Phase transition in dioctadecyldimethylammonium bromide and chloride vesicles prepared by different methods

The gel to liquid crystalline phase transition of the double-chained cationic dioctadecyldimethylammonium chloride and bromide (DODAX, X = Cl- or Br-) in aqueous vesicle dispersions prepared by non-sonication. sonication and extrusion has been investigated using high-sensitivity differential scanning calorimetry (DSC). The transition temperature (Tm) is a function of the preparation method, amphiphile concentration, vesicle curvature and nature of the counterion. DSC thermograms for DODAB and DODAC non-sonicated vesicle dispersions exhibit a single endothermic peak at Tm roughly independent of concentration up to 10 mM. Extrusion broadens the transition peak and shifts Tm downwards. Sonication, however, broadens slightly the transition peak and tends to shift Tm upwards suggesting that extrusion and sonication form vesicles with different characteristics. DODAC always exhibits higher Tm than DODAB irrespective of the preparation method. Tm changes as follows: Tm (sonicated) > or = Tm (non-sonicated) > Tm (extruded). Hysteresis of about 7 degrees C was observed for DODAB vesicle dispersions.     

Effect of staphylococcal delta-lysin on the thermotropic phase behavior and vesicle morphology of dimyristoylphosphatidylcholine lipid bilayer model membranes. Differential scanning calorimetric, 31P nuclear magnetic resonance and Fourier transform infrared spectroscopic, and X-ray diffraction studies

We investigated the effects of various concentrations of staphylococcal delta-lysin on the thermotropic phase behavior of large multilamellar dimyristoylphosphatidylcholine (DMPC) vesicles by differential scanning calorimetry (DSC), 31P nuclear magnetic resonance (NMR) and Fourier transform infrared (FTIR) spectroscopy, and X-ray diffraction. The DSC studies revealed that at all concentrations, the addition of delta-lysin progressively decreases the enthalpy of the pretransition of DMPC bilayers without significantly affecting its temperature or cooperativity. Similarly, the addition of smaller quantities of peptide has little effect on the temperature of the main phase transition of DMPC bilayers but does reduce the cooperativity and enthalpy of this transition somewhat. However, at higher peptide concentrations, a second phase transition with a slightly increased temperature and a markedly reduced cooperativity and enthalpy is also induced, and this latter phase transition resolves itself into two components at the highest peptide concentrations that are tested. Moreover, our 31P NMR spectroscopic studies reveal that at relatively low delta-lysin concentrations, essentially all of the phospholipid molecules produce spectra characteristic of the lamellar phase, whereas at the higher peptide concentrations, an increasing proportion exhibit an isotropic signal. Also, at the highest delta-lysin concentrations that are studied, the isotropic component of the 31P NMR spectrum also resolves itself into two components. At the highest peptide concentration that was tested, we are also able to effect a macroscopic separation of our sample into two fractions by centrifugation, a pellet containing relatively smaller amounts of delta-lysin and a supernatant containing larger amounts of peptide relative to the amount of lipid present. We are also able to show that the more cooperative phase transition detected calorimetrically, and the lamellar phase 31P NMR signal, arise from the pelleted material, while the less cooperative phase transition and the isotropic 31P NMR signal arise from the supernatant. In addition, we demonstrate by X-ray diffraction that the pelleted material corresponds to delta-lysin-containing large multilamellar vesicles and the supernatant to a mixture of delta-lysin-containing small unilamellar vesicles and discoidal particles. We also show by FTIR spectroscopy that delta-lysin exists predominantly in the alpha-helical conformation in aqueous solution or when interacting with DMPC, and that a large fraction of the peptide bonds undergo H-D exchange in D(2)O. However, upon interaction with DMPC, the fraction of exchangeable amide protons decreases. We also demonstrate by this technique that both of the phase transitions detected by DSC correspond to phospholipid hydrocarbon chain-melting phase transitions. Finally, we show by several techniques that the absolute concentrations of delta-lysin and the thermal history, as well as the lipid:peptide ratio, can affect the thermotropic phase behavior and morphology of peptide-lipid aggregates.     

Interaction of short-chain lecithin with long-chain phospholipids: characterization of vesicles that form spontaneously

Stable unilamellar vesicles formed spontaneously upon mixing aqueous suspensions of long-chain phospholipid (synthetic, saturated, and naturally occurring phosphatidylcholine, phosphatidylethanolamine, and sphingomyelin) with small amounts of short-chain lecithin (fatty acid chain lengths of 6-8 carbons) have been characterized by using NMR spectroscopy, negative staining electron microscopy, differential scanning calorimetry, and Fourier transform infrared (FTIR) spectroscopy. This method of vesicle preparation can produce bilayer vesicles spanning the size range 100 to greater than 1000 A. The combination of short-chain lecithin and long-chain lecithin in its gel state at room temperature produces relatively small unilamellar vesicles, while using long-chain lecithin in its liquid-crystalline state produces large unilamellar vesicles. The length of the short-chain lecithin does not affect the size distribution of the vesicles as much as the ratio of short-chain to long-chain components. In general, additional short-chain decreases the average vesicle size. Incorporation of cholesterol can affect vesicle size, with the solubility limit of cholesterol in short-chain lecithin micelles governing any size change. If the amount of cholesterol is below the solubility limit of micellar short-chain lecithin, then the addition of cholesterol to the vesicle bilayer has no effect on the vesicle size; if more cholesterol is added, particle growth is observed. Vesicles formed with a saturated long-chain lecithin and short-chain species exhibit similar phase transition behavior and enthalpy values to small unilamellar vesicles of the pure long-chain lecithin prepared by sonication. As the size of the short-chain/long-chain vesicles decreases, the phase transition temperature decreases to temperatures observed for sonicated unilamellar vesicles. FTIR spectroscopy confirms that the incorporation of the short-chain lipid in the vesicle bilayer does not drastically alter the gauche bond conformation of the long-chain lipids (i.e., their transness in the gel state and the presence of multiple gauche bonds in the liquid-crystalline state).     

Thermotropic behavior of dipalmitoylphosphatidylcholine vesicles reconstituted with the glycoprotein of vesicular stomatitis virus

The vesicular stomatitis virus glycoprotein reconstituted into dipalmitoylphosphatidylcholine (DPPC) vesicles exerts a profound effect upon the DPPC gel to liquid-crystalline phase transition. The glycoprotein was reconstituted into DPPC vesicles by octyl glucoside dialysis. The gel to liquid-crystalline phase transition of these vesicles was monitored by differential scanning calorimetry. Vesicles formed in the absence of glycoprotein (600--2100-A diameter) underwent the phase transition at 41.0 degrees C and had an associated enthalpy change of 8.0 +/- 1.6 kcal/mol. Increasing the mole ratio of glycoprotein to DPPC in the vesicles to 0.15 mol % reduced both the transition temperature and the transition enthalpy change. The enthalpy change as a function of the mole percent glycoprotein could be fit to a straight line by a least-squares procedure. Extrapolation of the results to the glycoprotein concentration where the enthalpy change was zero indicated one glycoprotein molecule bound 270 +/- 150 molecules of DPPC.     

The effect of potential-sensitive molecular probes on the thermal phase transition in dimyristoylphosphatidylcholine preparations

Differential scanning calorimetry (DSC) has been employed to determine the effect of five commonly employed extrinsic potential-sensitive probes on phase transitions of multilamellar suspensions of L-alpha-dimyristoylphosphatidylcholine (DMPC). At mol% values of less than five, the effect of these probes on the excess heat capacity curve in the vicinity of the gel to liquid crystal phase transition can be described by an equation based on the formation of ideal solutions in both phases. Even at up to 4 mol%, these dyes only moderately reduce the enthalpy change associated with this transition, but cause a marked decrease in the size of the cooperative unit parameter. The excess heat capacity profile for diS-C3-(5) is represented by the ideal solution equation, even at 12 mol%, whereas the suspensions with the other probes present at this level have profiles covering large temperature ranges. Multiple peaks appear at the higher levels for the negatively charged oxonols V and VI, and merocyanine 540, a result consistent with the presence of well-defined microdomains or even phase separation. The enthalpy change associated with the transition near 15 degrees C involving packing in the headgroup region is decreased significantly, indicating that the probes probably affect the lipid headgroup conformation, even at low levels. The cyanine probe diS-C3-(5) causes the heat capacity profile of small unilamellar vesicles to be transformed very rapidly into one similar to that of the vortexed lipid preparations, presumably by a dye-mediated vesicle fusion process, enhanced by the surface location of this probe. All our results are consistent with diS-C3-(5) being located on the surface of the bilayer in both phases, but a penetration of the other probes into the hydrocarbon region, at least in the liquid crystal phase.     

Dantrolene inhibits halothane-induced membrane reorganization A study using 31P-NMR and differential scanning calorimetry

The action of the relaxing agent dantrolene on dipalmitoylphosphatidylcholine (DPPC) model membranes in the presence and absence of the general anesthetic halothane has been investigated by DSC and 31P-NMR. Dantrolene has a weak effect on both the thermodynamic and NMR parameters of the pure model membrane. When halothane is present in the system, the relaxing agent acts to counterbalance the strong anesthetic-induced membrane perturbation. This is reflected in DSC experiments by a change of the enthalpy variation (delta H) and of the main gel-to-fluid phase transition temperature (Tc) towards the values of the pure lipid system. The amount of halothane-induced small tumbling vesicles, as detected by 31P-NMR by the superposition of an isotropic line on a lamellar-type powder spectrum, is considerably reduced upon dantrolene addition. This means that the relaxing agent "cures" the membrane de-structuring action promoted by halothane. Membranes first treated with dantrolene are also protected from the halothane perturbation. So, the relaxing agent is both "curative" and "preventative" against halothane. The optimum effect is obtained for 1 dantrolene molecule per ca 34 halothane molecules. The mechanisms of action were discussed in relation to membrane fluidity.     

Effect of cholesterol on the valinomycin-mediated uptake of rubidium into erythrocytes and phospholipid vesicles

Human erythrocytes have been treated with lipid vesicles in order to alter the cholesterol content of the cell membrane. Erythrocytes have been produced with cholesterol concentrations between 33 and 66 mol% of total lipid. The rate of valinomycin-mediated uptake of rubidium into the red cells at 37°C was lowered by increasing the cholesterol concentration of the cell membrane. Cholesterol increased the permeability to valinomycin at 20°C of small (less than 50 nm), unilamellar egg phosphatidylcholine vesicles formed by sonication. Cholesterol decreased the permeability to valinomycin at 20°C of large (up to 200 nm) unilamellar egg phosphatidylcholine vesicles formed by freezethaw plus brief sonication. It is concluded that cholesterol increases the permeability of small membrane vesicles to hydrophobic penetrating substances while above the transition temperature but has the opposite effect on large membrane vesicles and on the membranes of even larger cells.     

Effect of cholesterol on the valinomycin-mediated uptake of rubidium into erythrocytes and phospholipid vesicles.

Human erythrocytes have been treated with lipid vesicles in order to alter the cholesterol content of the cell membrane. Erythrocytes have been produced with cholesterol concentrations between 33 and 66 mol% of total lipid. The rate of valinomycin-mediated uptake of rubidium into the red cells at 37 degrees C was lowered by increasing the cholesterol concentration of the cell membrane. Cholesterol increased the permeability to valinomycin at 20 degrees C of small (less than 50 nm), unilamellar egg phosphatidylcholine vesicles formed by sonication. Cholesterol decreased the permeability to valinomycin at 20 degrees C of large (up to 200 nm) unilamellar egg phosphatidylcholine vesicles formed by freeze-thaw plus brief sonication. It is concluded that cholesterol increases the permeability of small membrane vesicles to hydrophobic penetrating substances while above the transition temperature but has the opposite effect on large membrane vesicles and on the membranes of even larger cells.     

Crystallization of water in multilamellar vesicles

The two-step crystallization of water in multilamellar vesicles (MLVs) of phosphatidylcholines has been investigated. The main crystallization occurs near -15 degrees C and involves bulk water. Contrary to unilamellar vesicles, a sub-zero phase transition is observed for MLVs at -40 degrees C that corresponds to the crystallization of interstitial water, as proved by Fourier transform infrared absorption and differential scanning calorimetry (DSC) experiments. Furthermore, by means of the DSC method and, more specifically, using the enthalpy change values Delta H(sub) at the sub-zero transition, the number of water molecules per 1,2-dipalmitoylphosphatidylcholine (DPPC) molecule giving rise to this transition has been estimated for different H(2)O/DPPC molar ratios. The curve of the molecular fraction of water molecules involved in the sub-zero transition versus the H(2)O/DPPC molar ratio exhibits a maximum for H(2)O/DPPC equal to 27 (40% in mass of water) and tends towards zero for H(2)O/DPPC ratio values approaching that of the swelling limit of the membrane. A smaller enthalpy value of the sub-zero transition is found for 1-oleoyl-2-palmitoyl-3-phosphatidylcholine (OPPC) than for DPPC. This may be explained by the decrease of interstitial water's quantity when the lipid contains an unsaturated chain. When troxerutin, a hydrophilic drug, is added to the DPPC multilayers, the decrease of Delta H(sub) and melting enthalpy of bulk water is attributed to a decrease of the entropy of the liquid phase owing to the network of water molecules surrounding troxerutin molecules. In all cases, the experiments revealed that the sub-zero transition occurs only in the presence of excess water with respect to the swelling limit of membranes. This evidence could be, at least qualitatively, related to an increase of membrane pressure on interstitial water subsequent to bulk water crystallization.     

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°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_β') 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.     

Membrane homeostasis: thermotropic behaviour of microsomal membrane lipids isolated from livers of rats fed cholesterol-supplemented diets

Differential scanning colorimetry (DSC) has been applied to study the phase transition properties of isolated lipids from liver microsomal membranes of rats fed high cholesterol diets with or without high levels of either saturated (coconut oil) or unsaturated (sunflower seed oil) fat. DSC of aqueous buffer dispersions of liver microsomal lipids exhibited two independent, reversible phase transitions. The dietary cholesterol treatments had their major effect on the temperature at which the lower phase transition (T1) occurred. This transition occurred at a lower temperature when cholesterol was added to the diet, irrespective of the nature of the fatty acid supplement. However the magnitude of decrease was more when cholesterol was fed with sunflower seed oil. Inclusion of cholesterol into the rat diets also lowered the enthalpy values for the lower phase transition (T1). No appreciable effect on the temperature of the higher phase transition (T2) was observed, however the enthalpy values were slightly decreased by cholesterol feeding. These results suggest that certain domains of microsomal lipids, probably containing some relatively higher melting-point lipids, independently undergo solidus or gel formation and this transition (T2) is not greatly affected by dietary cholesterol. On the other hand, domains representing the bulk of the microsomal lipids undergo a phase change (T1) at temperatures which are dependent on cholesterol content and fatty acid profiles of the membrane, which are in turn, modified by dietary cholesterol intake.     

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.     

Intermolecular interactions of lysobisphosphatidic acid with phosphatidylcholine in mixed bilayers

Lysobisphosphatidic acid (LBPA) can be regarded to represent a unique derivative of phosphatidylglycerol. This lipid is highly enriched in late endosomes where it can comprise up to 10-15 mol% of all lipids and in these membranes, LBPA appears to be segregated into microdomains. We studied the thermotropic behavior of pure dioleoyl-LBPA mono- and bilayers using Langmuir-lipid monolayers, electron microscopy, differential scanning calorimetry (DSC), and fluorescence spectroscopy. LBPA formed metastable, liquid-expanded monolayers at an air/buffer interface, and its compression isotherms lacked any indication for structural phase transitions. Neat LBPA formed multilamellar vesicles with no structural transitions or phase transitions between 10 and 80 degrees C at a pH range of 3.0-7.4. We then proceeded to study mixed LBPA/dipalmitoylphosphatidylcholine (DPPC) bilayers by DSC and fluorescence spectroscopy. Incorporating increasing amounts of LBPA (up to X(LBPA) (molar fraction)=0.10) decreased the co-operativity of the main transition for DPPC, and a decrease in the main phase transition as well as pretransition temperature of DPPC was observed yet with no effect on the enthalpy of this transition. In keeping with the DSC data for DPPC, 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC)/LBPA mixed bilayers were more fluid, and no evidence for lateral phase segregation was observed. These results were confirmed using fluorescence microscopy of Langmuir-lipid films composed of POPC and LBPA up to X(LBPA)=0.50 with no evidence for lateral phase separation. As late endosomes are eminently acidic, we examined the effect of lowering pH on lateral organization of mixed PC/LBPA bilayers by DSC and fluorescence spectroscopy. Even at pH 3.0, we find no evidence of LBPA-induced microdomain formation at LBPA contents found in cellular organelles.     

Comparison of the enthalpy state of vesicles of different size by their interaction with alpha-lactalbumin

The characteristics of small unilamellar, large unilamellar and large multilamellar vesicles of dimyristoylphosphatidylcholine and their interaction with alpha-lactalbumin are compared at pH 4. (1) By differential scanning calorimetry and from steady-state fluorescence anisotropy data of the lipophilic probe 1,6-diphenyl-1,3,5-hexatriene it is shown that the transition characteristics of the phospholipids in the large unilamellar vesicles resemble more those of the multilamellar vesicles than of the small unilamellar vesicles. (2) The size and composition of the lipid-protein complex formed with alpha-lactalbumin around the transition temperature of the lipid are independent of the vesicle type used. Fluorescence anisotropy data indicate that in this complex the motions of the lipid molecules are strongly restricted in the presence of alpha-lactalbumin. (3) The previous data and a comparison of the enthalpy changes, delta H, of the interaction of the three vesicle types with alpha-lactalbumin allow us to derive that the enthalpy state of the small unilamellar vesicles just below 24 degrees C is about 24 kJ/mol lipid higher than the enthalpy state of both large vesicle types at the same temperature. The abrupt transition from endothermic to exothermic delta H values around 24 degrees C for large vesicles approximates the transition enthalpy of the pure phospholipid.     

Thermodynamic elucidation of solute-induced lipid interdigitation phase: lipid interactions with hydrophobic versus amphipathic species.

Comparative thermodynamic studies on the interactions of aqueous dispersions of dipalmitoyl phosphatidylcholine (DPPC) bilayer vesicles with hydrophobic and amphipathic species were conducted to elucidate the nature of the solute-induced interdigitated lipid phase. Cyclohexanol, a strong hydrophobic species, lowers the temperature (tm) of the lipid main phase transition from the gel to the liquid-crystalline phase. Unlike ethanol (an amphipathic species), as reported previously, cyclohexanol does not exert a biphasic effect on tm (lowering tm at lower concentrations and raising tm at higher concentrations). At cyclohexanol greater than or equal to 15.4 mg/ml or 0.154 M, the thermogram of DPPC vesicles exhibits a small transition adjacent to the main phase transition but at a lower temperature. In contrast, ethanol does not promote such a small transition. Furthermore, the enthalpy (delta H) of the transition is increased in the presence of cyclohexanol. The sign of the enthalpy change (delta H-delta Ho) is positive and that of the free energy change (delta G-delta Go) is negative, a characteristic of solute-solute hydrophobic interaction. In contrast, DPPC bilayer vesicles exhibit both (delta H-delta Ho) and (delta G-delta Go) greater than 0 in the presence of ethanol in a concentration range where lipid vesicles exist in an interdigitated phase. To support the above distinct thermodynamic observations, fluorescence steady-state polarization (P) measurements were also performed. At the temperature below tm, the value of P decreases as cyclohexanol concentration increases, while a biphasic effect on P was found in the presence of ethanol. These findings support the postulation that the solute-induced interdigitated lipid phase requires the solute molecule to be amphipathic in nature.     

Differential scanning calorimetric study of the effect of the antimicrobial peptide gramicidin S on the thermotropic phase behavior of phosphatidylcholine, phosphatidylethanolamine and phosphatidylglycerol lipid bilayer membranes

We have studied the effects of the antimicrobial peptide gramicidin S (GS) on the thermotropic phase behavior of large multilamellar vesicles of dimyristoylphosphatidylcholine (DMPC), dimyristoylphosphatidylethanolamine (DMPE) and dimyristoyl phosphatidylglycerol (DMPG) by high-sensitivity differential scanning calorimetry. We find that the effect of GS on the lamellar gel to liquid-crystalline phase transition of these phospholipids varies markedly with the structure and charge of their polar headgroups. Specifically, the presence of even large quantities of GS has essentially no effect on the main phase transition of zwitterionic DMPE vesicles, even after repeating cycling through the phase transition, unless these vesicles are exposed to high temperatures, after which a small reduction in the temperature, enthalpy and cooperativity of the gel to liquid-crystalline phase transitions is observed. Similarly, even large amounts of GS produce similar modest decreases in the temperature, enthalpy and cooperativity of the main phase transition of DMPC vesicles, although the pretransition is abolished at low peptide concentrations. However, exposure to high temperatures is not required for these effects of GS on DMPC bilayers to be manifested. In contrast, GS has a much greater effect on the thermotropic phase behavior of anionic DMPG vesicles, substantially reducing the temperature, enthalpy and cooperativity of the main phase transition at higher peptide concentrations, and abolishing the pretransition at lower peptide concentrations as compared to DMPC. Moreover, the relatively larger effects of GS on the thermotropic phase behavior of DMPG vesicles are also manifest without cycling through the phase transition or exposure to high temperatures. Furthermore, the addition of GS to DMPG vesicles protects the phospholipid molecules from the chemical hydrolysis induced by their repeated exposure to high temperatures. These results indicate that GS interacts more strongly with anionic than with zwitterionic phospholipid bilayers, probably because of the more favorable net attractive electrostatic interactions between the positively charged peptide and the negatively charged polar headgroup in such systems. Moreover, at comparable reduced temperatures, GS appears to interact more strongly with zwitterionic DMPC than with zwitterionic DMPE bilayers, probably because of the more fluid character of the former system. In addition, the general effects of GS on the thermotropic phase behavior of zwitterionic and anionic phospholipids suggest that it is located at the polar/apolar interface of liquid-crystalline bilayers, where it interacts primarily with the polar headgroup and glycerol-backbone regions of the phospholipid molecules and only secondarily with the lipid hydrocarbon chains. Finally, the considerable lipid specificity of GS interactions with phospholipid bilayers may prove useful in the design of peptide analogs with stronger interactions with microbial as opposed to eucaryotic membrane lipids.     

A differential scanning calorimetry study of the interaction of lasalocid antibiotic with phospholipid bilayers

Interaction of lasalocid sodium salt (Las-Na) with dipalmitoylphosphatidylcholine (DPPC) as a membrane model was investigated by highly-sensitive differential scanning calorimetry (DSC). The insertion properties of the antibiotic were studied both in multilamellar suspensions and unilamellar vesicles, for Las-Na/DPPC molar ratios (r) ranging from 0.005 to 0.1. The effect of the antibiotic on the lipid thermotropic behavior is concentration dependent and drastically changes at a critical r of 0.04 in both model membranes. Below this ratio, Las-Na molecules interact with DPPC bilayers without disrupting the global organization of the membrane. In the multilamellar systems only the transition cooperativity is affected whereas for the mixed vesicles, a decrease in the enthalpy change suggests a different mode of insertion. Above this ratio, implantation of the antibiotic give rise to lateral phase separation in multilamellar systems. These structural modifications have repercussions on the formation of mixed LAS-Na/DPPC vesicles which seems limited to an r value of 0.04.     

Calorimetric studies of the effects of cholesterol on the phase transition of C(18):C(10) phosphatidylcholine.

Differential scanning calorimetry (DSC) has been employed to study the effects of cholesterol on the phase transition of C(18):C(10) phosphatidylcholine (C(18):C(10)PC). C(18):C(10)PC is an asymmetric mixed-chain phosphatidylcholine known to form mixed-interdigitated structures below the transition temperature and form partially interdigitated lipid bilayers above the transition. Three types of samples were used. The treated sample is the lipid dispersion that had undergone three freeze-thaw cycles and stored at 4 degrees C for more than 48 h. The untreated sample was made by vortexing the dry lipid in 50 mM KCl, without the above-mentioned pretreatment. The cold-treated sample was prepared by incubating the treated sample at -20 degrees C for 15 d. There is no apparent difference in the DSC curves between the treated and cold-treated samples. The data derived from the treated samples seem to be more reproducible. The DSC curves between the cholesterol/C(18):C(10)PC and cholesterol/symmetric diacylphosphatidylcholine mixtures are different in three aspects: overall appearance, the cholesterol dependence of delta H, and the effect of cholesterol on the maximal transition temperature Tm, the onset temperature To, and the completion temperature Tc. for both the treated and untreated samples, the total enthalpy change delta H of the phase transition of C(18):C(10)PC decreases with increasing cholesterol content, approaching zero at approximately 25 mol%. This level is lower than the total enthalpy changes reported previously for the cholesterol/symmetric diacylphosphatidylcholine mixtures. Both the heating and cooling thermograms show that Tm, To, and Tc decrease with increasing cholesterol content. The decreasing rates of these temperatures with cholesterol are in the neighborhood of -0.24 degree per mol% of cholesterol.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of the enthalpy state of vesicles of different size by their interaction with α-lactalbumin

The characteristics of small unilamellar, large unilamellar and large multilamellar vesicles of dimyristoylphosphatidylcholine and their interaction with α-lactalbumin are compared at pH 4. (1) By differential scanning calorimetry and from steady-state fluorescence anisotropy data of the lipophilic probe 1,6-diphenyl-1,3,5-hexatriene it is shown that the transition characteristics of the phospholipids in the large unilamellar vesicles resemble more those of the multilamellar vesicles than of the small unilamellar vesicles. (2) The size and composition of the lipid-protein complex formed with α-lactalbumin around the transition temperature of the lipid are independent of the vesicle type used. Fluorescence anisotropy data indicate that in this complex the motions of the lipid molecules are strongly restricted in the presence of α-lactalbumin. (3) The previous data and a comparison of the enthalpy changes, $\text{ΔH}$, of the interaction of the three vesicle types with α-lactalbumin allow us to derive that the enthalpy state of the small unilamellar vesicles just below 24°C is about 24 kJ/mol lipid higher than the enthalpy state of both large vesicle types at the same temperature. The abrupt transition from endothermic to exothermic $\text{ΔH}$ values around 24°C for large vesicles approximates the transition enthalpy of the pure phospholipid     

Protection by sugars against phase transition-induced leak in hydrated dimyristoylphosphatidylcholine liposomes

The disaccharides trehalose and sucrose have small effects on temperature and enthalpy of the pre- and main phase transition in hydrated DMPC bilayers. In contrast, these sugars cause a considerable retention of carboxyfluorescein when large unilamellar vesicles of DMPC are heated through the main transition. This effect is sugar specific, as the monosaccharides glucose and fructose are less effective and ethyleneglycol has no effect at all.