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Chromatography is the most powerful method for separating substances. It was first
developed by Tswett some 60 years ago, to separate and study the coloured components
of flowers. He used filter paper and different mixtures of solvents to achieve this
separation. Nowadays, chromatography has turned into sophisticated methods: coils of
capillary tubes more than 300 m long, lined with special polymers (huge molecules) are
used instead of paper as a support, and gases or pressurised liquids flow along them. In
paper chromatography a sample is spotted on a standardised filter paper and this paper
is placed inside a tank dipping in a small amount of a solvent. The solvent climbs up the
paper because of capillary action and sweeps the sample up. After some time, if the
sample is a mixture, the components will be separated because of their being swept at
different rates. These “sweeping rates” are different because the affinity of different
substances in the mixture to both the paper
and the solvent vary between wide ranges.
The coloured components are easily seen, but
in case a colourless substance has to be
located, it can be visualised by different
methods (UV lamps, iodine vapours, specific
reagents). Chromatography can also be used to
identify substances just as the melting or the
boiling point. If an unknown sample and
different standards are spotted one beside the
other on a horizontal line and the solvent is
allowed to climb. The standards will have
climbed up different distances from the
starting line and the unknown will lie on the
same horizontal line as the standard it is equal
The figure shows a chromatography tank with
a chromatogram with three different mixtures.
One mixture is missing the red components, a
second one is missing the green component and the third one the blue. Notice that the
individual colours are the same for the different mixtures (not two different greens or

Aims: to investigate the colours in several markers’ ink by means of paper


Experiment Number 1: Circular chromatography


Petri dish, filter paper (circular), scissors.

1- Cut two lines on a circular filter paper to produce a strip.

2- Bend the strip down so that it will dip into the water
when placed on the Petri dish.

3- Spot one drop of black ink at the centre of the paper.

4- Add 5 ml of distilled water to the dish

5- Place the filter paper atop of it with the strip bended so that it dips in water.

6- Wait until you notice no further separation. Dry and keep the chromatogram

Experiment Number 2: The colours of felt-tip pens


Beaker, chromatography paper, glass rod, stapler or clips, pencil, ruler


1- Cut a strip of chromatographic paper 4 cm wide. Be sure it is not “wedged”.

Using a pencil (no ink or ball-pen should be used) draw a horizontal line ½ cm
from one the narrow edges of the strip.

2- At 1 cm from the edge and on the pencil line, make a dot with one of the
markers chosen. Make a second and a third dot with other markers all of them
separated by a distance of about 1 cm. Blacks, browns and violets are most
adequate for observing separations because they are usually mixtures.

3- Make a loop at the other end of the paper and staple it so that you can push a
glass rod or a pencil through it. (See diagram). Fix with clips or staples

4- Put a little water in a beaker and hang the paper as shown in the diagram. It
should dip in the water but take care that water does not reach the spots or the
experiment will be spoiled.

5- Let the solvent climb up until it reaches near the top of the paper. Then remove
and let it dry.

6- Repeat the experiment using as solvents:

a- 5 % aqueous ethanoic (acetic) acid

b- 5% ammonia solution
c- 5 % sodium chloride

7- Glue the chromatograms in your report.

8- You can eventually repeat the experiments with different colour markers