Soma Yuki, Izumi Yoshihiro, Shimohira Takehiko, Takahashi Masatomo, Imado Yuri, Tominaga Saki, Tokito Kanako, Hata Kosuke, Shinadama Shoji, Oshiro Mana, Hayakawa Yoshihiro, Bamba Takeshi
Division of Metabolomics/Mass Spectrometry Center, Medical Research Center for High Depth Omics, Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
Laboratory for Synthetic Biology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, W5-729, 744 Motooka, Nishi-ku, Fukuoka 819-0395, Japan.
Metabolites. 2022 Aug 28;12(9):807. doi: 10.3390/metabo12090807.
Pre-column fluorescent derivatization has been used for the fast quantification of amino acids using high-performance liquid chromatography (HPLC) systems. However, it generally requires an offline in-vial derivatization process with multiple derivatization reagents. The offline derivatization requires the same number of reaction vials as the number of sample vials for use as a reaction chamber for the derivatization reaction in an autosampler. Therefore, the number of samples analyzed per batch using the pre-column derivatization method is halved. To benefit from the pre-column derivatization method, we transformed the derivatization process from an offline chamber process to an online in-needle process (in-needle Pre-column Derivatization for Amino acids Quantification; iPDAQ). Fluorescent derivatization in the injection needle obviated the need for vacant vials as reaction chambers. Consequently, the throughput per batch improved up to two times, and the consumption of derivatization reagents was reduced to less than one-tenth of that in the conventional vial method. We demonstrated to separate and quantify the amino acids in various biological samples. Herein, we presented a novel HPLC-based amino acid quantification method that enables the continuous analysis of a large number of samples. The iPDAQ facilitates accurate amino acid quantification due to the automation of derivatization and achieves improvement in the throughput and reduction of analysis labor.
柱前荧光衍生化已被用于使用高效液相色谱（HPLC）系统对氨基酸进行快速定量。然而，它通常需要使用多种衍生化试剂进行离线瓶内衍生化过程。离线衍生化需要与样品瓶数量相同的反应瓶，用作自动进样器中衍生化反应的反应室。因此，使用柱前衍生化方法每批分析的样品数量减半。为了受益于柱前衍生化方法，我们将衍生化过程从离线室过程转变为在线针内过程（用于氨基酸定量的针内柱前衍生化；iPDAQ）。进样针内的荧光衍生化无需使用空瓶作为反应室。因此，每批的通量提高了两倍，衍生化试剂的消耗量减少到传统瓶内方法的十分之一以下。我们证明了能够分离和定量各种生物样品中的氨基酸。在此，我们提出了一种基于HPLC的新型氨基酸定量方法，该方法能够连续分析大量样品。iPDAQ由于衍生化的自动化而有助于准确的氨基酸定量，并实现了通量的提高和分析工作量的减少。
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