FMOC-C1为柱前衍生化试剂对氨基酸的RP-HPLC定量分析方法的研究

Investigation of Quantitative Analysis Method of Amino Acids in HPLC With FMOC-C1 as a Pre-column Derivatising Agent

采用二元流动相程序洗脱高效液相色谱分离了蛋白质水解液中常见的18种氨基酸。除FMOC-ARG的色谱峰拖尾较为严重,分离效果不佳外,其它各种FMOC-氨基酸衍生产物均获得了较好的分离效果。应用FMOC-Cl为柱前衍生化试剂,对氨基酸的高效液相色谱定量分析方法进行了研究。FMOC-Cl与氨基酸的衍生反应速度快、重现性好、衍生产物稳定,在介质PH=8.5～9.5范围内衍生反应完全,符合绝对分析法对衍生反应的要求。因此,FMOC-Cl是一种较为理想的应用绝对分析法对氨基酸定量分析的柱前衍生化试剂。

The separation of 18 amino acids in hydrolysis solution of proteins has been conducted by a binary gradient in HPLC. Satisfactory separating results have been achieved except the FMOC-ARG. The quantitative analysis method of amino acids is investigated in HPLC with FMOC-C1 as a per-col-umn derivatising agent. It has been found by experiments that the derivatising reaction was fast, the reproducibility was good, most products were stable and the reaction was complete from pH 8. 2 to pH 9. 5. Therefore, FMOC-C1 is a candidate per-column derivatising agent for the quantitative analysis of amino acids in HPLC by absolute analysis method.