Specific GC–MS/MS stable-isotope dilution methodology for free 9- and 10-nitro-oleic acid in human plasma challenges previous LC–MS/MS reports

Tsikas, Dimitrios; Zoerner, Alexander A.; Mitschke, Anja; Homsi, Yahya; Gutzki, Frank‐Mathias; Jordan, Jens

doi:10.1016/j.jchromb.2008.12.062

Cited by 47 publications

(45 citation statements)

References 21 publications

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“…Consequently, the concentrations of fatty acid nitration products in tissues and fl uids have been the subject of controversy ( 34 ). Initial concentrations of free and esterifi ed NO 2 -FA were initially reported to be in the high nM range ( 14 ).…”

Section: Discussionmentioning

confidence: 99%

Characterization and quantification of endogenous fatty acid nitroalkene metabolites in human urine

Salvatore

Vitturi

Baker

et al. 2013

Journal of Lipid Research

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This article is available online at http://www.jlr.org biomolecule nitration ( 1 ). Nitroalkene substituents are electrophilic and promote Michael addition of fatty acids with biological nucleophiles such as cysteine and histidine. The extent, rate, and reversibility of these reactions will be dictated both by the concentration and reactivity of individual nucleophiles. In this regard, protein structure and compartmentalization affect the reactivity of individual nucleophilic centers and will defi ne the molecular targets of electrophilic fatty acids.While enzymatically-oxygenated unsaturated fatty acids typically transduce anti-infl ammatory actions via specifi c g protein-coupled receptor ligand activity ( 2, 3 ), transcriptional responses to electrophilic fatty acids reveal that a broader array of signaling events are instigated ( 4, 5 ). The basis for this pleiotropy resides in the facile Michael addition of electrophilic fatty acid derivatives with nucleophilic centers of proteins that regulate structure and function ( 6 ). Functionally-signifi cant protein targets of electrophilic fatty acids include the transcriptional regulatory protein complex nuclear factor kappa B (NFkB), the

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Section: Discussionmentioning

confidence: 99%

Characterization and quantification of endogenous fatty acid nitroalkene metabolites in human urine

Salvatore

Vitturi

Baker

et al. 2013

Journal of Lipid Research

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“…The presence of the endogenously formed NO 2 -FA was reported for the first time more than a decade ago [9]. Several LC-MS/MS and gas chromatography-MS/MS studies have been directed toward quantification of LaNO 2 and OaNO 2 in human blood plasma, and reported values between 300 pmol/L [9][10][11]13] and 600 nmol/L [14,15], whereas AaNO 2 remained undetected [10]. Recently, Bonnaci et al [12] showed that dietary cLa rather than bisallylic La is a preferential substrate for the nitration by ONOO -and • NO 2 , with endogenous levels 0.3 to 1.3 nmol/L in healthy human plasma.…”

Section: No 2 -Fa In the Cardiomyocyte Nitrosative Stress Modelmentioning

confidence: 99%

“…In vivo formation of nitrated fatty acids (NO 2 -FA) was supported by their detection in human blood plasma [9][10][11][12][13], membranes of red blood cells [14], and urine [15,16]. Recent evidence indicates increased nitroalkene production rates in various inflammatory models, such as a 20-fold higher cholesteryl nitrolinoleate level in murine J774.1 macrophages activated by an inflammatory stimulus [17].…”

Section: Introductionmentioning

confidence: 99%

Profiling and relative quantification of multiply nitrated and oxidized fatty acids

et al. 2015

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The levels of nitro fatty acids (NO2-FA), such as nitroarachidonic, nitrolinoleic, nitrooleic, and dinitrooleic acids, are elevated under various inflammatory conditions, and this results in different anti-inflammatory effects. However, other multiply nitrated and nitro-oxidized FAs have not been studied so far. Owing to the low concentrations in vivo, NO2-FA analytics usually relies on targeted gas chromatography-tandem mass spectrometry (MS/MS) or liquid chromatography-MS/MS, and thus require standard compounds for method development. To overcome this limitation and increase the number and diversity of analytes, we performed in-depth mass spectrometry (MS) profiling of nitration products formed in vitro by incubating fatty acids with NO2BF4, and ONOO(-). The modified fatty acids were used to develop a highly specific and sensitive multiple reaction monitoring LC-MS method for relative quantification of 42 different nitrated and oxidized species representing three different groups: singly nitrated, multiply nitrated, and nitro-oxidized fatty acids. The method was validated in in vitro nitration kinetic studies and in a cellular model of nitrosative stress. NO2-FA were quantified in lipid extracts from 3-morpholinosydnonimine-treated rat primary cardiomyocytes after 15, 30, and 70 min from stress onset. The relatively high levels of dinitrooleic, nitroarachidonic, hydroxynitrodocosapenataenoic, nitrodocosahexaenoic, hydroxynitrodocosahexaenoic, and dinitrodocosahexaenoic acids confirm the presence of multiply nitrated and nitro-oxidized fatty acids in biological systems for the first time. Thus, in vitro nitration was successfully used to establish a targeted LC-MS/MS method that was applied to complex biological samples for quantifying diverse NO2-FA. Graphical Abstract Schematic representation of study design which combined in vitro nitration of different fatty acids, MS/MS characterization and optimization of MRM method for relative quantification, which was applied to follow dynamic of fatty acid nitration in cellular model of SIN-1 treated cardiomyoctes.

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“…For the first time, the nonesterified nitro-oleic acid (NO 2 -OA) has been found by LC-MS/MS in human plasma, at a basal concentration of about 600 nM [5]. Recently, our group has reported on the development, validation, and application of a highly specific GC-MS/MS method for the quantification of NO 2 -OA in human plasma [13]. In order to avoid potential interferences we incorporated in this GC-MS/MS method a HPLC step for separation and isolation of NO 2 -OA.…”

Section: Introductionmentioning

confidence: 99%

Nitro‐fatty Acids Occur in Human Plasma in the Picomolar Range: a Targeted Nitro‐lipidomics GC–MS/MS Study

Tsikas

Zoerner

Mitschke

et al. 2009

Lipids

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First studies on the occurrence of nitrated fatty acids in plasma of healthy subjects revealed basal concentrations of 600 nM for free/nonesterified nitro-oleic acid (NO(2)-OA) as measured by liquid chromatography tandem mass spectrometry (LC-MS/MS). We recently showed by a gas chromatography tandem mass spectrometry (GC-MS/MS) method the physiological occurrence of two isomers, i.e., 9-NO(2)-OA and 10-NO(2)-OA, at mean basal plasma concentrations of 880 and 940 pM, respectively. In consideration of this large discrepancy we modified our originally reported method by replacing solid-phase extraction (SPE) by solvent extraction with ethyl acetate and by omitting the high-performance liquid chromatography (HPLC) step for a more direct detection and with the potential for lipidomics studies. Intra-assay imprecision and accuracy of the modified method in human plasma were 1-34% and 91-221%, respectively, for added NO(2)-OA concentrations in the range 0-3,000 pM. This method provided basal plasma concentrations of 306 +/- 44 pM for 9-NO(2)-OA and 316 +/- 33 pM for 10-NO(2)-OA in 15 healthy subjects. Nitro-arachidonic acid and nitro-linolenic acid were not detectable in the plasma samples. In summary, our studies show 9-NO(2)-OA and 10-NO(2)-OA as endogenous nitrated fatty acids in human plasma in the pM range; HPLC is recommendable as a sample clean-up step for reliable quantification of nitro-oleic acids by GC-MS/MS.

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Specific GC–MS/MS stable-isotope dilution methodology for free 9- and 10-nitro-oleic acid in human plasma challenges previous LC–MS/MS reports

Cited by 47 publications

References 21 publications

Characterization and quantification of endogenous fatty acid nitroalkene metabolites in human urine

Characterization and quantification of endogenous fatty acid nitroalkene metabolites in human urine

Profiling and relative quantification of multiply nitrated and oxidized fatty acids

Nitro‐fatty Acids Occur in Human Plasma in the Picomolar Range: a Targeted Nitro‐lipidomics GC–MS/MS Study

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