Establishment and Application of an Automated Chiral Two-dimensional High Performance Liquid Chromatography for Bio-analysis of D-Acidic Amino Acids
10.3724/SP.J.1096.2014.40010
- VernacularTitle:体内 D-酸性氨基酸分析的自动二维手性高效液相色谱法的建立及应用
- Author:
Hai HAN
;
Qiqin WANG
;
Huihui WU
;
Huan WANG
- Publication Type:Journal Article
- Keywords:
D-Acidic amino acids;
Enantiomers;
Full-automated;
Two-dimensional high performance liquid chromatography;
Senescence accelerated mouse prone 1
- From:
Chinese Journal of Analytical Chemistry
2014;(6):891-896
- CountryChina
- Language:Chinese
-
Abstract:
The physiological and bio-marker function of D-acidic amino acids is now becoming the hot topic on metabolomics study and new drug discovery. A fully automated two-dimensional high performance liquid chromatography (2D-HPLC) system was established by using monolithic ODS column as the first dimension column, acetonitrile-trifluoro acetic acid-water (9: 0. 05: 92, V/ V) as the mobile phase; micro Chiralpak QD-1-AX column as the enantiomer separation column, 10 mmol/ L citric acid in methanol-acetonitrile (50: 50, V/ V) as the mobile phase for the second dimension, 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) as the fluorometrical derivative reagent. The separation efficiency ( Rs > 2. 5), determination sensitivity ( LOD =1 fmol) of acidic amino acids enantiomers were higher than those of existing methods, and an online confirmation of the enantiomers amounts was also achieved using this system. The recoveries were around 97-104% , RSD values for intra-day and inter-day precision were less than 5% for the acidic amino acids enantiomers in the biological samples. Furthermore, by analyzing the aging model senescence accelerated mouse prone 1 (SAMP1) mice which have low immunocompetence, the amounts of D-aspartic acid in thymus and spleen were determined as (206±18) and (264±21) nmol/ g, respectively. It is the first time that an obvious trend of the increasement of D-aspartic acid (p<0. 01) was observed in thymus and spleen of SAMP1 mice compare to senescence accelerated mouse resistant 1 (SAMR1) mice.
