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. 2000 May;78(5):2459–2469. doi: 10.1016/S0006-3495(00)76790-7

Dimyristoylphosphatidylcholine/C16:0-ceramide binary liposomes studied by differential scanning calorimetry and wide- and small-angle x-ray scattering.

J M Holopainen 1, J Lemmich 1, F Richter 1, O G Mouritsen 1, G Rapp 1, P K Kinnunen 1
PMCID: PMC1300835  PMID: 10777742

Abstract

Ceramide has recently been established as a central messenger in the signaling cascades controlling cell behavior. Physicochemical studies have revealed a strong tendency of this lipid toward phase separation in mixtures with phosphatidylcholines. The thermal phase behavior and structure of fully hydrated binary membranes composed of dimyristoylphosphatidylcholine (DMPC) and N-palmitoyl-ceramide (C16:0-ceramide, up to a mole fraction X(cer) = 0.35) were resolved in further detail by high-sensitivity differential scanning calorimetry (DSC) and x-ray diffraction. Both methods reveal very strong hysteresis in the thermal phase behavior of ceramide-containing membranes. A partial phase diagram was constructed based on results from a combination of these two methods. DSC heating scans show that with increased X(cer) the pretransition temperature T(p) first increases, whereafter at X(cer) > 0.06 it can no longer be resolved. The main transition enthalpy DeltaH remains practically unaltered while its width increases significantly, and the upper phase boundary temperature of the mixture shifts to approximately 63 degrees C at X(cer) = 0.30. Upon cooling, profound phase separation is evident, and for all of the studied compositions there is an endotherm in the region close to the T(m) for DMPC. At X(cer) >/= 0.03 a second endotherm is evident at higher temperatures, starting at 32.1 degrees C and reaching 54.6 degrees C at X(cer) = 0.30. X-ray small-angle reflection heating scans reveal a lamellar phase within the temperature range of 15-60 degrees C, regardless of composition. The pretransition is observed up to X(cer) < 0.18, together with an increase in T(p). In the gel phase the lamellar repeat distance d increases from approximately 61 A at X(cer) = 0. 03, to 67 A at X(cer) = 0.35. In the fluid phase increasing X(cer) from 0.06 to 0.35 augments d from 61 A to 64 A. An L(beta')/L(alpha) (ripple/fluid) phase coexistence region is observed at high temperatures (from 31 to 56.5 degrees C) when X(cer) > 0.03. With cooling from temperatures above 50 degrees C we observe a slow increase in d as the coexistence region is entered. A sudden solidification into a metastable, modulated gel phase with high d values is observed for all compositions at approximately 24 degrees C. The anomalous swelling for up to X(cer) = 0.30 in the transition region is interpreted as an indication of bilayer softening and thermally reduced bending rigidity.

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