David J. Siminovitch

The quadrupolar spectrum of a spin I = 1 in a lipid bilayer in the presence of paramagnetic ions

Abstract The NMR signal from selectively deuterated molecules in a lipid bilayer where there are paramagnetic ions present in the aqueous region is influenced by both the nuclear quadrupole interaction and the dipolar interaction between the deuterium nuclei and the surrounding ions. The quadrupolar split powder pattern is no longer symmetric about the center of the spectrum. The spectra which result from the use of the quadrupole echo pulse sequence are quite complex since this sequence does not refocus the dephasing due to the dipolar interaction. A new pulse sequence which refocuses both the dipolar and quadrupolar interactions is suggested. Both 2H and 14N spectra from lipid molecules with the phosphatidylcholine headgroup obtained with the conventional quadrupole echo sequence and the new sequence are compared with predictions of density matrix theory calculations. There is excellent agreement between the experimental and simulated spectra.

Orientational order in the choline headgroup of sphingomyelin: A 14N-NMR study

An aqueous dispersion of fully hydrated bovine sphingomyelin was studied using 14N-NMR spectroscopy. Spectra were obtained as a function of temperature over the range 15-80 degrees C, in both the liquid crystal and gel phases. In the liquid crystal phase, powder pattern lineshapes were obtained, whose quadrupolar splitting slowly decreases with increasing temperature. The spectra are increasingly broadened as the temperature is lowered through the phase transition into the gel phase. The linewidths and the second moments of these spectra indicate that the onset of a broad phase transition occurs at approx. 35 degrees C, in agreement with previous calorimetric and 31 P-NMR measurements. There is no evidence from the lineshapes for an hexagonal phase in this system, and this conclusion is supported by X-ray diffraction measurements carried out on aqueous dispersions of sphingomyelin in both phases. Assuming that the static nitrogen quadrupole coupling constant is the same for both sphingomyelin and dipalmitoyl-L-alpha-phosphatidylcholine (DPPC), the decrease observed in the quadrupolar splitting of sphingomyelin compared to that of DPPC indicates that the orientational order of the choline headgroup in liquid crystalline sphingomyelin is not the same as that of its counterpart in DPPC. Preliminary relaxation time measurements of T1 and T2 are presented which suggest that there are also dynamic differences between sphingomyelin and DPPC in the choline headgroup.

The use of wide-line [14N]nitrogen NMR as a probe in model membranes

1. Introduction A major aim of bioiogi~al membrane research is to understand the relationship between membrane struc- ture and function at the molecular level. The presently accepted fluid mosaic model [ 1] indicates that the membrane is basically an anisotropic fluid composed of a lipid bilayer in which proteins are embedded. At the molecular level it is necessary to describe both the average conformation of the molecules and their molecular dynamics. There is both translation and rotation of the lipid and protein molecules within the bilayer structure. For the lipids in particular, there are many internal degrees of freedom. Of the many techniques [Z] being used to probe the molecular environment within membranes, nuclear magnetic resonance (NMR) [3,4] has proven to be one of the most useful methods for investigating the dynamical state of the membrane. Work has con- centrated on describing the dynamical behaviour of the lipid molecules and the interaction of these molecules with other membrane components such as various proteins [5,6] and cholesteroi [7,8]. To do this, it is essential to probe various positions in the lipid molecules. Because of the overabundance of hydrogen atoms, proton NMR spectra have been difficult to interpret [9,10] and give little information about the motion of a single segment of the lipid molecule. 3’P NMR [l I] has been successfully used to probe the head group region of phospholipids. Selective incorporation of ‘II [12], 13C [ 131 and tgF [14] into lipid molecules has been the most fruitful method of probing the molecular dynamics. However, considerable time and expertise are necessary for the synthesis of the labelled molecules required for a complete investigation of the lipid dynamics. The use of naturally abundant NMR probes is there- fore advantageous whenever possible. One naturally

Studies on the interaction of anesthetic steroids with phosphatidylcholine using 2H and 13C solid state NMR

The effects of the anesthetic steroid alphaxalone and its inactive analog delta 16-alphaxalone on model phospholipid membranes were studied using 13C and 2H solid-state nuclear magnetic resonance spectroscopy. Aqueous multilamellar dispersions of dipalmitoylphosphatidylcholine (DPPC) with specific 13C and 2H labels as endogenous probes at the carbonyl and the C-7 methylene groups, respectively, of the sn-2 chain were used to study the conformational and dynamical properties of the bilayer as a function of temperature. There were no significant changes between the 13C and 2H spectra of the DPPC preparation containing the inactive steroid and that of DPPC with no drug. However, the physiologically active steroid produces significant spectral 2H and 13C changes. These changes include a reduction of the main phase transition temperature and a broadening of that transition. Alphaxalone also increases the relative number of gauche conformers in the liquid-crystalline phase of DPPC and increases the rate of axial diffusion in both the gel and liquid-crystalline phase. The thermotropic properties of the above preparations, as monitored by differential scanning calorimetry, were congruent with the spectroscopic data.

Syntheses of 2H- and 13C-labeled 1,2-di-O-hexadecyl-sn-glycero-3-phosphoethanolamines and 1,2-di-O-hexadecyl-sn-glycero-3-phosphocholines

Abstract Improved syntheses of 1,2-di-O-hexadecyl-sn-glycero-3-phosphoethanolamine and 1,2-di-O-hexadecyl-sn-glycero-3-phosphocholine were performed in 71% and 57% overall yields, respectively. Reported reactions which have been greatly improved include the dialkylation of 3-O-benzyl-sn-glycerol, the hydrogenolysis of the 3-O-benzyl-sn-glycerol derivative, and the quaternization of the sn-glycero-3-phosphoethanolamine to the corresponding sn-glycero-3-phosphocholine. Several 2H-, and 13C-labeled analogs of 1,2-di-O-hexadecyl-sn-glycero-3-phosphocholine were then prepared by the appropriate modifications of our synthetic sequence.