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Abstract

For canonical lipid raft mixtures of cholesterol (chol), N -palmitoylsphingomyelin (PSM), and 1-palmitoyl-2-oleoyl- phosphatidylcholine (POPC), electron paramagnetic resonance (EPR) of spin-labeled phospholipids—which is insensitive to domain size—is used to determine the ternary phase diagram at 23 C. No phase boundaries are found for binary POPC/chol mixtures, nor for ternary mixtures with PSM content < 24 mol %. EPR lineshapes indicate that conversion from the liquid-disor- dered (L a ) to liquid-ordered (L o ) phase occurs continuously in this region. Two-component EPR spectra and several tie lines attributable to coexistence of gel (L b ) and fluid phases are found for ternary mixtures with low cholesterol or low POPC content. For PSM/POPC alone, coexistence of L a and L b phases occurs over the range 50 95.5 mol % PSM. A further tie line is found at 3 mol % chol with endpoints at 50 and R 77 mol % PSM. For PSM/chol, L b L o coexistence occurs over the range 10 38 mol % chol and further tie lines are found at 4.5 and 7 mol % POPC. Two-component EPR spectra indicative of fluid-fluid (L a L o ) phase separation are found for lipid compositions: 25% < PSM < 65%, 5% < chol < 30–35%, 65% > POPC > 10%, and confirmed by nonlinear EPR. Tie lines are identified in the L a L o coexistence region, indicating that the fluid domains are of sufficient size to obey the phase rule. The three-phase triangle is bounded approximately by the compositions 40 and 75 mol % PSM with 10 mol % chol, and 60 mol % PSM with 25 mol % chol. These studies define the compositions of raft-like L o phases for a minimal realistic biological lipid mixture.