A rapid method for the determination of free alpha-amino acids in pea (Pisum sativum) using ultra high performance liquid chromatography-high resolution accurate mass-mass spectrometry

Abstract

The analysis of alpha-amino acids in biological matrices usually requires time-consuming ion exchange liquid chromatography with post column derivatisation. Using ultra high performance liquid chromatography coupled to high resolution accurate mass-mass spectrometry (UHPLC-HRAM-MS) we describe a rapid, accurate and sensitive method for the analysis of the 21 common free amino acids in pea seeds (Pisum sativum). Total analysis time including extraction, separation by LC and detection was less than an hour per sample. Extraction efficiency and repeatability were assessed using surrogate standards and were found to be acceptable with recoveries of the standards between 84% and 109% and relative standard deviations of less than 8% between replicates. Mass accuracy of the ions detected (between 2 and 4 ppm), linearity of standards and sensitivity were excellent with limits of detection (LOD) less than 30 ng g⁻¹ and r² values greater than 0.96 for all amino acids tested.