Biological Nitrogen Fixation: How It Works

Many forms of biological nitrogen fixation occur in the soil. We're providing a layman's guide to this essential part of soil building!

An image of root nodules on legumes as a way of nitrogen fixation in the soil


Nitrogen is an essential nutrient that supports plant growth, and nitrogen fixation is one-way plants obtain it to grow. Nitrogen-fixing bacteria in the soil take nitrogen (N) gas and turn it into a usable form for plants. These forms come in several types, but symbiotic nitrogen fixation is the most common. This is the relationship legumes have with bacteria. 

Legumes are an important part of crop rotation and soil building. Because of their relationship with nitrogen-fixing bacteria, they add nitrogen for future crops in a rotation. If you’re curious about crop rotation, consider reading our article on that topic!

This article should give you an idea of nitrogen as a plant nutrient, information on nitrogen dynamics, and an understanding of the types of nitrogen present in the soil. Understanding biological nitrogen fixation helps you understand plant-microbe interactions. Understanding how fixed nitrogen works also helps gardeners understand how to utilize certain plants in a rotation. 

Nitrogen As A Plant Nutrient

Nitrogen is a critical component of plant growth. In general, plants need nitrogen in the greatest amount compared to other nutrients. In plants, nitrogen is used for chlorophyll. Chlorophyll is what makes plants green. It resides within photosynthetic chloroplasts. Nitrogen is critical in amino acids, which serve as the building blocks of proteins. 

Why are amino acids so important for plant physiology? Well, they produce chlorophyll which ties directly into photosynthesis. When a nitrogen-fixing microorganism has a healthy relationship with a plant, a plant-derived membrane called the thylakoid membrane gives chloroplasts an easier time absorbing light. Better light absorption leads to better water and plant nutrition uptake. 

Forms of Nitrogen 

There are several forms of nitrogen in the plant world. The two forms utilized by plants are nitrate and ammonium. Nitrate is a negatively charged nitrogenous compound composed of one nitrogen atom and three oxygen atoms. A positively charged compound with one atom of nitrogen and four atoms of hydrogen form ammonium. If plants have a choice between nitrate and ammonium, they prefer nitrate. However, they are both acceptable forms for plants in the nitrogen economy. 

There are a few other important forms of nitrogen involved with plants and the nitrogen cycle. Dinitrogen gas makes up 78% of the air around us. It’s made of two nitrogen atoms triple bonded together. These bonds are extremely hard to break. Another important gaseous nitrogenous compound is ammonia. One nitrogen and three hydrogen atoms form ammonia. This toxic gas is a part of the N cycle and can accumulate in certain agricultural settings. A final important nitrogenous compound to consider is dissolved organic nitrogen. These carbon-containing compounds are organic acids found in soils. 

The Nitrogen Cycle

The nitrogen cycle illustrated in a visual form
Nitrogen cycling happens in many forms. Source: University of Delaware

Before getting into nitrogen fixation as a whole, it is important to understand the basics of the N cycle. There are several parts to this cycle, and we will be covering the basics in an effort to give you an understanding of how nitrogen changes in this process. The components of the cycle include the soil, atmosphere, and living tissues. The easiest way of discussing this topic is by examining how nitrogen enters and exits the soil. 

Nitrogen Entering Soil

Nitrogen enters the soil via decomposition of organic matter. This can occur when gardeners add compost, decaying organisms, decomposing plant material, manure, and others to their gardens. Another way is through nitrogen fixation in the soil itself. This process entails specific nitrogen-fixing bacteria taking in dinitrogen gas and turning it into forms plants can utilize. There are three different types of nitrogen fixation: nitrogen-fixing symbiosis, heterotrophic fixation, and associative fixation). They all have the ability to break the triple bond in dinitrogen gas and, in the process, deposit nitrogen in the soil.  

Nitrogen Leaving Soil

Denitrification, ammonia volatilization, and leaching or runoff are the major ways nitrogen leaves the soil. Denitrification is the process by which nitrate in the soil is transformed into gas by anaerobic bacteria. Low oxygen concentrations create anaerobic conditions. Decomposable organic matter, nitrate, and warm temperatures conditions are also needed for denitrification. In this process, dinitrogen gas and dinitrogen monoxide are released into the air as atmospheric nitrogen. 

Dinitrogen monoxide (made of two nitrogen and one oxygen atom) is a greenhouse gas released in a much lower concentration than dinitrogen gas, which is not a greenhouse gas. Factors affecting the amount of dinitrogen monoxide released include soil pH and temperature. 

Another form of lost nitrogen occurs through its volatilization into ammonia gas. This happens when soils are dry, warm, and have a low cation exchange capacity – the soil’s capacity to hold positively-charged ions. The result of these conditions is the application of ammonium to the soil surface. 

One example of the volatilization process can be found in urea. Urea is a common organic form of N fertilizer used worldwide and is a byproduct of urine from humans or other animals. It can often lead to the volatilization of ammonia gas when conditions are right. As the ammonia vaporizes and rises into the atmosphere, it leaves the soil lacking the ammonia-based nitrogen compounds that had been added via the urea. Think of it as if it were a hot-air balloon whisking our nitrogen right out of the soil where it belongs!

A visual representation of ammonia volatilization
Imagine ammonia volatilization as the nitrogen being carried off into the sky. Source: University of Missouri Extension

Leaching and runoff are two other ways nitrogen leaves the soil, especially when it comes to cropping systems that receive regular chemical fertilizers. Often nitrogen is not sequestered in the soil well, especially as nitrate. It moves easily through the soil profile and eventually ends up in groundwater, which flows underground. The nitrogen then ends up in rivers, streams, and other bodies of water. 

This nutrient pollution causes eutrophication or a buildup of nutrient concentration in wetlands and waterways. Just as nitrogenous fertilizers promote growth in crops, their runoff causes excessive growth of plants in these areas, leading to limits in the amount of oxygen available to other organisms.  

Dead zones are another result of runoff, as algae blooms develop, removing oxygen from wildlife in oceans.

Nitrogen Immobilization

Another important thing to consider with nitrogen is immobilization or nitrogen that is unavailable to plants, primarily because the nitrogen is found in the tissues of free-living bacteria in the soil. Immobilization can occur more when compost and amendments add too much readily available carbon. Carbon serves as energy for microorganisms in the soil. These organisms use up nitrogen in the soil for tissues and proteins. Consider this when adding straw and wood mulches into a garden. As these are mostly carbon with little nitrogen, they can lead to immobilization. 

Gardeners need to add more N fertilizer and N fixing plants into the garden to counter the immobilization cycle. Compost can also lead to immobilization when it does not have the correct combined nitrogen to carbon ratio. An excess of nitrogen in the form of nitrate is marked by odors emitted from a compost pile. To rebalance the ratio and promote better ammonia assimilation add carbon. 

How Bacteria Fix Nitrogen

N gas gets “fixed” into the soil by rhizobial bacteria, but how? The triple bonds within dinitrogen are incredibly strong, making them hard to break. These nitrogen-fixing organisms utilize an enzyme, nitrogenase, to break this bond. The nitrogenase enzyme is found in rhizobial bacteria and nitrogen-fixing cyanobacteria. The nitrogenase complex turns the dinitrogen into ammonia, then reactions turn it into usable forms for the crop plant. 

Humans figured out how to break this triple bond at the start of the 20th century via the Haber bosch process. This industrialized method of taking in atmospheric nitrogen gas and transforming it into usable forms for plants alleviated the strain of procuring nitrogen fertilizers. Therefore, fertilizers made from this process come from inorganic compounds and cannot be used on certified organic farms. 

Symbiotic Nitrogen Fixation

The most common form of fixation is symbiotic nitrogen fixation. This is the relationship legumes and actinorhizal plants have with nitrogen-fixing bacteria in the soil. Crops that support N-fixing bacteria include beans, peas, peanuts, clover, vetch, alfalfa, and lupines. Some other species are leguminous but include some trees and shrubs. 

Most land plants do not have this symbiosis with bacteria. These host plants form root nodules that contain nitrogen-fixing microorganisms, including plant growth-promoting rhizobacteria. Symbiotic relationships benefit both parties: the rhizobium species receive sugars while the plant gets usable nitrogen compounds. When you apply N fertilizer, these nodules do not form because the plants do not need the help of nitrogen-fixing bacteria to supply nitrogen to plant mitochondria. 

Each legume species differs in the level of nodulation and efficiency. Common beans like green beans are not as good as grain legumes like peanuts, cowpeas, and soybeans. Perennials are even better at affixing nitrogen. These crops include clovers and alfalfa. There are many options for such plants that affix nitrogen. 

Nodule Formation

Root nodules on peanut
Pink and red coloring in these peanut roots indicate active nitrogen fixation. Source: Texas A&M Agrilife

Different crops have differently shaped nodules. The nodule formation process is actually an infection of N-fixing bacteria. Annual crops have pea-size nodules while perennials have more elongated nodules. But how do these nodules form? First, the plant-associated bacteria invades the plant host. Bacteria living in the soil enter the plant cells, residing within the root cortex. The bacteria remain within the intracellular region of plant tissues, eventually forming the nodules you see.  

I recommend gardeners go into the garden and pull up a mature nitrogen-fixing plant. You will see these nodules. Cut one open to see the inside. Active nodules appear red on the inside because of a compound similar to hemoglobin in human blood.

Heterotrophic Nitrogen Fixation

Heterotrophic nitrogen-fixing bacteria are unlike symbiotic ones. They do not have a relationship with plants to get carbon and other compounds they need. Instead, they support nitrogen fixation passively by consuming decaying matter in soil. 

This study conducted by Eckford and a team of researchers identified several heterotrophic bacteria within Antarctic soils containing hydrocarbon fuels. The study also emphasizes that nitrogen fixation occurs in soils all over the world. 

These bacteria gain energy by consuming other compounds in the soil, making them heterotrophic. Humans are heterotrophic because we must produce and consume food to survive. Plants, algae, and other photosynthetic or chemosynthetic organisms are autotrophs because they can produce the food necessary for survival within themselves. 

Associative Nitrogen Fixation

Associative fixation is similar to the other types in the sense that dinitrogen gas is fixed into the soil, but in a casual relationship with plants. While symbiosis involves bacteria living within plant tissues, association refers to bacteria within the plant microbiome. They are free-living soil bacteria that don’t rely on plants to do their work.

In this study, Roley and others examined the potential relationship bacteria have with switchgrass. They found that the perennial grass was often unresponsive to nitrogen fertilizer. They examined the soil surrounding roots to see the activity of bacteria. They found that this crop benefits from bacteria in the soil having an association with the plant to supply nitrogen while receiving carbon from the surrounding environment. 

How to Use Nitrogen Fixing Crops in Rotation

The best way to use nitrogen-fixing plants is to include them in a rotation. Food crops use varying amounts of nitrogen. Most are heavy nitrogen feeders, like sweet corn, pumpkins, squash, and peppers. Most plants within the garden and on the farm benefit from rotating nitrogen fixers. Nitrogen fixation takes place when gardeners plant nitrogen-fixing species of plants before planting heavy feeders. Doing so alongside annual crops can be beneficial for plant soil too. 

Interplanting legumes with the other crops can benefit both plants. It is important to note that nitrogen-fixing plants will only provide so much nitrogen to other plants when they are alive. It is still beneficial to plant them together, but legume nodules may not be capable of supplying all the nitrogen to another crop like peppers or tomatoes. 

When using nitrogen fixers before heavy feeders, remember that the plant roots (and above-ground tissues) should remain in the garden. Removing the plants will essentially take away the fixed nitrogen done by bacteria. Therefore, chopping and dropping or cutting down a nitrogen fixer at the end of its life will be more beneficial than pulling it up by the root. 

Consider picking crops that have the capability to provide more nitrogen fixed. Common beans provide less adequate nitrogen fixation than something like soybeans or peanuts. While nitrogen-fixing cereals like soybeans are not typically used in the garden, including edamame, fava, or peanuts could benefit the soil. Also including cereal crops and plants without nitrogen fixation genes in general in the rotation can help build organic matter and benefit the garden. 

Frequently Asked Questions

An image of root nodules on legumes as a way of nitrogen fixation in the soil
Root nodules on legumes store nitrogen in the soil. Source: DutraElliott

Q: What are nitrogen-fixing plants?

A: Nitrogen-fixing plants are those that act as a host plant for specific types of bacteria. The host plant itself is not fixing the nitrogen. Rather, the bacteria are in a symbiotic relationship with the plant species. 

Q: What trees fix nitrogen?

A: There are many species of tree known to affix nitrogen. Some of them include autumn olive, black locust, eastern redbud, and alder. 

Q: What vegetables are nitrogen-fixers?

A: Common beans affix nitrogen (although not as effectively as others). Common crop plants include green beans, black beans, and pinto beans. As well as peas like sugar snap and snow peas. Again, if you want to add adequate amounts of nitrogen to the soil, including better crops like peanuts or edamame could be good options. Edamame is similar to grain soybeans and does a better job of affixing nitrogen than common beans. Most other vegetables, like tomatoes, pepper, corn, eggplant, and squash, are not nitrogen fixers. 

Q: What flowers fix nitrogen?

A: Lupine is the most noteworthy flower that fixes nitrogen. Hyacinth beans are legumes that produce beautiful flowers. They are frequently used as ornamental plants. 

Q: Which plants fix the most nitrogen?

A: Perennial legumes like alfalfa and clover have the potential to fix the most nitrogen. However, grain legume plants like peanuts, fava beans, soybeans, and cowpeas also do a  good job of affixing nitrogen. Common beans like green beans and dried beans are not as good at fixing nitrogen. 

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