Quantitative stereochemical analysis of subnanogram amounts of 12-hydroxy-(5,8,10,14)-eicosatetraenoic acid by sequential chiral phase liquid chromatography and stable isotope dilution mass spectrometry.
The two enantiomers of 12-hydroxy-(5,8,10,14)-eicosatetraenoic acid (12-HETE) are products of different biosynthetic pathways and have distinct biologic actions. Conventional methods of stereochemical analysis of 12-HETE require multimicrogram amounts of material and cannot be applied to systems where the availability of tissue is limited and only trace quantities of 12-HETE are generated. We have developed a method capable of measuring subnanogram amounts of 12-HETE enantiomers which involves addition of racemic. 18O2-labeled 12-HETE as an internal standard, chiral phase HPLC of the pentafluorobenzyl ester derivative of 12-HETE, and stable isotope dilution gas chromatographic-negative ion chemical ionization mass spectrometric quantitation of the resolved stereoisomers. This method has been employed to determine the stereochemical composition of 12-HETE produced by isolated pancreatic islets.
Turk
J
J
Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Stump
W T
WT
Wolf
B A
BA
Easom
R A
RA
McDaniel
M L
ML
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006893:Hydroxyeicosatetraenoic Acids / Q000032:analysis*; D007515:Islets of Langerhans; D008297:Male; D013058:Mass Spectrometry; D008832:Microchemistry; D010103:Oxygen Isotopes; D051381:Rats; D011919:Rats, Inbred Strains; D013237:Stereoisomerism
D006893:Hydroxyeicosatetraenoic Acids; D010103:Oxygen Isotopes; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid