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At least 19 recordsLinked to original sources
PubMed2002

Investigation on lipid asymmetry using lipid probes: Comparison between spin-labeled lipids and fluorescent lipids.

Synthetic lipids with a nitroxide or a fluorescent probe have been extensively used during the last 30 years to determine the transmembrane diffusion of phospholipids in artificial or biological membranes. However, the relevance of data obtained with these modified lipids has sometimes been questioned. Beside possible artefacts introduced by the reporter probe, synthetic lipids used in cells often contain a short fatty acid chain in the sn-2 position, which gives them higher water solubility than naturally occurring lipids. In the present review, we have attempted to give a critical appraisal. Main strategies are recalled and important discoveries obtained with lipid probes on transmembrane lipid traffic in eukaryotic cells are briefly summarized. Examples of artefacts caused by lipid probes are given. Comparisons between data obtained by different techniques such as ESR and fluorescence allow us to emphasize the complementary character of the two approaches and more generally show the necessity to use several probes before drawing conclusions concerning endogenous lipids. In spite of these pitfalls, overall, lipid probes have provided a wealth of useful information that, to date, cannot be obtained with unlabeled lipids.

Animals↗
PubMed1984

Endotoxic properties of chemically synthesized lipid A part structures. Comparison of synthetic lipid A precursor and synthetic analogues with biosynthetic lipid A precursor and free lipid A.

Synthetic lipid A part structures corresponding structurally to a biosynthetic lipid A disaccharide precursor have been analyzed for endotoxic activity in several systems in vivo and in vitro. It was found that a synthetic beta-1,6-linked D-glucosamine disaccharide, which carries four molar equivalents of (R)-3-hydroxytetradecanoyl residues in positions 2, 3, 2' and 3' and phosphoryl groups in positions 1 and 4' (preparation 406), exhibited lethal toxicity, B lymphocyte mitogenicity, the capacity to engender prostaglandin formation in macrophages and to induce endotoxic tolerance, as well as serological lipid A antigenicity. On a weight basis, preparation 406 was of comparable activity to lipid A precursor and bacterial free lipid A. Preparation 406, like lipid A precursor, lacked, however, the ability to induce the local Shwartzman phenomenon and both preparations were of moderate pyrogenicity. Two further synthetic analogues which contained only one phosphoryl group (preparation 404 at C-4', preparation 405 at C-1) showed comparable or diminished activity depending on the test system employed, except in the capacity to inactivate complement where they exhibited, in contrast to preparation 406, significant activity. The results show that the endotoxic principle of lipopolysaccharides, as postulated previously is embedded in the lipid A component. Our results also suggest initial conclusions on the structural requirements for the expression of endotoxin activities.

Animals↗
PubMed1980

Application of highly purified anti-galactocerebroside antibody to the analysis of lipid-lipid or lipid-protein interactions. Significance of lipid matrix.

The significance of the lipid matrix in the reaction of liposomal antigen, antibody and complement (Ag-Ab-C) was analyzed using purified anti-glactocerebroside antibody and synthetic lipids and the following results were obtained. 1. For the optimal Ag-Ab-C reaction it was necesary that galactocerebroside (galxCMH) and lecithin molecules were well dispersed by virtue of cholesterol (Chol) and that the molar ratio of cholesterol to the sum of galactocerebroside and lecithin was more than one. 2. The Ag-Ab-C reactivity changed depending upon the chain length of lecithin, and the maximal reaction was observed in the case of dilauroylphosphatidylcholine. When the fatty acyl chain of lecithin was either shorter or longer than that of dilaurosylphosphatidylcholine, the reactivity was reduced. 3. The Ag-Ab-C reactivity was increased by elongation of the fatty acyl chain of galactocerebroside and an abrupt change was found to be around the carbon number 8 to 10 of the fatty acyl chain. 4. The Ag-Ab-C reactivity was elevated by the increase in unsaturation of fatty acyl moiety. 5. There is a tendency that an increase in the charge of the lipid matrix leads to the reduction of the Ag-Ab-C reactivity. 6. The results suggest that the physicochemical properties of lipids and especially the lipid-lipid interaction in the hydrophobic region of the lipid matrix play an important role in the Ag-Ab-C reaction.

Antibodies↗
PubMed2000

Fluorescence-based evaluation of the partitioning of lipids and lipidated peptides into liquid-ordered lipid microdomains: a model for molecular partitioning into "lipid rafts".

A fluorescence-quenching assay is described that can directly monitor the relative extents of partitioning of different but structurally homologous fluorescent molecules into liquid-ordered (l(o)) domains in lipid vesicles exhibiting liquid-ordered/liquid-disordered (l(o)/l(d)) phase coexistence. Applying this assay to a series of bimane-labeled diacyl phospholipid probes in cholesterol-containing ternary lipid mixtures exhibiting l(o)/l(d) phase separation, we demonstrate that partitioning into l(o)-phase domains is negligible for diunsaturated species and greatest for long-chain disaturated species. These conclusions agree well with those derived from previous studies of the association of lipids and lipid-anchored molecules with l(o)-phase domains, using methods based on the isolation of a detergent-insoluble fraction from model or biological membranes at low temperatures. However, we also find that monounsaturated and shorter-chain saturated species partition into l(o) phases with significant, albeit modest affinities, and that the level of partitioning of these latter species into l(o)-phase domains is significantly underestimated (relative to that of their long-chain saturated counterparts) by the criterion of low-temperature detergent insolubility. Finally, applying the fluorescence-quenching method to a family of lipid-modified peptides, we demonstrate that the S-palmitoyl/S-isoprenyl dual-lipidation motif found in proteins such as H- and N-ras and yeast Ste18p does not promote significant association with l(o) domains in l(o)/l(d)-phase-separated bilayers.

Binding Sites↗
PubMed1983

Lipid peroxidation and haemoglobin degradation in red blood cells exposed to t-butyl hydroperoxide. The relative roles of haem- and glutathione-dependent decomposition of t-butyl hydroperoxide and membrane lipid hydroperoxides in lipid peroxidation and haemolysis.

Red cells exposed to t-butyl hydroperoxide undergo lipid peroxidation, haemoglobin degradation and hexose monophosphate-shunt stimulation. By using the lipid-soluble antioxidant 2,6-di-t-butyl-p-cresol, the relative contributions of t-butyl hydroperoxide and membrane lipid hydroperoxides to oxidative haemoglobin changes and hexose monophosphate-shunt stimulation were determined. About 90% of the haemoglobin changes and all of the hexose monophosphate-shunt stimulation were caused by t-butyl hydroperoxide. The remainder of the haemoglobin changes appeared to be due to reactions between haemoglobin and lipid hydroperoxides generated during membrane peroxidation. After exposure of red cells to t-butyl hydroperoxide, no lipid hydroperoxides were detected iodimetrically, whether or not glucose was present in the incubation. Concentrations of 2,6-di-t-butyl-p-cresol, which almost totally suppressed lipid peroxidation, significantly inhibited haemoglobin binding to the membrane but had no significant effect on hexose monophosphate shunt stimulation, suggesting that lipid hydroperoxides had been decomposed by a reaction with haem or haem-protein and not enzymically via glutathione peroxidase. The mechanisms of lipid peroxidation and haemoglobin oxidation and the protective role of glucose were also investigated. In time-course studies of red cells containing oxyhaemoglobin, methaemoglobin or carbonmono-oxyhaemoglobin incubated without glucose and exposed to t-butyl hydroperoxide, haemoglobin oxidation paralleled both lipid peroxidation and t-butyl hydroperoxide consumption. Lipid peroxidation ceased when all t-butyl hydroperoxide was consumed, indicating that it was not autocatalytic and was driven by initiation events followed by rapid propagation and termination of chain reactions and rapid non-enzymic decomposition of lipid hydroperoxides. Carbonmono-oxyhaemoglobin and oxyhaemoglobin were good promoters of peroxidation, whereas methaemoglobin relatively spared the membrane from peroxidation. The protective influence of glucose metabolism on the time course of t-butyl hydroperoxide-induced changes was greatest in carbonmono-oxyhaemoglobin-containing red cells followed in order by oxyhaemoglobin- and methaemoglobin-containing red cells. This is the reverse order of the reactivity of the hydroperoxide with haemoglobin, which is greatest with methaemoglobin. In studies exposing red cells to a wide range of t-butyl hydroperoxide concentrations, haemoglobin oxidation and lipid peroxidation did not occur until the cellular glutathione had been oxidized. The amount of lipid peroxidation per increment in added t-butyl hydroperoxide was greatest in red cells containing carbonmono-oxyhaemoglobin, followed in order by oxyhaemoglobin and methaemoglobin. Red cells containing oxyhaemoglobin and carbonmono-oxyhaemoglobin and exposed to increasing concentrations of t-butyl hydroperoxide became increasingly resistant to lipid peroxidation as methaemoglobin accumulated, supporting a relatively protective role for methaemoglobin. In the presence of glucose, higher levels of t-butyl hydroperoxide were required to induce lipid peroxidation and haemoglobin oxidation compared with incubations without glucose. Carbonmono-oxyhaemoglobin-containing red cells exposed to the highest levels of t-butyl hydroperoxide underwent haemolysis after a critical level of lipid peroxidation was reached. Inhibition of lipid peroxidation by 2,6-di-t-butyl-p-cresol below this critical level prevented haemolysis. Oxidative membrane damage appeared to be a more important determinant of haemolysis in vitro than haemoglobin degradation. The effects of various antioxidants and free-radical scavengers on lipid peroxidation in red cells or in ghosts plus methaemoglobin exposed to t-butyl hydroperoxide suggested that red-cell haemoglobin decomposed the hydroperoxide by a homolytic scission mechanism to t-butoxyl radicals.

Antioxidants↗
PubMed1992

Cholesterol modulation of lipid intermixing in phospholipid and glycosphingolipid mixtures. Evaluation using fluorescent lipid probes and brominated lipid quenchers.

Carbazole- and indole-labeled phospholipids have been used to monitor the homo- or heterogeneity of lipid mixing in several types of lipid bilayers combining a brominated and a nonbrominated lipid with varying amounts of cholesterol. Experimental quenching curves (relating the normalized probe fluorescence intensity to the mole fraction of brominated lipid) show a characteristic smooth, monophasic form for homogeneous liquid-crystalline lipid mixtures. However, for mixtures exhibiting lipid lateral segregation, such curves show marked perturbations in form over the region of composition where segregation occurs. Using this approach, it is found that high mole fractions of cholesterol (40-50 mol %) promote the formation of apparently homogeneous solutions in mixtures of disaturated and monounsaturated phosphatidylcholines (PCs) that exhibit extensive thermotropic phase separations in the absence of sterol. At only slightly lower levels of cholesterol, however, these systems exhibit inhomogeneous lipid mixing over a wide range of relative proportions of the two PC components. Mixtures of cerebroside and monounsaturated PCs, even at high bilayer cholesterol contents, exhibit significant inhomogeneity in lipid mixing over a wide range of cerebroside/PC ratios. Phase-separating PC/PC and PC/cerebroside mixtures can readily form long-lived metastable solutions when the level of the higher-melting component in the liquid-crystalline phase exceeds its equilibrium solubility by as much as 20-30 mol %; this tendency is significantly increased by cholesterol. Cholesterol shows no significant ability to enhance lipid intermixing in a third type of phase-separating lipid system, combining a monounsaturated PC with a monounsaturated phosphatidic acid--calcium complex. Experiments using cleavable phospholipid conjugates, linking a fluorescent lipid to a brominated lipid, suggest that each fluorescent molecule probes a local lipid domain comprising approximately less than 40-50 nearby acyl chains.

Animals↗
PubMed2000

The mechanism of Fe(2+)-initiated lipid peroxidation in liposomes: the dual function of ferrous ions, the roles of the pre-existing lipid peroxides and the lipid peroxyl radical.

The mechanism of Fe(2+)-initiated lipid peroxidation in a liposomal system was studied. It was found that a second addition of ferrous ions within the latent period lengthened the time lag before lipid peroxidation started. The apparent time lag depended on the total dose of Fe(2+) whenever the second dose of Fe(2+) was added, which indicates that Fe(2+) has a dual function: to initiate lipid peroxidation on one hand and suppress the species responsible for the initiation of the peroxidation on the other. When the pre-existing lipid peroxides (LOOH) were removed by incorporating triphenylphosphine into liposomes, Fe(2+) could no longer initiate lipid peroxidation and the acceleration of Fe(2+) oxidation by the liposomes disappeared. However, when extra LOOH were introduced into liposomes, both enhancement of the lipid peroxidation and shortening of the latent period were observed. When the scavenger of lipid peroxyl radicals (LOO(.)), N,N'-diphenyl-p-phenylene-diamine, was incorporated into liposomes, neither initiation of the lipid peroxidation nor acceleration of the Fe(2+) oxidation could be detected. The results may suggest that both the pre-existing LOOH and LOO(.) are necessary for the initiation of lipid peroxidation. The latter comes initially from the decomposition of the pre-existing LOOH by Fe(2+) and can be scavenged by its reaction with Fe(2+). Only when Fe(2+) is oxidized to such a degree that LOO(.) is no longer effectively suppressed does lipid peroxidation start. It seems that by taking the reactions of Fe(2+) with LOOH and LOO(.) into account, the basic chemistry in lipid peroxidation can explain fairly well the controversial phenomena observed in Fe(2+)-initiated lipid peroxidation, such as the existence of a latent period, the critical ratio of Fe(2+) to lipid and the required oxidation of Fe(2+).

Animals↗
PubMed1980

Lipid-lipid and lipid-protein interactions in chromaffin granule membranes. A spin label ESR study.

The ESR spectra of six different positional isomers of a stearic acid and three of a phosphatidylcholine spin label have been studied as a function of temperature in chromaffin granule membranes from the bovine adrenal medulla, and in bilayers formed by aqueous dispersion of the extracted membrane lipids. Only minor differences were found between the spectra of the membranes and the extracted lipid, indicating that the major portion of the membrane lipid is organized in a bilayer arrangement which is relatively unperturbed by the presence of the membrane protein. The order parameter profile of the spin label lipid chain motion is less steep over the first half of the chain than over the section toward the terminal methyl end of the chain. This 'stiffening' effect is attributed to the high proportion of cholesterol in the membrane and becomes less marked as the temperature is raised. The isotropic hyperfine splitting factors of the various positional isomers display a profile of decreasing polarity as one penetrates further into the interior of the membrane. No marked differences are observed between the effective polarities in the intact membranes and in bilayers of the extracted membrane lipids. The previously observed temperature-induced structural change occurring in the membranes at approx. 35 degrees C was found also in the extracted lipid bilayers, showing this to be a result of lipid-lipid interactions and not lipid-protein interactions in the membrane. A steroid spin label indicated a second temperature-dependent structural change occurring in the lipid bilayers at lower temperatures. This correspond to the onset of a more rapid rotation about the long axis of the lipid molecules at a temperature of approx. 10 degrees C. The lipid bilayer regions probed by the spin labels used in this study may be involved in the fusion of the chromaffin granule membrane leading to hormone release by exocytosis.

Adrenal Medulla↗
PubMed1997

Immunobiological activities of chemically defined lipid A from Helicobacter pylori LPS in comparison with Porphyromonas gingivalis lipid A and Escherichia coli-type synthetic lipid A (compound 506).

Helicobacter pylori lipid A, characterised by a glucosamine beta (1-6) disaccharide 1-(2-aminoethyl)phosphate acylated by (R)-3-hydroxyoctadecanoic acid and (R)-3-(octadecanoyloxy)octadecanoic acid at the 2- and 2'-positions, respectively, exhibited no or very low endotoxic activities, i.e. lethal toxicity in galactosamine-loaded mice, pyrogenicity for rabbits and the activity of the Limulus test compared with Escherichia coli-type synthetic lipid A (compound 506), which possesses beta-(1-6)-linked glucosamine disaccharide 1,4'-bisphosphate, with two acyloxyacyl groups at the 2'- and 3'-positions and two 3-hydroxytetradecanoyl groups at the 2- and 3-positions. The endotoxic properties of H. pylori lipid A were also a little weaker than those of the low endotoxic lipid A of P. gingivalis, which has 1-phospho beta-(1-6)-linked glucosamine disaccharide with 3-hydroxy-15-methylhexadecanoyl and 3-hexadecanoyloxy-15-methylhexadecanoyl groups at the 2- and 2'-positions, respectively. Further, the mitogenic activity of H. pylori lipid A in murine splenic mononuclear cells was also less than those of P. gingivalis lipid A and compound 506. However, H. pylori lipid A induced comparable production of interleukin-6 (IL-6) by human peripheral blood mononuclear cells (PBMC) compared with P. gingivalis lipid A and compound 506. H. pylori lipid A also increased human natural killer cell activity, and strongly agglutinated rabbit erythrocytes. However, the lipid As of H. pylori and P. gingivalis showed lower activities in inducing tumour necrosis factor alpha (TNF-alpha) production by human PBMC and IL-8 production by human gingival fibroblasts than that of compound 506. The structural feature of H. pylori lipid A may be associated with low endotoxic properties and potent immunobiological activities.

Animals↗
PubMed2002

Relationship between lipid levels and clinical outcomes in the Long-term Intervention with Pravastatin in Ischemic Disease (LIPID) Trial: to what extent is the reduction in coronary events with pravastatin explained by on-study lipid levels?

BACKGROUND: The Long-Term Intervention with Pravastatin in Ischemic Disease (LIPID) trial showed that pravastatin significantly reduced mortality and coronary heart disease (CHD) events in 9014 patients with known CHD and total cholesterol 4.0 to 7.0 mmol/L at baseline. Secondary objectives included assessment of CHD event reduction according to lipid levels. METHODS AND RESULTS: We investigated the relationships of baseline and on-study lipids with subsequent CHD events in separate Cox models. Treatment effect on CHD event reduction was examined by baseline lipids and after adjustment for on-study lipid levels. Baseline lipids were significant predictors of CHD events. The adjusted relative risk per mmol/L (on placebo) was 1.24 (P=0.004) for total cholesterol, 1.28 (P=0.002) for low-density lipoprotein cholesterol, and 0.52 (P=0.004) for high-density lipoprotein cholesterol. Apolipoproteins A1 and B were strong predictors (each P=0.001). Pravastatin reduced the risk of the composite outcome of fatal CHD or nonfatal myocardial infarction by 24% (95% confidence interval [CI], 15% to 32%) and the expanded end point of fatal CHD, nonfatal myocardial infarction, unstable angina, or coronary revascularization by 17% (95% CI, 10% to 24%). Similar relative effects were observed for different categories of baseline lipids. The proportion of treatment effect explained by on-study lipid levels was 67% (95% CI, 27% to 106%) for the composite and 97% (95% CI, 49% to 145%) for the expanded end point. The most important lipids associated with event reduction were apolipoprotein B, low-density lipoprotein cholesterol, and the combination of total and high-density lipoprotein cholesterol. CONCLUSIONS: Changes in lipid levels can explain all or most of the observed benefit of pravastatin. Some treatment effect may also be mediated through nonlipid changes.

Adult↗
PubMed1995

Quantitative analysis of lipid-lipid and lipid-protein interactions in membranes by use of pyrene-labeled phosphoinositides.

The lateral and rotational dynamics of phosphoinositides and their interactions with proteins were characterized using pyrene-labeled lipid analogues. In these systems, the collision frequency of pyrene-labeled lipids was studied by monitoring the monomeric pyrene fluorescence yield as a function of their mole fraction in the membranes. From this dependence, the lateral diffusion coefficient and a repulsion factor between two pyrene phosphoinositides could be estimated by applying an extended form of the Milling Crowd model [Eisinger, J., Flores, J., & Petersen, W. P. (1986) Biophys. J. 49, 987-1001]. The repulsion appeared to be highly dependent on the amount of negative charge of the lipid headgroups. From experiments with dioleoylphosphatidylcholine vesicles containing band 3 protein, the fraction of lipid molecules bound to this protein and the minimum number of sites possessing affinity for phosphatidylinositol-4-phosphate could be approximately estimated. The results of this study indicate that phosphoinositides are located preferentially adjacent to band 3. Intramolecular excimer formation of dipyrene-labeled phosphatidylcholine, phosphatidylinositol, and phosphatidylinositol-4-phosphate yielded information about the acyl chain dynamics of lipids surrounding the protein and of lipids in the bulk membrane. Time-resolved measurements of the pyrene fluorescence anisotropy showed that in membranes of resealed erythrocyte ghost cells the rotational freedom of pyrene-labeled phosphatidylinositol-4,5-bisphosphate is smaller than that of pyrene-labeled phosphatidylcholine. In contrast, no significant differences could be detected when these pyrene lipids were dispersed in dioleoylphosphatidylcholine membranes. It is proposed that the nonrandom distribution of the phosphoinositides induced by lipid-lipid repulsion and protein-lipid attraction will have a profound effect on the phospholipase C-catalyzed hydrolysis of the phosphoinositides into second-messenger molecules.

Animals↗
PubMed2000

Intracellular transport of phosphatidic acid and phosphatidylcholine into lipid bodies: use of fluorescent lipids to study lipid-body formation in an oleaginous fungus.

Fluorescent phosphatidic acid and phosphatidylcholine were used to characterize lipid-transport pathways into lipid bodies in an oleaginous fungus, Mortierella ramanniana var. angulispora. Several characteristics of the lipid transport such as temperature dependence and ATP dependence were evaluated. The transport depicted by these fluorescent lipids was consistent with metabolism of radiolabelled lipids, indicating that fluorescent lipids are useful to study lipid-body formation in this fungus. The results dissect lipid transport of phosphatidic acid and phosphatidylcholine into lipid bodies and reveal regulatory steps for lipid-body formation in this fungus.

Adenosine Triphosphate↗
PubMed1991

In vivo and in vitro loading of lipid by artificially lipid-depleted lipophorins: evidence for the role of lipophorin as a reusable lipid shuttle.

Lipid transport in the hemolymph of Manduca sexta is facilitated by a high density lipophorin in the resting adult insect (HDLp-A, d approximately 1.109 g/ml) and by a low density lipophorin during flight (LDLp, d approximately 1.060 g/ml). Lipophorin presumably shuttles different lipids between sites of uptake or storage, and sites of utilization. In order to shuttle lipid, a lipid-depleted lipophorin should be able to reload with lipid. To test this hypothesis, we used HDLp-A particles that were artificially depleted of either phospholipid (d approximately 1.118 g/ml) or diacylglycerol (d approximately 1.187 g/ml) and subsequently radiolabeled in their protein moiety. Upon injection into adult moths, both particles shifted their density to that of native HDLp-A, indicating lipid loading. Also, upon subsequent injection of adipokinetic hormone, both particles shifted to a lower density (d approximately 1.060 g/ml) indicating diacylglycerol loading and conversion to LDLp. Both phospholipid and diacylglycerol loading were also studied using an in vitro system. The lipid-depleted particles were incubated with fat body that had been radiolabeled in either the phospholipid or the triacylglycerol fraction. Transfer of radiolabeled phospholipid and diacylglycerol from fat body to lipophorin was observed. During diacylglycerol loading, apoLp-III associated with lipophorin, whereas phospholipid loading occurred in the absence of apoLp-III. The results show the ability of lipid-depleted lipophorins to reload with lipid and therefore reaffirm the role of lipophorin as a reusable lipid shuttle.

Animals↗
PubMed2003

Effect of different lipid extraction methods on delta13C of lipid and lipid-free fractions of fish and different fish feeds.

For many ecological applications of stable carbon isotope techniques, it is necessary to separate the lipid and lipid-free fractions. The effect of different lipid extraction methods on the isotope signature of the remaining lipid-free matter as well as the lipid fraction was tested. A hot extraction form of the Soxhlet method using petrol-ether was compared with two liquid-liquid extraction methods for lipid determination described by Bligh and Dyer and Smedes. Solid samples of fish and different natural food items were subjected to extraction and the carbon isotope ratios in lipid and lipid-free matter determined by IRMS. All methods were suitable for lipid extraction from all samples analysed here and did not cause biologically relevant differences (> 1%) in carbon isotopic ratios, except the Bligh and Dyer extraction method using chloroform which caused systematic errors for delta13C when applied to diatoms.

Animal Feed↗
PubMed1980

Lipid-lipid and lipid-protein interactions in membranes.

Over the past decade spectroscopic methods (fluorescence, ESR, and NMR) have been used to provide new information about the molecular dynamics of lipid-lipid and lipid-protein interactions in membranes. The various methods of characterizing isotropic and anisotropic motion are described. Lipid bilayers are highly dynamic, with rapid acyl chain motion and rapid lateral diffusion in the plane of the membrane. In membranes where proteins penetrate through the bilayer, a large hydrophobic surface area exists in contact with the bilayer lipids. Lipids at the protein interface are in dynamic equilibrium with the remaining pools of bilayer. The protein has been shown spectroscopically to have some influence on the dynamics of the nearest neighbor lipids, leaving the rest of the bilayer relatively unperturbed. Evidence is summarized that, in some cases, the lipid composition in the interfacial region is influenced by the protein.

Chemical Phenomena↗
PubMed1997

Volatile anaesthetic effects on calcium conductance of planar lipid bilayers formed with synthetic lipids or extracted lipids from sarcoplasmic reticulum.

Volatile anaesthetics are known to increase leakage of calcium from the light fraction of skeletal sarcoplasmic reticulum (L-SR) which has no calcium release channels. To explore the role of the lipid environment, we have examined the effect of volatile anaesthetics on calcium conductance (gCa) of lipid membranes. Planar lipid bilayers were formed with a mixture of synthetic phospholipids and cholesterol, resembling the composition of SR membranes, or with lipids extracted from skeletal L-SR, gCa was estimated by calculating the calcium transference number (tCa) using diffusion potential measurements. Membranes formed with L-SR-extracted lipids had a higher gCa than membranes formed with synthetic lipids. Volatile anaesthetics increased total conductance and gCa in a dose-dependent manner, but did not affect tCa or membrane specific capacitance. In membranes formed with L-SR-extracted lipids, isoflurane induced the largest increase in gCa (1260 (SEM 304) % increase, n = 4, 0.94 mmol litre-1), followed by enflurane (264 (75)%, n = 5, 1.88 mmol litre-1) and halothane (53 (33)%, n = 5; 1.54 mmol litre-1). In membranes formed with synthetic lipids, volatile anaesthetic-induced increases in gCa followed the same trend but were larger. Volatile anaesthetics increased gCa without changing the ionic selectivity of membranes. However, the magnitude of the increase in gCa in the presence of volatile anaesthetics cannot account for the previously observed calcium leakage from L-SR vesicles. Therefore, the volatile anaesthetic-induced increase in calcium leakage in L-SR vesicles must be mediated via other pathways involving membrane proteins.

Anesthetics, Inhalation↗
PubMed1997

Structural elucidation of oxygenated storage lipids in cucumber cotyledons. Implication of lipid body lipoxygenase in lipid mobilization during germination.

At early stages of germination, a special lipoxygenase is expressed in cotyledons of cucumber and several other plants. This enzyme is localized at the lipid storage organelles and oxygenates their storage triacylglycerols. We have isolated this lipid body lipoxygenase from cucumber seedlings and found that it is capable of oxygenating in vitro di- and trilinolein to the corresponding mono-, di-, and trihydroperoxy derivatives. To investigate the in vivo activity of this enzyme during germination, lipid bodies were isolated from cucumber seedlings at different stages of germination, and the triacylglycerols were analyzed for oxygenated derivatives by a combination of high pressure liquid chromatography, gas chromatography/mass spectrometry, and nuclear magnetic resonance spectroscopy. We identified as major oxygenation products triacylglycerols that contained one, two, or three 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid residues. During germination, the amount of oxygenated lipids increased strongly, reaching a maximum after 72 h and declining afterward. The highly specific pattern of hydroperoxy lipids formed suggested the involvement of the lipid body lipoxygenase in their biosynthesis. These data suggest that this lipoxygenase may play an important role during the germination process of cucumber and other plants and support our previous hypothesis that the specific oxygenation of the storage lipids may initiate their mobilization as a carbon and energy source for the growing seedling.

Chromatography, High Pressure Liquid↗