Principles of Biochemistry (2+1)

Identification and Calculation

• Spray the chromatogram with the ninhydrin reagent using an automiser.
• Dry the paper for about 5 min at room temp followed by at 100ºC in an oven for 2-3min.
• Amino acids appear as purple sports; hydroxyproline and proline give yellow coloured sports.
• Mark all the spots and calculate their R_f values by the formula

          Distance (cm) moved by the solute from the origin

 R_f = -----------------------------------------------------------------

          Distance (cm) moved by the solvent from the origin

Identify the amino acids present in the sample by comparing

the Rf values with that of the standard amino acids co-

chromatographed.