Nitrate (NO3) & Nitrite (NO2)

Nitrogen is the most abundant element in Earth’s atmosphere, composing about 78% of the atmosphere. In the biotic factors (ie. living components) in any ecosystem, nitrogen serves as an essential component of amino acids, nucleic acids, proteins, chlorophyll, enzymes, and coenzymes. However, the majority of organisms, including all plants and animals, lack the ability to utilize atmospheric nitrogen gas (N2). This makes nitrogen the element in the shortest supply relative to the needs of the biotic factors existing within an ecosystem. 

So, if it’s so important, how do plants and animals find a way to consume nitrogen in usable ways? In essence, plants need ammonia (NH3) to build amino acids (animals get their amino acids from consuming the plants), yet they lack the ability to convert N2 to NH3. Therefore, they take up inorganic nitrogenous forms (No3- or NH4+) and convert this to NH3. The basics of this system are represented through the Nitrogen Cycle, or the biogeochemical cycle by which nitrogen circulates through atmosphere, marine, and terrestrial ecosystems in multiple chemical forms.

Nitrogen Fixation: During nitrogen fixation, bacteria turn N2 gas from the atmosphere into ammonia. Fixing bacteria utilize the enzyme “nitrogenase” to break apart the two N atoms and recombine the nitrogen with hydrogen, forming ammonia (NH3). Most nitrogen fixation occurs naturally in the soil by bacteria either living freely in soil or water, or existing symbiotically with plant roots. Because nitrogen fixing is a metabolic pathway, bacteria often require external sources of energy to complete the process. By inhabiting the root nodules of certain plants (more specifically, legumes), they receive the energy plants create via photosynthesis and use it to fix the nitrogen into usable forms. In essence, plants provide energy to bacteria in exchange for nitrogen that they can use to grow their own plant tissues. 

There also exist bacteria living freely in soil or water that have the capacity to fix nitrogen independently and release it into their surroundings. 

Additionally, a small amount of nitrogen can be fixed using the energy from lightning, forming NO and NO2. These forms of nitrogen then enter soils via rain or snow, and can be used in the next step of the nitrogen cycle. 

Humans have developed artificial fixing through the industrial process that creates fertilizer. By using high heat and pressure, atmospheric nitrogen and hydrogen are combined to form ammonia, which is processed further to form ammonium nitrate. Ammonium nitrate is a salt that can be added to soils and used by plants. While this form of nitrogen fixing may seem like a biological hack, excess nitrogen can have extremely negative effects on waterways and land.

Ammonification/mineralization: 

Mineralization is the process by which microorganisms in the soil convert organic forms of nitrogen, such as manure or dead organic material, are converted into inorganic forms of nitrogen that can be taken up by plants. Because all plants, save legumes, get their required nitrogen from the soil, it is essential that nitrogen is accessible in a useable form. 

Ammonification specifically refers to any chemical reaction in which the NH2 groups are converted into ammonia (NH3), which then reacts with water (H2O) in the soil to form ammonium (NH4) as an end product. This conversion is an essential metabolic pathway for many bacteria, as the directions provide them with the energy necessary to stay alive. When the ammonia produced by microorganisms is in excess, it becomes available within the soil to be assimilated by plants. One major benefit of ammonification is the recycling of nitrogen from a dead plant or animal back into a useful form rather than building up in the atmosphere or soil. 

Nitrification:

During nitrification, multiple soil bacteria turn the ammonia (NH3) produced during ammonification into nitrites (NO2-) and nitrates (NO3-). This is done in multiple steps requiring two different types of bacteria. First, soil bacteria convert ammonia into nitrogen dioxide, or nitrite (NO2-). Then, another type of soil bacteria adds a third oxygen to create nitrate (NO3-). It is important to note that while nitrites cannot be utilized by plants, they can be quickly converted into usable nitrates. By completing this process, the bacteria gain the energy from metabolizing nitrogen and oxygen.

Because nitrates are the usable form of nitrogen for plants, their availability is the limiting factor on plant growth. Common soil fertilizers are traditionally composed of nitrates, allowing farmers to bypass the necessity of nitrogen-fixing bacteria. 

Immobilization:

Immobilization, or reverse mineralization, is the process limiting the amount of nitrogen in the soil. When there is a limited amount of nitrogen available to microorganisms, they will actually turn to nitrogen in the form of nitrates (NO3-) or ammonium (NH4+) in the soil to power their own metabolic processes. Essentially this ties the nitrogen previously available to plant in the soil and ties it up in these microorganisms. While this may seem to be a limit on plant growth, it is a necessary balancing process in the Nitrogen Cycle to keep things in order. 

Denitrification:

Denitrification returns nitrogen to the atmosphere as nitrates (NO3-) are converted back into atmospheric nitrogen (N2) by bacteria. This prevents nitrogen from building up in soils to an unhealthy degree, maintaining balance within the system. 

So, while certain prokaryotic microbes can actually synthesize ammonia (NH3) and nitrate (NO3-) from N2, making it accessible to plants, others can turn NH3 and NO3 back into original atmospheric nitrogen. The balance of these two microbial pathways determines the accessibility of nitrogen to life-forms. 

Why do we test for Nitrates/Nitrites and not other forms of Nitrogen?

Since nitrites and nitrates are the most immediate and proximate forms of nitrogen utilized by plants, they are important indicators of ecosystem health in immediate relation to plants and soil composition. 

Impacts of excess Nitrates/Nitrites:

While nitrogen is essential to living organisms, there comes a point at which the environment cannot function safely. Through fertilizer runoff, leaky cesspools, sewage treatment plants, manure runoff, and car exhausts, nitrogen enters water sources with negative consequences. 

1. Disease: While nitrite is a relatively short-lived form of nitrogen that is quickly converted to nitrate, excess nitrites can produce a serious illness in fish for the short amount of time they exist. Known as “Brown Blood Disease”, nitrite enters the bloodstream of fish and combines with hemoglobin (on oxygen transporter in blood) to form methemoglobin, which cannot transport oxygen. Similarly, in human babies under 3 months of age, excess nitrogen will bond to hemoglobin in human blood and inhibit oxygen transport, known as “Blue Baby Disease”. 

2. Atmospheric pollutants: Excess atmospheric nitrogen can produce ammonia and ozone, two pollutants which limit visibility and affect plant growth (think back to the nitrogen cycle). When excess nitrogen comes back from the atmosphere, it can harm forest, waterway, and soil health.

3. Eutrophication: An abundance of nitrogen and other nutrients within a water source increases the amount of plant and algae growth, leading to algal blooms that block essential sunlight from reaching plants, and low oxygen (hypoxic) water that kills fish and other underwater habitats. When the excess algae dies and decomposes, it produces large amounts of carbon dioxide. This lowers the pH of water, and in estuaries and the ocean is known as ocean acidification. This puts strain on the fish, shellfish, and molluscs that survive the algal bloom.

Sources and Resources:

On NO2 & NO3:

• What is Eutrophication? from NOAA

• Nitrates in the Environment from Water Research Center

• Nitrogen and Water from USGS Water Science School

Additional Articles of Interest

• Corals Form Characteristic Associations with Symbiotic Nitrogen-Fixing Bacteria by Kimberley A. Lema, Bette L. Willis, David G. Bourne

• Nitrogen and Climate Change by Bruce A. Hungate et al.

• Nitrogen Excess in North American Ecosystems: Predisposing Factors, Ecosystem Responses, and Management Strategies by Mark E. Fenn et al.