Ion-Exchange Chromatography

Practice: What is the order of elution of the following proteins from a cation-exchange chromatography column? 

Net charges of Proteins: Protein A = +1              Protein B = -2                Protein C = -5                Protein D = +3.

Practice: In a cation-exchange column at neutral pH, which peptide would elute last?

Practice: Mixtures of amino acids can be analyzed by first separating the mixture into its components through ion exchange chromatography. Certain amino acids placed on a cation-exchange resin containing sulfonate groups (—SO₃^-) flow down the column slowly because of two factors that influence their movement: (1) ionic attraction between the sulfonate residues on the column and positively charged functional groups on the amino acids, and (2) hydrophobic interactions between amino acid R-groups and the strongly hydrophobic backbone of the polystyrene resin. For each pair of amino acids listed below, circle the amino acid that is eluted first from the cation-exchange column by a buffer at pH 7.

Practice: Give the order of elution of the following peptides when using cation-exchange chromatography at pH 7.2.

Peptide #1: A-D-G-H-E.           Peptide #2: K-L-M-R-A.           Peptide #3: M-D-L-I-V.             Peptide #4: I-L-R-P-M.

Order of Elution: _______________, _______________, _______________, _______________

                                 (1   ^st to elute)                                                                             (Last to elute)