Simultaneous quantitation of ceramides and 1, 2‐diacylglycerol in tissues by latroscan thin‐layer chromatography‐flame‐ionization detection

K Okumura, K Hayashi, I Morishima, K Murase… - Lipids, 1998 - Wiley Online Library
K Okumura, K Hayashi, I Morishima, K Murase, H Matsui, Y Toki, T Ito
Lipids, 1998Wiley Online Library
Abstract Ceramides and 1, 2‐diacylglycerol have been demonstrated in intracellular
signaling pathways. A method of simultaneous mass determination of ceramides and 1, 2‐
diacylglycerol in tissues was developed using the latroscan which combines thin layer
chromatography and flame ionization detection (TLC/FID) techniques. Because of relatively
low amounts of these components in tissues, the fraction of nonpolar lipids, which included
ceramides and glycerides, was eluted with chloroform/acetone mixture (3∶ 1, vol/vol) …
Abstract
Ceramides and 1,2‐diacylglycerol have been demonstrated in intracellular signaling pathways. A method of simultaneous mass determination of ceramides and 1,2‐diacylglycerol in tissues was developed using the latroscan which combines thin layer chromatography and flame ionization detection (TLC/FID) techniques. Because of relatively low amounts of these components in tissues, the fraction of nonpolar lipids, which included ceramides and glycerides, was eluted with chloroform/acetone mixture (3∶1, vol/vol) through a silicic acid column to eliminate the polar phospholipids. Development of Chromarods was carried out using three solvent systems in a four‐step development technique. The relationship of the peak area ratio to weight ratio compared with cholesteryl acetate added as an internal standard was linear. The amount of ceramides increased with incubation of rat heart homogenate and human erythrocyte membranes in the presence of sphingomyelinase (E.C. 3.1.4.12). The latroscan TLC/FID system provided a quick and reliable assessment of ceramides and 1,2‐diacylglycerol.
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