13C-NMR determination of the solubility and molecular dynamics of cholesteryl oleate in dimyristoylphosphatidylcholine vesicles

Abstract

Using 13C-NMR measurements of T1, T2 and the nuclear Overhauser enhancement factor at 50.29, 90.55 and 150.87 MHz, we have measured the dynamics of 1.5 mol% cholesteryl oleate in dimyristoylphosphatidylcholine (DMPC) small unilamellar vesicles (SUVs) at 37°C. Using the model-free approach of Lipari and Szabo [J. Am. Chem. Soc. 104: 4546–4559, 1982], we have found that the motion of the rigid steroid ring can be described by an equal contribution from two effective motions with correlation times of 15 and 0.38 ns. The motions of the C26 and C27 carbon atoms of cholesteryl oleate were found to have an effective correlation time of 7 ± 2 ps and a value for the square of the generalised order parameter of 0.03 ± 0.01. The corresponding values for the C25 carbon atom were 14 ± 9 ps and 0.17 ± 0.05, showing slower motion and greater order for this carbon atom, which is nearer to the rigid steroid ring. This establishes that the motion of the steroid ring of cholesteryl oleate in DMPC vesicles is significantly different to that previously measured for cholesterol in DMPC vesicles. On the other hand, the effective motion of the portions of the corresponding alkyl chains about the steroid ring is the same. On preparing vesicles, when the ester was at the limit of miscibility with the DMPC, the solubility of cholesteryl oleate in DMPC vesicles at 55°C was found to be 2.8 ± 0.3 mol%. Using aqueous Mn2+ as a paramagnetic relaxation enhancement agent, comparison of the appropriate T1 values indicated that the carbonyl group of the cholesteryl ester was the same distance from the aqueous phase as the C4 carbon of the steroid ring.