Chromatography

In the separation of plant pigment by paper chrom, Malunggay leaves was used as the plant sample. Acetone was added to the grinded leaf sample. Spotting was done five times to ensure visibility of the pigment separation. Three solvents were tested to know which one is best for plant pigment separation.

In the chromatogram where petroleum ether only was used as the solvent, the plant pigment spots were very close to each other and the colors were very hard to distinguish. In another chromatogram where 9:1 petroluem ether:acetone was used, the spots were somehow distanced from each other and its colors are well distinguished. And in the last chromatogram where 9:1:1 petroleum ether:diethyl eteher:acetone was used as the solvent, the spots are far and distinct from each other and the spots’ colors are well defined.

The best solvent chosen is the 9:1:1. As discussed in the introduction, Rf is the Retention factor. It is the ratio of the distance travelled by the substance to the distance travelled by the solvent. Rf value is always between 0 and 1 and has no unit. According to the rf values(see table 4.1) computed, the three spots from the chromatogram gave the largest values (near 1). While the two other chromatogram resulted to values which are small and near to each other. The suitable developing solvent may be governed partly by the observation that slower moving solvents produce rounder and less diffuse spots. The rate of the solvent movement is based among its properties like viscosity, surface tension and density.

It was not an objective of the exercise to identify the identity of the spot (colors: orange, yellow, yellow-green, and green). Although not needed, these are the possible identity of the spots from the resulting chromatogram: carotene, xanthophyll, chlorophyll a, and chlorophyll b. Visible light, or white light, is made up of the colors of the rainbow. Some of these colors are absorbed by pigments and others are reflected. And that is the...