Gogichaeva Natalia V, Alterman Michail A
Analytical Proteomics Laboratory, University of Kansas, Lawrence, KS, USA.
Office of Policy for Pharmaceutical Quality, Office of Pharmaceutical Quality, Center for Drug Evaluation and Research, US FDA, Silver Spring, MD, USA.
Methods Mol Biol. 2019;2030:17-31. doi: 10.1007/978-1-4939-9639-1_3.
Here we describe two different AAA protocols based on application of matrix-assisted laser desorption ionization time-of-flight (MALDI TOF) mass spectrometry (MS). First protocol describes a MALDI TOF MS-based method for a routine simultaneous qualitative and quantitative analysis of free amino acids and protein hydrolysates. Linear responses between the amino acid concentration and the peak intensity ratio of corresponding amino acid to internal standard were observed for all amino acids analyzed in the range of concentrations from 20 to 300 μM. Limit of quantitation varied from 0.03 μM for arginine to 3.7 μM for histidine and homocysteine. This method has one inherent limitation: the analysis of isomeric and isobaric amino acids. To solve this problem, a second protocol based on the use of MALDI TOF/TOF MS/MS for qualitative analysis of amino and organic acids was developed. This technique is capable of distinguishing isobaric and isomeric compounds. Both methods do not require amino acid derivatization or chromatographic separation, and the data acquisition time is decreased to several seconds for a single sample.
在此，我们描述了基于基质辅助激光解吸电离飞行时间（MALDI TOF）质谱（MS）应用的两种不同的氨基酸分析方法。第一种方法描述了一种基于MALDI TOF MS的方法，用于对游离氨基酸和蛋白质水解产物进行常规的同时定性和定量分析。在所分析的浓度范围为20至300μM的所有氨基酸中，观察到氨基酸浓度与相应氨基酸与内标物的峰强度比之间呈线性响应。定量限从精氨酸的0.03μM到组氨酸和同型半胱氨酸的3.7μM不等。该方法有一个固有局限性：对同分异构体和等压氨基酸的分析。为了解决这个问题, 开发了基于使用MALDI TOF/TOF MS/MS对氨基酸和有机酸进行定性分析的第二种方法。该技术能够区分等压和同分异构化合物。这两种方法都不需要氨基酸衍生化或色谱分离，并且单个样品的数据采集时间缩短至几秒钟。
