Structure and Dynamics of Lens Lipid Membranes Derived from a Single Porcine Donor: High Field EPR Study

To obtain correct EPR line shapes, spin-lattice relaxation times, and bimolecular collision rates between spin labels and oxygen, samples must be thoroughly deoxygenated or equilibrated with a controlled oxygen partial pressure. These measurements are conveniently carried out using a gas permeable plastic sample tube of small diameter that fits in a loop-gap resonator. Flow of gas over the tube allows easy deoxygenation or controlled oxygenation of the sample. In the initial design of the W-band (94 GHz) loop-gap resonator, samples were equilibrated with gas at room temperature outside the resonator, transferred to a quartz capillary, and positioned in the resonator. In a recently designed W-band loop-gap resonator, the sample is positioned in a gas-permeable Teflon tubing inside the resonator, which allows measurements of extremely small volume (∼30 nL) of sample. We used this new design to measure properties of lens lipid membranes derived from total lipids extracted from both lenses (single donor) of a 2-year-old porcine cortex and nucleus. Detailed profiles of membrane fluidity and oxygen transport parameter were obtained from saturation recovery EPR. Analysis of conventional spectra using the microscopic-order macroscopic-disorder (MOMD) model provided rotational diffusion coefficients (R(perpendicular) and R(parallel)) and order parameters. Three different types of motion of lipid spin labels n-PC, T-PC, and CSL (ASL) with, respectively, nitroxide z-axis, x-axis, and y-axis parallel to the bilayer normal, are discussed. Results demonstrate that EPR at W-band has the potential to be a powerful tool for studying samples of small volume, ∼30 nL, obtained from eye lenses of a single human donor.