynamics in dipalmitoylphosphatidylcholine bilayers was carried out and the efficiency was interpreted in terms of the cross-relaxation time and the spin-lattice relaxation time of protons in the rotating frame. Except for the subgel phase, the cross-relaxation time was shown to be orientation dependent and to reflect the fluctuation of the director of the polar head group. Temperature dependence of the spin-lattice relaxation time in the rotating frame showed characteristic changes at the sub-, pre-, and main transitions, respectively. The change of cross-relaxation time and chemical-shift anisotropy at the pretransition clearly showed that the pretransition significantly affects the dynamic state of the polar head group.

Thermotropic properties and molecular dynamics of cholesteryl ester rich very low density lipoproteins: effect of hydrophobic core on polar surface.

Cholesteryl ester rich very low density lipoproteins (CER-VLDL), isolated from the plasma of rabbits fed a hypercholesterolemic diet, have been studied by differential scanning calorimetry (DSC), 13C nuclear magnetic resonance (NMR), and spin-label electron paramagnetic resonance (EPR) to determine the temperature-dependent dynamics of cholesteryl esters in the hydrophobic core and of phospholipids on the polar surface. Intact CER-VLDL exhibit two DSC heating endotherms; these occur at 40-42 and 45-48 degrees C. Cholesteryl esters isolated from CER-VLDL also exhibit two DSC endotherms; these occur at 50.0 and 55.1 degrees C and correspond to the smectic----cholesteric and cholesteric----isotropic liquid-crystalline phase transitions. A model mixture containing cholesteryl linoleate, oleate, and palmitate in a ratio (0.21, 0.51, and 0.28 mol fraction) similar to that in CER-VLDL exhibited comparable DSC endotherms at 45.2 and 51.5 degrees C. CER-VLDL at 37 degrees C gave 13C NMR spectra that contained no resonances assignable to cholesteryl ring carbons but detectable broad resonances for some fatty acyl chain carbons, suggesting the cholesteryl esters were in a liquid-crystalline state. When the mixture was heated to 42 degrees C, broad ring carbon resonances became detectable; at 48 degrees C, they became narrow, indicating the cholesteryl esters were in an isotropic, liquid-like state. With increasing temperature over the range 38-60 degrees C, the resonances for cholesteryl ring carbons C3 and C6 in CER-VLDL narrowed differentially. Similar spectral changes were observed for the synthetic cholesteryl ester mixture, except they occurred at temperatures about 10 degrees C higher. These results indicate that the two DSC transitions in CER-VLDL do not directly correlate with the smectic----cholesteric and cholesteric----isotropic transitions exhibited by pure cholesteryl esters. (5-Doxylpalmitoyl)-phosphatidylcholine (5-DP-PC) and (12-doxylstearoyl)phosphatidylcholine (12-DS-PC) were used to probe the polar surface monolayer of CER-VLDL; the corresponding cholesteryl esters (5-DP-CE and 12-DS-CE) were used to probe the hydrophobic core. None of these probes in CER-VLDL detected an abrupt change in EPR order parameters, S, or maximum splitting, 2T max, over the temperature range 20-58 degrees C even though 12-DS-PC and 5-DP-PC can detect phase transitions in phospholipid bilayers and 12-DS-CE and 5-DP-CE can detect phase transitions in neat cholesteryl esters. However, 12-DS-CE and 5-DP-CE did detect a much greater acyl chain order for the neutral lipids of CER-VLDL than for those of normal triglyceride-rich VLDL.(ABSTRACT TRUNCATED AT 400 WORDS)

Study of a model for the main phase transition of lipid bilayers using the Kikuchi cluster method

The statistical mechanics of Doniach's two-state lattice model for the main gel – liquid crystal phase transition of phospholipid bilayers is treated in a similar manner to order–disorder transformations in binary alloys and magnetic systems, using the cluster variation method developed by Kikuchi. Indeed, the analogy holds better for the latter system, since the entropy difference between the two states gives rise to an effective temperature-dependent field. This effective field vanishes at the first-order phase transition, whose latent heat is associated with the discontinuity in the order parameter.We use Kikuchi's approximation with the inclusion of triangle bond correlations, and pair and site probabilities in the expression for free energy. We assume that the lipid chains only interact through nearest neighbour pair potentials and that triangle correlations are important for approximate counting of allowed states. Two long-range order parameters and a short-range order parameter are introduced in the formulation of the theory. Both long-range order parameters are discontinuous at the transition temperature. Numerical results for the physical quantities are presented and discussed with respect to earlier work.

The effects of water-soluble solutes on the phase transitions of phospholipids.

A previous treatment [Sturtevant, J.M. (1982) Proc. Natl. Acad. Sci. USA 79, 3963-3967], based on ideal solution theory, of the influence of small molecules on the main phase transitions of phospholipid bilayers is extended to include the effect of water solubility of the added solute. This effect is manifested primarily in a sharpening of the phase transition, resulting from a buffering by the aqueous phase of the concentration of the solute in the lipid phases.

Thermotropic behaviour of phospholipid vesicles reconstituted with rat liver microsomal cytochrome P-450

The interaction of rat liver microsomal cytochrome P-450 with phospholipids has been investigated employing differential scanning microcalorimetry. It is shown that the thermotropic behaviour of phospholipid vesicles reconstituted with cytochrome P-450 depends on liposome composition, protein concentration and the mode of reconstituted system preparation. From the ΔH dependence on protein concentration in proteoliposomes it was calculated that one cytochrome P-450 molecule influence 350±50 dimyristoylphosphatidylcholine (DMPC) molecules. The electrostatic interaction of cytochrome P-450 and negatively charged phospholipid, phosphatidylinositol (PI), mixed with DMPC involves the temperature stabilization of proteoliposomes at a phase transition of phospholipid bilayers. The thermal denaturation temperature is increased due to negatively charged PI added.

Interfacial preference of anesthetic action upon the phase transition of phospholipid bilayers and partition equilibrium of inhalation anesthetics between membrane and deuterium oxide

The half-height linewidth v 1 2 of the 1H-NMR spectra of dipalmitoylphosphatidylcholine vesicles changes abruptly at the phase transition temperature. In the absence of inhalation anesthetics, proton signals from the choline head group (hydrophilic interface) and acyl-chain tails (lipid core) change at the same temperature of 39.6°C. The present study compared the effect of four inhalation anesthetics, i.e., methoxyflurane, chloroform, halothane and enflurane, upon the ligand-induced phase transition of phosphatidylcholine vesicle membranes at 37°C. The anesthetics showed differential action upon the phase transition of the phospholipid vesicle membranes between the lipid core and the hydrophilic interface. The concentrations of anesthetics which induced the phase transition of the lipid core were about 2-fold greater than those required for the phase transition of the interfacial choline head groups. From the area under the proton signals of inhalation anesthetics in the NMR spectra, the maximum solubilities of methoxyflurane, chloroform and halothane in 2H 2O at 37°C were determined to be 0.671 · 10 -4 , 2.637 · 10 -4 and 1.398 · 10 -4 (expressed as mole fractions), or 3.35, 13.17 and 6.98 mmol/1 000 g 2H 2O, respectively. The solubilities of the anesthetic vapor in 2H 2O expressed as mole fractions according to Henry's law were 9.586 · 10 -4 , 6.432 · 10 -4 and 2.311 · 10 -4/ atm ( 1.013 · 10 5 Pa ) partial pressure, respectively. The presence of phospholipid vesicles in 2H 2O increased the solubility of the inhalation anesthetics. From difference between solubility in 2H 2O and a dipalmitoylphosphatidylcholine vesicle suspension, the partition coefficients of methoxyflurane, chloroform and halothane between the phospholipid vesicle membranes and 2H 2O were estimated. These values, calculated from the mole fractions, were 3364, 1660 and 3850, respectively at 37°C.

Landau phenomenological theory of the pressure-induced phase transition of phospholipid bilayers

A theoretical model is developed to describe the temperature and pressure-induced phase transitions of phospholipid bilayer membranes simultaneously. By means of this model the phase transition enthalpies,-temperatures,-pressures and volume changes of different phospholipid membranes are calculated and compared with the available experimental data. Thep−T diagrams of the membranes are obtained, too. Phase diagrams for binary mixtures with components differing in chain length by 2 and 4 carbons are determined at different pressures. It is found that diagrams shift without significant deformation in their shape towards higher temperatures with increasing pressure.

Non-electrostatic contribution to the titration of the ordered-fluid phase transition of phosphatidylglycerol bilayers

The ordered-fluid phase transition of phospholipid bilayers is of interest not only because it yields information on the nature of the fluid bilayer regions in biomembranes, but also because it may provide a means for triggering lateral phase separation between the ordered and fluid lipids in membranes [ 1,2]. It has been shown [3-61 that the phase transition of negatively charged lipid bilayers shows strong temperature shifts on titrating the phosphate group of the lipid polar head. This provides a way of triggering the phase transition at constant temperature by varying the pH. The free energy of the bilayer made of protonated lipids, G* = H* X.7*, changes upon deprotonation to G* GPr and this change results in a shift of the phase transition temperature from q = LUYF/LU~ to Tt , given by: