Nitrogen Cycle Processes and Human Impacts

Introduction

Here is an introduction video that sets up the tone for this blog: Basically, NITROGEN = LIFE

Throughout the history, human activities have had significant impacts on the Nitrogen cycle. Activities such as burning fossil fuels, utilization of Nitrogen-based fertilization, and other activities have lead to an increase in the total amount of biousable Nitrogen in ecosystems globally. Nitrogen availability is directly related to primary production in many terrestrial and aquatic ecosystems; thus, large changes in the availability of Nitrogen can result in extreme alterations in the Nitrogen cycle. Vitousek et al. 1997 (Vitousek, Mooney, Lubchenco, & Melillo, 1997) , discusses the implication of industrial Nitrogen fixation and how as a result of human activity, total amount of Nitrogen fixation has doubled since the 1940s, on a global scale.

Located at the 7^th position on the periodic table of elements; a colorless, odorless, and tasteless gas, Nitrogen is a critical primary nutrient, essential for life to proceed for all living organisms(Lide & Milne, 1995). Nitrogen was discovered in 1772 by Daniel Rutherford. By volume, Nitrogen fills 78% of air in earth’s atmosphere, compared to Mars, which this percentage is much lower, 2.6%. Regardless of its total contribution to the atmospheric air, its presence illustrates the important role it plays in the universe as a whole. Despite its massive abundance in earth’s atmosphere, in its natural form, Nitrogen is not biousable for many organisms, making it a scare resource and a limiting factor in many ecosystems globally. Farmers for instance, need to provide Nitrogen for their crops to maintain growth and expand economical trades. But how does this Nitrogen become available to organisms and what happens to Nitrogen levels as an outcome of human activities, is explored in this blog.

Nitrogen has many uses. Probably the coolest usage of nitrogen is in the making of light bulbs. An insert gas, nitrogen, is often used in light bulbs because it fills the light bulb because it prevents air from entering the light bulb – air and light bulb don’t work well together! Additionally, physical properties of nitrogen help to conduct energy, in the form of heat, from the glowing filaments to the glass bulb, Similar to a cargo if you imagine.

Image illustrating the usage of nitrogen as an inert gas in the making of light bulbs. In the old days they used to have a vacuum that sucked air out, but it was too much of a hassle and the lights bulbs were not as bright.

Another usage of nitrogen gas, which had lead to great discoveries in biological sciences, especially virology, is the usage of nitrogen gas in cryogenic electron microscopy, or cryo-EM. Regular electron microscopy has been aiding structural biologist in determining the 3D structures of various viruses and their proteins (Baker, Olson, & Fuller, 1999). Through these studies, many vaccines have been developed. However, regular EM often introduces bias information due to crystallization of the solvent. With the usage of liquid nitrogen, which exists at extremely low temperatures (-350-400 C), the samples used in cryo-EM are frozen so fast that the solvent, water in many cases, does not crystallize and therefore both the sample and the solvent stay in their most native forms. Discovery of nitrogen in this process has been a huge breakthrough in virology and cryo-EM.

Left - A 5 million Dollar worth cryogenic electron microscope - Right, The structure of gp140 - a protein on the surface of HIV.

In my opinion, not all uses of nitrogen are beneficial to humans; and I’m not talking about plants, I’m talking about BOMBS. Nitrogen triiodide for instance, an extremely sensitive explosive – can be set off by slightest contact. Even small amounts of nitrogen triiodide can generate massive energy in the form of sound (Boopathy & Kulpa, 1992). The trait that allows nitrogen to be suitable for explosives is that it is not radioactive, like Uranium for instance, and most of its explosive properties are triggered through chemical reactions. This is due to its stability and occurrence in only two isotopes, N14 and N-15.

Image showing the classic TNT explosives as we all remember it in Wiley Coyote. A very sensitive substance that through chemical reaction releases a massive amount of energy.

Before moving onto the natural processes of nitrogen and humans impacts, I’d like to discuss one last usage of nitrogen, loved by millions worldwide; the laughing gas!

Perhaps one of the things that make getting your teeth fixed is the usage of nitrogen in laughing gas. Nitrogen oxide or dinitrogen oxide is a colorless gas with a sweet smell and taste. Once inhaled, it causes disorientation euphoria, numbness and loss of motor coordination; in other words, it knocks you out and the best part is you’ll be laughing while it happens.

Now let’s move into the biological and ecological side of Nitrogen:

Nitrogen undergoes a number of alterations and exists in more than one form; each organism, depending on the nature of the ecosystem utilizes Nitrogen in fundamentally different forms. Nitrogen is mainly usable when it is converted from its natural form, N₂ (dinitrogen gas) into Ammonia (NH₃), which is used by primary producers. Nevertheless, Nitrogen exists in a number of other forms as well, either organic such as in amino acids, or nucleic acids, or in inorganic forms such as Ammonia and nitrate. Through this cycle, different forms of Nitrogen are used as nutrients for growth and energy by different living organisms. The major forms of Nitrogen discussed in this blog are the processes or Nitrogen fixation, nitrification, denitrification, anammox, and ammonification, as well as the transformation of Nitrogen into oxidation states, which is a critical step to productivity in the biosphere and extremely dependant on the activities of microorganisms, such as bacteria and fungi.

The diagram blow depicts the broad picture the Nitrogen cycle. Each individual stem illustrated in this cycle will be discussed detail.

Nitrogen Fixation

Perhaps the most studied process in the Nitrogen cycle is Nitrogen fixation, during which diNitrogen gas, N2, is converted into Ammonia (NH3). There are two ways that Nitrogen is converted to Ammonia naturally; by lightning, or by bacteria in plant root nodules .The most common process of Nitrogen fixation is done through microorganisms such as Nitrogen fixing bacteria and fungi. Below is the chemical reaction equation for the process of Nitrogen fixation. 1 unit of atmospheric Nitrogen gas in reduced to generate 2 units of biologically usable Ammonia and hydrogen gas.

As discussed above, Nitrogen is essential for synthesis of proteins, DNA, and their building blocks; nucleic acids and amino acids. Since Nitrogen has a triple bond (N2), it is very stable and the separation of these bonds requires a massive amount of energy. 8 electron and 16 ATP molecules are required for a typical bacteria or fungi to carry out this high energy demanding reaction.

This process is especially important for plant nutrition, which often involved other organisms. Through a symbiotic relationship, some bacteria are found predominantly in what’s called the Rhizosphere – the soil layer in which the plant root is embedded in. Others are decomposers which derive their nutrient from decaying organic material in the topsoil. Some of the most well studied microorganisms that fix Nitrogen are Cyanobacteria, Green sulfur bacteria, Azotobacteraceae, Rhizobia, and Frankia. These bacteria are often associated with the roots of various trees and shrubs. The bottom line is that plants require Ammonia to grow better and faster.

Human Impacts – Nitrogen Fixation

It can be concluded, that Nitrogen is necessary for farming and agriculture. The importance of Nitrogen in agriculture was finally understood in the late 1800; Desfosses was the first scientist to observe that Nitrogen has unique reactivity with certain chemicals. After the importance of Nitrogen was well understood, the push was towards finding ways to gain more Ammonia to facilitate agriculture. In fact, one of the main events that lead to huge human population growth since the early 1900s was new innovations that industrialized Nitrogen fixation. Early approaches of attaining biousable Nitrogen was in Chile, where they used Sodium Nitrate, and unstable source of Nitrogen to produce Ammonia – these resources were rapidly exhausted. It was finally Fritz Harber, who in 1909, invented the process of industrial implementation of the reaction of Nitrogen gas, an unusable form of Nitrogen, and hydrogen gas, which yield 2 moles of Ammonia.

Haber process

The process is conducted under 200 atm and between 400-450 degrees Celsius. Pure dinitrogenous gas (N₂) is mixed with 3 units of Hydrogen gas to produce 2 units of Ammonia(Appl, 2000). Often a catalyst is required to facilitate the reaction and to maintain the direction of this two way equation. It is estimated that 99% of global inorganic Nitrogen is produced in the Harber process.

Nature has limited Nitrogen in the atmosphere the limiting reagent for primary producers for a reason, and manipulation of the levels of bioavailable Nitrogen will affect many ecosystems. However, due to environmental effects of the Anthropogenic Nitrogen has introduced excess Nitrogen to a Nitrogen-limited environment and disrupted natural equilibrium. It has caused an increase in productivity in ecosystems and as a result biological life is threatened though different processes; some discussed below.

Human Population Growth As a Result of the Haber process

Since the year 1950, we have doubled our population from three to seven billion(Bloom, 2011) and this number is estimated to increase to ~10 billion by the year 2050! If it wasn't for major epidemics throughout human population history (i.e. plague; noted on the graph to the right), the human population could have been even higher than what it is today. The figure above illustrates the human growth since the time of pre-agriculture (Old Stone Age) to present.

The story of nitrogen and human population growth:

Nitrogen is an essential part of protein building block, for all amino acid contain a nitrogen group and therefore, without nitrogen, living organisms cannot sustain life and will die from proteins deficiency. All living organisms require nitrogen to grow and multiply in numbers.

People obtain their nitrogen through their nutrients – by eating green plants, meat, now days, protein bars. The bottom line is that both animals and plants require nitrogen to sustain life on this planet. But nitrogen does not come easy! Unfixed nitrogen is as useless to living organisms as vermiform appendix is to humans. It is the fixed nitrogen that can be taken up by animal and plant cells and converted into proteins and amino acids. But how did people figure this out?

Well they didn’t for a long time. Back at the dawn of civilization, at the beginning of the transformation of hunter-gatherers to farmers, it was observed by these early humans that crops such as rice, wheat and corn, only last for a limited amount of time; when soils got exhausted and depleted of nitrogen, these crops no longer grew (Boopathy & Kulpa, 1992). It was the legumes that shed light on the importance of nitrogen and agriculture; the hunter-gatherer people of course had no idea! Turns out that legume crops such as soy, peas, lentils, alfalfa, grew much long and iteratively over many years. These crops had roots associated with a master of nitrogen fixation, rhizobia. This bacterium fixes nitrogen at high rates and makes it available for plants to take up and utilize the biousable nitrogen. But they could only fix so much nitrogen in units of metric tons!

Ostensibly, with no excess vegetation globally, human populations were very small compared to today. It was the Haber-Bosch process, or Haber process that resulted in the explosion of human population on earth. Discovered in 1909 as discussed above, the Haber process was able to produce 300 metric tons of biousable nitrogen annually. The human population spiked after what’s known as the Green Revolution in the 1940s (Vitousek et al., 1997). The development or technology in agriculture between the years 1940s and 1970s, increased agriculture production on a global scale, particularly in the developing world, leading to massive increase in world population numbers.

Image of very advanced heavy machinery used in today's agriculture.

Excess Nitrogen and Nature

Nature has limited Nitrogen in the atmosphere resulting in Nitrogen to be the limiting reagent for primary producers. Therefore, manipulation of the levels of bioavailable Nitrogen will affect ecosystems. Because of environmental effects caused by anthropogenic activities, Nitrogen is being introduced in excess to a Nitrogen-limited environment which leads to the disruption of natural Nitrogen equilibrium. This excess Nitrogen has caused an increase in productivity in ecosystems and as a result biological life is threatened though different processes; some discussed below.

One of the ways by which release of Anthropogenic Nitrogen (excess Nitrogen) affects various ecosystems (i.e. Marine Ecosystems) is Eutrophication. Eutrophication, or commonly called hypertrophication is the feedback of additional, or artificial Nitrogen containing compounds that enter an ecosystem that under normal conditions would not. Substances such as Nitrates will find their way into aquatic systems through sewage, or agriculture byproducts (i.e. fertilizers).

Impacts of Eutrophication

Before discussing the impacts of Eutrophication, I would like to briefly explore some of the fundamental processes in relation to marine ecology, focusing on the coral reefs.

Coral reefs are the most primary productive habitats on earth, averaging between 2,500-5,000 grams of Carbon per meter square per year globally; even compared to tropical wetlands under direct impacts of the ITCZ, the net primary production is at least one third when compared to coral reef biomes. Perhaps it’s the historical established mutualisms between corals and other organisms that have lead coral reefs to be the most productive biome on earth. Zooxanthellaes that fix Carbon or the Crustaceae which protect coral from predation by various predators play an important role in the sustainability of coral reefs.

As I mentioned above, Nitrogen is essential for all living organisms. Same rule applies for marine organisms. Coral and Phytoplanktonic organisms utilize Nitrogen that is fixed by Cyanobacteria, which are considered to be “as old as life itself”. These cyanobacteria fix carbon by photosynthesis and fix Nitrogen by Nitrogen Fixation, where for every mole of Nitrogen gas they produce one mole of Ammonia, at the expense of 16 ATP molecules; massive investment of energy.

Significance

As mentioned above, for one mole of Nitrogen, 16 molecules of ATP are required. This suggests that nature has made this process extremely high energy demanding to perhaps establish a balance for the amount of Nitrogen that enters a particular biome (Capone, 2001; Capone & Montoya, 2001). What Eutrophication, as a result of anthropogenic activities there is an increase phytoplankton population; they grow better and rapid because there’s excess nutrition. This will result in decomposition of phytoplankton which will decrease Oxygen levels. Decreased Oxygen levels will result in hypoxic conditions (see picture below from NASA, the green regions are near the estuaries and illustrate an average accumulation of Nitrogenous compound input over time) which will have negative impacts on marine biodiversity as well as an increase in the susceptibility to invasive species (Bell, 1992). This will cause a decline in coral reef because habitats such as the coral reefs are nutrient-sensitive and require the lowest external inputs to trigger Eutrophication.

Hypoxic Zones (Visual Illustration)

Many sources of Eutrophication are illustrated in the focus below. These are artificial sources of Nitrogen added to the marine environments. Among these are:

Nitrogen compounds produced by cars
Discharge of untreated municipal sewage
Discharge of detergents and treated sewage
Runoff from streets, lawns, and construction lots

These external inputs of Nitrogen into the marine biomes change the ecology and sustainability of these environments which in turn will have an impact on species biodiversity and etc…

Nitrification

Nitrification is the process of ammonia conversion into nitrite and nitrate (an overall diagram of the process below). Under aerobic conditions, prokaryotes carry out this reaction; some fungi have been known to do this as well, but at much less numbers compared to bacteria. The first step of the reaction, conversion of ammonia into nitrite is done under aerobic conditions by prokaryotes known as ammonia oxidizer. The first step of nitrification is done exclusively by bacteria because they posses two enzymes, ammonia monooxygenase and hydroxylamine oxidoreductase , which convert ammonia into an intermediate, hydroxylamine (Rumer, Gupta, & Kaiser, 2009). This process generates small amounts of energy.

Dissimilar to Nitrogen fixation, which is carried out by a broad spectrum of different microbes, ammonia oxidation is more selective in terms of which microbes can carry the process out. Bacterial genera such as Nitrosomonas, Nitrosopira, and Nitrosococcus are experts in ammonia oxidation. Thus, not very many microbes are able to oxidize ammonia to be used in step two of the process; conversion of nitrite to nitrate.

The second step of the process nitrification is termed nitrite oxidation, where one mole of nitrite is oxidized via half a mole of oxygen gas, or (just O), to generate one mole of nitrate. This process is carried out by other exclusively selective prokaryotes known as nitrite-oxidizing Bacteria. Both ammonia-oxidizers and nitrite-oxidizers function under anaerobic conditions; the soil for instance, or in lakes and open-ocean regions. These prokaryotes are also useful in maintaining a healthy environment in the open-oceans. They facilitate the removal of toxic ammonium containing substances excreted in fish urine and feces (Johnson et al., 2010). Humans have, as always, have figured out a way to take advantage of such processes.

Human Impacts

As mentioned above in the agriculture and Eutrophication paragraphs, anthropogenic processing of nitrogen containing compounds will result in anoxic zones in the ocean; especially at estuaries (Treusch et al., 2005). One positive action done by humans is take advantage of such processes. Synthetic (meaning man-made) facilities help reduce or remove excess ammonium and prevent pollution of the waste receiving waters. Image below is one example of such facilities near estuaries in bay area off the coast of SF.

Anammox

This process seems very important to me because it has been associated to have direct impacts on nitrogen levels in the ocean and the global nitrogen cycle; however, it is not generally discussed in many websites that discuss the role of nitrogen. Including Wikipedia. In this part of the blog I would like to explore the role of Anammox.

Generally, nitrification occurs and is carried out under aerobic conditions; the presence of oxygen is essential. However, relatively new discoveries have shown that ammonia oxidation can also occur in anoxic environments (Strous et al., 1999). A process called Anammox, which stands for anaerobic ammonia oxidation, is carried out by a special type of prokaryotes called Planctomycetes. The process utilizes special enzymes which are still under study, to convert ammonia and nitrite into gaseous nitrogen and water. Due to its nature, this process occurs in oceanic zones with low oxygen, estuaries and freshwater lakes. Plant species in these areas are unable to grow vastly and are few in numbers because the biousable nitrogen is sucked out of the system (Kuypers et al., 2005). Some argue that this is due to denitrification (discussed below) rather than anammox. The overall chemical reaction of Anammox is depicted below.

Denitrification

Denitrification is the process of regenerating oxidized atmospheric nitrogen. It converts nitrate back into nitrogen gas and therefore diminishes the available biousable nitrogen, returning it to the atmosphere. Other gasses such as nitrous oxide (N₂O) are the intermediate by-products in the process of denitrification.

Dissimilar to nitrification discussed above, denitrification is an anaerobic process and mostly occurs in soils and sediments and anoxic zones in open oceans and lakes. Unlike, nitrification, denitrification is carried out by a number of prokaryotes; recent studies illustrate that denitrification can also be carried out by eukaryotes (Risgaard-Petersen et al., 2006). Denitrification is actually a good thing! The overall reaction of denitrification is depicted below.

Significance

Above, I discussed the impacts of nitrogen pollution and excess nitrogen influx into the ocean – resulting in dead zones. Denitrification is actually beneficial for oceanic creatures! It removes excess biousable nitrogen from estuaries, impeding efflux of nitrogen to be taken up by algae.

N2O is considered a greenhouse gas and therefore contributes to “air pollution”. I put air pollution in quotations because this process is done in nature and therefore doesn’t really sound like pollution as we know it. Regardless, it does react with the ozone and contributes to both global warming and ozone layer destruction. It is however crucial to keep in mind, that the amount of impact and input to greenhouse gasses is close to zero compared to other sources of air pollution and greenhouse effect; such as carbon dioxide.

Human Impacts

Humans have constructed a facility that converts biousable nitrogen back to its gaseous form to reduce Eutrophication. The Blue Pains Wastewater Treatment Facility is a perfect example. This facility utilizes methanol to enhance the efficiency the conversion nitrite or nitrate or ammonia into N₂. The images below are from the BPWTF.