Determination of minute amounts of D-leucine in various brain regions of rat and mouse using column-switching high-performance liquid chromatography

Tomomi Inoue, Kenji Hamase, Akiko Morikawa, Kiyoshi Zaitsu

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Abstract

A highly sensitive method for the determination of minute amounts of D-Leu in biological samples was developed. For accurate and sensitive determination, a column-switching system using a micro ODS column and a chiral column was adopted. After pre-column derivatization of D- and L-Leu with NBD-F, the derivatives of the enantiomers were purified on a micro ODS column as a DL mixture. The eluted DL-Leu was then introduced to the chiral column, and each enantiomer was determined. The calibration curve for D-Leu, which was constructed by adding known amounts of D-Leu to a rat hippocampus, was linear from 1 to 1000 fmol (r>0.999), and the detection limit of added D-Leu was 1 fmol (S/N=5). Within-day and day-to-day precisions of D-Leu determination using the same homogenate of rat hippocampus were 5.11 and 5.25% (RSD), respectively. The content of D-Leu in rat hippocampus was 0.69 nmol/g wet tissue (the percentage of D-enantiomer for total Leu was 0.97%), which was consistent with the reported value. The distribution of D-Leu in mouse brain was also investigated, and the presence of D-Leu in various regions of the mammalian brain was first observed. Copyright (C) 2000 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)213-219
Number of pages7
JournalJournal of Chromatography B: Biomedical Sciences and Applications
Volume744
Issue number1
DOIs
Publication statusPublished - Jul 7 2000

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Enantiomers
High performance liquid chromatography
Leucine
Rats
Brain
Switching systems
Calibration
Tissue
Derivatives

All Science Journal Classification (ASJC) codes

  • Chemistry(all)

Cite this

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title = "Determination of minute amounts of D-leucine in various brain regions of rat and mouse using column-switching high-performance liquid chromatography",
abstract = "A highly sensitive method for the determination of minute amounts of D-Leu in biological samples was developed. For accurate and sensitive determination, a column-switching system using a micro ODS column and a chiral column was adopted. After pre-column derivatization of D- and L-Leu with NBD-F, the derivatives of the enantiomers were purified on a micro ODS column as a DL mixture. The eluted DL-Leu was then introduced to the chiral column, and each enantiomer was determined. The calibration curve for D-Leu, which was constructed by adding known amounts of D-Leu to a rat hippocampus, was linear from 1 to 1000 fmol (r>0.999), and the detection limit of added D-Leu was 1 fmol (S/N=5). Within-day and day-to-day precisions of D-Leu determination using the same homogenate of rat hippocampus were 5.11 and 5.25{\%} (RSD), respectively. The content of D-Leu in rat hippocampus was 0.69 nmol/g wet tissue (the percentage of D-enantiomer for total Leu was 0.97{\%}), which was consistent with the reported value. The distribution of D-Leu in mouse brain was also investigated, and the presence of D-Leu in various regions of the mammalian brain was first observed. Copyright (C) 2000 Elsevier Science B.V.",
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