7-NITROBENZ-2-OXA-1,3-DIAZOLE-4-YL-LABELED PHOSPHOLIPIDS IN LIPID-MEMBRANES - DIFFERENCES IN FLUORESCENCE BEHAVIOR

Steady-state and time-resolved fluorescence properties of the 7-nitrob enz-2-oxa-1,3-diazole-4-yl (NBD) fluorophore attached either to the sn -2 acyl chain of various phospholipids (phosphatidylcholine, phosphati dylethanolamine, phosphatidylserine, and phosphatidic acid) or to the polar headgroup of phosphatidylethanolamine were studied after inserti on of these NBD-labeled lipid probes into unilamellar vesicles of phos phatidylcholine, phosphatidylglycerol, phosphatidic acid, and phosphat idylserine. The fluorescence response of the NBD group was observed to strongly depend on the chemical structure and physical state of the h ost phospholipids and on the chemical structure of the lipid probe its elf. Among the various fluorescence parameters studied, i.e., Stokes' shifts, lifetimes, and quantum yields, the quantum yields were by far the most affected by these structural and environmental factors, where as the Stokes' shifts were practically unaffected. Thus, depending on the phospholipid probe and the host phospholipid, the fluorescence emi ssion of the NBD group was found to vary by a factor of up to 5. Caref ul analysis of the data shows that for the various couples of probe an d host lipid molecules studied, deexcitation of the fluorophore was do minated by nonradiative deactivation processes. This great sensitivity of the NBD group to environmental factors originates from its well-kn own solvatochromic properties, and comparison of these k(nr) values wi th those obtained for n-propylamino-NBD in a set of organic solvents c overing a large scale of polarity indicates that in phospholipids, the NBD fluorophore experiences a dielectric constant of around 27-41, co rresponding to a medium of relatively high polarity. From these epsilo n values and on the basis of models of the dielectric transition that characterizes any water-phospholipid interface, it can be inferred tha t for all of the phospholipid probes and host phospholipids tested, th e NBD group is located in the region of the polar headgroups, near the phosphoglycerol moiety of the lipids.