EXPERIMENT 4

PAPER CHROMATOGRAPHY
QUESTIONS:
1. Why is the chromatogram developed in an essentially closed system?
2. What are the considerations in choosing a chromatographic solvent?
3. Compare and contrast between normal phase and reverse phase
chromatography.
4. Explain how iodine crystals help to visualize the other pigments.
5. Suggest one method that can be used to visualize highly conjugated
compounds but are not colored.
ANSWERS:
1. The chromatogram is developed in an essentially closed system because it
needs to be developed in an atmosphere saturated with solvent vapor. If it is
not closed, then evaporation of solvent may occur faster than it is supposed
to be, which replaces the capillary action that must take place. In this case,
separation of components may not be achieved.
2. Since the basic principle of chromatography is that different substances have
different partition coefficients between stationary and mobile phases, it is
important to choose a solvent that is polar enough to have different affinities
with the compound. The compound that interacts weakly with the stationary
phase will spend most of its time in the mobile phase and move rapidly while
compounds that interact strongly with the stationary phase will move very
slowly. As a consequence, each will move through a system with different
rates that leads to successful separations.
3. In normal phase, the stationary phase is polar and selectively retains the
polar substances in the mixture and the mobile phase is non polar that should
move non polar compounds in the system. On the other hand, reverse phase
chromatography has a non polar stationary phase and polar mobile phase.
4. The use iodine vapour in visualizing the other pigments depends on the
ability of the mixture to have a charge-transfer complexes with iodine.
Compounds that do not contain

or Lewis basic functions do not form

charge transfer complexes with iodine. Through weak electronic attraction,

iodine will collect on the spots and form a yellow or brown stain on most
organic compounds.
5. If the compounds are not colored, the paper is first sprayed with a solution of
sulfuric acid for better charring. Fluorescent materials can be visualized with
ultraviolet light. The developed plate is irradiated with a low density
unltraviolet lamp, and then the fluorescence of the dye being quenched by
the compounds on the plate produces a light blue background of the
fluorescing dye.
References:

D. Pasto, C. Johnson, M. Miller. 1992. Experiments and Techniques in Organic Chemistry. Prentice-Hall
International Ed.
Organic Chemistry. 2014 Ed. Institute of Chemistry University of the Philippines