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Effects of reducing reagents and temperature on conversion of nitrite and nitrate to nitric oxide and detection of NO by chemiluminescence

Fan Yang, Eric Troncy, Martin Francur, Bernard Vinet¹, Patrick Vinay², Guy Czaika and Gilbert Blaise^a

Laboratory of Anesthesia, Departments of Anesthesia,
¹ Biochemistry, and
² Medicine, Hôpital Notre-Dame, Université de Montréal, 1560 Sherbrooke St. East, Montréal, QC, Canada H2L 4M1.
^a Author for correspondence. Fax 514-896-4754; e-mail blaiseg{at}ere.umontreal.ca

To measure the concentration of nitrites and nitrates by chemiluminescence, we examined the efficiency of five reducing agents [V(III), Mo(VI) + Fe(II), NaI, Ti(III), and Cr(III)] to reduce nitrite (NO₂^-) and (or) nitrate (NO₃^-) to nitric oxide (NO). The effect of each reducing agent on the conversion of different amounts of NO₂^- and (or) NO₃^-(100–500 pmol, representing concentrations of 0.4 to 2 µmolar) to NO was determined at 20 °C for NO₂^- and at 80 °C for NO₃^-. The effect of temperature from 20 to 90 °C on the conversion of a fixed amount of NO₂^- or NO₃^- (400 pmol or 1.6 µmolar) to NO was also determined. These five reducing agents are similarly efficient for the conversion of NO₂^- to NO at 20 °C. V(III) and Mo(VI) + Fe(II) can completely reduce NO₃^- to NO at 80 °C. NaI and Cr(III) were unable to convert NO₃^- to NO. Increased temperature facilitated the conversion of NO₃^- to NO, rather than that of NO₂^- to NO. We evaluated the recovery of NO₂^- and NO₃^- from plasmas of pig and of dog. Recovery from plasma of both animals was reproducible and near quantitative.

Key Words: indexing terms: endothelium-derived relaxing factor • nitric oxide synthase • free radical • vasodilation • inflammation • thrombosis • immunology • neurotransmission

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