Diaphorase Can Metabolize
Some Vasorelaxants to NO and Eliminate NO Scavenging Effect of
2-Phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide (PTIO)
P. BARTÍK1, K. CHALUPSKÝ1,2, L. VAVRUŠKA1,
B. MULLER3, J.-C. STOCLET2, G. ENTLICHER1
1 Department of Biochemistry, Faculty of Science,
Charles University, Prague, Czech Republic, 2Laboratoire
de Pharmacologie et de Physiopathologie Cellulaires, CNRS URA,
Universite Louis Pasteur, Strasbourg, France and 3Laboratoire
de Pharmacologie de l‘UFR de Sciences Pharmaceutiques,
Universite Victor Segalen, Bordeaux, France
Received July 17, 2003
Accepted January 23, 2004
Summary
Diaphorase was studied as a possible oxidoreductase
participating in NO production from some vasorelaxants. In the
presence of NADH or NADPH, diaphorase can convert selected NO
donors, glycerol trinitrate (GTN) and formaldoxime (FAL) to
nitrites and nitrates with NO as an intermediate. This activity
of diaphorase was inhibited by diphenyleneiodonium (DPI)
(inhibitor of some NADPH-dependent flavoprotein
oxidoreductases), while it remained uninhibited by
NG-nitro-L-arginine methyl ester (inhibitor of NO synthase)
7-Ethoxyresorufin (inhibitor of cytochrome P-450 1A1 and
cytochrome P-450 NADPH-dependent reductase) inhibited the
conversion of GTN only. Existence of NO as an intermediate of
the reaction was supported by results of electron paramagnetic
resonance spectroscopy. In addition to its ability to affect the
above mentioned NO donors, diaphorase was able to reduce
2-phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide (PTIO)
and thus to eliminate its NO scavenging effect. This activity of
diaphorase could also be inhibited by DPI. The reaction of
diaphorase with GTN and PTIO was not affected by superoxide
dismutase (SOD) or catalase. Reaction of FAL with diaphorase was
lowered with SOD by 38 % indicating the partial participation of
superoxide anion probably generated by the reaction of
diaphorase with NADH or NADPH. Catalase had no effect.
Diaphorase could apparently be one of the enzymes participating
in the metabolism of studied NO donors to NO. The easy reduction
and consequent elimination of PTIO by diaphorase could affect
its use as an NO scavenger in biological tissues.
Key words
Formaldoxime metabolism • Glycerol trinitrate metabolism •
Diaphorase •
2-phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide (PTIO)
reduction
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