diphenylhexatriene and 1-palmitoyl-2-((2-(4-(6-phenyl-1-3-4-hexatrienyl)phenyl)ethyl)carbonyl)-3-phosphatidylcholine

Orientational order parameters of two diphenylhexatriene (DPH)-based fluorescent probes, 2-(3-(diphenylhexatrienyl)propanoyl)-1-hexadecanoyl-sn-glycero-3-p hosphocholine (DPHpPC) and 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), in dipalmitoylphosphatidylcholine (DPPC) Langmuir-Blodgett monolayers on quartz have been determined by total internal reflection fluorescence (TIRF). From these order parameters orientation distributions were reconstructed by the maximum-entropy method. For monolayers transferred from the liquid-condensed phase, preferential tilt angles with respect to the substrate normal around 14 degrees in the tail region and 5 degrees near the glycerol-acyl chain linkage were found, as reflected by the DPHpPC and TMA-DPH probes, respectively. The degree of ordering near the headgroup region seems to be larger than that further away from the surface. A substantial fraction of the TMA-DPH probes have a flat orientation and are probably located between the phospholipid headgroups and the substrate surface. Monolayers transferred from the liquid-expanded phase show a more random ordering, and most of the probe molecules (DPHpPC) are more or less flat on the surface. The results are consistent with earlier atomic force microscopy measurements on identical monolayers and are in reasonable agreement with previously published data on other organized phospholipid systems.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Biophysical Phenomena; Biophysics; Diphenylhexatriene; Fluorescent Dyes; Membrane Lipids; Microscopy, Atomic Force; Models, Chemical; Molecular Structure; Phosphatidylcholines; Phospholipids; Spectrometry, Fluorescence; Thermodynamics

We have used an extended Perrin equation which was in agreement with literature data for steady-state anisotropy (rSS) for a wide variety of artificial and isolated biological membranes labeled with various probes (Van der Meer et al. (1986) Biochim. Biophys. Acta 854, 38-44 to obtain the static component (r infinity) for the intact plasma membranes of living cells. We show that lipid structural order parameters can be obtained for DPH and TMA-DPH in the plasma membranes of intact cells. We have examined the relationship between 'fractional limiting hindered anisotropy', r infinity/r0, which is related to the lipid structural order parameter, of DPH, TMA-DPH, DPHpPC, and a series of depth-dependent probes (n-(9-anthroyloxy) fatty acids, with n = 2-16), using data from 19 cell types. There was a linear relationship between r infinity/r0 values of DPH and TMA-DPH, but the relationship between either of these probes was non-linear with respect to DPHpPC or the series of fatty acid probes. The relationship between r infinity/r0 values of DPHpPC and the series of fatty acid probes was linear, suggesting that they not only undergo similar motions in the membrane, but also experience similar types of restriction to motion, a type which is different from that experienced by DPH and TMA-DPH. We show that for the plasma membranes of living cells, 'second degree' order parameters can be estimated for DPH and TMA-DPH, and propose that the parameter r infinity/r0, or the 'fractional limiting hindered anisotropy', analogous to a 'first degree' order parameter, can be estimated for DPHpPC and the depth-dependent fatty acid probes to evaluate the density of membrane packing.

Topics: Cell Membrane; Cells, Cultured; Diphenylhexatriene; Fluorescence Polarization; Fluorescent Dyes; Humans; Kinetics; Phosphatidylcholines

We have investigated the reason for the sensitivity of the fluorescence excited-state lifetime of 1,6-diphenyl-1,3,5-hexatriene (DPH) and its phospholipid derivatives, 1-palmitoyl-2-[2-[4-(6-phenyl-trans-1,3,5- hexatrienyl)phenyl]ethyl)carbonyl)-3-sn-phosphatidylcholine (DPHpPC) and 1-palmitoyl-2-[2-[4-(6-phenyl-trans-1,3,5- hexatrienyl)phenyl]ethyl)carbonyl)-3-sn-phosphatidic acid (DPHpPA), to the concentration of these probes in dipalmitoylphosphatidylcholine (DPPC) multilamellar membranes (Barrow, D. A., and B. R. Lentz, 1985. Biophys. J. 48:221-234; Parente, R. A., and B. R. Lentz. 1985. Biochemistry. 24:6178-6185). We have interpreted self-quenching data, excitation and emission spectra, and phase and modulation lifetime data in terms of a model that envisions dimerization of these probes in a membrane bilayer. It is proposed that dimerization alters the symmetry of the DPH excited state so as to allow more rapid decay via the normally symmetry-disallowed route from the 1Ag* state. Global analysis of fluorescence phase shift and modulation ratio data for DPHpPC in terms of the dimerization model provided a good fit of these data as a function of both modulation frequency and probe concentration. Global analysis of a similar set of data for the charged phosphatide DPHpPA predicted that this probe was much less prone to dimerize than was the uncharged DPHpPC. This physically reasonable result provides support for the assumptions made in the development of our model. We conclude that the dimerization model allows rationalization of many of the anomalous photophysical properties of DPH and its derivatives in membranes.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Diphenylhexatriene; Fluorescent Dyes; Kinetics; Lipid Bilayers; Models, Theoretical; Phosphatidic Acids; Phosphatidylcholines; Polyenes; Spectrometry, Fluorescence