Low-Capacity Cation-Exchange Chromatography of Amino Acids Using a Novel Sulfoacylated Macroreticular Polystyrene-Divinylbenzene Column with Binary Gradient Elution

Abstract

This paper describes a versatile technique for amino-acid separation using a novel low-capacity sulfoacylated macroreticular polystyrene-divinylbenzene cation-exchange column with a simple binary high-pressure pH gradient elution. Proteinic 16 amino acids were well separated within 50 min using a H₃PO₄/Na₂HPO₄-CH₃CN eluent system, and the cycle time was about 70 min. The chromatography with postcolumn OPA fluorescent detection was reproducible with RSDs less than 1% for retention times, and was quantitative with RSDs less than 5% for area responses. A linear regression line with an r² value above 0.9990 was obtained for each analyte in concentration from 0.1 to 10 µM by 20 µL injection. The method was applicable to the separation and detection of urinary diagnostic amino acid due to inborn errors of metabolism, such as phenylketonuria. The analytical costs would be decreased by using the proposed method.