Small-angle neutron scattering study of lateral phase separation in dimyristoylphosphatidylcholine-cholesterol mixed membranes

Publisher Website
Google Scholar
Abstract
The small-angle neutron scattering (SANS) technique developed previously is used to study the lateral phase separation in dimyristoylphosphatidylcholine (DMPC)-cholesterol mixed vesicles in the L.alpha. (35.degree. C) and L.beta.'' (7.degree. C) phase of DMPC. To increase the sensitivity of the previous method, we apply the so-called inverse contrast variation technique where contrast matching is performed at a constant H2O/D2O ratio by varying the ratio of DMPC with deuterated and protonated hydrocarbon chains. Phase boundaries can be determined to an accuracy of .+-. 0.5 mol%. In parallel experiments phase separation in the L.beta.'' phase was also studied by freeze-fracture electron microscopy. For DMPC in the L.alpha. phase complete miscibility is clearly established up to cholesterol molar fractions of xc = 0.14. Strong evidence is provided that this is also the case up to xc .apprxeq. 0.45. Cholesterol is no longer soluble above this limit and precipitates as small crystallines. For the L.beta.'' phase (7.degree. C) phase boundaries are clearly established at xc1 = 0.08 and xc2 = 0.24, and very strong evidence is provided for two additional boundaries at xc3 = 0.435 and xc4 .apprxeq. 1.0. At 0 .ltoreq. xc .ltoreq. xc1 the mixture forms a tilted solid solution in both the L.beta.'' and P.beta.'' phase while at xc1 .ltoreq. xc .ltoreq. xc2 this phase coexists with a nontilted mixture containing 24 mol% cholesterol. At xc2 .ltoreq. xc .ltoreq. xc3 a second region exists where mixtures containing 24 and 43.5 mol% cholesterol coexist within the plane of the membrane. To fulfill the phase rule, xc2 must correspond to a stoichiometric mixture which would reconcile the concepts of phase separation and complex formation. Our freeze-fracture studies do not provide evidence for a decrease of the repeat distance of the ripple phase at xc .ltoreq. 0.08 and for a regular ripple phase with defined ripple distance within the first coexistence region.