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Updated: Jul 4, 2026

Measuring Nitrite and Nitrate, Metabolites in the Nitric Oxide Pathway, in Biological Materials using the Chemiluminescence Method
Published on: December 25, 2016
A rapid technique for determination of nitrate and nitric acid by acid reduction and diazotization at elevated
1Division of Veterinary Pharmacology & Toxicology, Sher-e-Kashmir University of Agricultural Sciences & Technology of Kashmir, FVSc & AH, Srinagar 190001, Jammu and Kashmir, India. mirsamir.19@rediffmail.com
Abstract:
A rapid technique for determination of nitrate by acid reduction and diazotization at elevated temperature has been standardized. The technique is based on quantitative diazotization of sulfanilamide by nitrate on incubation in boiling water bath for 3, 5 or 10 min in presence of high concentration of HCl, ca. 64.5%. The diazotized sulfanilamide is coupled at room temperature to N-1-(naphthyl)-ethylenediamine dihydrochloride, and the chromophore evaluated spectrophotometrically at 540 nm. The technique provides linear estimate of nitrate over the test range of 0.5 through 10 microg N mL(-1) sample with all test incubation time periods using alkali nitrate and nitric acid as sources of nitrate anion. Urea treatment enables selective determination of nitrate in presence of nitrite with overall 99+/-1% recovery, and without affecting nitrate determination (P>0.1) or its regression coefficient. The technique has obvious advantages over metal-reduction technique. It is simple, rapid, selective in presence of nitrite, and an inexpensive method for routine determination of nitrate with detection range 0.5-10 microg N mL(-1) sample. Besides, the technique provides opportunity to detect nitric acid as low as 35 microM even in presence of other acids.
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