Identification of 5-Keto-(7E,9E,11Z,14Z)-Eicosatetraenoic Acid as a Novel Nonenzymatic Rearrangement Product of Leukotriene A4

Abstract

Leukotriene A4 (LTA4), the reaction product of 5-lipoxygenase in human polymorphonuclear (PMN) leukocytes, is transformed both to LTB4 and a mixture of 5,6- and 5,12-dihydroxy-eicosatetraenoic acids (diHETE) via nonenzymatic hydrolysis. Evidence has been obtained that LTA4 is also converted to 5-keto-(7E,9E,11Z,14Z)-eicosatetraenoic acid (5-oxo-ETE). The compound was isolated from the products of the 5-lipoxygenase reaction and its structure elucidated by UV spectroscopy, LC-MS, two-dimensional [1H]NMR spectroscopy and chemical reduction to the corresponding alcohol. The 5-oxo-ETE represented about 14% of the LTA4 hydrolysis products as compared to 72 and 14% for the 5,12-diHETE and 5,6-diHETE, respectively. A similar profile of hydrolysis products was obtained after incubation of synthetic LTA4 in aqueous buffer. Human PMN leukocytes produced 5-oxo-ETE in an arachidonic acid-dependent and MK-886-inhibitable manner. The 5-oxo-ETE caused 50% inhibition of 5-lipoxygenase activity at 1 μM. These results demonstrate that the noneazymatic conversion of LTA4, in addition to the previously described hydrolysis products, yields 5-oxo-ETE during both the 5-lipoxygenase reaction and arachidonic acid oxidation by human PMN leukocytes. They indicate that allylic epoxides can rearrange in aqueous media at physiological pH to spontaneously form β,γ-unsaturated ketones.