Common Biochemical Tests in Microbiology: Nitrate Reduction Test

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BACKGROUND

Nitrate reduction test is used for differentiating organism on the basis of their capability to produce nitrate reductase enzyme that causes hydrolysis of nitrate to nitrite, which again depending on the various enzymes secreted by the organism will degrade to various other nitrogen products like nitrogen oxide, nitrous oxide and ammonia.¹

So, the basic objective of this test is to conduct the nitrate reduction test.

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REQUIREMENTS

Media:                Nitrate Broth with upturned Durham’s tube

Reagents:           Sulphalinic acid reagent,

     Alpha naphthylamine reagent

     Zinc dust

Miscellaneous:   Inoculating loop

      Burner

      Dropper

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PROCEDURE

Prepare nitrate broth in test tube and inoculate the test organism in this under aseptic conditions. Then incubate it at required temperature for about 24-48 hours. Then to broth add each sulfanilic acid 1 drop and alpha-naphthylamine one drop. If at this stage there is formation of red colouration, then it is nitrate reduction test positive and you can stop the test here. If there is no formation of red colour, then you can go to next step. Take small amount of zinc and add to each broth. Addition of zinc catalyzes the reduction of nitrate to nitrite. If red colour is formed then nitrate reduction test positive and no colour change then negative test results.^1,2

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CONCLUSION

Nitrate reduction test is used for differentiation of various bacterial species.

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REFERENCES

1. Chapin K, Lauderdale TL. 2007. Reagents, stains, and media: bacteriology, p. 339. In P. R. Murray, E. J. Baron, J. H. Jorgensen, M. L. Landry, and M. A. Pfaller (ed.), Manual of clinical microbiology, 9th ed. ASM Press, Washington, DC.
   
2. Committee on Laboratory Standards and Practices, American Public Health Association. 1975. Bacterial nitrate reduction test: suggestions for use of alternate (noncarcinogenic) reagents. ASM News 41:225.