- Purpose
Paper chromatography is a technique which is used to distinguish between the various components of a mixture. It helps to separate out the components of a solution and identify individual components. In Paper chromatography, filter paper is used as the primary material for conducting the experiment.
- Procedure
For the experiment, a beaker of 1L capacity is taken and filled with 100mL solvent. The beaker is known as chromatogram tank. Next, a filter paper around 12 inches long and 9 inches wide is taken. A light line which is 2cm apart from the bottom of the paper is drawn on the filter paper. Further, three tiny dots of amino acid and one of the unknown solution is put on the filter paper at a distance of 2.5cm from each other and then left undisturbed for 5min. and allowed to dry. Then the filter paper is rolled in the form of a cylinder, stapled in put in the tank. After a lapse of around 2 hours the filter paper is taken out and the solvent lines are sprayed on by ninhydrin solution. After the paper gets dried, the solvent line is marked and the distance travelled by it from the base line is recorded. Also, the distance covered by the amino acids from the base line are measured and recorded. The Rf values in the case of each amino acid is then calculated. The information collected about the Rf values and the colors of the known amino acids helps to determine the unknown acid used.
- Data analysis
Data collected while conducting the experiment is represented with the help of a table.
Rf=Distance travelled by the amino acid from the base lineDistance travelled by the solvent from the base line
Distance travelled by the solvent from the base line is found to bear a value of 7.7 cm.
- Error analysis
Percentage Error =Value obtained-Standard valueStandard value×100
Percentage Error =2.95-2.92.9×100
Percentage Error =1.724%
- Interpretation
Figure 1: Variations in the distance travelled by the known and unknown acids and their respective Rf values.
It has been observed that the distance travelled by the acids depends upon the solubility of the amino acid in the solvent and the attractive forces between the amino acids and the filter paper. Furthermore, the polar amino acids that are soluble in the solvent will move with the solvent much faster.
- Conclusion
Paper chromatography is a simple yet effective technique that can help identify the unknown amino acid under consideration. The effectiveness of this experiment lies in the fact that the amino acids have higher affinity with the filter paper.
Solutions
- Every amino acid bears an amine and an acid functional group.
- The neutralization reaction that takes place within an amino acid in aqueous solution, gives rise to a salt-like ion which is known as zwitterion. An example of the zwitterions is shown below.
H H O
H N+ C C O-
H R
- Isoelectric point refers to the particular pH at which a molecule bears no electric charge.
- As it can be inferred from the results, phenylalanine that contains the bulkiest side chain travels the highest distance while glycine with just a hydrogen atom in its side chain shows minimum distance travelled. Therefore, it can be deduced side chain (“R”) of an amino acid directly affects the distance it travels.
- Since the ink of the pen can conflict with the chemical, therefore instead of a pen pencils are used to mark the spots.
