Profiling of arachidonic acid metabolites in rabbit platelets by radio gas chromatography.
A method for profiling arachidonic acid metabolites by radio gas chromatography (GC) is described. The incubation mixture of rabbit platelets with [14C]arachidonic acid was purified on a Sep-Pak C18 cartridge and derivatized with diazomethane, O-methylhydroxylamine and dimethyl-isopropylsilylimidazole. The recovery of total 14C-radioactivity was 93.1 +/- 7.2%. Loss of radioactivity during derivatization was negligible. Baseline separations for [14C]arachidonic acid and its metabolites were obtained in a single run within 45 min by GC using a synchronized accumulating radioisotope detector (GC/SARD). The recovery of radioactivity from the GC column was virtually 100%. The chemical structures of the metabolites were confirmed by GC/mass spectrometry; peaks of arachidonic acid metabolites were assigned by comparison of the methylene unit values with those of radioactive peaks in GC/SARD analyses. The intra-assay coefficients of variation in GC/SARD analyses were less than 10%. The method was used to map the profile of arachidonic acid metabolites formed by rabbit platelets in the presence of indomethacin, baicalein or glutathione.
Akira
K
K
Tokyo College of Pharmacy, Japan.
Nakamura
T
T
Shinohara
Y
Y
Baba
S
S
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives / Q000032:analysis; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D001792:Blood Platelets / Q000737:chemistry* / Q000378:metabolism*; D002849:Chromatography, Gas / Q000379:methods*; D005231:Fatty Acids, Unsaturated / Q000032:analysis*; D006893:Hydroxyeicosatetraenoic Acids / Q000032:analysis; D015289:Leukotrienes / Q000032:analysis*; D011817:Rabbits
D005231:Fatty Acids, Unsaturated; D006893:Hydroxyeicosatetraenoic Acids; D015289:Leukotrienes; D016718:Arachidonic Acid; C008033:12-hydroxy-5,8,10-heptadecatrienoic acid; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; C022948:10-hydroxy-11,12-epoxyeicosa-5,8,14-trienoic acid; D015126:8,11,14-Eicosatrienoic Acid