6.1 Chromatography

Question 1

How do we use Chromatography to identify amino acids in a protein

Answer 1

1. Cut a chromatography paper so it fits in a beaker, handling the paper as little as possible, draw a line 25MM from the bottom with a pencil. Using a cocktail stick dot each amino acid under the line until its visible
2. Stand paper in beaker with the solvent
3. After the solvent has moved up to at least 75% of the way to the top remove the paper and draw around each spot of amino acid and sketch the line of solvent
4. Use a hairdryer to dry the paper
5. To identify the amino acids, Work out the Rf value: RF = distance moved by spot
   distance moved by solvent

Question 2

Why might not all of the amino acids in the mixture have appeared on your chromatogram

Answer 2

Some of them might be masked by other amino acids due to them having similar rf values

Question 3

Why should we handle the chromatogram as little as possible

Answer 3

To prevent contamination because our skin has amino acids on the surface which can cause incorrect results being identified