The nitrate reductase test is a test to differentiate between
bacteria
Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one biological cell. They constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria were among ...
based on their ability or inability to reduce
nitrate
Nitrate is a polyatomic ion
A polyatomic ion, also known as a molecular ion, is a covalent bonded set of two or more atoms, or of a metal complex, that can be considered to behave as a single unit and that has a net charge that is not zer ...
(NO
3−) to
nitrite
The nitrite polyatomic ion, ion has the chemical formula . Nitrite (mostly sodium nitrite) is widely used throughout chemical and pharmaceutical industries. The nitrite anion is a pervasive intermediate in the nitrogen cycle in nature. The name ...
(NO
2−) using
anaerobic respiration
Anaerobic respiration is respiration using electron acceptors other than molecular oxygen (O2). Although oxygen is not the final electron acceptor, the process still uses a respiratory electron transport chain.
In aerobic organisms undergoing re ...
.
Procedure
Various assays for detecting nitrate reduction have been described.
[V.B.D. Skerman, A guide to the identification of the genera of bacteria, The Williams & Wilkins Co., Baltimore, MD, p.218 - 220 (1967).] One method is performed as follows:
#
Inoculate
Inoculation is the act of implanting a pathogen or other microorganism. It may refer to methods of artificially inducing immunity against various infectious diseases, or it may be used to describe the spreading of disease, as in "self-inoculati ...
nitrate broth with an isolate and incubate for 48 hours.
#Add two nitrate tablets to the sample. If the bacterium produces
nitrate reductase
Nitrate reductases are molybdoenzymes that reduce nitrate (NO) to nitrite (NO). This reaction is critical for the production of protein in most crop plants, as nitrate is the predominant source of nitrogen in fertilized soils.
Types
Euka ...
, the broth will turn a deep red within 5 minutes at this step.
#If no color change is observed, then the result is inconclusive. Add a small amount of
zinc
Zinc is a chemical element with the symbol Zn and atomic number 30. Zinc is a slightly brittle metal at room temperature and has a shiny-greyish appearance when oxidation is removed. It is the first element in group 12 (IIB) of the periodi ...
to the broth. If the solution remains colorless, then both nitrate reductase and
nitrite reductase
Nitrite reductase refers to any of several classes of enzymes that catalyze the reduction of nitrite. There are two classes of NIR's. A multi haem enzyme reduces NO2− to a variety of products. Copper containing enzymes carry out a single elect ...
are present. If the solution turns red, nitrate reductase is not present.
References
Bacteriology
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