Ultrastructural analysis of black lipid membranes

Black films of egg phosphatidylcholine were investigated by electron microscopy of thin sections. A stabilizing method was developed based on a tricomplex reaction of the lipid, in order to avoid difficulties of interpretation pertaining to OsO₄ and KMnO₄-fixation techniques. Membranes were essentially trilaminar, as could be demonstrated by specimen tilting, and extensively undulated. Densitometer tracings of micrographs revealed a peak to peak width of 45 Å. Corrected for electron radiation damage this width would amount to 60Å. Taking into account the thickness of the polar layers, this would mean a total thickness of 69 Å of the native membrane

The distribution of cholesterol in bilayers of phosphatidylcholines as visualized by freeze fracturing

The crystallization behavior of bilayers of synthetic phosphatidylcholines in the presence of cholesterol was investigated by freeze fracturing. 1. 1. Below the lipid-phase transition, cholesterol is randomly distributed over the lateral plane of a bilayer of the single species dimyristoylphosphatidylcholine, as far as can be detected within the lateral resolution of freeze fracturing. 2. 2. In an equimolar mixture of dilauroyl- and dimyristoylphosphatidylcholine, which shows cocrystallization, cholesterol is distributed randomly in the bilayers below the phase-transition temperature. 3. 3. In an equimolar mixture of dipalmitoyl and 1-palmitoyl-2-oleoylphosphatidylcholine, which shows monotectic behaviour, cholesterol interacts preferentially with the liquid crystalline species 1-palmitoyl-2-oleoylphosphatidylcholine when the other component dipalmitoylphosphatidylcholine passes from the liquid crystalline to the gel state upon cooling the mixture

Influence of Ca2+ and Mg2+ on the thermotropic behaviour and permeability properties of liposomes prepared from dimyristoyl phosphatidylglycerol and mixtures of dimyristoyl phosphatidylglycerol and dimyristoyl phosphatidylcholine

1. 1. Calorimetric experiments showed a marked effect of Ca2+ and Mg2+ on the thermotropic behaviour of dimyristoyl phosphatidylglycerol. 2. 2. Concentrations of Ca2+ and Mg2+ lower than 1 ion to 2 molecules of phosphatidylglycerol produced a shift of the phase transition to higher temperatures and an increase in the enthalpy change which is consistent with a closer packing of the lipid molecules in the liposomes. 3. 3. Above the 1:2 ratio, freeze-fracture electron microscopy demonstrated typical “crystal” structures both in the presence of Ca2+ and Mg2+. In the presence of Mg2+ a metastable behaviour was noticed in the calorimetric experiments. 4. 4. A Ca2+- and Mg2+-induced shift in the transition temperature and an increase in the enthalpy change was also observed in a 1:1 mixture of dimyristoyl phosphatidylglycerol and dimyristoyl phosphatidylcholine. However, these mixed samples remained liposomal in structure at any concentration of the divalent ions. 5. 5. Liposomes prepared from a 1:1 mixture of dimyristoyl phosphatidylglycerol and dimyristoyl phosphatidylcholine in the absence of divalent cations are permeable in the range 10–50 °C. Bilayers of mixtures neutralized by Ca2+ or Mg2+ were demonstrated to be completely impermeable to K+, except in the vicinity of the phase transition. 6. 6. The leak of ions from liposomes of a 1:1 mixture of dimyristoyl phosphatidylglycerol and dimyristoyl phosphatidylcholine in the vicinity of the phase transition temperature was considerably less in the presence of Ca2+ than in the presence of Mg2+. 7. 7. It is concluded that there is a correlation between the calorimetric data and the permeability properties of dimyristoyl phosphatidylglycerol-containing bilayers with respect to the influence of Ca2+ and Mg2+

Calorimetric and freeze-etch study of the influence of Mg2+ on the thermotropic behaviour of phosphatidylglycerol

In the presence of Mg2+ ions phosphatidylglycerol shows supercooling which leads to the formation of a metastable gel phase. This contrasts with the behaviour of this negatively charged phospholipid in the presence of Ca2+ ions (Biochim. Biophys. Acta 339 (1974) 432). It is demonstrated that the heat content of this phospholipid is dependent on the ionic environment

Freeze-etching: Freezing velocity and crystal size at different locations in samples

Freezing velocity was studied experimentally as well as theoretically in relation to sample size and location within the sample. Crystal sizes in a freeze-etch replica displaying the complete fracture area of a sample were compared to the experimental and theoretical results concerning freezing velocities at different locations in such a sample. The freezing velocity was found to be higher in the center and near the surface than halfway between the center and the outer surface of the sample. Also, the crystals in the center and near the border appear to be small as compared to the crystals in the other fracture domain