Interference of 3-hydroxyisobutyrate with measurements of ketone body concentration and isotopic enrichment by gas chromatography-mass spectrometry.
Concentrations and 13C2 molar percentage enrichments of blood R-3-hydroxybutyrate and acetoacetate are measured by selected ion monitoring gas chromatography-mass spectrometry. Samples are treated with NaB2H4 to reduce unlabeled and labeled acetoacetate to corresponding deuterium-labeled RS-3-hydroxybutyrate species. Only the gas chromatographic peak for the tert-butyldimethylsilyl derivative of 3-hydroxybutyrate needs to be monitored. The various compounds are quantitated using an internal standard of RS-3-hydroxy-[2,2,3,4,4,4-2H6]-butyrate. Concentrations of ketone bodies are obtained by monitoring the m/z 159 to 163 fragments of tert-butyldimethylsilyl derivatives of labeled and unlabeled 3-hydroxybutyrate species. High correlations were obtained between ketone body concentrations assayed (i) enzymatically with R-3-hydroxybutyrate dehydrogenase and (ii) by gas chromatography-mass spectrometry. The limit of detection is about 10 nmol of substrate in blood samples. The current practice of monitoring the m/z 275 to 281 fragments overestimates the concentration of endogenous R-3-hydroxybutyrate, due to co-elution of 3-hydroxyisobutyrate, a valine metabolite. The method presented is used to measure ketone body turnover in vivo in 24-h-fasted dogs.
Des Rosiers
C
C
Department of Nutrition, University of Montreal, Quebec, Canada.
Montgomery
J A
JA
Desrochers
S
S
Garneau
M
M
David
F
F
Mamer
O A
OA
Brunengraber
H
H
D020155:3-Hydroxybutyric Acid; D000090:Acetoacetates / Q000097:blood; D000818:Animals; D002247:Carbon Isotopes; D004285:Dogs; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006885:Hydroxybutyrates / Q000097:blood; D007657:Ketone Bodies / Q000097:blood*
D000090:Acetoacetates; D002247:Carbon Isotopes; D006885:Hydroxybutyrates; D007657:Ketone Bodies; C016635:acetoacetic acid; D020155:3-Hydroxybutyric Acid