Our next will involve a technique called paper chromatography.
Below are some images of the type of activity that we will be doing. (scroll down)
The first image is something like the results we hope to get.
Then there is a drawing of the different types of pigments found in chloroplasts.
Chromatography is a method of separation of substances based on their different mobility in a given stationary support. If you have a mixture of substances, chromatography can allow you to separate them. For example, inks are usually formed by a mixture of dyes, with the chromatography you can separate them and then analyze and identify them. Chromatography is also used to obtain large quantities of a pure substance. There are many methods of chromatography. Among them there is paper chromatography, the thin layer chromatography (TLC), gas chromatography, the liquid phase chromatography, the column chromatography. Usually the mixture of substances to be separated is inserted in a solvent. The type of solvent and of stationary medium can be chosen in a wide range of possibilities. Usually, this choice is made according to the molecules to be separated. According to the situation, one can exploit the molecules different size, chemical affinity toward the support, the amount of ionization, pH, solubility in water, etc. This experiment is for home or school so we use solvents and supports that are easy to find. Test tubes are often used to contain the solvent vapors, keeping the paper from drying and not overexposing the experimenter. Later on we will give you some experiments for paper and thin layer chromatography.
Also calculate the value of Rf of each band isolated (with reference to the pencil line, Rf = distance moved by the solute / distance moved by the solvent).
In these tests, the mixture to be examined has to be in high concentration. You can prepare the mixture according to the preceding experiment. Cut a paper filter stripe narrow enough so that it can be inserted in a test tube without touching the walls (wider at the bottom), as shown in the figure 20. With a pencil trace a horizontal line at 25 mm, (1 inch) from the bottom of the stripe. Place a little drop of mixture on this line and let it dry. Place another drop on the previous drop and let it dry. Repeat this operation to obtain a small, very concentrated spot. With a pin, fix the stripe to the bottom of the tube plug. Pour several cc of solvent in a test tube. There needs to be enough so the solvent is about half way between the bottom of the paper and the 25mm (1 inch) mark on the paper.
Insert the stripe in the tube. The smaller part of the stripe has to plunge into the solvent without touching the bottom of the tube. The pencil line and the spot have to be at about one cm above the surface of the solvent. For capillary action, the solvent will be absorbed by the fibers of the paper and, when it reaches the spot, it will start to carry the substances present in the mixture. According to their characteristics, these substances will travel faster or slower among the fibers of cellulose and the faster one will pull a head of the slower and show as separate bands on the paper. Remove the stripe before the solvent reaches the end.
With a pencil, immediately mark the position attained by the solvent and let the stripe dry