Effect of ionic liquid on the retention and separation behavior of various amino acids in reversed phase chromatography

The chromatographic retention behaviors of various amino acids were investigated using ionic liquid, 1-butyl-3-methylimidazolium tetrafluoroborate ([bmim][BF4]), as a mobile phase additive and a reversed phase C-18 media as a stationary phase. The amino acids investigated include L-tyrosine, N-CBZ-L-tyrosine, L-phenylalanine, N-CBZ-L-phenylalanine, L-methionine, and N-CBZ-L-methionine. The effect of ionic liquid on N-CBZ-L-amino acids was found to be different from that on L-amino acids. As the concentration of ionic liquid in mobile phase increased, the retention factors of L-amino acids showed a decreasing trend, whereas those of N-CBZ-L-amino acids exhibited a minute change. The selectivity between L-amino acid and N-CBZ-L-amino acid was, thus, improved by increasing the content of ionic liquid. In addition, the peak shapes of L-amino acids, if asymmetric, were transformed into symmetric ones by increasing the ionic liquid content. The increase of feed concentration causes the overloaded condition, leading to a decrease in the retention factor. If such effect of feed concentration is coupled with the effect of using ionic liquid, one can achieve a better separation between L-amino acid and N-CBZ-L-amino acid.