Nitrogen cycle

Nitrogen

Biological availability of N, P and K is of considerable economic importance, since they are the major plant nutrients derived from the soil. Of the three, N stands out as the most susceptible one to microbial transformations. This element is the key building block of the protein molecule upon which all life is based, it is an indispensable component of the protoplasm of plants, animals and microorganism.

Molecular N₂ constitutes about 78% of the earth’s atmosphere but it is chemically inert and cannot be utilized by more living organism, plant animals and microorganisms therefore depend on a source of combined N such as ammonia, nitrate or organic N compounds for their growth.

Nitrogen undergoes a number of transformations that involve organic, inorganic and volatile forms of nitrogen. A small part of the large reservoir of N₂ in the atmosphere is converted to organic compounds by certain free living microorganisms or by plant microbe association that makes the element available to plant growth. The nitrogen present in the proteins or nucleic acids of plant tissue is used by animals. In the animal body, the N is converted to other simple and complex compounds. Upon the death, plants and animals undergo microbial decay and organic N is released as ammonium, which is then utilized by vegetation or is oxidized to nitrate by microorganisms. The nitrate from of N is mostly used by the plants or may be lost by bacteria reduced to gaseous N₂, which escapes to atmosphere, there by completing the cycle. The Nitrogen cycle mainly includes transformations such as

Nitrogen mineralization : In which N containing organic complexes are decomposed and converted into inorganic compounds for use by plants
N immobilization : In which N containing inorganic compounds are assimilated

N₂ in the atmosphere is acted on by certain microorganisms sometimes in symbiosis with a higher plant, which can use it is as a N source for growth. This process nitrogen fixation, results in the accumulation of new organic compounds in the cells of responsible microorganisms. The N₂ thus fixed reenters general circulation when the newly formed cells are in turn mineralized.

By means of these reactions the subterranean microflora regulates the supply and governs the availability and chemical nature of N in soil.

The Nitrogen cycle involves various major processes

Ammonification
Nitrification
Denitrification
Nitrogen fixation

in which ammonification is the process of mineralization

I. Nitrogen mineralization

The conversion of organic N to the more mobile, inorganic state is known as nitrogen mineralization. As a consequence of mineralization, ammonium is generated and organic N disappears. This takes place in two distinct microbiological steps.

Ammonification

It is the process of mineralization in which proteins, nucleic acids and other organic components are degraded by microorganisms with the eventual liberation of ammonia. This is called ammonification. A part of the liberated ammonia is assimilated by the microorganisms themselves. Ammonification occurs by two step process.

A. Proteolysis

B. Deamination

A. Proteolysis : Proteolysis the process of enzymatic breakdown/hydrolysis of proteins ,nucleic acids and other organic nitrogenous compounds into amino acids by the microorganisms with the help of proteolytic enzymes . The breakdown of proteins is completed in two stages:

I. Proteins are converted into peptides or polypeptides by enzyme proteinases and

II. Polypeptides / Peptides are further broken down into amino acids by the enzyme peptidases.

Proteins → Peptides → Amino Acids

Proteinases Peptidases

The most active microorganisms responsible for elaborating the proteolytic enzymes (Proteinases and Peptidases) are Pseudomonas, Bacillus, Proteus, Clostridium histolyticum, Micrococcus, Alternaria, Penicillium etc.,

B. Deamination: After Proteolysis, the amino compounds are delaminated to yield ammonia with the activity of deaminase enzymes.

CH₃-CHNH₂-COOH + Alanine + ½ O₂→ CH₃COCOOH + Pyruvic acid + NH₃

deaminase

Break down of nitrogenous substance is brought about by the activity of a multitude of microbial species. Almost all bacteria, actinomycetes and fungi can bring about proteolysis and the amino acids produced are utilized for the growth of these organisms.

Putrefaction: Ammonification usually occurs under aerobic conditions, while under anaerobic conditions protein decomposition leads to conversion of ammonia into amines and related compounds (eg) Clostridium. The anaerobic decomposition of protein called as putrefaction. These amines are subsequently oxidized in the presence of O₂ to release ammonia.

Nitrification

The biological oxidation of ammonium salts (in soil) to nitrites and the subsequent oxidation of nitrites to nitrates is called as nitrification. i.e. the biological conversion of N in soil from a reduced to a more oxidized state, called nitrification. Soil conditions such as well aerated soils rich in calcium carbonate, a temperature below 30 ° C, neutral pH and less organic matter are favorable for nitrification in soil.

Nitrification is a two stage process and each stage is performed by a different group of bacteria.

Process of conversion of Ammonia into nitrate, called as Nitrosification

2 NH₃ + 1½ H₂O₂ → NO₂^- + 2H+H₂O-Nitrosofication

This process has been carried out by Chemoautotrophic Nitrifiers (Ammonia oxidizing bacteria)-Nitrosomonas, Nitrosolobus, Nitrosococus, Nitrosospira. These bacteria obtain their energy requirement by the oxidation of NH₄⁺ to NO^-₂. Among the nitrifiersNitrosomonas are more prevalent in soils.

Heterotrophic Nitrifiers are also involved in the process. -Streptomyces, Nocardia, Arthrobacter, Aerobacter.

In second step,Nitrite is further oxidized to nitrate

HNO₂ + ½ O₂ → HNO₃.

Organisms :Nitrobacter, Nirococcusmobilis, Aspergillus, Penicillium, Cephalosporium

Factors influencing the growth of nitrifying bacteria in soil:

Levels of ammonia and nitrite, aeration, moisture, temperature, pH and organic matter. In acid soils – nitrification is poor. In Waterlogged soils – deficient in O₂ – not congenial for nitrification.
When N mineralization is low or N immobilization is high nitrification rates will be low.
All nitrifiers are aerobic organisms, occur only in thin aerobic layer in submerged soil
Nitrification inhibitors: commercial compounds such as pyridines, pyrimidines, amino triazoles, and sulfur compounds like ammonium thiosulfate will inhibit the activity of nitrifying bacteria
Neem oil slows down the nitrification process
Nitrogen is lost from ecosystems mainly after its conversion to NO₃ and prior to plant uptake, so keeping N in the NH₄ form keeps it from being lost by nitrate leaching and denitrification, the two principal pathways of unintentional N loss in most ecosystems.

Denitrification

The convention of nitrate and nitrite into molecular N₂ or nitrous oxide through microbial processes is known as denitrification. Certain bacteria are capable of using nitrate as the terminal electron acceptor under anaerobic conditions. This is called nitrate respiration. i.e. anaerbic conversion of nitrate into molecular nitrogen. As a consequence of nitrate respiration, NO₃ is reduced to N₂ gas or nitrous oxide. Denitirifcation leads to the loss of N from the soil. It depletes N, and therefore it is not a desirable reaction.

Denitirfication occur mostly in waterlogged anaerobic soils with a high organic matter contents. Denitrification of bound nitrogen to gaseous N is mediated by numerous species of bacteria, which use nitrates and nitrites as terminal electron accepter under anaerobic condition.

Bacterial genera which bring about denitrification Pseudomonas, Achromobacter, Bacillus, Micrococcus

2NO^-₃ +10 H → N₂ + 4H₂O+ 2OH^- (or)

2NO^-₂ +6 H → N₂ +2H₂O +2OH^- (or)

N₂O + 2H → N₂ + H₂O

Since nitrates are used as a source of electron acceptor, there is a net loss of N from soil. This process is termed also as dissimilatory nitrate reduction.

Many soil bacteria like Thiobacillus denitrificans oxidizes (chemoautotrophically) and also reduce nitrate to nitrogen.

5S + 6 KNO₃ + 2 H₂O → 3N₂ + K₂SO₄ + 4KHSO₄ (or)

5 K₂S₂O₃ + 8 KNO₃ + H₂O → 4N₂ + 9 K₂SO₄ + H₂SO₄

General pathway of denitrification:

Nitrate is first reduced to nitrite, which is then transformed to nitrous oxide (NO). The nitrous oxide is converted to N₂ with N₂O as an intermediate.

2 HNO₃ → 2HNO₂ → 2 NO → N₂O → N₂

The enzymes involved in the above sequence are 1. Nitrate reductase; 2. Nitrite reductase ; 3. Nitric oxide reductase; 4. Nitrous oxide reductase

Advantages of Denitrification process:

Fallow soils flooded with water are more congenial for denitrification than well drained and continuously cropped soils.
Though it is an undesirable reaction in point of view of plant nutrition, but have ecological importance. Because without denitrification the supply of N on the earth would have got depleted and NO₃ would have been accumulated.
High concentration of NO₃ are toxic, denitrification is a mechanism by which some of the N is released back to the atmosphere.
From a management perspective, denitrification is advantageous when it is desirable to remove excess NO₃from soil prior to its movement to ground or surface waters.
Sewage treatment often aims to remove N from waste streams by managing nitrification and denitrification.

Nitrate reduction

Several heterotrophic bacteria (E. coli, Azospirillum) are capable of converting nitrates to nitrites and nitrites to ammonia. Complete reversal of nitrification is known as nitrate reduction. Nitrate reduction normally occurs under anaerobic soil conditions (water logged soils) and the overall process is as follows:

HNO₃ + Nitrate + 4 H₂→NH₄ + 3H₂0

Nitrate reduction leading to production of ammonia is called “assimilatory nitrate reduction" since some of the microorganisms assimilate ammonium, through which they synthesis of proteins and amino acid this process is called as assimilatory nitrate reduction.

Nitrogen fixation

Nitrogen gas (N₂) is the most stable form of N and is a major reservoir for N on Earth. However, only a relatively small number of prokaryotes are able to use N₂ as a cellular N source by nitrogen fixation. N₂ + 8H → 2NH₃ + H₂. The N recycled on Earth is mostly already “fixed N”; that is, N in combination with other elements, such as in ammonia (NH₃) or nitrate (NO₃). In many environments, however, the short supply of fixed N puts a premium on biological nitrogen fixation, and in these habitats, nitrogen-fixing bacteria flourish. The example organisms are Azotobacter, Cyanobacteria, Clostridium, purple and green phototrophic bacteria, Methanobacterium (Archaea), Rhizobium, Bradyrhizobium, Frankia, Azospirillum.

Nitrogen immobilization

The process of microbial assimilation of inorganic nitrogen is referred as immobilization. In contrast to mineralization, microbial immobilization leads to the biosynthesis of the complex molecules of microbial protoplasm from ammonium and nitrate. Immobilization results in a marked reduction of nitrogen availability to the plants.

The mineralization of organic N and the microbial assimilation of inorganic ions proceeds simultaneously. Both mineralization and immobilization takes place regardless of the % of N in the organic N in organic matter. On the death of microorganisms, the immobilized N is however released through mineralization. It is also a loss of nitrogen. NO₃ when accumulated in microbial protoplasm it is also referred as assimilatory NO₃ reduction.

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