Bilayers of Arachidonic Acid Containing Phospholipids Studied by ²H and ³¹P NMR Spectroscopy

The configurational properties and dynamics of the arachidonic acyl chains of phospholipid bilayers have been investigated for the first time by solid-state ²H NMR techniques, with the goal of achieving a better understanding of the biological roles of polyunsaturated phospholipids. Vinyl perdeuterated arachidonic acid (20:4^5,8,11,14-d₈) was prepared from eicosatetraynoic acid (ETYA) and was esterified with 1-palmi-toyl-sn-glycero-3-phosphocholine to yield 1 -palmitoyl-2-vinylperdeuterioarachidonoyl-sn-glycero-3-phos-phocholine [(16:0)(20:4-d₈)PC]. ³¹P NMR spectra of aqueous dispersions of (16:0)(20:4-d₈)PC as well as 1 -perdeuteriopalmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine [(per-²H-16:0)(20:4)PC] were characteristic of the lamellar liquid-crystalline state. The dispersions had similar ³¹P chemical shift anisotropics, with little apparent motional averaging of the lineshapes due to macroscopic reorientation of liposomes or lateral diffusion of phospholipids about their curved surfaces. Comparison to other phosphatidylcholines indicated that both samples comprised the fully hydrated L_a phase plus excess water. However, the dispersion of (16:0)(20:4-d₈)PC yielded relatively narrow powder-type ²H NMR spectra, compared to (per-²H-16:0)(20:4)PC in the liquid-crystalline state. The differences in the ²H NMR powder patterns thus reflect differences in the configurational properties of the polyunsaturated sn-2 arachidonic acyl chain compared to the saturated sn-1 palmitic chain. When the powder-type ²H NMR spectra of the (16:0)(20:4-d₈)PC bilayer were dePaked (θ = 0°), they showed three kinds of deuterons upon integration: one with a large splitting (~25-35 kHz), two with intermediate splittings (=10-15 kHz), and the remainder with smaller splittings (~0.3–5 kHz). The residual quadrupolar couplings of the vinylic >C=C< deuterons were less than those of most C²H₂ groups of the saturated acyl chain. It is concluded that some of the >C=C< segments are inequivalent and exhibit differences along the arachidonic acyl chain, as found previously for the C²H₂ groups of the polymethylene chains of saturated phospholipid bilayers. In addition, the ²H spin-lattice relaxation rates, R_1Z, of the vinylic >C=C< deuterons of the sn-2 arachidonic acyl chain were larger relative to their quadrupolar splittings than for most C²H₂ segments of the saturated sn-1 chain. Simple models for interpretation of the findings are briefly mentioned and discussed. The results indicate clearly that the structural and dynamic properties of polyunsaturated acyl chains differ from those of saturated and monounsaturated chains in phospholipid bilayers.