Nitrite and Nitrate

This is one of the most common applications for segmented flow analyzers and there are quite a few ways of doing it. The one important caveat is that there is no way to directly determine soluble nitrates, all approaches first reduce the nitrate to nitrite and then analyze for total nitrite. The old standard for this test is nitrate reduction by metallic cadmium, a variation on this method uses fluorometric detection which is useful for eliminating artifacts commonly seen when analyzing salt water, another well documented method uses hydrazine to reduce the nitrate, a "new" method that uses vanadium trioxide has the advantage of less toxic reagents and, finally, there is an enzymatic method that works well on the discrete analyzer.

Nitrate/Nitrite by Cadmium Reduction

This method is used for the determination of nitrite or nitrate plus nitrite in drinking, surface and saline waters, domestic and industrial wastes, plants and soils. This method is applicable to several ranges.

Summary

Nitrate is reduced quantitatively to nitrite by cadmium metal in the form of an open tubular cadmium reactor (OTCR). The nitrite thus formed plus any originally present in the sample is determined as an azo dye at 520 nm following its diazotization with sulfanilamide and subsequent coupling with N-1-naphthylethylenediamine. These reactions take place in acidic solution. Nydahl provides a good discussion of nitrate reduction by cadmium metal, while the specific details of OTCR's are given by Patton. The information concerning mechanisms and kinetics of the color forming reactions can be found in (citation needed).

Limitations

The imidazole buffer eliminates interferences from iron, copper or other metals. Turbid samples should be filtered before determination. Samples with background absorbance at an analytical wavelength may interfere. Samples containing oil or grease must be extracted with an organic solvent prior to analysis. Samples containing sulfide cannot be determined by this method without first removing the sulfide by precipitation with cadmium salts. Norwitz and Keliher (citation needed) have compiled a comprehensive study of interferences in the spectrophotometric analysis of nitrite. 

System Method # Range(s)
Flow System A170 & A173 0.05 - 50 mg/l as N
Astoria 2 A170 & A173 0.05 - 50 mg/l as N
rAPID-T N/A  

Nitrate/Nitrite by Hydrazine Reduction

This method is used for the determination of nitrate and nitrite nitrogen in drinking and surface waters, domestic and industrial wastes, plants and soils and is applicable to many ranges.

Summary

Nitrate is reduced quantitatively to nitrite by hydrazine sulfate in the presence of copper. The nitrite thus formed, plus any originally present in the sample, is determined as an azo dye and measured at 520 nm following its diazotization with sulfanilamide and subsequent coupling with N-1-naphthylethylenediamine. (1) These reactions are effected in acidic solution. Kamphake, et al, provides a good discussion of nitrate reduction by hydrazine sulfate. (2) Information concerning mechanisms and kinetics of the color forming reactions can be found in References 3 and 4.

Limitations

Turbid samples should be filtered before determination. Samples with background absorbance at the analytical wavelength may interfere. Samples containing sulfide cannot be determined by this method without first removing the sulfide by precipitation with cadmium salts. Norwitz and Keliher have compiled a comprehensive study of interferences in the spectrophotometric analysis of nitrite. (5,6,7) Samples containing calcium or magnesium in high concentrations will precipitate at the high pH of the initial stage of the analysis.

System Method # Range(s)

Flow System

A174 0.01-5 mg/l, 0.5-200 mg/l as N
Astoria 2 A174 0.01-5 mg/l, 0.5-200 mg/l as N
rAPID-T N/A  

Nitrate/Nitrite in Sea Water

This method is used for the determination of nitrate+nitrite in seawater and freshwater. The applicable range of this method is 0.05 to 5 µM.

Summary

Sample nitrate is reduced quantitatively to nitrite by cadmium metal in the form of a open tube cadmium reactor (OTCR). The nitrite reduced plus any nitrite originally present undergoes a diazotization with fluorosceinamine in an acidic medium. After sufficient incubation the reaction is completed by addition of a strong base. The resulting dye is measured fluorometrically (480nm excitation and 520nm emission) and is directly proportional to concentration.

Limitations

Iron, manganese, bromate and sulfite were determined to cause some interference at 100 fold excess or more. 

System Method # Range(s)
Flow System A179 0.05-5 μM
Astoria 2 A179 0.05-5 μM
rAPID-T    

 

Nitrate/Nitrite by Vanadium Trioxide Reduction

This method describes the determination of nitrate in drinking, ground, surface, domestic and industrial waters.

Summary

Nitrate is reduced quantitatively to nitrite by vanadium in a dilute acidic solution. The nitrite formed, plus any originally present in the sample, is determined as an azo dye at 540 nm, following its diazotization with sulfanilamide and subsequent coupling with N-1-naphthylethylenediamine. The information concerning mechanisms and kinetics of the color forming reactions can be found in (citation needed).

System Method # Range(s)
Flow System    
Astoria 2    
rAPID-T D179  0.02–5.0 mg/l, 0.002–0.200 mg/l as N

 

 

Nitrate/Nitrite by Enzymatic Reduction

This method describes the determination of nitrate in drinking, ground, surface, domestic and industrial waters.

Summary

Nitrate is reduced quantitatively to nitrite enzymatically using the nitrate reductase enzyme (NaR) with its natural reducing agent, reduced nicotinamide dinucleotide (NADH). The nitrite thus formed plus any originally present in the sample is determined as an azo dye at 540 nm following its diazotization with sulfanilamide and subsequent coupling with N-1-naphthylethylenediamine. These reactions take place in acidic solution.

Limitations

High concentrations of metals can cause low results; EDTA is used to complex metal ions and eliminate their interference. Turbidity can interfere with results and is avoided by filtering samples with a 0.45 µm membrane filter. Occasionally, commercially prepared standards contain azides or chlorinated solvents that can bind with the NaR enzyme and inhibit reduction. Always check commercially prepared standards against a laboratory prepared second source.

System Method # Range(s)
Flow System    
Astoria 2    
rAPID-T D17  0.01–1.0 mg/l as N