Nitrogen – Biogeochemical Cycle

Nitrogen is uniquely positioned in nature’s biogeochemical cycle of elements because there are so many microorganisms that have the ability to alter forms of nitrogen. The existence of life has an absolute need for the many end products of nitrogen metabolism by microorganisms;

however, there are many problems that also result. A major problem occurs when denitrifying bacteria remove fertilizing nitrates from the soil. Another problem occurs where nitrite is used as a corrosion inhibitor and microorganisms convert it to nitrate. Undesirable transformations occur in natural and man-made environments, for example, waste treatment processes producing unpleasant odors, corrosive gases, and pH modifications. Many products of nitrogen transformations are very corrosive to metals and concrete. All of these are of interest. The following steps provide a summary of these microorganisms and the transformations they have the ability to perform.

Step 1. Fixation of Molecular Nitrogen

The natural reservoir for the N2 in our environment is the atmosphere. No substantial amounts

are found in geological deposits. Atmospheric N2 is not readily available to biological systems,

Nitrogen fixation is a process that requires a substantial amount of energy input. Small amounts of combined nitrogen are introduced to the ecosphere by volcanic activity, by atmospheric lightning discharges, and ionizing radiation. Chemical fixation of molecular N2 by man (Haber-

Bosch synthesis) is limited but assuming an increasingly important role in the global nitrogen cycle.

The primary mechanism of biogeochemical cycling of the element Nis highly dependent on the activities of microorganisms. The first step involves nitrogen fixation by a very specific group of bacteria, usually found in soil environments (rhizosphere) or in aquatic environments (fresh or seawater). These bacteria include Rhizobium sp. (which accounts for the largest contribution of combined N2) in terrestrial habitats. In aquatic habitats, cyanobacteria (blue-green algae) such as Anabena sp. and Nostoc sp. are the most important in nitrogen fixation processes.

Step 2. Ammonification

The first biologically active compound resulting from N2 fixation is the formation of organic

nitrogen (NH2), which is immediately converted to NH3 and incorporated into amino acids and

other nitrogen-containing biochemicals, usually essential to microbiological metabolism and subsequently to the metabolism of higher life forms. Many microorganisms, plants, and animals are capable of ammonification through the reaction:

2(NH2) + CO + H2O + urease enzyme system Æ 2NH3 + CO2

The incorporated amino group can be transferred through transamination to form other amino acids, proteins, and other nitrogen-containing compounds by one group of organisms, and is subsequently used as sources of carbon, nitrogen, for synthesis and energy by other groups of organisms.

Free ammonia in solution, or as a gas, will exist as a byproduct of this process. The amount depends on the ability of the environment to contain or trap the ammonia. When the environment is favorable for containment, many of the potential problems mentioned earlier can take place. Step 3. Nitrification/Nitrosification

Nitrification is a process performed by autotrophic/chemolithotrophic bacteria that utilize nitrite (NO2) or ammonia (nitrosification) as an energy source, and CO2 as the primary carbon source.

The reaction is:

Nitrite-oxidizing bacteria (nitrifying) Æ NO2 + ½ O2 Æ NO3-

• Nitrobacter sp. (also heterotrophic) • Nitrospina sp.

Chemistry-Related Problems in Closed Cooling Water Systems

Both nitrite- and ammonia-oxidizing groups are frequently called nitrifying bacteria. These microorganisms live in a wide range of environments and often show a tendency to attach to surfaces (sessile) forming tight clusters of cells commonly called cysts, and some produce biofilm. They are aerobic or micro-aerophilic bacteria.

Ammonia-Oxidizing Bacteria (nitrosifying bacteria)

Ammonia-oxidizing bacteria in contained environments (for example, CCW systems with minimal amounts of O2)can produce either alkaline or acidic byproducts that contribute to

increased potential for MIC, especially with the corrosion of non-ferrous metals such as copper alloys.

Nitrosomonas sp. Nitrosovibrio sp.

→ NH3 + O2 → HNO3 Acidic byproducts

Nitrosococcus sp. → NH3 (low O2) → NO2 Neutral/weak acidic byproducts Nitrosomonas sp.

Nitrosopira sp. Nitrosolobus sp.

→ NH3 + O2 → NO/NH2OH Alkaline byproducts

Step 4. Nitrate Reduction/Denitrification

This process includes at least three different metabolic pathways. The microorganisms that are involved with these reactions are chemosynthetic/heterotrophic (require an organic energy source). They are diversified by the characteristic that some are aerobic, micro-aerophilic, facultatively anaerobic, or anaerobic and, thus, have optional metabolism pathways. The metabolic pathways are defined by the chemical metabolites produced. Microorganisms often associated with MIC assimilate nitrate nitrogen to organic nitrogen/ammonia. These include (those in bold print are significant contributors to MIC):

Aeromonas sp. Edwardsiella sp.

Arthrobacter sp. Clostridium sp.

Enterobacter sp. Escherichia sp.

Assimilation of Nitrate Nitrogen

A metabolic pathway is: Bacteria Æ NO3 + metalloprotein/reduced cofactors/catalysts Æ NO2 Æ

NO intermediates (?) Æ NH2OH Æ NH3 Æ amino acids Æ proteins/ATP Æ microbiological

metabolism.

This pathway is always associated with a significant increase in the amount of biomass produced and might result in biofouling problems. This pathway is also very common with metabolism of yeast, fungi, and all higher plants.

Dissimilation of Nitrate Nitrogen

A metabolic pathway is: Bacteria Æ NO3 + reductase enzyme catalyzed reduction Æ NO2 Æ ?

and possibly ammonification.

This is a pathway that can occur under both aerobic and anaerobic conditions. It is an alternative pathway to O2 respiration for some microorganisms. In most cases, this process is associated

with an increase in NO2 concentration, often NH3, and biomass. Although not directly involved

with corrosion, the metabolic pathway increases the potential for MIC. Typical microflora include: Aeromonas sp. Enterobacter sp. Arthrobacter sp. Eschericia sp. Bacillus sp. Micrococcus sp. Citrobacter sp. Nocardia sp. Denitrifying Bacteria

These microorganisms are responsible for the actual removal of nitrogen compounds, as such, from the microenvironment. The byproducts of this metabolic pathway are usually metabolites for other pathways or a release of gas that contributes to gassing problems in closed loop cooling systems. The process is essentially an anaerobic function at a site with excessive deposition of nitrate sludge and other organic biomass, usually in a system where nitrite oxidation by

microorganisms had occurred for an extended time. The gassing problem can be justification for bleed and feed or flush and fill procedures. Denitrification does not directly increase the potential for corrosion (MIC); however, it does increase difficulties in maintaining stable operating and control conditions.

A metabolic pathway is:

Chemistry-Related Problems in Closed Cooling Water Systems

Typical microflora include:

Alcaligenes sp. Hyphomicrobium sp.

Bacillus sp. Pseudomonas sp.

Nitrosomonas sp. Thiobacillus sp.

Step 5. Recycling and/or Short Cuts

It is important to remember that this biogeochemical cycle is a continuously recycling process. It does not typically reach an endpoint. Some stages are limited by the accumulation of byproducts that either affect the metabolism of the microorganisms or change the microenvironment. It is a dynamic process where several stages of the cycle can occur simultaneously. Several of the stages are reversible and there are short cut pathways that bypass the dominant pathways. The recycling process ensures that all living organisms are supplied with a form of nitrogen necessary for their existence. This is particularly significant when one considers that the primary nitrogen reservoir exists as a non-metabolizable compound, N2.

This diversity contributes to potential problems in both terrestrial and aqueous environments. The diversity has provided situations where great benefits are obtained but, when not managed, the nitrogen biogeochemical cycle can contribute to problems such as environmental pollution, MIC, plugging and fouling, biofouling, and loss of heat transfer in cooling water systems.

Suggested Reading

Microbial Ecology, Fundamentals and Applications – Fourth Ed.. By R. M. Atlas and R. Bartha,

Chapter 11, Biogeochemical Cycling: Nitrogen, Sulfur, Phosphorus, Iron, and Other Elements. Pub. Benjamin/Cumming Science. Imprint - Addison Wesley Longman. Inc., Menlo Park, CA, 1998.