Trace analysis of β-hydroxybutyrate in human plasma by derivatization and high-performance liquid chromatography

Abstract

KeyWordsColumn liquid chromatography DerTvatizatTon fi-Hydroxybutyrate in plasma SummaryA sTmple and sensitTve liquid chromatographTc method is described for determTnation of fi-hydroxybutyrate, as a highly sensitive derivative, from human plasma. Plasma spiked wth/F hy droxybutyrate was extracted with acetonitrile and derivatized with 4 bromomethyl / methox ycoumarin, in a homogeneous system, using ]_5 crown 5 as activator. The resulting derivative was separated on a LiChroCART Rp C18 column wth 40% methanol in water as mobile phase and 2,2' dinitrobiphenyl as internal standard. Conditions affecting the extraction and deriva tization of/F hydrox,/butyrate from spiked plasma were investigated. For/F hydroxybutyrate de termTnatTon the detection limit (S/N-3; sample sTze, 10 M,L) was approxTmately 5 nmol mL 1; the relatTve standard deviatTon was <9% for Tntra-day assay (n-6) and < 10% for Tnter-day assay (n -6).All recoveries were > 98%.