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  • Isolevuglandins covalently modify phosphatidylethanolamines in vivo: detection and quantitative analysis of hydroxylactam adducts.

Isolevuglandins covalently modify phosphatidylethanolamines in vivo: detection and quantitative analysis of hydroxylactam adducts.

Free radical biology & medicine (2009-09-16)
Wei Li, James M Laird, Liang Lu, Sanjoy Roychowdhury, Laura E Nagy, Rong Zhou, John W Crabb, Robert G Salomon
RESUMEN

Levuglandins (LGs) and isolevuglandins (isoLGs, also called "isoketals" or "isoKs") are extraordinarily reactive products of cyclooxygenase- and free radical-induced oxidation of arachidonates. We now report the detection in vivo and quantitative analysis of LG/isoLG adducts that incorporate the amino group of phosphatidylethanolamines (PEs) into LG/isoLG-hydroxylactams. Notably, LC-MS/MS detection of these hydroxylactams is achieved with samples that are an order of magnitude smaller and sample processing is much simpler and less time consuming than required for measuring protein-derived LG/isoLG-lysyl lactams. A key feature of our protocol is treatment of biological phospholipid extracts with phospholipase A(2) to generate mainly 1-palmitoyl-2-lysoPE-hydroxylactams from heterogeneous mixtures of phospholipids with a variety of acyl groups on the 2 position. Over 160% higher mean levels of LG/isoLG-PE-hydroxylactam (P<0.001) were detected in liver from chronic ethanol-fed mice (32.4+/-6.3 ng/g, n=6) compared to controls (12.1+/-1.5 ng/g, n=4), and mean levels in plasma from patients with age-related macular degeneration (5.2+/-0.4 ng/ml, n=15) were elevated approximately 53% (P<0.0001) compared to those of healthy volunteers (3.4+/-0.1 ng/ml, n=15). Just as LG/isoLG-protein adducts provide a dosimeter of oxidative injury, this study suggests that LG/isoLG-PE-hydroxylactams are potential biomarkers for assessing risk for oxidative stress-stimulated diseases.

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16:0 PE-d62, Avanti Research - A Croda Brand 860372P, powder