The Alchemy of Nitrogen-Fixation

By Kevin Hancock

With an ever-increasing world population comes an increased demand for food, fuel, and fiber. Land, water, and energy resources are becoming scarcer. Nitrogen is abundant worldwide, and is needed for the growth of most plant species. The majority of the world’s nitrogen is in the gaseous form, which cannot be utilized by most plants. This means that most plants must rely on additions of synthetic fertilizers to supply the needed nutrients.

There are very few plant species that are capable of fixing atmospheric, N2 gas, converting it into a usable form like ammonia, and storing it in root tissue. These plants are referred to as nitrogen-fixing. Symbiotic nitrogen-fixation (snf), which occurs naturally in some leguminous crops, can play a vital role in transforming atmospheric nitrogen gas into ammonia that can be utilized by these plants.

Biological reduction to ammonia can only be performed by prokaryotes and is a highly oxygen-sensitive process. Symbiotic interactions between prokaryote partners occur in two groups of soil bacteria — rhizobia in symbioses in legumes and Frankia bacteria in actinorhizal symbioses.

Snf is highly important in the production of biofuel feedstocks. Many current plants which produce abundant amounts of biofuels such as oil palm, canola, and corn are not nitrogen-fixers and consequently they rely on inorganic nitrogen fertilizers. Every step in the production, delivery and application of nitrogen fertilizer requires fossil fuels. Even though the formation of fossil fuels occurs naturally through anerobic decomposition of plants and animals, they are not considered renewable sources of energy.  Decomposition takes millions of years to form large enough quantities of fossil fuels. Those reserves are being depleted at a much higher rate than they are being formed.

The problem with the use of synthetic fertilizers is that plants only absorb a small percentage of applied fertilizer at any one time. The remainder of the applied fertilizer (30-50%) is subject to runoff, volatilization, and are leached beyond the root zone or denitrified. In many areas this can create algae blooms and eutrophication – a condition of high concentration of nutrients, but low oxygen levels. For example, Lake Okeechobee in Florida is experiencing this due to nutrient runoff from adjacent croplands.

Snf reduces the plant’s dependence on inorganic nitrogen sources and can provide a substitute for nitrogen fertilizers, thus reducing costs and helping the environment at the same time. Biological nitrogen fixation has been estimated to produce approximately 200 million tons of nitrogen annually.

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Leguminous root nodules – PC: NMSU

It would be very beneficial to humanity as well as the environment if all agriculturally important plants were capable of fixing atmospheric nitrogen.  Although a lot is still unknown, a lot of work has been conducted to better understand the intricacies involved in symbiotic nitrogen fixation. Nitrogen-fixation is composed of 3 components; first, the formation of nodules which provide the correct environment for nitrogen-fixing bacteria; second, the regulation of nodule numbers by both internal and external factors, and third, the actual conversion of atmospheric nitrogen into ammonia by the invading bacteria using the nitrogenase enzyme complex.

Nitrogen-fixing plants are not capable of extracting N2 gas directly from the atmosphere, they work in concert with common soil bacteria called Rhizobium. Rhizobia attached to root hairs, induce a pronounced curling of root hair cells. The root hair becomes deformed and the bacteria enter the plant by a newly formed infection thread growing through it. At this same time, cortical root cells are mitotically activated giving rise to the nodule primordium. Infection threads will grow towards the primordium and the bacteria are released into the cytoplasm of the host cells. The bacteria become encapsulated in the small compartment formed by the curling. The bacteria enter the plant’s root system and form nodules along the root pathway. The plant supplies all the essential nutrients as well as energy to the bacteria. Within a week after infection, nodules will become visible by the naked eye. Under field studies, nodules appear within 2-3 weeks. The nodules allow the plant to absorb the N2 gas that is present in the soil, and the plant converts it into ammonia that enters into a biochemical pathway producing both organic and inorganic forms before reverting it to N2 gas. Nitrogen-fixing bacteria need high calcium levels to work efficiently. Three micro-metabolic elements, iron, molybdenum, and cobalt are essential for the nitrogen-fixing process in bacteria.

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Nodule Formation Cycle in Pea Plant – PC: Pearson Education

Most legumes form symbiotic relationships with a select few Rhizobium, however Pongamia pinnata is able to produce snf relationships with various strains of Rhizobia as well as Bradyrhizobium. In areas of India the results clearly demonstrate the major advantage of the leguminous nature of Pongamia when compared to the Jatropha tree, another plant feedstock being evaluated as a source of biofuel energy.

Since the presence of oxygen can inactivate the process of nitrogen-fixing, it is important to know that legumes can regulate the gas permeability in their nodules allowing enough oxygen to maintain respiration without deactivating the nitrogenase enzyme. Nodules contain a heme protein called leghemoglobin. Leghemoglobin is present in the cytoplasm of infected cells at high concentrations (700 uM in soybean nodules). This protein gives the nodule a pink color.

The mystery of the symbiotic relationship is that it only occurs through a complex exchange of signals between specific genes of the plant host and symbiont. Infection and nodule organogenesis occurs simultaneously during root nodule formulation. The symbiotic relationship between legume and bacteria is not obligatory. It is quite possible for a seedling to live out its life cycle without becoming associated with a symbiont.

Among many compelling characteristics, the reduction of dependence on commercial, nitrogen fertilizers, the reduction of runoff and minimizing other environmental concerns all show the benefits of the snf process inherent in Pongamia pinnata.



Majda, W. (2014). How to increase the rate of biological nitrogen fixation. Retrieved from

Meyer, S. B., Anderson, D. B., Bohning, R. H., & Fratianne, D. G. (1973). Introduction to plant physiology (2nd ed.). New York, NY: D. Van Nostrand Company.

Rhoades, H. (2017). Nitrogen nodules and nitrogen fixing plants. Retrieved from

Taiz, L., & Zeiger, E. (2002). Plant physiology (3rd ed.). Sunderland, MA: Sinauer Associates, Inc.

Wikipedia. (2017). Nitrogen fixation. Retrieved from

Flynn, R, & Idowu, J (2015) Guide A129 Nitrogen Fixation by Legumes. Retrieved from

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