Glycerol penetration profile in phospholipid bilayers measured by ESEEM of spin-labelled lipids

Abstract

Electron spin echo envelope modulation (ESEEM) spectroscopy of phospholipids systematically spin-labelled along the sn-2 chain was used to detect the penetration of a deuterium-substituted water–glycerol mixture H2O/C3D5(OH)3 (1:1 v/v) into dipalmitoyl phosphatidylcholine (DPPC) bilayer membranes. The results provide a detailed profile of the penetration of glycerol across the membrane. The ESEEM amplitude induced by the hyperfine interaction with the deuterium nuclei drops substantially beyond the 10th label position. Beyond the 12th label position, the amplitude was nearly constant. A quantitative assessment of the glycerol concentration in the middle of the bilayer resulted in a value of ∼0.4 M (∼3% v/v). For comparison, analogous measurements were performed for the penetration of pure deuterium water D2O. In both cases, the ESEEM amplitudes varied similarly across the bilayer, except for the terminal 16th position. At this position, the amplitude for pure water was small and close in magnitude to the noise level.