Identification of L-3-hydroxybutyrate as an original ketone body in rat serum by column-switching high-performance liquid chromatography and fluorescence derivatization.
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 3.98 microM (3.61%) and 106.20 microM (96.39%), respectively. Using the present analytical method, the dubious interpretation of the existence of L-3HB was clarified.
Tsai
Yih-Chiao
YC
Department of Pharmaceutical Analysis, School of Pharmacy, Taipei Medical University, No. 250, Wu-Hsing St., 110, Taipei, Taiwan, ROC.
Liao
Tzu Hsin
TH
Lee
Jen-Ai
JA
D020155:3-Hydroxybutyric Acid / Q000097:blood* / Q000737:chemistry; D000818:Animals; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D005453:Fluorescence; D007657:Ketone Bodies / Q000097:blood* / Q000737:chemistry; D008958:Models, Molecular; D051381:Rats; D017207:Rats, Sprague-Dawley; D012680:Sensitivity and Specificity; D013237:Stereoisomerism
D007657:Ketone Bodies; D020155:3-Hydroxybutyric Acid