D-amino acids have distinct roles from their l-enantiomer. In particular, some D-amino acids function as agonists or antagonists of neuronal receptors and are involved in higher brain functions. Thus, it is important to precisely measure the levels of these amino acid enantiomers in cells and tissues. Various quantification methods have been developed for measurements of chiral amino acids. However, each method has advantages and disadvantages. Additionally, measuring the amino acid enantiomers in crude biological samples requires a higher selectivity. In this study, we developed a quantification method for amino acid enantiomers using derivatization with N^α-(5-Fluoro-2,4-dinitrophenyl)-l-leucinamide (l-FDLA) followed by liquid chromatography–tandem mass spectrometry (LC/MS/MS) with a conventional reversed-phase column. We simultaneously identified 10 chiral amino acids. Furthermore, we applied this method to investigate murine tissue samples and examined the effect of aging on the amino acid levels in aged brain regions. We found that aging decreased the levels of both D-serine and D-aspartate in the hippocampus. In addition, D-Phenylalanine in the thalamus significantly increased with age. In conclusion, our method is suitable for the quantification of the D-amino acids in crude biological samples and may contribute to elucidating the biological roles of chiral amino acids.

中文翻译：

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通过LC / MS / MS与Nα-（5-氟-2,4-二硝基苯基）-1-亮氨酸酰胺（1-FDLA）衍生化相结合的LC / MS / MS同时测量老年小鼠大脑样品中的氨基酸对映体

D-氨基酸与它们的l-对映异构体具有不同的作用。特别地，一些D-氨基酸充当神经元受体的激动剂或拮抗剂，并参与更高的脑功能。因此，重要的是精确测量细胞和组织中这些氨基酸对映体的水平。已经开发了用于定量手性氨基酸的各种定量方法。但是，每种方法都有优点和缺点。另外，测量粗生物样品中的氨基酸对映体需要更高的选择性。在这项研究中，我们开发了使用衍生化的氨基酸对映异构体的量化方法Ñ ^α-（5-氟-2,4-二硝基苯基）-1-亮氨酸酰胺（1-FDLA），然后使用常规反相色谱柱进行液相色谱-串联质谱（LC / MS / MS）。我们同时鉴定了10个手性氨基酸。此外，我们将这种方法用于研究鼠类组织样品，并研究了衰老对老年大脑区域氨基酸水平的影响。我们发现衰老降低了海马体中D-丝氨酸和D-天冬氨酸的水平。此外，丘脑中的D-苯丙氨酸会随着年龄的增长而显着增加。总之，我们的方法适用于定量粗生物样品中的D-氨基酸，并且可能有助于阐明手性氨基酸的生物学作用。