Phospholipid liquid crystalline phase

Pressure was applied in this study to fine tune the lipid chain-lengths and conformation and to select specific lamellar phases. For example, the phospholipid bilayer thickness increases by 1 A/kbar in the liquid-crystalline phase, and up to six gel phases have been found in fully hydrated DPPC dispersions in the pressure-temperature phase space up to 15 kbar and 80 °C, respectively. NMR spectral parameters were used to detect structural and dynamic changes upon incorporation of the polypeptide into the lipid bilayers. 

There may be limitations in applying the above model to other systems. For instance, the initial surfactant often exists as a lamellar phase as for phospholipids, so that there are no interfaces between various liquid crystalline phases whose velocities can be measured and used to determine effective dif-fusivities as in the AOT analysis above. As a result, the base of the myelinic figures must approach the base of the vertical cell as the volume of the lamellar phase shrinks, and the assumption made above that the composition of the myelins is independent of time may not be valid. 

The appearance of tubular myelin-like structures in swollen lecithin was observed by light microscopy well before the systematic investigation of liposomes [351-352]. Similarly, it was also demonstrated some time ago that the addition of calcium ions converted phospholipid liposomes to cochleate cylinders [353]. Subsequent studies have, however, revealed that the system is extremely complex. For example, examination of the phase-transition behavior of synthetic sodium di-n-dodecyl phosphate [(C12H2sO)2PO2Na+ or NaDDP] and calcium di-n-dodecyl phosphate [Ca(DDP)2] showed the presence of many diverse structures [354]. In particular, hydrated NaDDP crystals were shown to form lyotropic liquid-crystalline phases which transformed, upon heating to 50 °C, to myelin-like tubes. Structures of the tubes formed were found... 

Liquid crystals, liposomes, and artificial membranes. Phospholipids dissolve in water to form true solutions only at very low concentrations ( 10-10 M for distearoyl phosphatidylcholine). At higher concentrations they exist in liquid crystalline phases in which the molecules are partially oriented. Phosphatidylcholines (lecithins) exist almost exclusively in a lamellar (smectic) phase in which the molecules form bilayers. In a warm phosphatidylcholine-water mixture containing at least 30% water by weight the phospholipid forms multilamellar vesicles, one lipid bilayer surrounding another in an "onion skin" structure. When such vesicles are subjected to ultrasonic vibration they break up, forming some very small vesicles of diameter down to 25 nm which are surrounded by a single bilayer. These unilamellar vesicles are often used for study of the properties of bilayers. Vesicles of both types are often called liposomes.75-77... 

Several investigators have used radioactive tracer methods to determine diffusion rates. Bangham et al. (32) and Papahadjopoulos and Watkins (33) studied transport rates of radioactive Na+, K+, and Cl" from small particles or vesicles of lamellar liquid crystal to an aqueous solution in which the particles were dispersed. Liquid crystalline phases of several different phospholipids and phospholipid mixtures were used. Because of uncertainties regarding particle geometry and size distribution, diffusion coefficients could not be calculated. Information was obtained, however, showing that the transport rates of K+ and Cl" in a given liquid crystal could differ by as much as a factor of 100. Moreover, relative transport rates of K+ and Cl" were quite different for different phospholipids. The authors considered that ions had to diffuse across platelike micelles to reach the aqueous phase. 

Upon heating, very pure uniform phospholipids show an additional small transition below the crystalline to liquid crystalline phase transition. According to Janiak et al. [104], this is due to a structural change in the head groups. The occurrence of pretransition is mainly determined by the structure of the head groups and their environment. 

An additional example for the usefulness of IR spectroscopy in studying drug interactions with phospholipid vesicles is the quantitative determination of acyl chain conformation in gramicidin-DPPC mixtures [63]. The technique provides a quantitative measure of the extent to which membrane-spanning peptides induce disorder of phospholipid gel phases and order in liquid crystalline phases. 

A later publication from the same group [153] reporting on 31P- and 2H-NMR and DSC studies on zwitterionic and anionic phospholipids in the absence and presence of doxombicin (adriamycin) described different results [152], Doxorubicin had a stronger disordering effect on the membrane of lipid mixtures enriched with anionic lipids. However, extensive segregation of DOPE and DOPA or DPS was not observed even under conditions of Hlrphase formation . According to the authors, the reason for this discrepancy was that in the earlier paper the phase separation was obtained with membranes subject to gel-liquid crystalline phase transition , which was, however, discounted in the first paper. 

How does this shift from fluid to gel state during desiccation cause damage to the membrane, and how does the presence of trehalose or sucrose—water substitutes—prevent this damage As the anecdote about baking technique implies, the crux of the problem occurs when dried cells are rehydrated. It is known from studies of model membranes that when phospholipid bilayers pass through the temperature range over which the gel phase is replaced by the liquid crystalline phase, transient changes in membrane permeability occur (Crowe et al., 1997). The precise mechanism responsible for this transient breakdown in the permeability barrier is not entirely clear, but it... 

Figure 7.20. Effects of temperature on phase and static order of the membrane bilayer. Reductions in temperature from the physiological temperature of the organism (cell), that is, temperatures near those of adaptation or acclimation, lead to formation of gel-phase regions, which may separate from lipids that remain in the liquid-crystalline phase, the phase characteristic of most of the bilayer at physiological temperatures. At sufficiently low temperatures, the bulk of the membrane phospholipids enter the gel phase.

Analysis of the results and comparison with the lipid phase transition observed iq the bulk lipid/water systems allows to conclude that the lowest temperature during heating at which measurable diffusion occurred correlates with the onset of formation of the lamellar Ln liquid crystalline phase of the given phospholipid. Therefore, the data support a correlation between the surface and bulk phase transitions. This was confirmed in recent studies where the lipid surface phase transition was successfully measured for the first time in foam film by independent means involving the detailed investigations of the temperature dependences of the W(C) curve for the foam film and its thickness. 

The C=0 stretching band in the IR absorption spectrum is equally useful. Phase transition shifts this band from 1738 cm towards lower wavenumbers it is found around 1733 cm as phospholipid vesicles undergo a gel to liquid crystalline phase transition Tm. Mantsch interpreted the shift towards lower wavenumbers of the overall band contour peak as resulting from the intensity increase of the component at 1727 cm upon a change in hydration (by hydrogen bonding) during the transition (Mantsch and Me Elhaney, 1991). 

Vibrational spectroscopy shows that inclusion of cholesterol in phospholipid bilayers tends to decrease the fluidity of the hydrophobic region above the main transition point Tm and to increase it below Tm. The presence of cholesterol in DPPC or DMPC muti-layered vesicles does not affect the transition point but simply broadens the transition by decreasing the CH2-stretching wavenumber in the liquid crystalline phase and by increasing it in the gel-like phase (Lippert and Peticolas, 1971 Spiker and Levin, 1976 Casal and Mantsch, 1984). There is also evidence that lipid-cholesterol interaction increases the amount of bound water in the headgroups (Levin et al., 1985). 

The sharpness or cooperativity of the gel-to-liquid-crystalline phase transition can also be evaluated from the DSC trace. The sharpness of the phase transition is often expressed as the temperature width at half-height, AT1/2, or as the temperature difference between the onset or lower boundary of the phase transition, Ts, and the completion or upper boundary, Ti, or AT = Ti — Tg. The AT1/2 values may range from <0.1°C for very pure synthetic phospholipids to as much as 10-15°C for biological membranes. From the Tm and AT1/2 values determined for a particular phase transition, the van t Hoff enthalpy, AHvh, can be approximately determined from the relationship ... 

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