Nitrification

Nitrification: Nitrification is a two steps process of oxidation of ammonium first to nitrite (nitritation) and then nitrate (nitratation) carried out by highly specified specialized autotrophic bacteria. So, nitrification is an oxidative process. In the first step, nitrosomonas (and nitrosococcus & to some extent by nitrosogoea, nitrosocystic) participates in the oxidation of ammonium to nitrite, whereas the second step, i.e. the oxidation of nitrite to nitrate is carried out by Nitrobactor. This type of nitrification is also known as autotrophic nitrification. However, recent literature reviews show that apart from autotrophic nitrification, the heterotrophic nitrification, carried out by heterotrophic bacteria and fungi, also takes place in soil. In this case, there is no formation of nitrite, i.e. ammonium is directly converted to nitrate. The heterotrophic process of nitrification is of little agronomic importance since only small amount of nitrate is formed through this process. The first step of reaction in nitrification is, Nitrosomas +

NO2- + H2O + 2H+ +66Kcal

NH4 + 1 ½ O2 Bacteria

This step is a direct oxidation not involving any cytochrome system or auxiliary carriers. The second step of oxidation of nitrite to nitrate by nitrobactor undoubtedly involves a cytochrome (respiratory enzyme) and their oxidation cannot take place by the addition of atmospheric oxygen to nitrate. It can be expressed in following way: NO2-.H2O + 2Cy+.Fe++ 2Cy+.Fe++ +2H+ + ½ O2

NO3- +2Cy+Fe2+ +2H+ 2Cy+.Fe+++ + H2O

The overall autotrophic process of nitrification can be shown simply as: NH3 H+ NH4+ H+

NO2nitrosomas

NO3nitrobactor

Pathway of nitrification: The six electron transfer accompanying the oxidation of + NH4 (oxidation state of -3) to NO2- (oxidation state of +3) by nitrosomas suggests of least two intermediates, the most likely candidates being hydroxylamine (NH2OH) and nitroxyl (NOH). NH4+ +1/2 O2 NH2OH NOH +1/2 O2 NO-2 -H+ -H+ Ammonium Hydroxyl amine Nitroxyl Nitrite Energy is released by the reaction (65Kcal/mol) is used by the organisms for carrying out its life activities. Several studies have indicated that N2O is a by-product + of NH4 oxidation. This gas may arise by chemical dismutation of nitroxyl (NOH) and through the action of nitric reductase. NH4+

2e-1

NH2OH 2e- [NOH]

2e-

NO2-

Chemical Dismutation

Denitrifying Nitrite reductase

N2O

The oxidation of NO2- to NO3- by nitrobactor involves a two-electron change in the oxidation state of N (from +3 to +5) with release of 17.8 Kcal/mole of energy: 2eNO2- + ½ O2 NO3Nitrite (+3)

nitrate (+5)

NO intermediates are suspect. The reaction is facilitated by an NO oxidase system with the elements being carried to O2 via a cytochrome system, with generation of ATP. 2

Why nitrite does not usually accumulate in soil? If do, then under what condition nitrite accumulate in soil? NO2- does not usually accumulate in soil but when it does, it can adversely affect plants and microorganisms. Ammonium also does not accumulate in soil. But, under same condition, it does .

----- Temperature is very low ----- Water saturated soil condition When NH4+ accumulates in soil , a portion of it is utilized by microorganisms and plants. Another portion undergoes volatilization. According to J.P Robertson nitrification is NH4+ limited; the amount of NH4+ accumulated, is immediately transformed to nitrate (NO3-) by nitrifiers. From this, we can say that microbial bacteria, although they are small in number, they are efficient (Nitrosomonas, nitrobactor, nitrosococcus, etc. bacteria convert NH4+ to NO3-). But when pH is above 9.0, then some nitrite can accumulate in soil. Thus, we can conclude that nitrifiers are much more efficient than ammonifiers. Which organism is more active in the transformation of nitrite to nitrate? Bacteria Bacteria---- 1500 μg/ml NO2—N or NO3--N Fungi------ 30 μg/ml NO3—N or NO3—N Chemical composition and functional group of humic substances: Elemental composition – C,O2, N, H mainly In humic acid rich in carbon 41-57%, H2 and n contents O2--------33-46% N2------2-5% In fulvic acid lower carbon content, H2 & N2 content is also lower. O2------44-54% N2-----07-2.6% Functional groups: 1. Carboxyl groups (acid groups) :R-COOH 2. Hydroxyl groups (total hydroxyl, phenolic OH-groups and alcoholic OH-group)-R-OH-R-O3. Carboxyl groups (ketonic or quinoid)- R=O-R-O-