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It is widely accepted that unesterified polyunsaturated ω‐6 and ω‐3 fatty acids (PUFA) are converted through various lipoxygenases, cyclooxygenases, and cytochrome P450 enzymes to a range of oxygenated derivatives (oxylipins), among which the polyhydroxides of unesterified PUFA have recently been recognized as cell signaling molecules with anti‐inflammatory and pro‐resolving properties, known as specialized pro‐resolving mediators (SPMs). This study investigates the mono‐, di‐, and trihydroxy 16:0/PUFA‐GPCs, and the corresponding 16:0/SPM‐GPC, in plasma lipoproteins. We describe the isolation and identification of mono‐, di‐, and trihydroxy AA, EPA, and DHA‐GPC in plasma LDL, HDL, HDL3, and acute phase HDL using normal phase LC/ESI‐MS, as previously reported. The lipoproteins contained variable amounts of the polyhydroxy‐PUFA‐GPC (0–10 nmol/mg protein), likely the product of lipid peroxidation and the action of various lipoxygenases and cytochrome P450 enzymes on both free fatty acids and the parent GPCs. Polyhydroxy‐PUFA‐GPC was hydrolyzed to variable extent (20%–80%) by the different secretory phospholipases A2 (sPLA2s), with Group IIA sPLA2 showing the lowest and Group X sPLA2 the highest activity. Surprisingly, the trihydroxy‐16:0/PUFA‐GPC of APHDL was largely absent, while large amounts of unidentified material had migrated in the free fatty acid elution area. The free fatty acid mass spectra were consistent with that anticipated for branched chain polyhydroxy fatty acids. There was general agreement between the masses determined by LC/ESI‐MS for the polyhydroxy PUFA‐GPC and the masses calculated for the GPC equivalents of resolvins, protectins, and maresins using the fatty acid structures reported in the literature. [ABSTRACT FROM AUTHOR]

Plasma lipoproteins are carriers of various glycerophospholipids including diacyl, alkenyl/acyl, and alkyl/acyl glycerophosphocholines (GPCs), which become distributed among cells and tissues during metabolism. For metabolic function, these phospholipids require hydrolysis by phospholipases, but the responsible enzymes have not been identified. We had previously shown that after complete digestion of lipoprotein diacyl‐ and oxo‐diacyl‐GPCs, degradation of residual alkyl/acyl and alkenyl/acyl GPCs continues, despite the fact that ether lipids are resistant to hydrolysis by Ca2+‐activated secretory PLA2s and require the presence of the Ca2+‐independent PLA2. In the course of further investigation, we came across a report by Khaselev and Murphy in which the autoxidative degradation of plasmalogens in the presence of 2,2′‐azobis(2‐amidinopropane) dihydrochloride (AAPH) proceeded beyond the formation of dihydroperoxides, hydroxides and epoxides, and led to an attack on the enyl bond of the plasmalogen, resulting in formation of 1‐OH/2‐20:4‐GPC and 1‐formyl/2‐20:4‐GPC. Our preliminary investigation indicated that lipoprotein 16:0p/20:4ω6‐GPC yielded the same autoxidation products as those reported for synthetic 16:0p/20:4ω6‐GPC in the presence of AAPH. Such autoxidative degradation of lipoprotein plasmalogens had not been previously reported with or without AAPH. Subsequent study led to the conclusion that this reaction was not limited to arachidonates, but extended to other polyunsaturated eicosanoids, docosanoids, and tetracosanoids, as well as oligounsaturated octadecanoids. These observations led to a hypothesis that the autoxidative cleavage of the lipoprotein plasmalogens proceeded under the influence of apo‐protein‐derived free radicals as intermediates of oxidative processes. [ABSTRACT FROM AUTHOR]

A study was conducted to analyze the inhibition of exogenous phospholipase A2 (PLA2) activity by new PLA2 inhibitors chosen from a group of compounds characterized by phosphatidylethanolamine (PE) covalently linked to polymeric carriers. Reaction conditions were selected to yield hydrolysis of 5-10% of the phospholipid substrates in the absence of inhibitors. Results indicated that the external polymers of cel-impermeable PLA2 inhibitors mimic cell surface proteoglycans.

This study was prompted by recent reports that epoxyeicosatrienoic (EET) and epoxyeicosatetraenoic (EEQ) acids accelerate tumor growth and metastasis by stimulation of angiogenesis, while eicosapentaenoic (EPA) and epoxydocosapentaenoic (EDP) acids inhibit angiogenesis, tumor growth, and metastasis. Cytochrome P450 epoxygenases convert arachidonic to EET, eicosapentaenoic acid to EEQ, and docosahexaenoic acid to EDP, which are found both in free form and esterified to glycerophosphocholine (GPC). Both free and esterified epoxy (EP) acids are also formed during lipid autoxidation. For biological activity, the GPC‐EP requires hydrolysis, which we presumed could occur by sPLA2s located in proximity of lipoproteins carrying the lipid epoxides. The plasma lipoproteins were isolated by ultracentrifugation and analyzed by LC/ESI‐MS. The GPC‐EPs were identified by reference to standards and to retention times of phospholipid masses. The GPC‐EP monoepoxides (corrected for isobaric ether overlaps) in stored human LDL, HDL, HDL3, or APHDL ranged from 0 to 1 nmol/mg protein, but during 4‐h incubation at 37°C increased to 1–5 nmol/mg protein. An incubation of autoxidized LDL, HDL, or HDL3 with 1 μg/ml of group V or X sPLA2 resulted in complete hydrolysis of diacyl GPC epoxide esters. Group IIA sPLA2 at 1 μg/ml failed to produce significant hydrolysis in 4 h, but at 2.5 μg/ml in 8 h yielded almost 80% hydrolysis, which represented complete diacyl GPC‐EP hydrolysis. The present study shows that group IIA, V, and X sPLA2s are capable of extensive hydrolysis of PtdCho epoxides of autoxidized plasma lipoproteins. Therefore, all three human sPLA2s were potentially capable of inducing epoxide biological activity in vivo. [ABSTRACT FROM AUTHOR]