Separation of Racemic Mixtures of Amino Acids Using Chiral Eluents

A commercially available non-chiral RP-C8 chromatographic column is used with an aqueous chiral mobile phase containing N,N-dimethyl-L-phenylalanine and copper in a ratio 20 : 1 for the enantiomeric resolution of the following amino acids: DL-valine (Val), DL-methionine (Met), DL-leucine (Leu), DL-phenylalanine (Phe) and DL-tyrosine (Tyr). The influence of organic modifier, temperature and flow-rate on the capacity factors and on the selectivity is experimentally determined and the separation behavior is discussed. The capacity factors of some of the amino acids remained constant with changing the composition of the mobile phase, i.e. increasing methanol amount. With increase of the temperature the retention times decreased and the selectivities were slightly reduced. In a study of the effect on retention and selectivity, linear van’t Hoff plots were obtained giving H and S values of the solute transfer from the mobile to the stationary phase. These were found to be within the range obtained for hydrophobic compounds in RP-HPLC systems. Using van Deemter plots high separation efficiency and low apparent axial dispersion coefficients for all amino acids could be determined.