Chiral lipidomics of monoepoxy and monohydroxy metabolites derived from long-chain polyunsaturated fatty acids

A chiral lipidomics approach was established for comprehensive profiling of regio- and stereoisomeric monoepoxy and monohydroxy metabolites of long-chain polyunsaturated fatty acids as generated enzymatically by cytochromes P450, lipoxygenases, and cyclooxygenases and, in part, also unspecific oxidations. The method relies on reversed-phase chiral liquid chromatography coupled with electrospray ionization/tandem mass spectrometry. Applications revealed partially opposing enantioselectivities of soluble and microsomal epoxide hydrolases. Ablation of the soluble epoxide hydrolase gene resulted in specific alterations in the enantiomeric composition of endogenous monoepoxy metabolites. For example, the (R,S)/(S,R)-ratio of circulating 14,15-EET changed from 2.1:1 in wild-type to 9.7:1 in the sEH knockout mice. Studies with liver microsomes suggested that cytochrome P450/microsomal epoxide hydrolase interactions play a primary role in determining the enantiomeric composition of monoepoxy metabolites during their generation and release from the endoplasmic reticulum. Analysis of human plasma showed significant enantiomeric excess with several monoepoxy metabolites. Monohydroxy metabolites were generally present as racemates; however, Ca2+-ionophore stimulation of whole blood samples resulted in enantioselective increases of lipoxygenase (12S-HETE and 17S-HDHA) and cyclooxygenase (11R-HETE) derived metabolites. Our chiral approach may provide novel opportunities for investigating the role of bioactive lipid mediators that generally exert their physiological functions in a highly regio- and stereospecific manner