Oxidation of unsaturated phospholipids in membrane bilayer mixtures is accompanied by membrane fluidity changes

Steady-state and time-resolved fluorescence spectroscopy has been used to o btain information on oxidation processes and associated dynamical and struc tural changes in model membrane bilayers made from single unilamellar vesic les (SUV's) of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) mixed with i ncreasing amounts of 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (SAPC). The highly unsaturated arachidonoyl chain containing four double bo nds is prone to oxidation, Lipid oxidation was initiated chemically by a pr oper oxidant and could be followed on line via the fluorescence changes of an incorporated fluorescent lipophilic fatty acid: 4,4-difluoro-5-(4-phenyl -1,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic acid (BP-C11). The oxidation rate increases with an increasing amount of SAPC. Size measur ements of different SUV's incorporated with a trace amount of a phosphatidy lcholine analogue of BP-C11 using fluorescence correlation spectroscopy hav e demonstrated that an increase of lipid unsaturation results in smaller si zed SUV's and therefore to a larger curvature of the outer bilayer leaflet. This suggests that the lipid-lipid spacing has increased and that the unsa turated fatty acyl chains are better accessible for the oxidant. Oxidation results in some characteristic physical changes in membrane dynamics and st ructure, as indicated by the use of specific fluorescence probes. Fluoresce nce measurements of both dipyrenyl- and diphenylhexatriene-labelled PC intr oduced in non-oxidised and oxidised DOPC-SAPC membranes clearly show that t he microfluidity (local fluidity at the very site of the probes) significan tly decreases when the oxidised SAPC content increases in the lipid mixture . A similar effect is observed from the lateral diffusion experiments using monopyrenyl PC in the same membrane systems: the lateral diffusion is dist inctly slower in oxidised membranes. (C) 2000 Elsevier Science B.V. All rig hts reserved.