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• W2170053363 abstract "About one-fourth the phosphatidylcholines (PC) from retina photoreceptor rod outer segment (ROS) membranes contain docosahexaenoic acid (22:6n-3) at sn-2 and a very long chain polyunsaturated fatty acid (VLCPUFA) (C24 to C36) at the sn-1 position of the glycerol backbone. In order to study the thermotropic behavior of these PCs, subfractions and molecular species of PC (16:0/22:6, 18:0/22:6, 22:6/22:6, 32:5/22:6, 32:6/22:6, 34:5/22:6), were isolated from bovine ROS, and liposomes containing different proportions of these PCs and dimyristoyl-PC (DMPC) or dipalmitoyl PC (DPPC) were compared using the fluorescence probes Laurdan and 1,6-diphenyl-1,3,5-hexatriene (DPH). With both probes, the 22:6n-3 containing PCs from ROS, in all proportions tested, decreased the transition temperature (Tt) of both DMPC and DPPC. Below the transition temperature, coexistence of phases was evidenced in all cases. Liposomes formed with 100% of any of these PCs did not show phase transitions in the temperature range studied (8°C to 50°C). At physiological temperatures, as it is likely to be the case in ROS membranes, all of these PC species were in the liquid-crystalline state. With Laurdan, all dipolyunsaturated PCs seemed to behave similarly: despite the large number of double bonds per molecule, all of them decreased the Tt of DPPC less than did the hexaenoic PCs. With DPH, an ample difference was detected between the dipolyunsaturates, 22:6/22:6-PC and VLCPUFA/22:6-PCs, and between the latter and hexaenoic PCs throughout the temperature range studied.This difference is consistent with the interpretation that the largest “disorder” produced by PCs containing a VLCPUFA like 32:6n-3 at the sn-1 position occurs toward the center of the membrane. About one-fourth the phosphatidylcholines (PC) from retina photoreceptor rod outer segment (ROS) membranes contain docosahexaenoic acid (22:6n-3) at sn-2 and a very long chain polyunsaturated fatty acid (VLCPUFA) (C24 to C36) at the sn-1 position of the glycerol backbone. In order to study the thermotropic behavior of these PCs, subfractions and molecular species of PC (16:0/22:6, 18:0/22:6, 22:6/22:6, 32:5/22:6, 32:6/22:6, 34:5/22:6), were isolated from bovine ROS, and liposomes containing different proportions of these PCs and dimyristoyl-PC (DMPC) or dipalmitoyl PC (DPPC) were compared using the fluorescence probes Laurdan and 1,6-diphenyl-1,3,5-hexatriene (DPH). With both probes, the 22:6n-3 containing PCs from ROS, in all proportions tested, decreased the transition temperature (Tt) of both DMPC and DPPC. Below the transition temperature, coexistence of phases was evidenced in all cases. Liposomes formed with 100% of any of these PCs did not show phase transitions in the temperature range studied (8°C to 50°C). At physiological temperatures, as it is likely to be the case in ROS membranes, all of these PC species were in the liquid-crystalline state. With Laurdan, all dipolyunsaturated PCs seemed to behave similarly: despite the large number of double bonds per molecule, all of them decreased the Tt of DPPC less than did the hexaenoic PCs. With DPH, an ample difference was detected between the dipolyunsaturates, 22:6/22:6-PC and VLCPUFA/22:6-PCs, and between the latter and hexaenoic PCs throughout the temperature range studied. This difference is consistent with the interpretation that the largest “disorder” produced by PCs containing a VLCPUFA like 32:6n-3 at the sn-1 position occurs toward the center of the membrane. Most of the lipid classes of the disk membranes that are tightly packed into the outer segments of bovine retinal rods contain docosahexaenoic acid (22:6n-3). About one-fourth of the molecular species of such phospholipids, which closely interact with rhodopsin, are “dipolyunsaturated” (1Miljanich G.P. Sklar L.A. White D.L. Dratz E.A. Disaturated and dipolyunsaturated phospholipids in the bovine retinal rod outer segment disk membrane.Biochem. Biophys. Acta. 1979; 552: 294-306Google Scholar) or “supraenoic” (2Aveldaño M.I. Bazan N.G. Molecular species of phosphatidylcholine, -ethanolamine, -serine, and -inositol in microsomal and photoreceptor membranes of bovine retina.J. Lipid Res. 1983; 24: 620-627Google Scholar), named that way to reflect the fact that they have PUFAs at both the sn-1 and the sn-2 positions of the glycerol backbone. Further studies showed that a considerable proportion of the dipolyunsaturated molecular species of the lipids present in rod outer segment (ROS) membranes contain 22:6n-3 as one of the acyl chains, the other one being a PUFA of the n-3 or the n-6 series with long (C20, C22) or with very long chains (C24 to C36) (3Aveldaño M.I. A novel group of very long chain polyenoic fatty acids in dipolyunsaturated phosphatidylcholines from vertebrate retina.J. Biol. Chem. 1987; 262: 1172-1179Google Scholar, 4Aveldaño M.I. Sprecher H. Very long chain (C24 to C36) polyenoic fatty acids of the n-3 and n-6 series in dipolyunsaturated phosphatidylcholines from bovine retina.J. Biol. Chem. 1987; 262: 1180-1186Google Scholar). Whereas all bovine ROS phospholipid classes contain dipolyunsaturated molecular species (2Aveldaño M.I. Bazan N.G. Molecular species of phosphatidylcholine, -ethanolamine, -serine, and -inositol in microsomal and photoreceptor membranes of bovine retina.J. Lipid Res. 1983; 24: 620-627Google Scholar), the fatty acid distribution among them differs (5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar). Phosphatidylserine, phosphatidylinositol, and phosphatidylethanolamine dipolyunsaturated species contain 20, 22, or 24 carbon PUFA, whereas phosphatidylcholine (PC), besides these, is made up of significant proportions of dipolyunsaturated species that contain PUFA with up to 36 carbon atoms [very long chain (VLC)PUFA] (3Aveldaño M.I. A novel group of very long chain polyenoic fatty acids in dipolyunsaturated phosphatidylcholines from vertebrate retina.J. Biol. Chem. 1987; 262: 1172-1179Google Scholar, 4Aveldaño M.I. Sprecher H. Very long chain (C24 to C36) polyenoic fatty acids of the n-3 and n-6 series in dipolyunsaturated phosphatidylcholines from bovine retina.J. Biol. Chem. 1987; 262: 1180-1186Google Scholar, 5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar). These PCs are thus unique in that they display not only the highest degree of unsaturation but also the largest total number of carbon atoms yet reported for mammalian glycerophospholipids. Except for didocosahexaenoyl-PC, in most dipolyunsaturated PCs from ROS, having two PUFA of different length per molecule, 22:6n-3 tends to be located at the sn-2 position of the glycerol backbone, while the VLCPUFA tend to locate predominantly at the sn-1 position (5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar). Thus, at least in this positional “preference”, the VLCPUFA containing species of ROS PC resemble the more ubiquitous and widely known “hexaenoic” species of PC, like 16:0/22:6-PC or 18:0/22:6-PC, in which the saturated fatty acid and 22:6n-3 are generally found at sn-1 and at sn-2, respectively. The fact that the same kind of VLCPUFA-containing molecular species occur in a glycerophospholipid specifically located in the visual cells of animal species so distant in evolution as fish, birds, and various mammals suggested that such molecules probably fulfill an important requirement for the function of photoreceptor membrane proteins including rhodopsin (3Aveldaño M.I. A novel group of very long chain polyenoic fatty acids in dipolyunsaturated phosphatidylcholines from vertebrate retina.J. Biol. Chem. 1987; 262: 1172-1179Google Scholar). Docosahexaenoate-containing lipids may be expected to provide a fluid bilayer, but at physiological temperatures this condition could be fulfilled with much simpler lipids, such as oleic or linoleic acid containing species. In fact, reconstitution experiments have indicated that fluidity of the lipid bilayer is a necessary, though not sufficient, condition for rhodopsin functionality. Thus, the rates of rhodopsin photochemical transformations are reduced when the protein is reconstituted in liposomes of dimyristoyl-PC, even at temperatures above the Transition midpoint temperature (Tm) (6Baldwin P.A. Hubbell W.L. Effects of lipid environment on the light-induced conformational changes of rhodopsin. 1. Absence of metarhodopsin II production in dimyristoylphosphatidylcholine recombinant membranes.Biochemistry. 1985; 24: 2624-2632Google Scholar, 7Mitchell D.C. Kibelbeckand J. Litman B.J. Rhodopsin in dimyristoylphosphatidylcholine-reconstituted bilayers forms metarhodopsin II and activates Gt.Biochemistry. 1991; 30: 37-42Google Scholar), perhaps because these PCs have “too short” acyl chains compared with those of the lipids present in the native membrane. The equilibrium concentration of metharhodopsin II formed after rhodopsin photolysis is improved in the presence of phospholipids with one or more 22:6 acyl chains (8Litman B.J. Mitchell D.C. A role for phospholipid polyunsaturation in modulating membrane protein function.Lipids. 1996; 31: S193-S197Google Scholar, 9Brown M.F. Modulation of rhodopsin function by properties of the membrane bilayer.Chem. Phys. Lipids. 1994; 73: 159-180Google Scholar). Moreover, phospholipids with 22:6 acyl chains are required for the optimal kinetic functioning of meta II – transducin coupling (10Mitchell D.C. Niu S-L. Litman B.J. Optimization of receptor-G protein coupling by bilayer lipid composition I: kinetics of rhodopsin-transducin binding.J Biol. Chem. 2001; 276: 42801-42806Google Scholar, 11Niu S-L. Mitchell D.C. Litman B.J. Optimization of receptor-G protein coupling by bilayer lipid composition II: formation of metarhodopsin II-transducin complex.J. Biol. Chem. 2001; 276: 42807-42811Google Scholar, 12Litman B.J. Niu S-L. Polozova A. Mitchell D.C. The role of docosahexaenoic acid containing phospholipids in modulating G protein-coupled signaling pathways: visual transduction.J. Mol. Neurosci. 2001; 16: 237-242Google Scholar). No studies have yet been designed to investigate the possible function of PCs with VLCPUFA in photoreceptor membranes. These species of PC are not the major ones, but are not negligible either: as a group, they amount to about one-third of all the PCs present in bovine ROS (5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar). With the current knowledge about the organization of lipids in naturally occurring membranes, one of the most intriguing yet still unanswered questions that arises is how is it possible that phospholipid species like the described PCs (e.g., 32:6/22:6-PC), having two acyl chains at the same time so highly unsaturated and so uneven in length, accommodate in the thickness of the photoreceptor membrane. Another question that has not yet been addressed, and that we tried to approach in the present paper, is what are the physical properties of these peculiar PCs, and how they differ, if they do, from the more abundant - and much more ubiquitous - species also having 22:6n-3 at sn-2 but a saturate at sn-1. Whereas the introduction of a single cis double bond into a saturated acyl chain is known to result in a large decrease in molecular order in the liquid crystalline phase, the effects of higher levels of unsaturation are not as widely agreed upon, and appear to vary, depending on the specific location of the double bonds, on the location of the first double bond counting from the carboxyl ester end of the fatty acid, and on whether one or both phospholipid acyl chains contain unsaturations (13Cevc G. How membrane chain-melting phase-transition temperature is affected by the lipid chain asymmetry and degree of unsaturation: an effective chain-length model.Biochemistry. 1991; 30: 7186-7193Google Scholar). In PCs containing two equal polyenoic chains (18:2/18:2-PC and 20:4/20:4-PC), it has been shown that double bonds in excess of two per fatty acid chain do not substantially change the transition temperature (14Keough K.M. Kariel N. Differential scanning calorimetric studied of aqueous dispersions of phosphatidylcholines containing two polyenoic chains.Biochem. Biophys. Acta. 1987; 902: 11-18Google Scholar), but this type of measurements has not been done in dipolyunsaturated species in which one of the chains is much longer than the other. Thus, one cannot extrapolate from what is currently known what could be the behavior of dipolyunsaturated species of ROS PC, more heterogeneous as a group than previously considered. With the above questions in mind, we explored the physical characteristics of some of the molecules isolated from subfractions of ROS PC, and the effects they produce when added to liposomes made of dimiristoyl- or dipalmitoyl-PC (DMPC, DPPC), whose thermotropic behavior is very well characterized. We have used the amphiphilic fluorescent probes Laurdan (6-dodecanoyl-2-(−dimethylami-no) naphthalene) (15Antollini S. Barrantes F.J. Disclosure of discrete sites for phospholipid and sterols at the protein-lipid interface in native acetylcholine receptor-rich membrane.Biochemistry. 1998; 37: 16653-16662Google Scholar), and 1,6-diphenyl-1,3,6-hexatriene (DPH). Laurdan is considered to have a uniform lateral and transbilayer distribution, thus making it a good reporter molecule to sense molecular dynamics of solvent dipoles in the membrane as a whole (16Parasassi T. De Stasio G. d'Ubaldo A. Gratton E. Phase fluctuation in phospholipid membranes revealed by Laurdan fluorescence.Biophys. J. 1990; 57: 1179-1186Google Scholar). The main dipoles sensed by Laurdan in membranes are water molecules. Differences in water content in the hydrophilic/hydrophobic interface region of the membrane correlate with differences in solvent dipolar relaxation, and thus indirectly correlate with variations in membrane lipid fluidity (17Yu W. So P.T. French T. Gratton E. Fluorescence generalized polarization of cell membranes: a two-photon scanning microscopy approach.Biophys. J. 1996; 70: 626-636Google Scholar). DPH has been successfully used to study lipid bilayer structure over the past three decades (18Martinez-Senac M.M. Corbalan-Garcia S. Gomez-Fernandez J.C. The structure of the C-terminal domain of the pro-apoptotic protein bak and its interaction with model membranes.Biophys. J. 2002; 82: 233-243Google Scholar, 19Jacobson K. Papahadjopoulos D. Phase transitions and phase separations in phospholipid membranes induced by changes in temperature, pH, and concentration of bivalent cations.Biochemistry. 1975; 14: 152-161Google Scholar). Fluorescence polarization measurements provide a measure of the rotation diffusion of fluorophores. The rate of rotation of this probe is thought to reflect the viscous hindrance imposed by its immediate environment (20Lackowicz J.R. Prendergast F.G. Hogen D. Differential polarized phase fluorometric investigations of diphenylhexatriene in lipid bilayers. Quantitation of hindered depolarizing rotations.Biochemistry. 1979; 18: 508-527Google Scholar). Using these tools, in this paper we describe the thermal behavior of liposomes formed with some of the 22:6n-3 containing PCs of ROS membranes, and explore how the presence of these PCs affects the thermotropic behavior of liposomes of DPPC. Despite the indirect nature of the information provided by the fluorescence probes employed, they reveal differential characteristics of the 22:6 containing PC present in photoreceptor membranes. Laurdan was purchased from Molecular Probes (Eugene, OR). DPH and all other drugs were obtained from Sigma Chemical Co., St. Louis, MO. All solvents used were of high pressure liquid chromatography (HPLC) grade (J. T. Baker, Phillipsburg, NJ). The gas chromatography (GC) and HPLC equipments used were from Varian, Inc. Bovine eyes were obtained from a local abattoir and placed on ice within 10 min of the animal's death. The retinas were excised on ice, under dim red light, and the rod outer segments (ROS) were isolated therefrom using a discontinuous gradient of sucrose. Briefly, retinas were removed, shaken in a 40% sucrose solution containing 1mM MgCl2, 1mM DTT, 0.1mM PMSF in 70mM sodium phosphate buffer (pH 7.2) and the retinal ROS were separated from the remains of retinas by centrifugation at 2,200 g for 4 min. The supernatants containing ROS were diluted with sucrose-free buffer and centrifuged at 35,000 for 30 min. The pelleted ROS were gently resuspended and purified on a discontinuous gradient of sucrose (21Kühn H. Light-regulated binding of proteins to photoreceptor membranes and its use for the purification of several rod cell proteins.Methods Enzymol. 1982; 81: 556-564Google Scholar). The ROS band was isolated from the 0.84/1.00 M interface, washed with sucrose-free buffer, and pelleted. Lipids were extracted from the ROS pellets according to the procedure of Bligh & Dyer (22Bligh E.G. Dyer W.J. A rapid method of total lipid extraction and purification.Can. J. Biochem. Physiol. 1959; 37: 911-917Google Scholar), resolved into classes by preparative TLC on silica gel G-plates, located under UV light after spraying with dichlorofluorescein, and eluted from the support using the solvents described by Arvidson (23Arvidson G.A.E. Structural and metabolic heterogeneity of rat liver glycerophospholipids.Eur. J. Biochem. 1968; 4: 478-486Google Scholar). For TLC, chloroform-methanol-acetic acid-0.15 M NaCl (50:25: 8:2.5, v/v/v/v) (24Brown E.R. Subbaiah P.V. Differential effects of eicosapentaenoic acid and docosahexaenoic acid on human skin fibroblasts.Lipids. 1984; 29: 825-829Google Scholar) was used as the first developing solvent. Phos-phatidylcholine tended to separate into three bands, each containing different groups of molecular species, which were separately recovered. After elution, each of these bands was subjected to a second TLC using chloroform-methanol-ammonia (65:25:5, v/v/v). The lipid content was determined by phosphorus analysis (25Rouser G. Fleischer A. Yamamoto A. Two-dimensional thin-layer chromatography for the isolation and analysis of trace amounts of polar lipids and determination of polar lipids by phosphorus analysis of spots.Lipids. 1970; 5: 494-496Google Scholar). The fatty acid composition was determined by gas-chromatography (5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar) after preparation of fatty acid methyl ester derivatives. The latter were usually purified before GC on TLC plates that had been pre-washed with a polar solvent (methanol-ether, 75:25, v/v), by means of hexane-ether (95:5, v/v). Molecular species of PC were isolated from the PC bands resolved by TLC by means of reverse-phase HPLC. The separation was performed at room temperature using a short column (3.5 cm × 0.4 cm ID) of stainless steel packed with spherical, 5 μm particles of silica covered with octadecylsilane (C18) (Zorbax ODS, Dupont). Adequate aliquots of the extracts containing PC, previously filtered to remove traces of particulate matter, were injected. For elution, a solvent gradient was applied, consisting of aqueous 1 mM phosphate buffer, pH 7.4, (component A) and methanol (component B). Eluents were degassed before use. The columns were equilibrated with 93%B and the flowrate was 1 ml/min throughout. Gradient elution was performed in three steps: 93%B for the first 15 min, 95%B between 16 min and 40 min, and 96%B thereafter. The bands were detected with a variable-wavelength UV spectrophotometer, placed at 205 nm. The collected PC bands were recovered after evaporation of the solvent to dryness and re-dissolved in chloroform methanol (C:M) 2:1 (v/v). The separated PC molecular species were quantified by phosphorus analysis and identified by GC of their fatty acids. For fluorescence measurements, the isolated PC molecular species were prepared as multilamellar liposomes. Aliquots of the isolated PCs, alone or in combination with an adequate proportion of DPPC or DMPC in C:M (2:1), were mixed with an aliquot of the fluorescent probes used (Laurdan in ethanol or DPH in THF) to reach a lipid-fluorescent probe ratio of 100:1. The mixtures were evaporated in 1 h in the dark under N2, resuspended in buffer A (20 mM HEPES buffer, 150 mM NaCl, and 0.25 mM MgCl2, pH 7.4) and sonicated for 30 min. Each sample was diluted with buffer A to have a final lipid concentration of 100 μM in the (10 × 10 mm) quartz cuvettes that were used. In order to rule out the possibility of probe perturbation of the lipid bilayer and/or probe-probe interactions, we did a control experiment with a probe-lipid ratio of 1:300 and a final lipid concentration of 150 μM in the cuvettes. No significant differences were observed with respect to the present results. All fluorimetric measurements were performed in a SLM model 4800 fluorimeter (SLM Instruments, Urbana, IL) using the vertically polarized light beam from a Hannovia 200 W Hg/Xe arc obtained with a Glan-Thompson polarizer (4 nm excitation and emission slits). Emission spectra were corrected for wavelength-dependent distortions. The temperature was set with a thermostatted circulating water bath. Excitation generalized polarization (GP) (16Parasassi T. De Stasio G. d'Ubaldo A. Gratton E. Phase fluctuation in phospholipid membranes revealed by Laurdan fluorescence.Biophys. J. 1990; 57: 1179-1186Google Scholar, 26Parasassi T. De Stasio G. Ravagnan G. Rush R. Gratton E. Quantitation of lipid phases in phospholipid vesicles by the generalized polarization of Laurdan fluorescence.Biophys. J. 1991; 60: 179-189Google Scholar) was calculated according to the expression (Eq. 1)exGP = (I434 − I490) / (I434 + I490) where I434 and I490 are the emission intensities at the characteristic wavelength of the gel phase (434 nm) and the liquid-crystalline phase (490 nm), respectively. exGP values were obtained from the emission spectra at different excitation wavelengths (320–410 nm) or at only one excitation wavelength (360 nm). Emission GP was calculated according to the following formalism: (Eq. 2)emGP = (I410 − I340) / (I410 + I340) where I410 and I340 are the excitation intensities at the wavelengths corresponding to the gel (410 nm) and the liquid-crystalline (340 nm) phases, respectively (27Parasassi T. Loiero M. Raimondi M. Ravagnan G. Gratton E. Absence of lipid gel-phase domains in seven mammalian cell lines and in four primary cell types.Biochem. Biophys. Acta. 1993; 1153: 143-154Google Scholar). The emGP values were obtained from the excitation spectra at different emission wavelengths (420–500 nm). The excitation and emission wavelengths used were 365 and 425 nm, respectively. Fluorescence polarization measurements were done in the T format with Schott KV418 filters in the emission channels and corrected for optical inaccuracies and for background signals. The polarization value, P, was obtained as follow (28Shinitzky M. Barenholz Y. Fluidity paramenters of lipid regions determined by fluorescence.Biochem. Biophys. Acta. 1978; 515: 367-394Google Scholar) (Eq. 3)P = [(Iv/Ih)v − (Iv/Ih)h]/[ (Iv/Ih)v + (Iv/Ih)h] where (Iv/Ih)v and (Iv/Ih)h are the ratios of the emitted vertical or horizontally polarized ligth to the exciting, vertical or horizontally polarized, light, respectively. Polarization values can range between −0.33 and 0.5, the higher values denoting the higher structural lipid order. The transition temperatures (Tt) were obtained by calculation of the second derivative of the experimental values (GP or polarization vs. temperature). We considered that the Tt correspond to a second derivative value of zero. Partial resolution of ROS lipid classes into subfractions containing different groups of molecular species was first described by Miljanich et al. (1Miljanich G.P. Sklar L.A. White D.L. Dratz E.A. Disaturated and dipolyunsaturated phospholipids in the bovine retinal rod outer segment disk membrane.Biochem. Biophys. Acta. 1979; 552: 294-306Google Scholar), who observed that the dipolyunsaturated molecular species of all ROS phospholipids including PC migrated ahead of other species by TLC or column chromatography. Previous studies from our laboratory have shown that the dipolyunsaturated molecular species of ROS PC that contained C24-C36 polyenes, as a group, tended to migrate ahead of the rest of the PCs when subjected to TLC (5Aveldaño M.I. Phospholipid species containing long and very long polyenoic fatty acids remain with rhodopsin after hexane extraction of photoreceptor membranes.Biochemistry. 1988; 27: 1229-1239Google Scholar, 29Aveldaño M.I. Dipolyunsaturated species of retina phospholipids and their fatty acids.in: Léger C.L. Béreziat G. Biomembranes and Nutrition. Publications de l'Institute National de la Santé et de la Recherche Médicale (INSERM), Paris, France1989Google Scholar). This is because the large number of carbon atoms per molecule at the acyl chains hinders or reduces interactions between the polar part of the lipid and the polar adsorbent surface silanol groups. An almost clear-cut separation of ROS PC into three groups of species can be achieved at appropriate silica support-lipid ratios (termed PCdown, PCmiddle, and PCup in Table 1), mainly made up by disaturated, hexaenoic, and dipolyunsaturated groups of species, respectively, as shown by the fatty acid composition. The data in Table 1 show that the group of species migrating ahead on TLC contain, in addition to 22:6n-3, virtually all of the VLCPUFA of ROS PC.TABLE 1Typical fatty acid composition (wt%) of the three main groups of molecular species from ROS phosphatidylcholine as separated by TLCFatty AcidPCdownPCmiddlePCup14:01.415:00.8 0.10.116:070.2 22.11.217:01.0 0.80.118:06.6 22.63.024:00.1 0.010.116:15.4 1.60.217:10.6 0.2t18:18.0 6.20.720:1t 0.318:2n-61.2 1.30.318:3n-60.2 0.50.320:4n-60.1 4.41.422:4n-60.1 0.40.224:4n-60.1 1.51.830:4n-60.632:4n-62.832:5n-61.018:3n-30.1 0.10.218:4n-30.1 0.50.320:5n-3 0.50.322:5n-30.1 1.91.122:6n-32.0 32.646.524:5n-30.2 1.42.624:6n-30.1 0.31.126:5n-3t 0.31.226:6n-3tt0.328:5n-30.628:6n-30.230:5n-32.430:6n-30.632:5n-310.632:6n-311.434:5n-33.634:6n-31.736:5n-32.336:6n-31.7The fatty acid composition was analyzed by gas chromatography. t, percentages lower than 0.05%. Open table in a new tab The fatty acid composition was analyzed by gas chromatography. t, percentages lower than 0.05%. The six 22:6n-3 containing molecular species of PC used in this study were isolated from the PCup and PCmiddle bands by means of reverse phase HPLC. The dipolyunsaturated species obtained from the PCup fraction were didocosahexaenoyl (22:6/22:6-PC) and three species with VLCPUFA and 22:6: 32:5/22:6-PC, 32:6/22:6-PC, and 34:5/22:6-PC. From the more abundant PCmiddle fraction two “hexaenoic” species were isolated: 16:0/22:6-PC and 18:0/22:6-PC The thermal behavior of the three main subfractions of ROS PC obtained by the TLC separation was measured by determining the GP of Laurdan as a function of temperature. The behavior of liposomes prepared from PCtotal, PCdown, PCmiddle, and PCup was compared with that of two well-characterized systems, DMPC and DPPC (Fig. 1). DMPC and DPPC by themselves presented a pronounced change of GP in a narrow range of temperature, characteristic of a phase transition. Both PCmiddle and PCup showed GP values lower than those of DMPC and did not exhibit a phase transition, showing that, even at the lowest temperature tested, these lipids were forming a liquid-crystalline phase. On the contrary, the curve obtained with PCdown was displaced to the right of that of DMPC, and rather closer to that of DPPC (Fig.1A). PCtotal presented an intermediate behavior, quite close to that of DMPC but displaced to the left. Thus, the PCtotal curve lay between the curve of PCdown (richer in saturates than PCtotal) and those of PCmiddle and PCup (richer in polyenes than PCtotal). When liposomes were prepared with 50% DMPC and 50% of either PCdown, PCmiddle or PCup (Fig. 1B), the resulting curves became closer to that of 100% DMPC. In Fig. 1C the behavior of 100% DPPC is compared with that of combinations of 50% DPPC and 50% of the three types of lipids from ROS-PC. In all three cases, the composite liposomes presented a Tt lower than that of DPPC alone. Figure 2displays the values of Tt obtained from the curves in Fig. 1. Whereas the presence of PCdown increased the Tt of DMPC, there was almost no difference between the Tt of DPPC and that of PCdown, consistent with the fact that this fraction contained mostly saturated fatty acids longer than C14. The presence of both PCmiddle and PCup decreased the Tt of both DMPC and DPPC, consistent with the fact that these subfractions of PCs are polyunsaturated. The actual Tt of the 22:6 containing species studied here are all below zero. Using calorimetry, some dipolyenoic (18:2/18:2, 20:4/20:4, and 22:6/22:6) PCs exhibited endothermic transitions ranging from about −80°C to −30°C (30Kariel N. Davidson E. Keough K.M. Cholesterol does not remove the gel-liquid crystalline phase transition of phosphatidylcholines containing two polyenoic acyl chains.Biophys. Biochem. Acta. 1991; 1062: 70-76Google Scholar), and using Raman spectroscopy, the phase transit" @default.
• W2170053363 created "2016-06-24" @default.
• W2170053363 creator A5079635755 @default.
• W2170053363 creator A5090250392 @default.
• W2170053363 date "2002-09-01" @default.
• W2170053363 modified "2023-10-09" @default.
• W2170053363 title "Thermal behavior of liposomes containing PCs with long and very long chain PUFAs isolated from retinal rod outer segment membranes" @default.
• W2170053363 cites W1040799746 @default.
• W2170053363 cites W1539365968 @default.
• W2170053363 cites W1543772563 @default.
• W2170053363 cites W1566497779 @default.
• W2170053363 cites W1567065116 @default.
• W2170053363 cites W1633176701 @default.
• W2170053363 cites W1966652107 @default.
• W2170053363 cites W1967326841 @default.
• W2170053363 cites W1967972955 @default.
• W2170053363 cites W1970440189 @default.
• W2170053363 cites W1972701314 @default.
• W2170053363 cites W1976907613 @default.
• W2170053363 cites W1978991073 @default.
• W2170053363 cites W1979735910 @default.
• W2170053363 cites W1988680661 @default.
• W2170053363 cites W1989784582 @default.
• W2170053363 cites W1993052912 @default.
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