How to Get Nitrogen Out of the Tank  

The most common bacteria found in aquatic environments are two genera named
Nitrosomonas sp. and Nitrobacter sp. Which are aerobic (needing oxygen),  
chemolithotrophic (living on rocks, or on the bottom, and feeding on chemicals from the
water as a source of energy) autotrophic (self feeding) nitrifying (changing ammonia into
nitrate or nitrate into nitrite) bacteria that multiply rapidly when the conditions are right
helping the aquatic environment balance and recycle the element nitrogen.
Together they are called nitrifying bacteria because they change ammonia (NH3) into
nitrates (NO2) and then nitrite (NO3), and this conversion is needed as a source of energy
for these bacteria. What conditions do they need to thrive?
1) Proper range of pH.
The pH of a tank is the “potential for Hydrogen” or how much hydroxide (H3O) is in the
water (H2O). This is measured on a scale of 0-14 with 7 being neutral. The higher the pH,
the more hydroxide (alkalinity) in the water. The lower the pH, the higher the hydronium
(H3O) content of the tank. Hydronium ions are what make the water more acid since they
will yield a Hydrogen ion (H+) and water (H2O). At pH 7 there two types of atoms are
balanced which means for every HO ion there is a extra H+ ion attached to water, when
these are added together you get 2H2O, which is, by itself pH 7.0, neutral. The nitrifying
bacteria need hydroxide, but in the right amounts. A pH of 7.0 will support the growth of
nitrifying bacteria, which prefer a pH of 6.3 to 9.0. pH. Low pH values inhibit nitrification
by providing a limiting amount of bicarbonate, the preferred carbon source for nitrifying
and hetrotrophic bacteria (see below). A variety of compounds, including bicarbonates,
salts of weak acids, and hydroxides contribute to alkalinity. When ammonia is oxidized
during nitrification, hydroxides contribute to alkalinity. When ammonia is oxidized during
nitrification, hydrogen ions (H+) (producing hydronium ions H3O) are liberated from
ammonia. Alkalinity is needed to neutralize these hydrogen ions. In fact, 8.64 mg/l of
alkalinity are consumed for each mg/l of ammonia that is oxidized. Without sufficient
alkalinity, the pH of the system will drop, and nitrification will slow down. Nitrification
works best when the pH is between 6.5 and 8.5. The process slows considerably at pH
values outside this range. (This is why there is a peat bog at the bottom of Spirit Lake at
Mt. Saint Helens.) The type of fish and plants in the aquarium often determine the pH
range. This tank uses a high pH because of these factors. The pH is raised by adding
common table salt, or baking soda, and other minerals.
2) They need to have their TAN.
In this case, TAN is Total Ammoniated Nitrogen. The nitrifying bacteria feed on
ammonia. If there is none in the tank, they will die. Ammonia comes mostly from the gills
of fish as a waste product of their respiration, then from decaying organic matter both
from plans and animal waste. The greater the biological mass (biomass) in the tank
(mainly the fish and the plants), the more ammonia is produced and the more nitrifying
bacteria are needed. (This balance can be disturbed by the addition of chemicals that kill
bacteria such as Methylene Blue or antibiotics, or by the sudden addition of too many
fish.) Once the biological filter is working and in balance, the number of bacteria will vary
according to the amount of ammonia in the tank until the total available surface area is
occupied by bacteria. See how well balanced the system is? It is unusual that Ammonia ss
the limiting factor, usually it is Oxygen.
3) They need their DO
Usually the limiting factor in adequate biofilters is a low dissolved oxygen (DO) level
which results form either a lack of adequate oxygenation by mechanical means or lack of
plant/light ratio which produces adequate oxygenation and, at the same time the plants use
up nitrogen, phosphorus, potassium, sulfur, iron, magnesium, etc., etc. Since the bacteria
filter uses oxygen to convert ammonia to nitrate, inadequate oxygenation is a critical
factor is the establishment of a biofilter. Inadequate oxygenation also produces
undesirable sulfur compounds from the deterioration of organic matter that foul the water
and give off unpleasant odors (see “Plants Need Nitrogen” below). Without a bubble filter,
where do we get the oxygen? This comes from the plants during photosynthesis. So,
keeping the plants strong balances the tank. Healthy plants, healthy fish.
4) Temperature.
Water temperature is important for bacterial growth and propagation and therefore, for the
health of the entire system. If the water is too cold, the bacteria will not reproduce fast
enough to adjust to changing ammonia levels, however, this is rarely a factor in an
established aquarium since the temperature is set for the needs of the tropical fish. This
fish likes its water warm, about 82 degrees. Yet cooling the water also slows the
production of the ammonia.
5) They need space.
They need large surface areas to live on. The question is, what will give the greatest
surface area for the mass in the filter system? Small filaments, hair-like structures have the
greatest surface area to mass ratio. A cheap source of this is common Fiber-Fill found in
sewing and craft stores. Packed into the canister, this gives large surface area and tight
passages which allow the bacteria to remove the ammonia efficiently. This is not the only
substance that can be used, but it is cost and surface area efficient.
When these factors are met, then an active biological filter is slowly established.
This process is referred to as the nitrogen cycle. Ammoniafication, as listed above, starts
in several places (fish and water as well as bacteria) but end up in the water where it is
potentially harmful to the fish.
Again, ammonia (NH3) is oxidized by the Nitrosomonas bacteria to Nitrate (NO2)which
is subsequently oxidized by Nitrobacter to Nitrite (NO3). The entire process is called
nitrification. Nitrite (NO3) can be used directly by cells as a source of nitrogen
(assimilatory nitrate reduction).
This entire process happens on most surface areas you can see and in the canister filter
under the tank. But this is only one side of the nitrogen cycle. The other side also occurs in
the tank. But where? You ask.
The Other Half of the Nitrogen Cycle
God sets up organic living systems that use everything. (This makes more sense than
saying everything is balanced for no reason.) Nitrogen is used by the plants as well as by
the bacteria. Certain bacteria can reduce (remove oxygen) nitrite during a process called
anaerobic respiration, where nitrate is used in place of oxygen as a terminal electron
acceptor for a process similar to aerobic respiration (using oxygen), in short, this is where
they get their energy. This is the opposite of what id describes above. In the case of
anaerobic respiration, NO3 is first reduced to NO2, which is subsequently reduced to N2
or NH3, all three of which are gasses which are soluble in water.
This process is called denitrification and it occurs in anaerobic environments where
nitrates are present and oxygen is not. While this process occurs in the aquarium, it is not
without merit. Even though we want most nitrogen out of the tank there are uses for it in
the tank first, as above, for the nitrifying bacteria, but also for plant growth.
Denitrification supplies the plants with the nitrogen needed for growth. Note that this
happens in an anaerobic environment, one with little or no oxygen present. This is why we
do not use an under-gravel filter. If this filter were present, then water would be drawn
through the gravel making it an aerobic environment and the denitrifying bacteria would
take on aerobic oxidation instead of anaerobic oxidation (yes, they can do that) starving
the plants of nitrogen (in this case ammonia), which would in turn starve the plants and
reducing the oxygen killing off much of the nitrifying bacteria and the entire system
would fail, or parts of it would fail and need support
Hetrotrophic saprophytic bacteria break organic material down into carbon dioxide and
water. The word hetrotrophic means they eat two ways. They can attach directly to
organic compounds such as fish waste or dead plant materials and consume small parts by
“eating” and decomposing them, or, like the autotrophs, they can absorb dissolved
nutrients directly from the water. Saprophytic means they live off of decaying material.
These bacteria breaking down organic matter into carbon dioxide and water. Carbon
dioxide is needed by the plants to form sugars during photosynthesis. The plants release
oxygen from this process, thus completing the oxygen cycle in the tank. This is one reason
adequate light is needed. Light is the energy source used in photosynthesis to capture an
electron that is needed to make the sugars which are used both for energy and for
structures in the plant. All plant fibers are made from sugars of different types.
Facultative denitrifiers such as Bacillus licheniformis, Bacillus megaterium Bacillus
subtilis, and Bacillus polymyxa produce digestive enzymes to break down organic waste
of fish and plants, and, ultimately feeding the nitrifying bacteria. These are also interesting
in that, when oxygen levels drop below critical levels they can burn nitrite and nitrate for
energy, which, of course, returns it to ammonia.
Vibrios (curved rod-like shape similar to a comma) are common bacteria in aquatic
environments like fish tanks. Pseudomonadaceae  and Vibrionaceae are families of
bacteria that are facultative bacteria, that is, they facilitate the breakdown of organic
materials. They have polar flagella (move by use of a tail-like appendage), and are
oxidase-positive (meaning they use oxygen to oxidize sugars as an energy source). These
help to break down organic matter into simpler compounds. In aquatic habitats they
overlap with the in their ecology, although pseudomonads favor fresh water and vibrios
prefer salt water.
Nitrogen fixation is the actual beginning of the nitrogen cycle but this is beyond the scope
of this paper as it accounts for less than 1% of the nitrogen in an aquarium.
The Oxygen Cycle
As above, during plant type oxygenic photosynthesis (as opposed to bacteria type) plants
take in carbon dioxide (CO2) and water with energy captured from the available light to
form sugars and give off oxygen (O2). During aerobic respiration, as in fish, this cycle is
reversed. The fish use the oxygen to burn sugars and convert the energy into stored energy
two nucleotide molecules (ATP and GTP) giving off carbon dioxide and water.
So autotrophic bacteria and plants produce oxygen from water and carbon dioxide while
heterotrophic bacteria and animals use oxygen and produce carbon dioxide and water.
The Carbon Cycle
Only organisms can reduce (remove oxygen) from carbon dioxide (CO2), so, organic
chemistry is the study of organic molecules, or those molecules that have reduced carbon.
Carbon forms the very basis for life. All “organic” molecules (though not necessarily all
molecules in a given organism) contain carbon. Carbon dioxide can be viewed either as
organic or inorganic.
Some bacteria that break down organic matter are called methanogens, or, methane
generating bacteria. Methane is sometimes called swamp gas. Methanogens have an
incredible type of metabolism that can use H2 as an energy source and carbon dioxide
(CO2) as a carbon source for growth. (Since these are absorbed directly from the
environment and not consumed pre se by the bacteria, they are also autotrophs, that is, self
feeding.) In the process of making cell material from H2 and CO2, the methanogens
produce methane (CH4) in a unique energy-generating process. The end product, methane
gas, accumulates in their environment. When the biological filter is filly developed small
amounts of methane are released from the gravel bed. When too much is being released
(as evidenced by bubbles being released from the gravel bed without agitation), it is an
indicator that the gravel needs to be cleaned a little more than usual to reduce the organic
matter. But this takes most of the biological energy and converts it into methane. There
should be some way to capture this energy and convert it into useful carbon dioxide. There
is.
Methanotrophs are bacteria that can consume methane (as much as 90% of it from the tank
in a well-developed biofilter) and oxygen and produce carbon dioxide and bicarbonate.
This reaction can happen at any place in the aquarium but rarely in the gravel bottom
which tends to be anaerobic. Bicarbonate acts as a buffer to pH and is the primary carbon
source for some bacteria. These are known to contain powerful enzymes (pMMO and
sMMO) that are know to attack many chlorinated organic compounds as well as many
other compounds returning them to the water for use by other bacteria.
The Methylococcaceae Methylomonas methanica and Methylosinus trichosporium are two
such bacteria that stay in the aerobic zone at the top of the gravel and above, including in
the canister filter. In salt water Methylosphaera hansonii partially fills this niche. They
oxidize formate (formic acid, from formaldehyde) and carbon monoxide to carbon
dioxide. This regenerates the reducing process and is a source of energy for the
microorganisms.
Interest was heightened in these bacteria after the Exxon Valdeze incident in the Prince
William Sound where the areas that were left alone to deal with the oils spill recovered
better after 2 years than did those areas where extensive intervention by man occurred.
As we have seen, autotrophs, such as plants, algae, photosynthetic bacteria, lithotrophs,
and methanogens, use carbon dioxide as the source of carbon for growth, and therefore
reduce from its gaseous form it include it into cell material. Heterotrophs require organic
carbon for growth, and usually energy, then, when they deteriorate, the carbon is
converted back to carbon dioxide.
This is the carbon cycle. This is why a balanced aquarium needs both autotrophic and
hetrotrophic, both nitrifying and ammoniafying bacteria to balance the entire system as
well as plants and fish in balance.
There are also lithotrophic bacteria that can oxidize carbon monoxide (CO) into carbon
dioxide (CO2), but their significance in the aquarium is unknown and their presence in the
aquarium does not seem to be of major importance at this time (but things change).
The methanogen bacteria are unique in their roll in the carbon cycle since they use carbon
dioxide in two ways. About 5% is used for cellular material and 95% is used to produce
methane gas (CH4), which makes them so unique. However, this is a good reason to clean
the gravel periodically to reduce the food sources for them, this preserving more future
carbon for making carbon dioxide which is more useful in the aquatic environment. But
this brings us to the methanomorphs. Methanomorphs take up methane, methanol
(oxidized methane, CH4O) or formaldehyde (CH2O) and use these as forms of energy and
therefore are a part of the biodegredation in the carbon cycle as well.
This is the place most people consider bacteria during the degradation of biological
components, the decomposition of, in this case, fish and plants in the aquarium. However,
as we have already seen their combined efforts are needed to balance the water habitat. It
is by the efforts of these bacteria that the large molecules (polymers) of an organism,
either plant or animal are broken into polymer subunits (parts of the polymer) that can, by
oxidation be recycled into the system as water, carbon dioxide, hydroxide, hydronium,
ammonia, sulfides, and other molecules and atoms.
The Sulfur Cycle
Even though we don't normally see or smell sulfur, it is a component of most biological
systems since it is a component of a number of vitamins, amino acids (cysteine and
methionine, which is needed to start building proteins) and may have other uses in other
organic molecules.
Anoxygenic photosynthetic bacteria (such as the various chemo-litho-autotrophic
Rhodobacter or Alcaligenes species) that oxidize sulfide (H2S) sulfur and sulfur to sulfate
(SO4) just like the nitrifying bacteria handle ammonia and nitrate. This is bacteria
photosynthesis that differs from plant, or the oxidative form of photosynthesis. There are
the purple and green sulfur bacteria that sometimes populate aquariums.
Another group are called the “colorless sulfur bacteria” such as the Riftia pachyptila
endosymbiont or various Thiobacillus species which oxidize sulfide and sulfur as a source
of energy. In either case, the organisms can usually mediate the complete oxidation of
sulfide (H2S) to sulfate (SO4). Both are useful in plants and animals.
Usually these are seen in sulfur vents such as in Yellowstone Park, but they may also be
found in the tropical aquarium but only in the anaerobic parts such as the gravel substrate
at the bottom of the tank. These would be found for example in abundance at the bottom
of Spirit Lake under Mount St. Helens.
Plants and Animals need Iron
The Iron cycle is like the sulfur cycle. There are bacteria that reduce (take oxygen from)
iron and those that oxidize iron. Oxidation of ferrous iron causes iron to be fixed or
chelated often forming red colored slime in pipes (hydrated ferric oxide), or rivers where
the reducing bacteria can then use it to form black colored slimes or soil deposits.
Our friend above the Thiobacillus has relatives like Thiobacillus ferrooxidans which is an
acidophile (loving acid) heterotrophs (both consuming nutrient and absorbing nutrient.
They can also oxidize Magnesium.
Leptothrix discophora (and others) cause oily films on the surface of ponds and streams
where they live. Is this actually oil? Yes, it is. They synthesize large amounts of fatty acids
which keep them afloat where they can get more oxygen and can be used for metabolic
purposes. These too oxidize both iron and magnesium.
Other iron eating bacteria include:
Leptothrix ochracea, Leptothrix sp.A, Leptothrix cholodnii, Siderocapsa cf.treubii,
Siderocystis sp., Siderocystis confervarum, and Gallionella ferruginea.
Our tank uses an iron rich clay like gravel called Fluorite mixed into the bottom to
introduce iron into the aquarium. That is the small red gravel at the bottom of the tank.
Other metals are handled in a similar way by these and other bacteria in oceans, rivers,
ponds, and, yes, the aquarium. These are all sensitive to chlorine in tap water. This is why
you need to make frequent water changes of 20%-30% and treat the water as you change it
so you protect these bacteria and the others.
This is a small part of the well designed system that is the world around us.
How I Set Up the Aquarium
The previous set up included a gravel bottom and the canister filter which we had not
changed for several weeks. This allowed for the build up of organic waste on the outside
of the filter media can which housed activated charcoal. First I changed only the internal
canister by removing the charcoal and replacing it with nylon scratch pads of differing
corseness.

(Left, filter materials goin into the filter. Middle, leaves in the bottom. Right, sand over
the leaves.

I had not deep cleaned my tank for several weeks allowing the detritus (debris in the
bottom) in the rock to built up. When I moved plants around this detritus is pulled into the
filter.
Then I planted the aquarium heavily, added some table salt and some phosphorus,
potassium salts, Epsom salt, zinc, sulfur, and a few other sources of minerals to feed the
bacteria and plants. I removed the defuser on the filter input to prevent the surface of the
water from being agitated. This allows the carbon dioxide to accumulate in the tank for
healthier plant growth, which, in turn, produces oxygen; it also allows bacteria to form at
the top, which helps regulate the gasses in the tank. I moved the input away from the
output pipe to create a more natural flow of water through the tank.
The minimum light requirements for a tank are about 1.5 watts per gallon of water in the
tank, so, I added a second florescent light to add energy to the tank for good plant growth.
A Brief Picture of the Aquarium Biofilter
1) At the surface of the water, light and oxygen are plentiful, carbon dioxide is fixed by
bacteria and oxygen is taken directly from the air into the water. At the surface bacteria
take gasses from the air directly and oxidize metals for use by other organisms.
2) In the canister filter, on the sides of the tank, the plants and the top of the gravel,
organic matter is captured and aerobic bacteria use plant and fish waste to produce carbon
dioxide, sulfide, ammonia, nitrate and nitrite which can be used by plants.
3) In the gravel at the bottom of the tank, detritus builds up and oxygen is soon used up,
then the anaerobic bacteria break down large organic particles, and produce carbon
dioxide and ammonia which are use by the plants. Some methane is also produced which
can act as an indicator of excessive plant material and can be used by other methane eating
bacteria to produce carbon dioxide.
4) Plants take up these nutrients and create oxygen for the aerobic bacteria and fish,
removing nitrogen and phosphorus from the water.
5) Light produced oxygen. Blue light which is more available at the surface produced
bushier growth in the plants. Red light which is slightly more dominant at the bottom of
the tank makes plants grow longer so they can reach the surface.
All together this creates a stable aquarium that has less disease, better gas regulation, and
is less expensive to run than mechanical chemical filters.
Yes, this took some thought. It took intelligence and design to use the natural systems
available to us in setting up this aquarium. The more complicated the system is to set up,
the higher intelligence is required to create it. This is a basic rule of engineering. How
much more intelligence did it take to create the life forms in the tank? One strand of DNA
from one bacterium in this tank is thousands of times more complicated than this entire
document you have just read. God was the writer of that blueprint.

 “And God saw every thing that he had made, and, behold, it was very good. And the
evening and the morning were the sixth day.” Genesis 1:31