1,2-Dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)-rich domain formation in binary phospholipid vesicle membranes:...

Decades of study have probed phase transitions in model phospholipid bilayers and vesicles, especially in the context of the equilibrium phase diagram. Critical to the response of vesicles to environmental triggers, to the ultimate domain morphology, and to the approach to equilibrium (or not), we present here a study of domain formation in vesicles, focusing on a mechanism by which the cooling rate, tension, and composition affect the first appearance (nucleation) and subsequent growth of solid membrane domains. Employing a popular mixed membrane model based on DOPC and DPPC (1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, respectively), we examined phase separation in giant two-component vesicles that were cooled from the one-phase fluid (Lα) region of the phase diagram into a region of fluid (Lα)-solid coexistence. At moderate and low membrane tensions, cooling produced solid DPPC-rich domains appearing as compact patches or irregular hexagons and likely with a Pβ' (ripple) arrangement. (The compact solid domains in this study differed distinctly from striped domains in vesicles of the same composition, in terms of molecular organization and conditions of first appearance during cooling.) The amounts of these solid domains were shown to adhere to the lever arm rule for a tie line on the phase diagram, with a solid composition near 95 mol % DPPC. The nucleation of the compact solid domains occurred in a short period, followed by rapid addition of ordered molecules to the nucleated domains, excluding tracer dye. The two-dimensional nucleation density of these compact solid domains (in the range of 10(-2)-10(-1) μm(-2)) was found to increase with the cooling rate (equivalent to the quench depth) with a greater than linear dependence. The 2-D nucleation density was also seen to decrease with membrane tension, presumably because membrane tension increases the line tension around a domain that opposes nucleation. A sigmoidal dependence of the nucleation density on the DPPC concentration was also found. With cooling rates in excess of ∼1 °C/min, solid domains persisted down to room temperature, likely passing from a preferred equilibrium to a local equilibrium with continued cooling. As a result of the persistence of the originally nucleated domains and the conservation of DPPC in the membrane, we observed an increasingly greater number of smaller domains with increased cooling rates. The domains in these vesicles were compact or hexagonal-shaped in contrast to flower-shaped dendritic domains in the same membrane system in a supported membrane configuration.