High-performance Liquid Chromatography of Amino Acids -Electrochemical Detection with a Copper Electrode-

Abstract

A channel thin-layer cell with a copper working electrode has been used as a directly amperometric detector for amino acids in high-performance liquid chromatography. All essential amino acids except cysteine gave the increase of the anodic dissolution current at the copper electrode by the reaction of copper(II) ions with amino acids at -20 m V vs. Ag/AgCl. The amperometric response of amino acids is dependent on their properti es and correlated to the acid dissociation constant of amino group of amino acids. This evidence indicates that the cell reaction is controlled by the rate of the complex formation of copper(II) ion with amino acid. A favorable S/N ratio was obtained with the cell geometry of the channel height and width of O.1 mm and O.5 mm, respectively, at the flow rate of 0.5 ml/min using 0.1 mol·dm^-3 phosphate buffer (pH 7.0)/methanol 9: 1 as the mobile phase. Although all essential amino acids were not well separated with a reverse phase column, the wide linear dynamic range more than three orders was obtained. The detection limit (S/N=2) was 20 pmol for histidine, 2 nmol for proline, and subnanomole order for other amino acids. The proposed electrochemical method is straightforward and simple compared with other detection methods.