Determination of free and glucuronide conjugated 20-hydroxyarachidonic acid (20-HETE) in urine by gas chromatography/negative ion chemical ionization mass spectrometry

20-Hydroxy-arachidonic acid (20-HETE) was determined in urine by an isotope dilution assay using gas chromatography/mass spectrometry (GC/MS). After addition of 18O2-internal standard, 20-HETE was extracted from urine with hexane either directly or after treatment with glucuronidase. 20-HETE was derivatized to the pentafluorobenzylester and the sample was applied to thin layer chromatography with iso-octane/iso-propanol 9:1 (v/v) as the developing solvent. The corresponding zone was extracted and 20-HETE was hydrogenated. After derivatization to the trimethylsilylether, 20-HETE was determined by GC/MS using the [M-pentafluorobenzyl]- -ion in the negative ion chemical ionization mode. Excretion rates of free and glucuronide conjugated 20-HETE was determined in healthy children and in children with hyperprostaglandin-E-syndrome/antenatal Bartter syndrome (HPS/aBS) with or without indomethacin treatment. Compared to the controls, the HPS/aBS children showed higher excretion rates of 20-HETE, which were suppressed to normal values under indomethacin medication. Free and glucuronide conjugated 20-HETE do not correlate with PGE2 excluding any participation in HPS/aBS.