Synthetic Ceramides Research Articles

Molecular arrangements of sphingolipids. The monolayer behaviour of ceramides

A series of synthetic ceramides have been studied at the air-water interface by recording the surface pressure-area isotherms at continuous compression.Ceramides that contain a 4,5-trans-double bond in the long chain base were found to condense into a close-packed arrangement with vertical chains already at a very low surface pressure. The corresponding saturated compounds adopt a similar close packed arrangement only at high surface pressure. At 30 dynes/cm, a lateral pressure representative of biological membranes, the area per molecule and compressibility was further found to depend on the number and configuration of the hydroxyl groups. The presence of a 2-D-hydroxyl group in the fatty acud generally promotes the condensation. A similar effect is observed if the long chain base contains a 4-D-hydroxyl group. Cis-double bonds or methyl branches in the fatty acid chain, which increase the space requirement, limit the lateral interaction of the polar group. However the 15-cis-double bond of nervonic acid can be accommodated without any distortion of the close-packing arrangement.

Molecular arrangements in sphingolipids Conformation and hydrogen bonding of ceramide and their implication on membrane stability and permeability

The preferred conformation of the ceramide part of sphingolipids has been deduced from single crystal structures of a series of sphingolipid constituents: N- tetracosanoylphytosphingosine , glycosylphytosphingosine hydrochloride, sphingosine hydrochloride, triacetylsphingosine, dl-2-hydroxytetradecanoic acid and N- stearoylethanolamine . The amide group of the ceramide, which serves as a link between the hydrocarbon chains, has a basic significance for the conformation of the entire molecule. This rigid group, which comprises six atoms in a planar conformation, adopts a perpendicular orientation towards the axes of the two hydrocarbon chains. The carbonyl oxygen thereby turns into an eclipsed position with the hydrogen atom at carbon atom 2 of the sphingosine. A parallel chain stacking is achieved by a sharp perpendicular bend of the fatty acid. This bend is produced by a sequence of two −60° rotations about the C-C bonds at both sides of the α-carbon atom. The orientation of the hydrogen bond donors and acceptors of the amide group and the hydroxyl groups allow lateral interaction with other lipid molecules. The proposed models are supported by infrared spectra, thin-layer chromatographic behaviour and monolayer studies of synthetic model ceramides. The functional role of the hydrogen bonding groups in the ceramide part of sphingolipids is emphasized and their significance for the formation of lateral hydrogen bonds within the membrane layer and thereof arising effects on membrane stability and permeability are dicussed.

Identification and quantitation of free ceramides in human platelets

Free ceramides were isolated from human platelets. Their structures were unequivocally determined by gas-liquid chromatography-mass spectrometry of the trimethylsilyl ether derivatives. The major components were N-(palmitoyl) sphingosine, N-(stearoyl) sphingosine, N-(eicosanoyl) sphingosine, N-(docosanoyl) sphingosine, N-(tetracosanoyl) sphingosine, and N-(tetracosenoyl) sphingosine. Sphinganine-and sphingadienine-containing ceramides as well as ceramides containing other unsaturated acids were also present. The amount of ceramides was determined by quantitative gas-liquid chromatography, using radioactive ceramide as internal standard and synthetic crystalline ceramides for comparison of peak areas. The concentration of ceramides was found to be 1.31 micro g/10(9) platelets or 0.47 micro g/mg of platelet protein.

Identification and quantitative determination of ceramides in human plasma.

Ceramides in human plasma have been isolated, identified and quantitated, using silicic acid chromatography, thin-layer chromatography, argentation chromatography of the ceramide acetates, and gas-liquid chromatography—mass spectrometry of ceramide trimethylsilyl ethers. Synthetic ceramides were prepared and used as references. Only ceramides with normal fatty acids were detected. Sphingosine was the main long-chain base, but ceramides with hexadecasphingosine, heptadecasphingosine, sphingadienine, sphinganine, and nonadecasphingosine were also found. Ceramides with long chain fatty acids dominated. The total amount of ceramides was 5.64 ± 1.51 μg/ml among 27 apparently healthy subjects.

Gas–liquid chromatography–mass spectrometry of synthetic ceramides containing 2-hydroxy acids

Ceramides containing either sphingosine or sphinganine and one of the 2-hydroxy acids, 14h:0, 16h:0, 18h:0, 20h:0, 22h:0, 24h:0, and 26h:0 were prepared and separated by gas chromatography as the 1,3,2'-tri-O-tri-methylsilyl derivatives. Mass spectrometric analyses of these derivatives showed that the ions formed on electron impact can be used to determine unequivocally the structures of the long-chain base and the fatty acid residue in the ceramide. Proposed structures of ions and the mechanisms of reaction of their formation are supported by mass spectra of homologous derivatives, by deuterium labeling experiments, and by high-resolution on mass spectrometry.