Lipid magnetic resonance imaging contrast agent interactions: A spin-labeling and a multifrequency EPR study

The interactions of two Lipophilic magnetic resonance imaging paramagnetic contrast agents, gadolinium complexes of 1,4,7,10-tetraazacyclododecane-N-(n-pentyl)-N',N'',N'''-triacetic acid (Gd-DOTAP) and 3,6,9-triaza-3,6,9-tris(carboxymethyl)-4-(4-ethoxybenzyl)undecandicarboxylic acid (Gd-EOB-DTPA), with model multilamellar liposomes prepared from 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) were assessed in three sets of EPR experiments. The first two experiments were carried out with phospholipids selectively labeled with a series of spin-labeled doxyl stearic acids. By means of counting the collisions of molecular oxygen with a spin-labeled site, local oxygen permeability across the bilayer was measured with the contrast agents present and compared with control experiments in which contrast agents were absent. The maximum increase of 6.5 +/- 0.3% for oxygen permeability at 20 mM Gd-DOTAP concentration was observed at 30.8 degrees C for the label located at the twelfth carbon position of the acyl chain. This result implied that Gd-DOTAP partitions within the DMPC bilayer, its preferred location being close to the bilayer center, and that the overall effect of Gd-DOTAP on the structural organization of the membrane is rather small. In contrast, no significant changes in local oxygen permeability were observed for Gd-EOB-DTPA. In the second set of spin-labeling experiments, the broadening of the spin-label spectra in the presence of Gd-DOTAP was measured as a function of position of the label across the bilayer. Again, maximum broadening was observed for the center of the bilayer, confirming the preferred location of this contrast agent. The third set of experiments utilized EPR spectroscopy of solutions of Gd3+ complexes at multiple high magnetic fields (corresponding microwave frequencies from 35 to 249 GHz). At these fields, Gd3+ serves as a useful probe to report on the microenvironment. Partitioning of Gd-DOTAP in DMPC liposomes resulted in partially resolved EPR spectra at 35 and 94.3 GHz. EPR experiments at multiple high fields (35, 94.3, and 249 GHz) show that the variation in spectral resolution observed across this frequency range arises from, first, line narrowing due to a decreased relative contribution of the zero field splitting (ZFS) in the spin Hamiltonian and, second, the shift of the resonance signal due to remaining ZFS effects. The frequency dependence of the apparent g-factor at multiple high magnetic fields as analyzed with third-order perturbation theory demonstrates a relationship between the observed shifts and the ZFS parameter. The analysis shows that Delta, the square root of the trace of the squared ZFS matrix, increases to 8.1 rad GHz when Gd-DOTAP partitions in the lipid phase of DMPC liposomes, up from 5.65 rad GHz for the aqueous phase. The high-field EPR method provides a direct measure of the populations of Gd complexes in various environments as well as an estimate of ZFS parameters in solutions.