| Autor: |
Ozaki M; Research and Development Department, Purification Section, Nacalai Tesque, Inc., Shimotsuma M; Research and Development Department, Purification Section, Nacalai Tesque, Inc., Kuranaga T; Department of System Chemotherapy and Molecular Sciences, Division of Medicinal Frontier Sciences, Graduate School of Pharmaceutical Sciences, Kyoto University., Kakeya H; Department of System Chemotherapy and Molecular Sciences, Division of Medicinal Frontier Sciences, Graduate School of Pharmaceutical Sciences, Kyoto University., Hirose T; Research and Development Department, Purification Section, Nacalai Tesque, Inc. |
| Abstrakt: |
D-Amino acids, which are present in small amounts in living organisms, are responsible for a variety of physiological functions. Some bioactive/biomolecular peptides also contain D-amino acids in their sequences; such peptides express different functions than peptides composed only of L-form amino acids. Among the 20 amino acids that make up proteins, threonine (Thr) and isoleucine (Ile) have two chiral carbons and thus have two enantiomers and diastereomers. These stereoisomers have been previously analyzed through HPLC using chiral columns or chiral resolution labeling reagents. However, the separation and identification of these stereoisomers are highly laborious and complicated. Herein, we propose an analytical method for the separation and identification of Ile stereoisomers through LC-MS using our original chiral resolution labeling reagent, 1-fluoro-2,4-dinitrophenyl-5-L-valine-N,N-dimethylethylenediamine-amide (L-FDVDA) and a PBr column packed with pentabromobenzyl-modified silica gel. Twenty DL-amino acids including Thr stereoisomers (41 amino acids including glycine) were separated and identified using C 18 column. Ile stereoisomers could be separated using not a C 18 column but a PBr column. Additionally, we showed that peptides containing Thr and Ile stereoisomers can be accurately detected through labeling with L-FDVDA. |
