diphenylhexatriene and 1-2-distearoyllecithin

Differential scanning calorimetry (DSC) and fluorescence spectroscopy are useful techniques for investigating the phase transitions of phospholipid bilayers. In this study, these methods have been extended to determine the effects of ethanol on DSPC and DSPC/2 mol.% cholesterol bilayers. The biphasic effect of the main transition was observed on the DSC heating scans above 0.60 M ethanol. In addition, the concentration at which the biphasic effect occurs is not significantly changed in the presence of 2 mol.% cholesterol. For the fluorescence studies, 1,6-diphenyl-1,3,5-hexatriene (DPH) has been incorporated into the bilayer to monitor the phase transitions through the displacement of DPH. This fluorescent probe is used to directly determine the onset of interdigitation in the bilayer systems as indicated by a large decrease in the DPH fluorescence intensity. The addition of cholesterol lowered and broadened the transition temperatures of the phosphatidylcholine (PC) system. However, 2 mol.% cholesterol did not have a significant effect on the induction of the interdigitated phase in DSPC as observed from the small difference in ethanol threshold concentration for the two systems. This suggests that DSPC forms a more stable interdigitated gel phase than other PCs with shorter acyl chains.

Topics: Calorimetry, Differential Scanning; Cholesterol; Diphenylhexatriene; Ethanol; Gels; Lipid Bilayers; Phosphatidylcholines; Spectrometry, Fluorescence; Temperature

Although it is now well established that the fully interdigitated phase is induced in saturated like-chain phosphatidylcholines (PCs) by a variety of amphipathic molecules including alcohols, no systematic study of the properties of the inducing molecules has been reported. To elucidate the stereochemical features that lead to the alcohol induction of interdigitation in PCs, we have investigated the induction of interdigitation in distearoylphosphatidylcholine (DSPC) by a series of alcohols. Our previously established DPH (1,6-diphenyl-1,3,5-hexatriene) fluorescence intensity method has been expanded (P. Nambi, E. S. Rowe, and T. M. McIntosh (1988), Biochemistry 27:9175-9182) and used to determine which of the alcohols induce interdigitation and to determine the threshold concentrations for each. We have found that each of the n-alcohols up to heptanol and several branched alcohols are capable of inducing interdigitation in DSPC; octanol and nonanol do not appear to induce interdigitation by these criteria. The threshold concentrations for interdigitation for each of these alcohols up to heptanol were found to be correlated with the membrane: buffer partition coefficients. The mole fraction of bound alcohol at the threshold concentration was similar for each of the alcohols up to pentanol. These results are discussed in terms of a general mechanism of the formation of the interdigitated phase.

Topics: Alcohols; Biophysical Phenomena; Biophysics; Diphenylhexatriene; In Vitro Techniques; Macromolecular Substances; Molecular Conformation; Phosphatidylcholines; Spectrometry, Fluorescence; Stereoisomerism; Structure-Activity Relationship; Temperature

Four fluorescent diphenylhexatriene derivatives were considered as membrane probes, namely two ammonium compounds, 3-(diphenylhexatrienyl)propyltrimethylammonium (TMAP-DPH) and 22-(diphenylhexatrienyl)docosyltrimethylammonium (LcTMA-DPH), and two phospholipids, 1-palmitoyl-2-[3-(diphenylhexatrienyl)propanoyl]-sn-glyc ero-3-phosphocholine (DPHpPC) and 1-palmitoyl-2-[21-(diphenylhexatrienyl)henicosanoyl]-sn-phos phocholine (LcDPHpPC). For each pair, the molecules differ by the length of the polymethylenic spacer between the fluorescent moiety and the polar head, so one pair comprises two short chain molecules (C3 spacer) and the other two long chain molecules (C21 or C22 spacer). The partitioning of these probes between gel and liquid crystalline phases of multilamellar vesicles with binary composition (DEPC and DSPC) was measured by a method based on fluorescence anisotropy. The partitioning was shown to depend strongly on the length of the spacer. Short chain probes preferably partition into fluid phases (Kf/s = 1.7 +/- 0.3 for TMAP-DPH; 2.6 +/- 0.11 for DPHpPC), whereas long chain probes show a strong preferential partitioning for gel phases of the vesicles (Kf/s = 0.12 +/- 0.06 for LcTMA-DPH; 0.22 +/- 0.11 for LcDPHpPC). This strong partitioning may be explained by the interdigitation of the long polymethylenic chains across the mid-point of the lipid bilayer (I.E. Mehlhorn et al. (1988) Biochim. Biophys. Acta 939, 151-159), which is enhanced by the better packing provided by a gel phase.

Topics: Diphenylhexatriene; Fluorescence Polarization; Fluorescent Dyes; Liposomes; Membrane Fluidity; Models, Biological; Molecular Structure; Phosphatidylcholines; Structure-Activity Relationship

2,2-Bis(p-chlorophenyl)-1,1-dichloroethylene (DDE) interaction with model and native membranes was studied by means of fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH), probing the bilayer core, and by intramolecular excimerization of 1,3-di(1-pyrenyl) propane (Py(3)Py), probing the outer regions of the bilayer. In the gel phase of DMPC bilayers, DDE induces concentration-dependent fluidizing effects into the hydrophobic core, but no effects are detected in the outer regions of the membrane, as evaluated by DPH and Py(3)Py, respectively. Regarding the fluid phase, DDE has no apparent effect on the bilayer center, but it induces a limited ordering effect on the outer regions. Similar effects are described for bilayers of DPPC and DSPC. Unlike DPH, Py(3)Py is very sensitive to DPPC and DSPC pretransitions, not abolished by DDE (50 microM), as opposite to the effects observed with lindane (Antunes-Madeira, M.C., Almeida, L.M. and Madeira, V.M.C. (1990) Biochim. Biophys. Acta 1022, 110-114), but similar to those observed with DDT (Antunes-Madeira, M.C., Almeida, L.M. and Madeira, V.M.C. (1991) Pestic. Sci. 33, 347-357). DDE inhibits to some extent the cholesterol-induced ordering in DMPC bilayers and high cholesterol concentrations (> or = 30 mol%) do not prevent DDE interaction, as evaluated by DPH. On the other hand, the effects of DDE reported by Py(3)Py depend on temperature and cholesterol contents of DMPC bilayers. For cholesterol levels ranging from 10 to 50 mol% and temperatures below the phase transition of DMPC, Py(3)Py fails to detect any significant effect. Nevertheless, above the phase transition, Py(3)Py detects either ordering effects of DDE at low cholesterol contents (< 30 mol%) or fluidizing effects at high cholesterol levels (> or = 30 mol%). The results in native membranes correlate reasonably with those obtained in models of synthetic lipids. Thus, DPH does not detect any apparent effect of DDE in relatively fluid native membranes of sarcoplasmic reticulum, but detects moderate disordering effects in membranes of brain microsomes and erythrocytes, i.e., membranes with high cholesterol. On the other hand, Py(3)Py reports ordering effects of DDE in fluid membranes of sarcoplasmic reticulum, an effect similar to that observed in fluid systems of synthetic lipids without or with low cholesterol. Additionally, as described for models, Py(3)Py detects disordering effects of DDE in cholesterol rich membranes, namely, brain micros

Topics: 1,2-Dipalmitoylphosphatidylcholine; Animals; Dichlorodiphenyl Dichloroethylene; Dimyristoylphosphatidylcholine; Diphenylhexatriene; Fluorescence Polarization; Hot Temperature; Membrane Fluidity; Membranes; Models, Biological; Phosphatidylcholines; Pyrenes

Topics: Chlorfenvinphos; Diphenylhexatriene; Fluorescence Polarization; Fluorescent Dyes; Insecticides; Liposomes; Membrane Fluidity; Membrane Lipids; Models, Biological; Phosphatidylcholines; Spectrometry, Fluorescence

Fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) was used to study the interaction of DDT with model and native membranes. DDT decreases the phase transition midpoint temperature (Tm) of liposomes reconstituted with dimyristoyl-, dipalmitoyl- and distearoylphosphatidylcholines (DMPC, DPPC and DSPC), and broadens the thermotropic profile of the transition. The effects of DDT are concentration dependent and are more pronounced in bilayers of short-chain lipids, e.g., DMPC. The insecticide fails to alter DPH polarization in the fluid phase of the above lipids. Similar effects were observed in binary mixtures of DMPC plus DPPC. Furthermore, DDT alters the single broad transition of the equimolar mixture of DMPC plus DSPC into a biphasic transition. The lower temperature component has a midpoint at 25 degrees C, i.e., a value close to the Tm of DMPC. DDT inhibits to some extent the cholesterol-induced ordering in DMPC bilayers and high cholesterol concentrations (greater than or equal to 30 mol%) do not prevent insecticide interaction, conversely to the effect observed for lindane (Antunes-Madeira, M.C. and Madeira, V.M.C. (1989) Biochim. Biophys. Acta 982, 161-166). Apparently, the bilayer order is not disturbed by DDT in fluid native membranes of mitochondria and sarcoplasmic reticulum, but moderate disordering effects are noticed in membranes enriched in cholesterol, namely, brain microsomes and erythrocytes.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Cholesterol; DDT; Dimyristoylphosphatidylcholine; Diphenylhexatriene; Erythrocytes; Fluorescence Polarization; Lipid Bilayers; Liposomes; Membrane Fluidity; Microsomes; Models, Biological; Phosphatidylcholines; Phospholipids; Sarcoplasmic Reticulum; Temperature

The effect of melittin on different binary mixtures of phospholipids has been studied by polarization of DPH fluorescence in order to determine if melittin can induce phase separation. Since the interaction between lipids and melittin is sensitive to both electrostatic and hydrophobic forces, we have studied the effect of the acyl chain length and of the polar head group of the lipids. In spite of the difference of the chain length between dipalmitoylphosphatidylcholine (DPPC) and distearoylphosphatidylcholine (DSPC), no phase separation occurs in an equimolar mixture of these lipids in presence of melittin. However, when the charged lipid dipalmitoylphosphatidylglycerol (DPPG) is mixed with either DPPC or DSPC, the addition of melittin leads to phase separation. The DSPC/DPPG/melittin system, which shows a very complex thermotropism, has also been studied by Raman spectroscopy using DPPG with deuteriated chains in order to monitor each lipid independently. The results suggest that the higher affinity of melittin for DPPG leads to a partial phase separation. We propose the formation of DPPG-rich domains perturbed by melittin and peptide-free regions enriched in DSPC triggered by the head group charge and chain-length differences.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Bee Venoms; Diphenylhexatriene; Fluorescence Polarization; Melitten; Phosphatidylcholines; Phosphatidylglycerols; Phospholipids; Spectrum Analysis, Raman; Temperature; Thermodynamics

Liposomes have been prepared from dipalmitoylphosphatidylcholine containing small amounts of a synthetic photochromic phospholipid, 'Bis-Azo PC'. In the dark, these are stable at room temperature, and contents do not significantly leak over weeks. Photoisomerisation results in immediate release of trapped marker, and in liposome fusion to form larger structures. Fusion has been detected using a fluorescence polarisation assay, and confirmed by electron microscopy. In mixtures, fusion occurs between 'photochromic' liposomes and those of pure lipid. Bis-Azo PC contains two photochromic acyl chains; analogues bearing a single photochromic chain appear to have little effect on bilayer permeability after isomerisation. Photo-induced leakage and liposome fusion suggest possible applications for localised drug delivery as an adjunct to phototherapy. The ability to non-invasively trigger fusion processes should be useful in fundamental studies of membrane interactions. We believe this to be the first report of photo-induced fusion to date.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Diphenylhexatriene; Fluoresceins; Fluorescence Polarization; Fluorescent Dyes; Isomerism; Light; Liposomes; Membrane Fusion; Microscopy, Electron; Phosphatidylcholines; Photochemistry; Temperature

In the microsomal fraction of thyroid glands, the temperature dependence of DPH fluorescence polarization showed a discontinuity in the range of 29-33 degrees C. The transition temperatures of DMPC, DPPC and DSPC are near to the observed for the microsomal fraction. So that, thyroid peroxidase (TPO) was incorporated into liposomes made with these phospholipids. When DPH was incorporated in this peroxidase-liposome complex, a less pronounced phase transition was observed in the profiles of temperature dependence of DPH polarization, and the incorporation of the enzyme decreased the Tc. Arrhenius plots of TPO incorporated into liposomes showed discontinuities at similar temperatures observed by fluorescence polarization. The decrease of transition temperature of liposomes induced by thyroid peroxidase incorporation suggests that this enzyme seems to need a fluid medium for its enzyme activity.

Topics: Animals; Cattle; Dimyristoylphosphatidylcholine; Diphenylhexatriene; In Vitro Techniques; Liposomes; Peroxidases; Phosphatidylcholines; Pulmonary Surfactants; Thyroid Gland