Profiling and relative quantification of multiply nitrated and oxidized fatty acids.

I Milic, E Griesser, V Vemula, N Ieda, H Nakagawa, N Miyata, JM Galano, C Oger, T Durand, M Fedorova

Research output: Contribution to journalArticle

Abstract

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.
Original languageEnglish
Pages (from-to)5587–5602
JournalAnalytical and Bioanalytical Chemistry
Volume407
Issue number19
DOIs
Publication statusPublished - Jul 2015

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Mass spectrometry
Fatty Acids
Nitration
Tandem Mass Spectrometry
Mass Spectrometry
Acids
Cardiac Myocytes
Biological systems
Anti-Inflammatory Agents
Gases
Rats
Lipids
Kinetics
Monitoring
Liquids
In Vitro Techniques

Cite this

Milic, I ; Griesser, E ; Vemula, V ; Ieda, N ; Nakagawa, H ; Miyata, N ; Galano, JM ; Oger, C ; Durand, T ; Fedorova, M. / Profiling and relative quantification of multiply nitrated and oxidized fatty acids. In: Analytical and Bioanalytical Chemistry. 2015 ; Vol. 407, No. 19. pp. 5587–5602.
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abstract = "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.",
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Milic, I, Griesser, E, Vemula, V, Ieda, N, Nakagawa, H, Miyata, N, Galano, JM, Oger, C, Durand, T & Fedorova, M 2015, 'Profiling and relative quantification of multiply nitrated and oxidized fatty acids.', Analytical and Bioanalytical Chemistry, vol. 407, no. 19, pp. 5587–5602. https://doi.org/10.1007/s00216-015-8766-3

Profiling and relative quantification of multiply nitrated and oxidized fatty acids. / Milic, I; Griesser, E; Vemula, V; Ieda, N; Nakagawa, H; Miyata, N; Galano, JM; Oger, C; Durand, T; Fedorova, M.

In: Analytical and Bioanalytical Chemistry, Vol. 407, No. 19, 07.2015, p. 5587–5602.

Research output: Contribution to journalArticle

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T1 - Profiling and relative quantification of multiply nitrated and oxidized fatty acids.

AU - Milic, I

AU - Griesser, E

AU - Vemula, V

AU - Ieda, N

AU - Nakagawa, H

AU - Miyata, N

AU - Galano, JM

AU - Oger, C

AU - Durand, T

AU - Fedorova, M

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AB - 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.

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