Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD

Yoshiaki H. Hitomi; Junna Okuda; Hirohito Nishino; Yasuhiro Kambayashi; Yuri Hibino; Kei Takemoto; Tomoko Takigawa; Hideki Ohno; Naoyuki Taniguchi; Keiki Ogino

doi:10.1093/jb/mvm050

Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD

Yoshiaki H. Hitomi, Junna Okuda, Hirohito Nishino, Yasuhiro Kambayashi, Yuri Hibino, Kei Takemoto, Tomoko Takigawa, Hideki Ohno, Naoyuki Taniguchi, Keiki Ogino

Research output: Contribution to journal › Article › peer-review

14 Citations (Scopus)

Abstract

3-Nitrotyrosine (NTyr) is considered as a biomarker of the generation of reactive nitrogen species (RNS). However, it is still difficult to determine its concentration in biological samples. To develop a reliable and high-throughput method, we optimized the conditions for high performance liquid chromatography and electrochemical detection (HPLC-ECD). The best separation of NTyr was achieved using a highly acidic mobile phase (pH 2.5). The concentration of protein-bound NTyr in plasma protein was 593.6 ± 53.8 fmol/mg in rats treated with lipopolysaccharide (LPS) and 114.4 ± 27.6 fmol/mg in control. After intravenous administration of in vitro-nitrated plasma protein, NTyr concentration decreased; the half-life was 63.4 ± 16.8 h. Consistently, protein-bound NTyr concentration in plasma after LPS treatment declined gradually, but was detectable for 1 week. Our protocol is reproducible and suitable for analysing multiple clinical samples to study RNS production in vivo.

Original language	English
Pages (from-to)	495-502
Number of pages	8
Journal	Journal of biochemistry
Volume	141
Issue number	4
DOIs	https://doi.org/10.1093/jb/mvm050
Publication status	Published - Apr 2007
Externally published	Yes

Keywords

3-nitrotyrosine
Electrochemical detection
HPLC
Lipopolysaccharide
Nitric oxide

ASJC Scopus subject areas

Biochemistry
Molecular Biology

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10.1093/jb/mvm050

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title = "Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD",

abstract = "3-Nitrotyrosine (NTyr) is considered as a biomarker of the generation of reactive nitrogen species (RNS). However, it is still difficult to determine its concentration in biological samples. To develop a reliable and high-throughput method, we optimized the conditions for high performance liquid chromatography and electrochemical detection (HPLC-ECD). The best separation of NTyr was achieved using a highly acidic mobile phase (pH 2.5). The concentration of protein-bound NTyr in plasma protein was 593.6 ± 53.8 fmol/mg in rats treated with lipopolysaccharide (LPS) and 114.4 ± 27.6 fmol/mg in control. After intravenous administration of in vitro-nitrated plasma protein, NTyr concentration decreased; the half-life was 63.4 ± 16.8 h. Consistently, protein-bound NTyr concentration in plasma after LPS treatment declined gradually, but was detectable for 1 week. Our protocol is reproducible and suitable for analysing multiple clinical samples to study RNS production in vivo.",

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author = "Hitomi, {Yoshiaki H.} and Junna Okuda and Hirohito Nishino and Yasuhiro Kambayashi and Yuri Hibino and Kei Takemoto and Tomoko Takigawa and Hideki Ohno and Naoyuki Taniguchi and Keiki Ogino",

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doi = "10.1093/jb/mvm050",

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pages = "495--502",

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T1 - Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD

AU - Hitomi, Yoshiaki H.

AU - Okuda, Junna

AU - Nishino, Hirohito

AU - Kambayashi, Yasuhiro

AU - Hibino, Yuri

AU - Takemoto, Kei

AU - Takigawa, Tomoko

AU - Ohno, Hideki

AU - Taniguchi, Naoyuki

AU - Ogino, Keiki

PY - 2007/4

Y1 - 2007/4

N2 - 3-Nitrotyrosine (NTyr) is considered as a biomarker of the generation of reactive nitrogen species (RNS). However, it is still difficult to determine its concentration in biological samples. To develop a reliable and high-throughput method, we optimized the conditions for high performance liquid chromatography and electrochemical detection (HPLC-ECD). The best separation of NTyr was achieved using a highly acidic mobile phase (pH 2.5). The concentration of protein-bound NTyr in plasma protein was 593.6 ± 53.8 fmol/mg in rats treated with lipopolysaccharide (LPS) and 114.4 ± 27.6 fmol/mg in control. After intravenous administration of in vitro-nitrated plasma protein, NTyr concentration decreased; the half-life was 63.4 ± 16.8 h. Consistently, protein-bound NTyr concentration in plasma after LPS treatment declined gradually, but was detectable for 1 week. Our protocol is reproducible and suitable for analysing multiple clinical samples to study RNS production in vivo.

AB - 3-Nitrotyrosine (NTyr) is considered as a biomarker of the generation of reactive nitrogen species (RNS). However, it is still difficult to determine its concentration in biological samples. To develop a reliable and high-throughput method, we optimized the conditions for high performance liquid chromatography and electrochemical detection (HPLC-ECD). The best separation of NTyr was achieved using a highly acidic mobile phase (pH 2.5). The concentration of protein-bound NTyr in plasma protein was 593.6 ± 53.8 fmol/mg in rats treated with lipopolysaccharide (LPS) and 114.4 ± 27.6 fmol/mg in control. After intravenous administration of in vitro-nitrated plasma protein, NTyr concentration decreased; the half-life was 63.4 ± 16.8 h. Consistently, protein-bound NTyr concentration in plasma after LPS treatment declined gradually, but was detectable for 1 week. Our protocol is reproducible and suitable for analysing multiple clinical samples to study RNS production in vivo.

KW - 3-nitrotyrosine

KW - Electrochemical detection

KW - HPLC

KW - Lipopolysaccharide

KW - Nitric oxide

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ER -

Disposition of protein-bound 3-nitrotyrosine in rat plasma analysed by a novel protocol for HPLC-ECD

Abstract

Keywords

ASJC Scopus subject areas

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