Identification of 5,6-trans-epoxyeicosatrienoic acid in the phospholipids of red blood cells.
A novel eicosanoid, 5,6-trans-epoxy-8Z,11Z,14Z-eicosatrienoic acid (5,6-trans-EET), was identified in rat red blood cells. Characterization of 5,6-trans-EET in the sn-2 position of the phospholipids was accomplished by hydrolysis with phospholipase A(2) followed by gas chromatography/mass spectrometry as well as electrospray ionization-tandem mass spectrometry analyses. The electron ionization spectrum of 5,6-erythro-dihydroxyeicosatrienoic acid (5,6-erythro-DHET), converted from 5,6-trans-EET in the samples, matches that of the authentic standard. Hydrogenation of the extracted 5,6-erythro-DHET with platinum(IV) oxide/hydrogen resulted in an increase of the molecular mass by 6 daltons and the same retention time shift as an authentic standard in gas chromatography, suggesting the existence of three olefins as well as the 5,6-erythro-dihydroxyl structure in the metabolite. Match of retention times by chromatography indicated identity of the stereochemistry of the red blood cell 5,6-erythro-DHET vis à vis the synthetic standard. High pressure liquid chromatography-electrospray ionization-tandem mass spectrometry analysis of the phospholipase A(2)-hydrolyzed lipid extracts from red blood cells revealed match of the mass spectrum and retention time of the compound with the authentic 5,6-trans-EET standard, providing direct evidence of the existence of 5,6-trans-EET in red blood cells. The presence of other trans-EETs was also demonstrated. The ability of both 5,6-trans-EET and its product 5,6-erythro-DHET to relax preconstricted renal interlobar arteries was significantly greater than that of 5,6-cis-EET. In contrast, 5,6-cis-EET and 5,6-trans-EET were equipotent in their capacity to inhibit collagen-induced rat platelet aggregation, whereas 5,6-erythro-DHET was without effect. We propose that the red blood cells serve as a reservoir for epoxides which on release may act in a vasoregulatory capacity.
Jiang
Houli
H
Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.
McGiff
John C
JC
Quilley
John
J
Sacerdoti
David
D
Reddy
L Manmohan
LM
Falck
John R
JR
Zhang
Fan
F
Lerea
Kenneth M
KM
Wong
Patrick Y-K
PY
D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D001158:Arteries / Q000473:pathology; D001792:Blood Platelets / Q000378:metabolism; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D003094:Collagen / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D007668:Kidney / Q000098:blood supply; D008055:Lipids / Q000737:chemistry; D013058:Mass Spectrometry; D008956:Models, Chemical; D010741:Phospholipases A / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism*; D010974:Platelet Aggregation; D051381:Rats; D017207:Rats, Sprague-Dawley; D021241:Spectrometry, Mass, Electrospray Ionization; D013997:Time Factors
D008055:Lipids; D010743:Phospholipids; C040776:5,6-epoxy-8,11,14-eicosatrienoic acid; D003094:Collagen; D010741:Phospholipases A; D015126:8,11,14-Eicosatrienoic Acid