Do you know which is the most abundant gas in the air? It is Nitrogen \(\left( {{{\rm{N}}_{\rm{2}}}} \right)\) which is nearly \({\rm{78\% }}\) in the air. Did you know that plants and animals cannot take this gas directly for their biological uses? Nitrification is one of the important steps of the nitrogen cycle. The nitrogen cycle is one such process that involves an interchange of nitrogen in its different forms that are utilized by plants and animals.

What is the Process of Nitrification?

Nitrification can be defined as the process of biological oxidation of nitrite and further into nitrates. Nitrification is an aerobic process. This is accomplished with the help of certain microorganisms.

Reactions Involved in Nitrification Process

Nitrification is a biological process mainly carried out by certain autotrophic nitrifying bacterias and involves oxidation of nitrogen compounds, main ammonia to nitrite and nitrate, that can be used by living organisms.
Nitrification occurs in mainly two steps:

Conversion of Ammonia to Nitrite

This step is carried forward by ammonia-oxidizing bacteria, also known as nitrifying bacteria, that oxidize ammonia to nitrite.
\({\rm{N}}{{\rm{H}}_{\rm{3}}}{\rm{ + }}{{\rm{O}}_{\rm{2}}} \to {\rm{N}}{{\rm{O}}_{{{\rm{2}}^{\rm{ – }}}}}{\rm{ + 3}}{{\rm{H}}^{\rm{ + }}}{\rm{ + 2}}{{\rm{e}}^ – }\)
During this reaction, ammonia reacts with intermediate hydroxylamine in the presence of two enzymes ammonia oxygenase and hydroxylamine oxidoreductase to form ammonium hydroxide. Ammonium hydroxide further reacts with water and forms nitrite. Nitrosomonas is the most frequently identified bacteria associated with this step. Other genera such as Nitrosospira, Nitrosococcus and certain subgenera like Nitrosolobus and Nitrosovibrio can also autotrophically oxidize ammonia. Aspergillus flavus, a common fungus, can also convert ammonia into nitrite.

Oxidation of Nitrate from Nitrite

The nitrite that is formed is oxidised to nitrate with the help of bacterias, mostly Nitrobacter, during this second step. Below is the chemical equation for the reaction that takes place here:
\({\rm{NO}}_{\rm{2}}^{\rm{ – }}{\rm{ + }}{{\rm{H}}_{\rm{2}}}{\rm{O}} \to {\rm{NO}}_{\rm{3}}^{\rm{ – }}{\rm{ + 2}}{{\rm{H}}^{\rm{ + }}}{\rm{ + 2}}{{\rm{e}}^ – }\)
Few other genera of bacteria also help in this process. Nitrocystis, Nitrospirae, Nitrospinae are often associated with the conversion of nitrite into nitrate. Aspergillus flavus, Penicillium are some common fungi, which can also convert ammonia into nitrite. The oxidation of nitrite to nitrate is an essential step as nitrate is the chemical form of nitrogen used by most plants from soil or water.

Process of Nitrification in the Nitrogen Cycle

Nitrogen in the air is mostly present in its elemental form that is chemically inert and cannot be used by the majority of organisms. Hence, it needs to be converted to forms that can be used by plants from soil or water.
This conversion is mainly carried out by certain nitrogen-fixing bacteria such as Azotobacter (freely in soil), Rhizobium (found in root nodules of leguminous plants), etc. Besides bacteria, several Blue-green algae or Cyanobacteria also help in nitrogen fixation. Nostoc, Anabaena etc., can fix atmospheric nitrogen both in-living and symbiotic forms.
These bacterias convert atmospheric nitrogen into water-soluble nitrates. The process of biofixation of nitrogen is called Nitrogen Fixation.  The key to biofixation is the enzyme nitrogenase which catalyzes the splitting of \({{\rm{N}}_{\rm{2}}}{\rm{.}}\)
During lightning, the nitrogen present in the atmosphere reacts with the oxygen and forms nitric acid, which comes down to earth as rainwater. The nitrates thus formed are used by the plants for the preparation of organic matter such as proteins. These proteins are consumed by animals when they consume plants as food. Now, nitrogenous compounds produced by plants are used to form new animal proteins and other cell components in the animal body.

When animals excrete nitrogenous wastes or nitrogenous compounds in the remains of their dead bodies are again converted into ammonia but certain bacterias. Plants directly use these ammonium ions or nitrifying bacterias to fix them into nitrate by nitrification. Thus, this never-ending cycle is repeated all over again.

Fig: Nitrification and Denitrification in Nitrogen Cycle

The main processes involved in the nitrogen cycle as follows:
1. Ammonification: The formation of Ammonia from nitrogen is called Ammonification.
2. Nitrification: The formation of Nitrate from Ammonia with the help of specialized microorganisms is called Nitrification.
3. Denitrification: A few microorganisms such as Pseudomonas denitrificans, Thiobacillus denitrificans, Micrococcus denitrificans, etc., convert nitrates into nitrogen or into some oxides. The conversion of nitrate salts into nitrogen gas is called Denitrification. These organisms use nitrates as the source of oxygen and obtain energy by reducing them. Denitrification depletes the nitrogen content of the soil.
The oxides that are formed are taken up and utilized by the plants, and the nitrogen returns back to the atmosphere.

Microorganisms Involved in Nitrogen Fixation

Bacterias play an important role in natural or industrial nitrogen fixation. Nitrogen cycle depends upon at least four different kinds of bacteria known as the decay causer, nitrifiers, denitrifiers and lastly, nitrogen fixers.
Microbes, like bacteria, fungus, actinomycetes, cyanobacteria help in the conversion of nitrogen and formation of ammonia. This can be categorised like:
1. Non-symbiotic cyanobacteria like-living forms can fix atmospheric nitrogen. Nostoc, Anabaena help in this process.
2. Symbiotic cyanobacteria can also help in nitrogen fixation. Coralloid roots of Cycas have Nostoc, water fern Azolla has Anabaena in the leaves, Frankia is associated with the leaves and stem of Alnus.
3. Non-symbiotic bacteria like-living soil-borne bacteria also help in nitrogen fixation. Azotobacter, Beijerinckia are-living, aerobic nitrogen-fixing bacteria. Clostridium, Bacillus, Klebsiella are-living, anaerobic nitrogen-fixing bacteria. Rhodospirillum is-living, photosynthetic nitrogen fixing bacteria.
4. Symbiotic bacteria contribute a major share in nitrogen fixation. Rhizobium is symbiotic to the roots of leguminous plants. Rhizobium leguminosarum, R. phaseoli are important species that form root nodules.

Find some microorganisms and their roles in the nitrogen cycle.

Microorganism	Role played by them
Azotobacter	These help in nitrogen fixation in soil.
Rhizobium	Help in nitrogen fixation in root nodules of plants like legumes.
Blue-green algae	These are non-symbiotic bacteria that play an important role in nitrogen fixation in Soil.
Putrefying bacteria	Ammonification
Fungi	Ammonification
Nitrosomonas	Convert ammonia into nitrites.
Nitrobacter	Convert nitrites into nitrates
Pseudomonas	Denitrification

The nitrogen-fixing bacterium of root nodules, Rhizobium leguminosarum, are aerobic bacteria that require some oxygen for their survival. Root nodules of leguminous plants have a haemoglobin-like protein called leg haemoglobin that is used by these bacterias. They require \(16\) molecules of ATP  for each molecule of nitrogen that is fixed. In the soil microorganism, e.g., Klebsiella pneumoniae a total of \(17\) special genes called nif genes are seen to be responsible for nitrogen fixation.

Fig: Root Nodules of Leguminous Plants

Factors Affecting Nitrification Process

Though nitrification is a natural process that occurs regularly in ecosystems, in order to increase plant yield, it is also induced artificially. Factors that lead to a decrease in the rate of nitrification are called nitrification inhibitors. Below are five factors that may influence the rate of nitrification:
1. Water Content in Soil: Increase or decrease in water content reduces the nitrogen fixation in soil.
2. Acidity or Alkalinity: Increase in pH reduces the nitrogen fixation as most aquatic life cannot tolerate approximately a pH of \(6-9,\) whereas if pH exceeds, then there will be a change in the nitrate concentration of dissolved carbon concentration.
3. Light Intensity: Increase in light intensity increases the rate of nitrogen fixation.
4. Oxygen Tension: Increase in oxygen tension decreases the rate of nitrogen fixation.
5. Drought: Increase in drought decreases the rate of nitrogen fixation.

Functions of Nitrification

Below are some functions of the nitrification process:
1. Nitrogen is an important constituent of protein, DNA, RNA, and enzymes. Since elemental or pure nitrogen cannot be directly utilized by most living organisms.
2. Nitrification is the process by which nitrogen becomes available for plants.
3. Nitrification plays an important role in agricultural fields to increase the yield of essential crops like rice, wheat and many leguminous plants.
4. Researchers in biotechnology are attempting to transfer nif genes from microorganisms like Pseudomonas to crop plants to get a better yield of crops.
5. Nitrification also holds an important function in the elimination of nitrogen from municipal wastewater.

Summary

The Nitrogen cycle is also known as a perfect cycle in the biosphere. This is because it maintains the total amount of nitrogen present in the environment, soil and water. Nitrogen is part of amino acids and proteins, which are the building blocks of life. Atmospheric nitrogen needs to be converted into its usable forms through nitrification, ammonification and denitrification regularly to maintain life on the planet.

Frequently Asked Questions (FAQs) on Nitrification

Q.1. What conditions are needed for nitrification?
Ans: The conditions needed for nitrification are:
a. Moisture content
b. pH between \(7-9\)
c. Temperature between approximately \({\rm{10 – 3}}{{\rm{5}}^ \circ }{\rm{C}}\)
d. Soil Retention Time(SRT)
e. Aeration
f. Soil matrix

Q.2. Why does nitrification lower pH?
Ans: Nitrification lowers the pH because most aquatic life cannot tolerate a pH of \(6-9.\) If the pH exceeds, then there will be a change in the nitrate concentration of dissolved carbon concentration.

Q.3. What is meant by nitrification and denitrification?
Ans: Nitrification is a process where ammonia is converted into nitrite and followed by further oxidation of nitrite to nitrate with the help of nitrifying bacteria. Whereas in denitrification a few microorganisms such as Pseudomonas convert nitrates into nitrogen or into some oxides.

Q.4. Which bacteria causes nitrification?
Ans: Nitrification is mainly carried out by certain nitrogen-fixing bacteria such as Azotobacter (freely in soil), Rhizobium (found in root nodules of leguminous plants), etc. These bacterias convert atmospheric nitrogen into water-soluble nitrates.

Q.5. What is the process of nitrification?
Ans: Nitrification occurs in mainly two steps: The conversion of Ammonia to Nitrite and Oxidation of Nitrate from Nitrite.

Q.6. Does nitrification require oxygen?
Ans: Yes, nitrification requires oxygen for the oxidation of ammonia to nitrites and then for nitrites to nitrates. These nitrates are then utilized by plants for growth and development..

Q.7. Name some denitrifying bacteria.
Ans: Pseudomonas denitrificans, Thiobacillus denitrificans, Micrococcus denitrificans are some common denitrifying bacteria.

Q.8. Name some Nitrifying bacteria.
Ans: Nitrosomonas, Nitrococcus convert ammonia into nitrite. Nitrobacter, Nitrocystis convert nitrite into nitrate.

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