Evaporative light-scattering detection coupled to microcolumn liquid chromatography for the analysis of underivatized amino acids: Sensitivity, linearity of response and comparisons with UV absorbance detection

Abstract

Microcolumn chromatography (1.0 mm to 0.3 mm column i.d.) of underivatized amino acids coupled to evaporative light-scattering (ELS) detection demonstrated good baseline stability using perfluorinated carboxylic acid ion-pairing agents of varying alkyl chain lengths (C₂–C₈) and concentrations (1–8 mM). Under the gradient elution conditions used, the baseline stability and the limits of detection for amino acids were better with ELS detection than those obtained with UV absorbance detection. Microcolumn chromatography enabled good mass sensitivity to be achieved with the ELS detector with as little as 0.5 ng of amino acid being detectable on-column with a 0.3 mm i.d. column under gradient conditions. Excellent linearity was obtained for calibration standards of four different amino acids over three orders of magnitude (0.5–500 ng on-column; R²=0.98 or better). From the data presented it can be concluded that ELS detection can be advantageously linked with microcolumn chromatography to give better sensitivity and improved linearity compared with conventional chromatography. © 2001 John Wiley & Sons, Inc. J Micro Sep 13: 169–175, 2001