Quantifying Nanomolar Levels of Nitrite in Biological Samples by HPLC-Griess Method: Special Reference to Arterio-Venous Difference in vivo
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- Ishibashi Takaharu
- Department of Pharmacology, School of Nursing, Kanazawa Medical University Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Miwa Tomoko
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Shinkawa Ikumi
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Nishizawa Naoki
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Nomura Mihoko
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Yoshida Junko
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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- Kawada Tomie
- Department of Clinical Pharmacy, Faculty of Pharmacy, Musashino University
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- Nishio Matomo
- Department of Pharmacology, School of Medicine, Kanazawa Medical University
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Description
Nitrite (NO2-) is assumed to play an important role in regulation of vascular tone as a reservoir of nitric oxide (NO). To examine its physiological contribution, however, a sensitive method is required for determination of the true level of NO2- in biological samples. To this end, practical consideration to avoid NO2- contamination through the quantification procedure is important. We present here a highly sensitive and accurate method for determining NO2- in plasma by improving the HPLC-Griess system with minimal NO2- contamination in the samples. The system achieved high sensitivity (detection limit of 2 nM and sensitivity to 1 nM) and complete separation of the NO2- signal peak by modifying the system setup and mobile phase. Using this method, we achieved acceptable quantification of low NO2- levels in plasma. Deproteinization by ultrafiltration and exposure to atmosphere before measurement were identified as the major sources of NO2- contamination during sample processing. We addressed these issues by the use of methanol for deproteinization and gas-tight caps. These countermeasures allowed us to detect small arterio-venous NO2- differences in rabbit plasma that may indicate kinetic difference of NO2- in a small number of samples (n = 6). This difference became prominent when NO2- or a NO releasing agent, NOR1, was intravenously applied. Our results indicate that application of a sensitive method with careful handling is important for accurate determination of NO2- and that our method is applicable for further examination of the kinetic features of NO2- in vivo.
Journal
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- The Tohoku Journal of Experimental Medicine
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The Tohoku Journal of Experimental Medicine 215 (1), 1-11, 2008
Tohoku University Medical Press
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Details 詳細情報について
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- CRID
- 1390282679217558656
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- NII Article ID
- 10024167728
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- NII Book ID
- AA00863920
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- ISSN
- 13493329
- 00408727
- http://id.crossref.org/issn/00408727
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- Text Lang
- en
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- Data Source
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- JaLC
- Crossref
- CiNii Articles
- KAKEN
- OpenAIRE
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- Abstract License Flag
- Disallowed