The effect of buffer concentration and cation type in the mobile phase on retention of amino acids and dipeptides in hydrophilic interaction liquid chromatography

The current work is focused on exploring the effect of buffer cation type and its concentration on retention of amino acids, dipeptides and their blocked analogues on two stationary phases, i.e., bare silica and amide-based in hydrophilic interaction liquid chromatography. Five different buffers of pH 4.0 composed of Tris/acetic acid, triethylamine/acetic acid, ammonium/acetic acid, Li⁺/acetic acid and Ba²⁺/acetic acid were used in various concentrations. Interestingly, an increase of the buffer concentration caused increasing, decreasing or stable retention of analytes, according to the cation type in the buffer. The buffers containing barium cations provided the highest retention of all the analytes in comparable mobile phases, i.e., buffers with the same ionic strength and pH on both columns. Moreover, using buffers with barium cation different selectivity for dipeptides was observed. The chromatographic systems with buffers consisting of triethylamine behaved differently compared to others.