Salazar Carolina, Armenta Jenny M, Cortés Diego F, Shulaev Vladimir
Waters Corporation, Golden, CO, USA.
Waters Corporation, Beverly, MA, USA.
Methods Mol Biol. 2019;2030:191-206. doi: 10.1007/978-1-4939-9639-1_15.
Amino acid analysis is a powerful tool in life sciences. Current analytical methods used for the detection and quantitation of low abundance amino acids in complex samples face intrinsic challenges such as insufficient sensitivity, selectivity, and throughput. This chapter describes a protocol that makes use of AccQ•Tag chemical derivatization combined with the exceptional chromatographic resolution of ultra-performance liquid chromatography (UPLC) and the sensitivity and selectivity of tandem mass spectrometry (MS/MS). The method has been fully implemented and validated using different tandem quadrupole detectors and thoroughly tested for a variety of samples such as P. falciparum, human red blood cells, and Arabidopsis thaliana extracts. Compared to currently available methods for amino acid analysis, the AccQ•Tag UPLC-MS/MS method presented here provides enhanced sensitivity and reproducibility and offers excellent performance within a short analysis time and a broad dynamic range of analyte concentration. The focus of this chapter is the application of this improved protocol for the compositional amino acid analysis in Arabidopsis thaliana leaf extracts using the Xevo TQ for mass spectrometric detection.
氨基酸分析是生命科学中的一种强大工具。目前用于检测和定量复杂样品中低丰度氨基酸的分析方法面临着诸如灵敏度不足、选择性差和通量低等内在挑战。本章介绍了一种方案，该方案利用AccQ•Tag化学衍生化技术，结合超高效液相色谱（UPLC）出色的色谱分辨率以及串联质谱（MS/MS）的灵敏度和选择性。该方法已使用不同的串联四极杆检测器全面实施并验证，并针对多种样品进行了充分测试，如恶性疟原虫、人类红细胞和拟南芥提取物。与目前可用的氨基酸分析方法相比，本文介绍的AccQ•Tag UPLC-MS/MS方法具有更高的灵敏度和重现性，并且在短分析时间和宽分析物浓度动态范围内表现出色。本章的重点是使用Xevo TQ进行质谱检测，将这种改进的方案应用于拟南芥叶提取物中的组成氨基酸分析。
