Measurement of 12(S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid and its metabolite 12-oxo-5Z,8E,10E-heptadecatrienoic acid in human plasma by gas chromatography/negative ion chemical ionization mass spectrometry
- Dr. Margarete Fischer-Bosch-Institut fuer Klinische Pharmakologie, Stuttgart (Germany, F.R.)
Thromboxane A2, the predominant product of arachidonic acid metabolism in the blood platelet, is a potent vasoconstrictor and platelet agonist. During its biosynthesis from cyclic endoperoxide, 12(S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid (HHT) is formed in equal amounts. The further metabolism of HHT, catalyzed by 15-hydroxyprostaglandin dehydrogenase, leads to 12-oxo-5Z,8E,10E-heptadecatrienoic acid (Oxo-HT). Sample workup procedures are described which allow for the sensitive and reproducible determination of these two arachidonic acid metabolites in human plasma by gas chromatography-mass spectrometry in the presence of deuterated analogues as internal standards. HHT is derivatized to the pentafluorobenzyl ester tert-butyldimethylsilyl ether. In order to enable quantification of low concentrations of about 10 pg/ml in nonstimulated human plasma, the samples have to be purified by HPLC. Oxo-HT is derivatized to the pentafluorobenzyl ester, which is purified by HPLC, and then derivatized to the trimethylsilyloxime. The method allows quantification of Oxo-HT in concentrations down to 10 pg/ml plasma. The reported methods have been used to measure HHT and Oxo-HT in stimulated platelet rich plasma and to quantify HHT in nonstimulated plasma. Determination of endogenous levels of these two arachidonic acid metabolites may give new insights into the overall biosynthesis of thromboxane A2 in man.
- OSTI ID:
- 6056280
- Journal Information:
- Analytical Biochemistry; (USA), Vol. 189:2; ISSN 0003-2697
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ARACHIDONIC ACID
METABOLISM
METABOLITES
CHEMICAL ANALYSIS
PROSTAGLANDINS
BIOSYNTHESIS
BLOOD PLASMA
BLOOD PLATELETS
DEUTERIUM
GAS CHROMATOGRAPHY
LIQUID COLUMN CHROMATOGRAPHY
MAN
MASS SPECTROSCOPY
TRACER TECHNIQUES
ANIMALS
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CARBOXYLIC ACIDS
CHROMATOGRAPHY
HYDROGEN ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MAMMALS
MATERIALS
MONOCARBOXYLIC ACIDS
NUCLEI
ODD-ODD NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PRIMATES
SEPARATION PROCESSES
SPECTROSCOPY
STABLE ISOTOPES
SYNTHESIS
VERTEBRATES
550501* - Metabolism- Tracer Techniques