Improved nitric oxide detection using 2,3-diaminonaphthalene and its application to the evaluation of novel nitric oxide synthase inhibitors

Naoki Nakatsubo, Hirotatsu Kojima, Kuniko Sakurai, Kazuya Kikuchi, Hiroshi Nagoshi, Yasunobu Hirata, Takaaki Akaike, Hiroshi Maeda, Yasuteru Urano, Tsunehiko Higuchi, Tetsuo Nagano

Research output: Contribution to journalArticlepeer-review

31 Citations (Scopus)

Abstract

A specific and sensitive detection method for nitric oxide (NO) in living cells and tissue culture systems is required in the search for novel NO synthase (NOS) inhibitors. We have improved a fluorometric determination with 2,3-diaminonaphthalene (DAN) by the addition of 2-phenyl-4,4,5,5- tetramethylimidazoline-3-oxide-1-oxyl (PTIO) as an oxidant to form NO2 from NO. This method is 3 times more sensitive than that without PTIO, and is suitable for examining the NOS-inhibitory activity of large numbers of test compounds using a 96-well microplate reader. The improved method was applied to N-monomethyl-L-arginine (L-NMMA) as a known inhibitor and the derivatives of 2-phenyl-1,2-benzisoselenazol-3(2H)-one as teat compounds in order to investigate the effect of these compounds on NO production from activated rat aortic smooth muscle cells. The results obtained indicate that this method is suitable for the rapid assay of large numbers of test compounds.

Original languageEnglish
Pages (from-to)1247-1250
Number of pages4
JournalBiological and Pharmaceutical Bulletin
Volume21
Issue number12
DOIs
Publication statusPublished - 1998
Externally publishedYes

Keywords

  • Diaminonaphthalene
  • Fluorescence
  • Inhibitor
  • Nitric oxide
  • Nitric oxide synthase
  • Probe

ASJC Scopus subject areas

  • Pharmacology
  • Pharmaceutical Science

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