Identification of L-3-hydroxybutyrate as an original ketone body in rat serum by column-switching high-performance liquid chromatography and fluorescence derivatization

Yih Chiao Tsai, Tzu Hsin Liao, Jen A. Lee

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

L-3-Hydroxybutyrate (L-3HB), the enantiomer of D-3-hydroxybutyrate (D-3HB), has traditionally been regarded the "unnatural" ketone body in mammals, although there is suspicion that it is a more-favorable energy fuel for mammalian tissues than D-3HB. In this study, we demonstrated that L-3HB is an original substance in rat serum by applying fluorescence derivatization and a column-switching high-performance liquid chromatography system as the analysis technique. Racemic 3HB in rat serum derivatized using 4-nitro-7-piperazino-2,1,3-benzoxadiazole was first separated by an ODS column and was further confirmed by verifying the disappearance of the racemic 3HB peak after pretreating rat serum with D-3-hydroxybutyryl dehydrogenase. A switching valve was used to simultaneously introduce isolated racemic 3HB to the enantiomeric separation by two CHIRALCEL OD-RH columns connected in tandem. An L isomer was found to accompany the D isomer, which were quantified to be 3.98μM (3.61%) and 106.20μM (96.39%), respectively. Using the present analytical method, the dubious interpretation of the existence of L-3HB was clarified.

Original languageEnglish
Pages (from-to)34-41
Number of pages8
JournalAnalytical Biochemistry
Volume319
Issue number1
DOIs
Publication statusPublished - Aug 1 2003

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Ketone Bodies
3-Hydroxybutyric Acid
High performance liquid chromatography
Rats
Fluorescence
High Pressure Liquid Chromatography
Serum
Isomers
Mammals
Enantiomers
Oxidoreductases
Tissue

Keywords

  • Cellulose-based chiral column
  • Column-switching HPLC
  • Derivatization
  • Enantiomeric separation
  • L-3-Hydroxybutyrate
  • NBD-PZ

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

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title = "Identification of L-3-hydroxybutyrate as an original ketone body in rat serum by column-switching high-performance liquid chromatography and fluorescence derivatization",
abstract = "L-3-Hydroxybutyrate (L-3HB), the enantiomer of D-3-hydroxybutyrate (D-3HB), has traditionally been regarded the {"}unnatural{"} ketone body in mammals, although there is suspicion that it is a more-favorable energy fuel for mammalian tissues than D-3HB. In this study, we demonstrated that L-3HB is an original substance in rat serum by applying fluorescence derivatization and a column-switching high-performance liquid chromatography system as the analysis technique. Racemic 3HB in rat serum derivatized using 4-nitro-7-piperazino-2,1,3-benzoxadiazole was first separated by an ODS column and was further confirmed by verifying the disappearance of the racemic 3HB peak after pretreating rat serum with D-3-hydroxybutyryl dehydrogenase. A switching valve was used to simultaneously introduce isolated racemic 3HB to the enantiomeric separation by two CHIRALCEL OD-RH columns connected in tandem. An L isomer was found to accompany the D isomer, which were quantified to be 3.98μM (3.61{\%}) and 106.20μM (96.39{\%}), respectively. Using the present analytical method, the dubious interpretation of the existence of L-3HB was clarified.",
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AU - Lee, Jen A.

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