Lysine, Aspartate, Threonine, Leucine
a) A student performed chromatography of the four amino acids and the results were shown in the chromatogram. If an anion exchange column (column is positively charged) was used in a neutral buffer, assign each amino acid to the corresponding peak in the chromatogram.
b) Write 2 – 3 sentences to describe the interactions that will occur for each amino acid with the anion exchange column and its effect on column retention.
c) Write 2 – 3 sentences to describe how the results would differ if a cation exchange column (negatively charged column) were used.
a) To assign each amino acid to the corresponding peak in the chromatogram obtained from the anion exchange column, you would need to take into consideration the charge and hydrophobicity of each amino acid. Lysine has a positive charge at neutral pH, so it would not bind to the positively charged anion exchange column and thus appear as the first peak in the chromatogram. Aspartate has a negative charge and would bind strongly to the column, resulting in a late eluting peak. Threonine has a neutral charge and would have weak interactions with the column, resulting in an intermediate elution. Leucine is hydrophobic and lacks a charged side chain, so it would not bind to the column and elute early.
b) Lysine, being positively charged, would repel the positively charged column and have minimal interaction with it, resulting in a fast elution. Aspartate, being negatively charged, would strongly interact with the positively charged column due to electrostatic attractions, leading to relatively slow elution. Threonine, having no charge, would have weak interactions with the column and therefore have moderate retention. Leucine, having a hydrophobic side chain, would not form strong interactions with the column and would elute early.
c) If a cation exchange column (negatively charged column) were used instead, the interactions with the amino acids would be reversed. Lysine, being positively charged, would now bind strongly to the negatively charged column, resulting in a late elution. Aspartate, being negatively charged, would repel the negatively charged column and have minimal interaction, leading to fast elution. Threonine, having no charge, would have weak interactions and moderate retention. Leucine, being hydrophobic and lacking a charged side chain, would still have minimal interactions with the column and elute early. Therefore, the elution order and retention times of the amino acids would be different compared to the anion exchange column.