Nitrated fatty acids: synthesis and measurement

Woodcock, Steven R.; Bonacci, Gustavo; Gelhaus, Stacy L.; Schöpfer, Francisco J.

doi:10.1016/j.freeradbiomed.2012.11.015

Cited by 55 publications

(43 citation statements)

References 32 publications

(35 reference statements)

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“…Nitroalkene Fatty Acid and Adduct Solutions-10-Nitrooctadec-9-enoic acid (nitrooleic acid, NO 2 -OA), 9-and 12-nitrooctadeca-9,11-dienoic acids (9-and 12-NO 2 -CLA), or purified isomer solutions were synthesized as described (51,52). For kinetic experiments, a 1:1 mixture of 9-and 12-NO 2 -CLA or purified isomer solutions were prepared (2 mM in methanol) and kept at Ϫ80°C until use.…”

Section: Methodsmentioning

confidence: 99%

The Chemical Basis of Thiol Addition to Nitro-conjugated Linoleic Acid, a Protective Cell-signaling Lipid

Turell

Vitturi

Coitiño

et al. 2017

Journal of Biological Chemistry

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Edited by F. Peter GuengerichNitroalkene fatty acids are formed in vivo and exert protective and anti-inflammatory effects via reversible Michael addition to thiol-containing proteins in key signaling pathways. Nitro-conjugated linoleic acid (NO 2 -CLA) is preferentially formed, constitutes the most abundant nitrated fatty acid in humans, and contains two carbons that could potentially react with thiols, modulating signaling actions and levels. In this work, we examined the reactions of NO 2 -CLA with low molecular weight thiols (glutathione, cysteine, homocysteine, cysteinylglycine, and ␤-mercaptoethanol) and human serum albumin. Reactions followed reversible biphasic kinetics, consistent with the presence of two electrophilic centers in NO 2 -CLA located on the ␤-and ␦-carbons with respect to the nitro group. The differential reactivity was confirmed by computational modeling of the electronic structure. The rates (k on and k off ) and equilibrium constants for both reactions were determined for different thiols. LC-UV-Visible and LC-MS analyses showed that the fast reaction corresponds to ␤-adduct formation (the kinetic product), while the slow reaction corresponds to the formation of the ␦-adduct (the thermodynamic product). The pH dependence of the rate constants, the correlation between intrinsic reactivity and thiol pK a , and the absence of deuterium solvent kinetic isotope effects suggested stepwise mechanisms with thiolate attack on NO 2 -CLA as rate-controlling step. Computational modeling supported the mechanism and revealed additional features of the transition states, anionic intermediates, and final neutral products. Importantly, the detection of cysteine-␦-adducts in human urine provided evidence for the biological relevance of this reaction. Finally, human serum albumin was found to bind NO 2 -CLA both non-covalently and to form covalent adducts at Cys-34, suggesting potential modes for systemic distribution. These results provide new insights into the chemical basis of NO 2 -CLA signaling actions.

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Section: Methodsmentioning

confidence: 99%

The Chemical Basis of Thiol Addition to Nitro-conjugated Linoleic Acid, a Protective Cell-signaling Lipid

Turell

Vitturi

Coitiño

et al. 2017

Journal of Biological Chemistry

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“…Oleic, linoleic, and conjugated linoleic acids were obtained from Nu-Check Prep, Inc. (Elysian, MN), 15-oxo-PGE 2 and 13,14-dihydro-15-oxo-PGE 2 were purchased from Cayman Chemical (Ann Arbor, MI), and nitroalkenes were synthesized as described previously (4,34). Nitroalkene concentration was measured daily in all synthetic stocks by using the appropriate extinction coefficients: NO 2 ).…”

Section: Methodsmentioning

confidence: 99%

Modulation of Nitro-fatty Acid Signaling

Vitturi¹,

Chen²,

Woodcock³

et al. 2013

Journal of Biological Chemistry

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Background: Nitroalkenes are electrophilic anti-inflammatory mediators that signal via Michael addition and are metabolized in vivo. Results: Prostaglandin reductase-1 is identified as a nitroalkene reductase. Conclusion: Prostaglandin reductase-1 reduces fatty acid nitroalkenes to nitroalkanes, inactivating electrophilic reactivity. Significance: A mammalian enzyme is identified that metabolizes fatty acid nitroalkenes in vivo to silence their signaling reactions.

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“…These radiolabeling experiments were conducted at Huntingdon Life Sciences (Cambridgeshire, and extra virgin olive oil (15,16). Notably, plasma and urinary NO 2 -FA concentrations in humans are increased following oral supplementation with NO 2  , nitrate (NO 3  ), and conjugated linoleic acid (CLA) (17,18 18 ]OA) were synthesized by direct alkene nitroselenation of the corresponding native FAs, as previously described (21). Nitro-CLA (NO 2 -CLA) was synthesized according to a previously described method for conjugated diene nitration (11 4 (obtained samples before and after hydrolysis, respectively.…”

Section: Quantitative Whole-body Autoradiography Analysismentioning

confidence: 99%

“…by nucleophilic substitution of [ 15 N]labeled sodium nitrate and 1-bromononane-6,6,7,7-d 4 ) and a nine-carbon aldehyde-ester via a previously described nitroaldol procedure (21,22 (21). DMEM, FBS, HBSS, and antibiotic-antimycotic solutions were from Corning Cellgro (Herndon, VA).…”

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Nitro-fatty acid pharmacokinetics in the adipose tissue compartment

Fazzari

Khoo

Woodcock

et al. 2017

Journal of Lipid Research

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inflammatory and metabolic stress (1-3). In particular, the reaction between unsaturated FAs and the nitric oxide and nitrite (NO 2  )-derived radical species nitrogen dioxide (·NO 2 ) generates electrophilic FA nitro-alkene byproducts (NO 2 -FAs) (4). These molecules are endogenously detected and have been observed to mediate salutary responses in models of inflammatory injury and oxidative stress. Current data supports that NO 2 -FAs predominantly signal via posttranslational protein modifications, specifically of functionally significant Cys residues of nuclear transcription factor erythroid 2-related factor 2 (Nrf2), soluble epoxide hydrolase (sEH), p65 subunit of nuclear factor kappa B (NF-kB), transient receptor potential cation channel subfamily V member 1 (TRPV1), and heat shock factor-1 (HSF-1) (5-9).NO 2 -FAs have been detected in a variety of species, including mammals and plants, as both free acid and as complex lipid-esterified species (3, 10, 11). For example, NO 2 -FAs are detected at micromolar concentrations in rodent cardiac mitochondria subjected to an episode of ischemia-reperfusion and at nanomolar concentrations in healthy human plasma and urine (12)(13)(14). Additionally, [D 4 ]octadecenoic acid; Nrf2, nuclear transcription factor erythroid 2-related factor 2; OA, oleic acid; PC, phosphatidylcholine; QWBA, quantitative whole-body autoradiography; sEH, soluble epoxide hydrolase; TAG, triacylglyceride. The designations "9-NO 2 -" and "10-NO 2 -"OA are used herein to describe the position of the nitro group in the fatty acid chain and do not refer to IUPAC nomenclature. MED Foundation (M.F.), National Institutes of Health Grants R01-AT006822 (F.J.S.), R01-HL64937, R01-HL132550, and P01-HL103455 (B.A.F.), and American Heart Association Grant 14GRNT20170024 (F.J.S.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the1 To whom correspondence should be addressed. e-mail: freerad@pitt.edu (B.A.F.); fjs2@pitt.edu (F.J.S.) The online version of this article (available at http://www.jlr.org) contains a supplement.

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Nitrated fatty acids: synthesis and measurement

Cited by 55 publications

References 32 publications

The Chemical Basis of Thiol Addition to Nitro-conjugated Linoleic Acid, a Protective Cell-signaling Lipid

The Chemical Basis of Thiol Addition to Nitro-conjugated Linoleic Acid, a Protective Cell-signaling Lipid

Modulation of Nitro-fatty Acid Signaling

Nitro-fatty acid pharmacokinetics in the adipose tissue compartment

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