Kato Megumi, Kato Hisashi, Eyama Sakae, Takatsu Akiko
Bio-Medical Standards Section, Organic Analytical Chemistry Division, National Metrology Institute of Japan, National Institute of Advanced Industrial Science and Technology, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8563, Japan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2009 Oct 1;877(27):3059-64. doi: 10.1016/j.jchromb.2009.07.027. Epub 2009 Jul 24.
Amino acid analysis that is based on the use of hydrophilic interaction liquid chromatography (HILIC) coupled with isotope dilution mass spectrometry (IDMS) has been developed for the accurate quantification of underivatized amino acids from hydrolyzed protein/peptide. Sufficient separation of amino acids on a zwitterion chromatography (ZIC)-HILIC column was achieved after removal of chloride ions in the hydrolyzate. The detection limits and quantification limits as concentration of the four amino acids ranged from 0.003 to 0.04pmol microL(-1) and from 0.01 to 0.1pmol microL(-1), respectively. The analytical results for the certified reference materials, angiotensin I and bovine serum albumin (BSA), were satisfactory. Furthermore, the quantitative results by this method were compared with those by the commercially available precolumn method, derivatizd with aminoquinolylhydroxysuccinimidyl carbamate (AQC method), and better recovery and more precise data were obtained with this method.
基于亲水作用液相色谱（HILIC）与同位素稀释质谱（IDMS）联用的氨基酸分析方法已被开发出来，用于准确测定水解蛋白质/肽中的未衍生化氨基酸。在去除水解产物中的氯离子后，两性离子色谱（ZIC）-HILIC柱上实现了氨基酸的充分分离。四种氨基酸的检测限和定量限（以浓度计）分别为0.003至0.04pmol μL⁻¹和0.01至0.1pmol μL⁻¹。对认证参考物质血管紧张素I和牛血清白蛋白（BSA）的分析结果令人满意。此外，将该方法的定量结果与市售前柱衍生化方法（用氨基喹啉基羟基琥珀酰亚胺基氨基甲酸酯衍生化，即AQC方法）的结果进行了比较，该方法获得了更好的回收率和更精确的数据。
