Development of an off-line heart cutting two-dimensional HPLC system for enantioselective analysis of serine, threonine and allo-threonine in human physiological fluids

Mai Oyaide, Aogu Furusho, Chiharu Ishii, Takeyuki Akita, Masashi Mita, Tomomi Ide, Kenji Hamase

研究成果: ジャーナルへの寄稿学術誌査読

抄録

A highly-selective two-dimensional high-performance liquid chromatographic (2D-HPLC, off-line heart cutting mode) system was developed for the determination of serine (Ser), threonine (Thr) and allo-threonine (aThr) enantiomers in human physiological fluids. Ser, Thr and aThr have a hydroxy group in their side chains, and the development of a simultaneous analytical method with a practically sufficient enantio/chemo-selectivity has been required to clarify their amounts in human physiological fluids. The amino acids in the samples were derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole and were isolated by a reversed-phase column (Singularity RP18, 1.0 x 250 mm) in the first dimension. After the target amino acids were collected, the fractions were manually introduced into an enantioselective column in the second dimension and were detected by their fluorescence. For the second dimension, a Pirkle-type chiral stationary phase (Singularity CSP-013S, 1.5 x 250 mm) was used. The resolution values of the enantiomers obtained by the Singularity CSP-013S column were 7.64 for Ser, 7.58 for Thr and 4.71 for aThr by using the mixture of methanol and acetonitrile containing formic acid as the mobile phases. The developed method was validated and applied to human plasma and urine. In the plasma, the obtained %D values (the percentage of D-form to total amino acid) were 1.7 for Ser, and trace levels of D-aThr and D-Thr were observed. In the urine, the %D values were 48.0 for Ser, 1.6 for Thr and 8.0 for aThr (calculated using D-aThr and L-Thr).

本文言語英語
論文番号114807
ジャーナルJournal of Pharmaceutical and Biomedical Analysis
217
DOI
出版ステータス出版済み - 8月 5 2022

!!!All Science Journal Classification (ASJC) codes

  • 分析化学
  • 薬科学
  • 創薬
  • 分光学
  • 臨床生化学

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