Thin Layer Chromatography - An MS Hybrid Thin Layer Plate Transport Interface for Peptide-Like Materials 2


The plate was the aluminum-backed silica gel plate manufactured by Merk (Cat. No. 1.0555) but in a modified form. Silica was scraped from one side of the plate in such a manner that caused the layer to taper over a horizontal distance of 1 cm. By doing this, the thickness gradually decreased from the original 250 mm to zero over the 1 cm distance, thus providing a gentle change from one coating to the next. About 5 g of the desorption medium containing 2,4,6 trihydroxyacetophenone or 2,5-dihydroxy benzoic acid was dispersed in a mixture of toluene and ethyl ether (60:40 for 2,4,6 trihydroxyacetophenone and 80:20 for 2,5-dihydroxy benzoic acid) and was sprayed onto the exposed plate backing.


The silica gel layer was appropriately shielded from the spray by a microscope slide. The coating was carried out after the separation had been developed. The samples used to test the device were Valinomycin, cyclosporin, and the protected peptide BOC-Phe-Lue-Phe-Lue--Phe. The TLC separation was carried out in two stages, first using chloroform that only moved the BOC-Phe-Lue-Phe-Lue--Phe and then ethyl ether-toluene that only moved Valinomycin. This procedure resulted in three separate spots suitably separated. The other side of the plate was then coated with the desorption material as described above and the plate rotated through 90˚. The solute spots were eluted into the desorption zone by the solvent mixture chloroform-acetic acid-methanol containing 0.1% trifluoro-acetic acid, (1.5:1:1). After transfer of the spots by elution each spot area was treated with methanol-water mixture (1:1) to improve analyte-desorbing agent co-crystallization.