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Two cationic phospholipid derivatives with asymmetric hydrocarbon chains were synthesized: ethyl esters of oleoyldecanoylethylphosphatidylcholine (C18:1͞C10-EPC) and stearoyldecanoylethylphosphatidylcholine (C18:0͞C10-EPC). The former was 50 times more effective as a DNA transfection agent (human umbilical artery endothelial cells) than the latter, despite their similar chemical structure and virtually identical lipoplex organization. A likely reason for the superior effectiveness of C18:1͞C10-EPC relative to C18:0͞C10-EPC (and to many other cationic lipoids) was suggested by the phases that evolved when these lipoids were mixed with negatively charged membrane lipid formulations. The saturated C18:0͞C10-EPC remained lamellar in mixtures with biomembranemimicking lipid formulations [e.g., dioleoyl-phosphatidylcholine͞ dioleoyl-phosphatidylethanolamine͞dioleoyl-phosphatidylserine͞ cholesterol at 45:20:20:15 (wt͞wt)]; in contrast, the unsaturated C18:1͞C10-EPC exhibited a lamellar-nonlamellar phase transition in such mixtures, which took place at physiological temperatures, Ϸ37°C. As is well known, lipid vehicles exhibit maximum leakiness and contents release in the vicinity of phase transitions, especially those involving nonlamellar phase formation. Moreover, nonlamellar phase-forming compositions are frequently highly fusogenic. Indeed, FRET experiments showed that C18:1͞C10-EPC exhibits lipid mixing with negatively charged membranes that is several times more extensive than that of C18:0͞C10-EPC. Thus, C18:1͞C10-EPC lipoplexes are likely to easily fuse with membranes, and, as a result of lipid mixing, the resultant aggregates should exhibit extensive phase coexistence and heterogeneity, thereby facilitating DNA release and leading to superior transfection efficiency. These results highlight the phase properties of the carrier lipid͞cellular lipid mixtures as a decisive factor for transfection success and suggest a strategy for the rational design of superior cationic lipid carriers.lipofection ͉ lipoplex ͉ mesophase I mportant therapeutic procedures, such as gene transfection and gene silencing, require efficient delivery of genetic material to cells. Synthetic cationic lipoids, which form complexes (lipoplexes) with polyanionic DNA, are promising gene carriers (1). Understanding the mechanism of lipid-mediated DNA delivery (lipofection) is essential for the successful application and rational design and synthesis of novel cationic lipoid compounds for enhanced gene delivery. Although considerable improvement in the transfection properties of cationic lipoids has come from the synthesis of new kinds of cationic amphiphiles or from the inclusion of noncationic helper lipids, an effective alternative strategy was recently described: The combination of two cationic lipid derivatives having the same headgroup but different hydrocarbon chains can synergistically enhance transfection (2). For example, the optimal combination of the long chain͞medium chain lipoids, dioleoyl-and dilauroyl-ethylphosphatidylcholines, de...

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By means of differential scanning calorimetry and from a review of published data we demonstrate in this work that low-molecular weight kosmotropic substances (water-structure makers) of different chemical structure such as disaccharides, proline, and glycerol have identical effects on the phase behavior of several kinds of phospholipids and glycolipids. These substances favor formation of the high-temperature inverted hexagonal phase (H(II)) and the low-temperature lamellar crystalline (L(c)) and gel (L( β )) phases at the expense of the intermediate lamellar liquid-crystalline phase (L( α )). The latter phase may completely disappear from the phase diagram at high enough solute concentration. By contrast, chaotropic substances (water-structure breakers) such as sodium thiocyanate and guanidine hydrochloride expand the existence range of L( α ) at the expense of the adjacent L( β ) and H(II) phases. Moreover, chaotropes are able to induce the appearance of missing intermediate liquid-crystalline phases in lipids displaying direct L( β )→H(II) transitions in pure water. In previous publications we have considered the influence of chaotropic and kosmotropic substances on the lipid phase behavior as a manifestation of their indirect (Hofmeister) interactions with the lipid aggregates. For a quantitative characterization of this effect, here we derive a general thermodynamic equation between lipid phase transition temperature and solute concentration, analogous to the Clapeyron-Clausius equation between transition temperature and pressure. It provides a clear description in physical quantities of the disparate effects of kosmotropic and chaotropic substances on the relative stability of the lipid-water phases. According to this equation, the magnitude of the solute effect is proportional to the hydration difference of the adjacent lipid phases and inversely proportional to the transition latent heat. The sign and magnitude of the transition shifts depend also on the degree of solute depletion (for kosmotropes) or enrichment (for chaotropes) at the interfaces, in comparison to the solute concentration in bulk water.