Phase studies of model biomembranes: Macroscopic coexistence of Lα+Lβ, with light-induced coexistence of Lα+Lo Phases

Abstract

Phase diagrams of 3-component lipid bilayer mixtures containing cholesterol reveal major differences among the different types of lipids. Here we report that mixtures of cholesterol together with POPC and a high-melting temperature PC or sphingomyelin show different phase behavior from similar mixtures that contain DOPC or di-phytanoyl-PC instead of POPC. In particular, only one region of macroscopic phase coexistence occurs with POPC, a region of coexisting liquid disordered and solid phases, {Lalpha+Lbeta}. Fluorescence microscopy imaging is useful for these studies, but is subject to artifactual light-induced domain formation, as reported by Ayuyan and Cohen [A.G. Ayuyan, F.S. Cohen, Lipid peroxides promote large rafts: Effects of excitation of probes in fluorescence microscopy and electrochemical reactions during vesicle formation, Biophys. J. 91 (2006) 2172-2183.]. This artifact can be attenuated by decreased illumination and low dye concentration. The use of the free radical scavenger n-propyl gallate can reduce the artifact, but this molecule enters the bilayer and itself perturbs the phase behavior. We suggest that the light-induced domain separation artifact might actually arise from pre-existing lipid clusters that are induced to coalesce, and therefore indicates highly nonrandom mixing of the lipid components.

Critical Concentration of Lecithin Enhances the Antimicrobial Activity of Eugenol against Escherichia coli

“…Several studies have revealed the formation of temperature-and concentrationdependent dense domains or "patches" in phospholipid bilayers surrounding lipid-rich aggregates (29)(30)(31)(32). These patches were reported to alter the structure of biological cell membranes (33) and potentially regulate cell signaling.…”

Phase Separation in Lipid Membranes

“…Also of interest for the behavior of cell membranes, even single phase nonideal lipid mixtures can be induced to increase the size of their heterogeneities. Here, a case that plagues the use of fluorescence microscopy is the introduction of artifactual visible domains by intense illumination (Zhao et al 2007b). This might be caused by free radical initiated polymerization that starts from the reactive excited singlet state (Ayuyan and Cohen 2006).…”

Cholesterol Packing around Lipids with Saturated and Unsaturated Chains: A Simulation Study

“…4 The particular contribution cholesterol makes in the structure of rafts is to facilitate the formation and maintenance of a liquid-ordered (L o ) membrane domain. 5,6 While the majority of experimental studies have involved dipalmitoyl or dimyristoyl phosphatidylcholine (DPPC or DMPC) [7][8][9][10] there are also experimental studies on the effect of cholesterol on palmitoyl-oleyol phosphatidylcholine (POPC). [11][12][13][14][15][16][17][18] Hyslop et al 11 utilized fluorescence depolarization to estimate that cholesterol molecules in a 1:1 POPC:cholesterol bilayer remain separated with a minimum separation distance of ∼10 Å.…”

“…POPC has also been investigated as part of ternary bilayer mixtures with sphingomeylin and cholesterol 8,20,21 or ceramide. 22 Heerklotz and co-workers used calorimetry to compare the interactions of cholesterol with large unilamellar vesicles of POPC plus 30% cholesterol, sphingomyelin plus 30% cholesterol, and 1:1 sphingomyelin:POPC plus 30% cholesterol.…”

“…20 They find that sphingomyelin-cholesterol interactions are favored by 7 kJ/m in sphingomyelin-cholesterol mixtures and by 13 kJ/m in 1:1 sphingomyelin-POPC mixtures over pure POPC-cholesterol mixtures. Zhao et al 8 find using flourescence imaging that ternary mixtues of POPC, sphingomyelin, and cholesterol are qualitatively different from mixtures of DOPC, sphingomyelin,and cholesterol. POPC-based ternary mixtures do not exhibit the same micrometer scale domains over a wide range of concentrations, compared to DOPC-based mixtures.…”