Enantiomeric separations of amino acids with inductively coupled plasma carbon emission detection

Abstract

A chiral crown ether column with a pH 1.9 perchloric acid buffered aqueous mobile phase is used to separate amino acid enantiomers by high performance liquid chromatography. An inductively coupled plasma atomic emission spectrometer coupled to the chromatographic system is used as a detector by monitoring the carbon atomic emission line at 193.09 nm. Seven underivatized amino acids are separated and detected resulting in an average mass detection limit of 5 ng (2.5 ng carbon). The chiral crown ether column resolves compounds with a primary amino group near the chiral center by forming a complex between the crown ether and an ammonium ion moiety from the sample. The d-form amino acid always elutes faster than its antipode. The carbon emission detector provides nearly identical sensitivities and similar detection limits for any compounds with comparable mass percents of carbon. Quantification is performed on unknown ratios of amino acids using an internal standard without the need for a calibration curve. Summing the calculated amounts of d and l amino acid and comparing to the known mixture quantity results in an average error of 1.0% for the seven amino acids separated.