LABORATORY PRACTICE 3

PAPER CHROMATOGRAPHY
Introduction:
Chromatography is a technique that is used to separate and to identify components of a mixture. This analytical
technique has a wide range of applications in the real world since many substances are mixtures of chemical
compounds. In this lab, you will explore two applications of chromatography - identification of an unknown
ink sample and the separation of food colorings. In paper chromatography, the sample mixture is applied to a
piece of filter paper, the edge of the paper is immersed in a solvent, and the solvent moves up the paper by
capillary action. Components of the mixture are carried along with the solvent up the paper to varying degrees,
depending on the compound's preference to be adsorbed onto the paper versus being carried along with the
solvent. The paper is composed of cellulose to which polar water molecules are adsorbed, while the solvent is
less polar, usually consisting of a mixture of water and an organic liquid. The paper is called the stationary
phase while the solvent is referred to as the mobile phase. Performing a chromatographic experiment is
basically a three-step process: 1) application of the sample, 2) "developing" the chromatogram by allowing the
mobile phase to move up the paper, and 3) calculating Rf values and making conclusions. In order to obtain a
measure of the extent of movement of a component in a paper chromatography experiment, we can calculate
an "Rf value" for each separated component in the developed chromatogram.

Aims:
 Describe how chromatography works and what happens during this process to black or colored ink.
 Explain why being able to separate solutions into their components is important to environmental
engineering and water quality.

Materials and methods:

 Filter paper
 Red, blue, purple and black markers.
 Alcohol
 Recipes
 Masking tape
 Pencils
Directions:

1. Cut 4 strips of the porous paper that is about 4 cm wide and a little taller than the glass.
2. Take the strips of chromatography paper and mark them 1 cm from the top using the black marker
3. Draw a horizontal line in other end of the chromatography paper at 1cm distance.
4. Paint a spot with a black, red, blue or purple markers on the free end of the strip, about 2 cm from the edge.
Make sure it is intense, but doesn't take up much space.
5. Roll one end around a pencil using tape, the other end reaches the bottom of the glass. Make sure the strip
does not touch the sides of the container.
6. Pour alcohol in 2 containers and water in the other 2. Do it in a height of approximately 1 cm. THE INK
SPOT MUST BE ABOVE THE SOLVENT LEVEL
5. Wait 3 minutes and watch what happens. As the alcohol rises along the strip, it carries with it the various
pigments contained in the ink stain. As not all of them are dragged with the same speed, after a while you see
colored stripes.

Discussion:

1. What were the two solvents used?

2. Black ink is more attracted to which solvent? How do you know?

3. What colors are present in black ink?

4. What do these colors represent?

5. How might the ability to separating solutions into their components be important to environmental engineers
looking into water quality issues?

6. Calculate the Rf of each strip.

Conclusions: Write your conclusions.