DIFFUSION ORDERED 2D-NMR SPECTROSCOPY OF PHOSPHOLIPID-VESICLES - DETERMINATION OF VESICLE SIZE DISTRIBUTIONS

Diffusion ordered 2D NMR (DOSY) has been used to characterize large unilamellar phospholipid (LUV) vesicles. Entrapped sucrose served as an NMR active label for the vesicles and permitted diffusion coefficients to be measured for vesicle preparations with average diameters ranging from 30 to 100 nm and total lipid concentrations ranging from 1 to 30 mM. Distributions of diffusion coefficients were obtained by analysis with the program CONTIN. Average diameters and distributions of diameters were then obtained from the vesicle diffusion coefficients after appropriate corrections by means of the Stokes-Einstein equation. Electron microscopy and dynamic light scattering measurements were also performed on the LUV's, and the results were found to be consistent with those obtained with DOSY. The apparent vesicle diffusion coefficients obtained with the DOSY experiment from phospholipids in the bilayers were up to 2 times larger than for the entrapped sucrose. This difference is a feature of polydispersity that results from size-dependent inhomogeneous magnetic dipole broadening of the resonances for the phospholipids but not the entrapped sucrose. The difference vanishes for small sonicated vesicles. DOSY has major advantages for the study of complex mixtures since molecules and assemblies of all sizes can be observed. For example, in this study both entrapped and free sucrose were resolved at the same chemical shifts. Thus, chemical equilibria involving molecules with different diffusion rates are accessible to study by DOSY.