linoleic-acid and 4-oxo-2-nonenal

Angiotensin (Ang) II is a major bioactive peptide of the renin/angiotensin system and is involved in various cardiovascular functions and diseases. Ang II type 1 (AT

Topics: Aldehydes; Angiotensin II; Ascorbic Acid; Carbon Isotopes; Cell Line; Copper Sulfate; Endothelial Cells; Humans; Isotope Labeling; Linoleic Acid; Lipid Peroxidation; Oxidative Stress; Receptor, Angiotensin, Type 1

Oxidative stress-induced lipid peroxidation leads to the formation of cytotoxic and genotoxic 2-alkenals, such as 4-hydroxy-2-nonenal (HNE) and 4-oxo-2-nonenal (ONE). Lipid-derived reactive aldehydes are subject to phase-2 metabolism and are predominantly found as mercapturic acid (MA) conjugates in urine. This study shows evidence for the in vivo formation of ONE and its phase-1 metabolites, 4-oxo-2-nonen-1-ol (ONO) and 4-oxo-2-nonenoic acid (ONA). We have detected the MA conjugates of HNE, 1,4-dihydroxy-2-nonene (DHN), 4-hydroxy-2-nonenoic acid (HNA), the lactone of HNA, ONE, ONO, and ONA in rat urine by liquid chromatography-tandem mass spectrometry comparison with synthetic standards prepared in our laboratory. CCl(4) treatment of rats, a widely accepted animal model of acute oxidative stress, resulted in a significant increase in the urinary levels of DHN-MA, HNA-MA lactone, ONE-MA, and ONA-MA. Our data suggest that conjugates of HNE and ONE metabolites have value as markers of in vivo oxidative stress and lipid peroxidation.

Topics: Acetylcysteine; Aldehydes; Animals; Biomarkers; Carbon Tetrachloride; Chromatography, Liquid; Lactones; Linoleic Acid; Lipid Peroxidation; Mass Spectrometry; Models, Chemical; Oxidative Stress; Rats; Rats, Inbred F344

Substantial work has been carried out to elucidate the nature of protein modification by 4-hydroxy-2-nonenal (HNE) and its relatives. Its keto cousin, 4-oxo-2-nonenal (ONE), which arises from linoleic acid oxidation independently of HNE, was previously reported to form Michael adducts with His and Cys that can subsequently, in part, condense with Lys residues to give imidazolylpyrrole cross-links. Despite mass spectrometric evidence also for ONE-Lys Michael adducts, the latter do not accumulate in solution. A long-lived adduct that has the same mass as the ONE Lys Michael adduct is suggested instead to be the isomeric 4-ketoamide that arises, along with other adducts, from the reversibly-formed ONE Lys Schiff base. The Lys-ketoamide and His-Lys imidazolylpyrrole cross-links appear to be unusually prominent markers of stable protein modification by ONE.

Topics: Aldehydes; Kinetics; Linoleic Acid; Mass Spectrometry; Models, Molecular; Molecular Conformation; Proteins; Schiff Bases

There has been significant recent interest in the protein reactivity and biological activity of 4-oxo-2-nonenal (ONE), the 4-keto cousin of HNE. Despite the ability of HNE to form at least metastable Michael adducts with protein Lys residues, mass spectrometric evidence for the existence of apparent ONE-Lys Michael adducts is not supported by preparative solution studies. We herein show that the m+154 adducts on Lys residues that persist upon incubating with ONE represent the isomeric 4-ketoamides. The same type of 4-ketoamide represents the apparent Michael adduct on Lys residues formed by the carboxy-terminating ONE-like 4-oxo-2-enal arising along with ONE from the oxidation of linoleic acid.

Topics: Aldehydes; Amides; Animals; Borohydrides; Cattle; Lactoglobulins; Linoleic Acid; Lysine; Molecular Structure; Stereoisomerism; Time Factors

Lipid peroxidation generates a variety of reactive products that covalently modify DNA, yielding several types of adducts with nucleobases. In the present study, we characterized the modification of nucleobases during peroxidation of linoleate and found that 2'-deoxycytidine (dC) could be a major target of the modification by lipid peroxidation reactions. Upon incubation with oxidized linoleate, dC and 2'-deoxyguanosine (dG) were significantly modified among four 2'-deoxynucleosides. The major product in dG/linoleate was identical to the 2-oxo-heptyl-substituted 1,N(2)-etheno-dG that had been previously identified as a 4-oxo-2-nonenal (ONE)-dG adduct. On the basis of spectroscopic and chemical characterization, we identified the major product in dC/linoleate as the 2-oxo-heptyl-substituted 3,N(4)-etheno-dC. The same adduct was also produced upon reaction of dC with ONE, suggesting that ONE might represent the major reactive species that modifies DNA during lipid peroxidation. Indeed, this proposition was supported by the observation that ONE was far more reactive with dC and dG than other genotoxic aldehydes, such as 4-hydroxy-2-nonenal. More strikingly, we found that, in contrast to the similar reactivity of ONE toward free nucleobases (dC and dG), ONE preferentially reacted with dC residues in double-stranded DNA. These results suggest that ONE and other 4-oxo-2-alkenals may possess by far the strongest electrophilic potential vs. dC and that the formation of 4-oxo-2-alkenal-adducted dC may thus serve as one mechanism for oxidative damage to DNA in vivo.

Topics: Aldehydes; Chromatography, High Pressure Liquid; Deoxycytidine; Deoxyguanosine; DNA; Free Radicals; Linoleic Acid; Lipid Peroxidation; Mass Spectrometry

trans-4-Oxo-2-nonenal (ONE) has recently been demonstrated to be a direct product of lipid peroxidation. In earlier studies to elucidate the structure of the trans-4-hydroxy-2-nonenal (HNE)-derived fluorescent Lys-Lys cross-link, we showed that ONE was capable of both oxidative and nonoxidative cross-linking of amines. A more comprehensive study on nonoxidative modification of protein nucleophiles by ONE is described here, focusing on the initial Michael addition of imidazole, thiol, and amine groups to C2 or C3 to give 4-keto aldehydes that can then condense with amines to form nucleophile-substituted pyrroles. 2,3-Substituted pyrroles (major) and 2,4-substituted pyrroles (minor) were distinguished by 2D NMR techniques, and N(tau)-substitution is preferred over N(pi)-substitution in the Michael addition of histidine. Mechanisms of both nonoxidative and oxidative side chain reactions of ONE are discussed, as is the relative propensity (ONE > HNE) to induce cross-linking of the model proteins ribonuclease A and beta-lactoglobulin.

Topics: Acetylcysteine; Aldehydes; Chromatography, High Pressure Liquid; Cross-Linking Reagents; Electrophoresis, Polyacrylamide Gel; Histidine; Imidazoles; Linoleic Acid; Lysine; Magnetic Resonance Spectroscopy; Oxygen; Proteins; Pyrroles; Ribonucleases; Stereoisomerism; Structure-Activity Relationship