A new type of two-dimensional thin-layer chromatography (TLC mapping) for analysis of acidic glycosphingolipid molecular species.

A new approach to the analysis of acidic glycosphingolipid molecular species has been developed by using two-dimensional thin-layer chromatography: TLC mapping. TLC mapping was accomplished by use of two types of high-performance thin-layer chromatographic (HPTLC) plates. A NH2-silica gel HPTLC plate developed with chloroform:methanol:1% diethylamine (50:47:15) was used for the first-dimension chromatography. Acidic glycosphingolipids on the plate were transferred to a normal silica gel HPTLC plate by developing the plate with chloroform:methanol:ammonia (4:10:6) after the NH2 plate was placed face to face on top of the silica gel HPTLC plate. Acidic glycophingolipids on the HPTLC plate were then developed with chloroform: methanol:0.2% CaCl2 (60:40:10) for the second-dimension chromatography. The use of the TLC-mapping method allowed us to compare detailed membrane constituents, including minor molecular species with regard to acidic glycophingolipids. The method is rapid and simple and is feasible for determining acidic glycosphingolipid from biological samples.