PATHWAYS TO 4-HYDROXYNONENAL

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The long-term objective of this project is to elucidate the basis of aldehyde formation during lipid peroxidation, and thus advance our understanding of the etiology of atherogenesis and other diseases associated with oxidative stress. 4-Hydroxy-nonenal (HNE) and related aldehydes are among the most important reactive agents formed from polyunsaturated fatty acids during autoxidation yet the reactions underlying their synthesis are not at all well understood. We have now developed a detailed hypothesis implicating inter-molecular peroxyl radical reactions as a main pathway leading to HNE and related aldehydes. This is associated with cross-molecular epoxidations that can generate additional novel and bioactive products. The proposed mechanisms will be examined utilizing identification of intermediates by HPLC, UV, mass spectrometry, and NMR, and using stable isotope labeling to follow the reaction pathway and study the mechanisms of oxygen transfer. The Specific Aims are to investigate: 1. Inter-molecular mechanisms of 4-hydroperoxy-nonenal formation: peroxyl radical-dependent pathways 2. Aldehyde and epoxide formation during autoxidation of natural substrates 3. Biological implications of novel lipid peroxidation-derived products An important component of these studies in Aims 1 and 2 is the proposed isolation of individual species of cross-linked fatty acids and phospholipids, their precise structural and stereochemical characterization, and a detailed analysis of their degradative products, postulated to include the HNE-related aldehydes. The activity of novel epoxy-hydroperoxide products of lipid peroxidation will be assessed as PPAR ligands, and vitamin C-HNE adducts will be characterized and quantified (Aim 3). Elucidation of the biosynthesis of these reactive aldehydes has practical applications in defining the parameters that influence the production of aldehydes, identification of novel and reactive chemical species involved in the synthetic pathways, and improving the assessment of peroxide tone/oxidative stress in tissues.