C. Separation of pigments

The condensed extract of each layer was spotted separately as a line, 2.5 cm from the edge of a 20 × 10 cm glass plate coated with an 0.50 mm thick layer of Merck Silica gel.

The plate was developed with a solvent containing one part methanol to five parts petroleum ether. The standard method of ascending thin-layer chromatography was used. The plates were air dried and the coloured bands then removed and eluted with either petroleum ether, methanol, ethanol, water or acetone, etc...., depending on the solubility of the pigments. The pigments so obtained were stored in the dark because the coloured bands on the plates decolourize very rapidly in the light. The eluted pigments were chromatographed on other plates using different solvents in case they were not homogenous.