The chamber should be saturated with the developing solvent to achieve high resolution.
Discussion: When the leaves of green plants are extracted, a complex mixture of components is obtained. The components that are obtained include anthocyanin, chlorophyll a and b, carotenes, and xanthophylls. If you try to extract these components from the green leaves by using water, the extraction is rather ineffective. This is because water is a polar compound and these components are nonpolar compounds. Therefore, water can’t effective dissolve them out of the leaves (remember: like dissolves like). A less polar solvent will be used for the extraction: acetone.
Likewise, if you attempt to run a paper chromatography of these components by using water as the eluent (the liquid that will carry the components up the paper) you will find that the water will do an adequate job of separating the components from each other. Instead, lighter fluid, which is nonpolar, more volatile, and faster moving, will separate them.
The experiment is carried out inside a closed container to eliminate evaporation of the eluting solution from the paper. (Evaporation would affect the Rf). This process of separation of the components is referred to as eluting (or developing) the chromatogram. The experiment is allowed to proceed uninterrupted until the solvent begins to approach the top of the paper. It must not be allowed to reach the top of the paper, otherwise separated components may again catch up with each other and recombine at the top. To stop the chromatogram, the paper is simply removed from its container and dried to remove the mobile phase.
In paper chromatography, when you place a colored chemical sample on a filter paper, you can get the colors to separate from the sample by placing one end of the paper in a solvent. As the solvent diffuses up the paper, it dissolves the various molecules in the sample according to the polarities of the molecules and the solvent. If the sample contains more than one color, that means it must have more than one kind of molecule. Because of the different chemical structures of each kind of molecule, the chances are very high that each molecule will have at least a slightly different polarity, and thus, a different solubility in the solvent.
The unequal solubilities cause the various color molecules to leave solution at different places as the solvent continues to move up the paper. The more soluble a molecule is, the higher it will migrate up the paper. If a chemical is very polar it will not dissolve at all in a very nonpolar solvent. The same is true for a very nonpolar chemical and a very polar solvent.
In this lab you will use paper chromatography to separate colorant molecules in pens and plants. In the process, you will be able to identify differences in polarity of pigment (color) molecules of different kinds of markers. You will use vis a vis and permanent markers as well as grass and leaves. The solvents you will use are (in order from most to least polar) water, ethyl alcohol, and hexane.
As the liquid moves up the paper, the molecules of the sample mixture will also move. Polar molecules within the sample will spend most of their time bound to the polar surface of the paper and will therefore not move very much. Nonpolar molecules, however, will spend most of their time dissolved in the liquid as it moves up the paper. When the liquid reaches the top of the paper, these molecules will also have traveled most of the way to the top. The two types of molecules (polar and nonpolar) are now separated.
The choice of the eluent or solvent is the most difficult task. Choosing the right polarity is critical because this determines the level of separation that will be achieved ( this degree of separation helps one to identify the constitute molecules). Common solvents used in chromatography, in order of increasing polarity, are: petroleum, ether or hexanes, cyclohexanes, toluene, chloroform, ethyl ether, acetone, ethanol, methanol, and water.