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Ammonia is one of
the most studied chemicals in aquaria, and the details of
its uptake, excretion and mechanisms of toxicity continue
to occupy many research scientists. Because of its high toxicity,
it is critically important in both freshwater and marine systems.
In fact, it is one of the few important chemical issues that
marine and freshwater aquaria share. Nevertheless, misunderstandings
abound about ammonia's sources, nature and toxicity, which
may not be recognized by many reef aquarists
Ordinarily, established reef aquaria do not have issues with
ammonia because their large amounts of bacteria and algae
serve to rapidly eliminate it from the water. However, new
aquaria, hospital tanks, quarantine tanks, shipping bags and
reef aquaria containing something that has died can have ammonia
levels that rapidly rise to toxic levels. In fact, I believe
that, along with low oxygen, ammonia can be the primary immediate
cause of many tank crashes. Something dies, it decays and
releases ammonia, the ammonia kills something else, and the
system spirals out of control, with ammonia being the main
factor.
This article provides a detailed basis for understanding
ammonia in seawater - where it comes from, how it is toxic
and at what concentrations, and how to deal with ammonia "emergencies."
It is beyond the scope of this article to provide size and
other guidance regarding filters designed to facilitate the
nitrogen cycle, and it deals primarily with reef aquaria where
no such filters are used, or at least where they are not the
focus of any concerns.
The sections are:
Introduction
Ammonia (NH3) can exist
in two primary forms in water. One is free ammonia, and the
second is an ammonium ion (NH4+). An
ammonium ion is formed when a proton in solution combines
with ammonia:
NH3
+ H+ à
NH4+
Water always contains protons, and the lower the water's
pH, the more protons it has. In fact, a drop of one pH unit
means exactly a tenfold increase in the number of protons.
So at lower pH, where there is a large amount of H+,
the equilibrium is shifted away from ammonia and toward ammonium.
This shift is actually very important in understanding its
toxicity, as NH3 and NH4+
may have different rates of passage across fishes' gills.
This exchange between ammonia and ammonium is incredibly
fast. It can happen in a billionth of a second, and for a
single ammonia molecule, the conversion can happen a billion
times a second.1 So on a human time scale it is
often inappropriate to think of them as different species,
but rather it is more appropriate to think of any individual
ammonia molecule as spending a portion of its time as free
ammonia, and a portion of its time as ammonium ion, with those
relative time portions being a function of pH.
Ammonia and Ammonium as a Function
of pH
The pKa of ammonium in seawater is
about 9.3 (some references state 9.5, with the difference
possibly relating to the different pH scales sometimes used
for seawater and freshwater).2 That pKa means that
at pH 9.3 the water has equal concentrations of ammonium and
ammonia. At pH values below this level, as is always the case
in reef aquaria, ammonium predominates. Figure 1 shows a plot
of the relative fractions of ammonia and ammonium as a function
of pH in seawater. At pH 8.2 only about 7% of the ammonia
is present as free ammonia, with 93% present as ammonium.
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Figure 1. The
fraction of free ammonia (NH3) and ammonium
ion (NH4+) present in seawater
as a function of pH.
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Because many authors attribute ammonia's toxicity primarily
to free ammonia (whether this is correct or not, see below),
Figure 2 shows an expanded view of Figure 1 for the free ammonia
concentration over the pH range of usual interest in reef
aquaria. The amount of free ammonia present at pH 7.8 is about
one-fourth the amount present at pH 8.5.
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Figure 2. The
fraction of free ammonia (NH3)
present in seawater as a function of pH over the range
of most interest to reef aquarists. This figure reproduces
Figure 1 on an expanded scale.
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Reef aquarists also are often interested in pH values below
those typically present in actual reef aquaria. A shipping
bag containing fish, for example, often drops substantially
in pH over the course of hours to days, as a result of expelled
carbon dioxide. That change in pH can convert even more of
the free ammonia to ammonium, and Figure 3 shows the fractions
of ammonia and ammonium on a log scale, making it clear that
the free ammonia continues to drop in concentration as the
pH drops, even when it is already present as a very small
fraction.
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Figure 3. The
fraction of free ammonia (NH3)
and ammonium ion (NH4+)
present in seawater as a function of pH. This figure
shows the fraction on a log scale over a wider pH range
than Figure 1.
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Units of Measure of Ammonia and
Ammonium
A variety of different units have
been used to report ammonia concentrations. The unit "ppm
NH3-N" represents the parts per million of
nitrogen in the sample's free ammonia. The unit "ppm
NH4+-N" represents the parts per
million of nitrogen in the sample's ammonium. The unit "ppm
total NH4-N"
is often used to represent the sum of nitrogen in both ammonia
and ammonium in the sample.
To convert ppm NH3 to ppm NH3-N, multiply
by 0.82, because ammonia is 82% nitrogen by weight. To convert
ppm NH4+ to ppm NH4+-N,
multiply by 0.77, because ammonium is 77% nitrogen by weight.
There can be no standard conversion between ppm total NH4-N
and ppm total NH4 because the conversion would
depend on the relative amounts of free ammonia and ammonium
ion. One ppm in seawater is close to one mg/L (actually 1.03
mg/L), so for most purposes in the context of ammonia in seawater,
mg/L and ppm are interchangeable units.
Many chemical oceanography papers report concentrations in
units of molar (M), mM or mM. One
M NH3 is equivalent to one mole/L, or 17 grams
per L, which equals 17,000 mg/L. One mM
NH3 is equivalent to one millimole/L, or 17 milligrams
per L. One M NH3 is equivalent to one micromole/L,
or 17 micrograms per L, which equals 0.017 mg/L.
Ammonia Concentration in the Ocean
The concentration of ammonia in the
ocean varies substantially, from less than 0.002 ppm to as
much as 0.7 ppm total NH4-N,
but is usually very low in surface seawater (<0.02 ppm
total NH4-N).3
For example, the seawater intake at the Hawaii Institute of
Marine Biology (on Coconut Island, Oahu, HI; 150 feet from
shore and 20 feet down) was found to have an ammonia level
that ranged over 0.0025 ± 0.0021 ppm total NH4-N.3
Remote ocean surface waters are reported to have 0.006 ±
0.004 ppm total NH4-N.3
Sources of Ammonia in Reef Aquaria:
Salt Mixes
There are a variety of sources of
ammonia in reef aquaria. Minor sources include: 1) tap water
(especially if it contains chloramine
and is not treated with a deionizing
resin) and 2) impurities in salt mixes and other additives.
It
has previously been shown that the total NH4-N
ranged from 0.55 to 11.9 micromole/kg (0.008 to 0.17 ppm total
NH4-N)
in an analysis of eight brands of artificial seawater mixes.
At the higher end of the scale, those levels will be detected
with an ammonia test kit and can present potential toxicity
concerns if fish are kept at those levels (see below). These
levels of ammonia may be introduced from impurities in calcium
chloride and magnesium chloride, where ammonia is a well known
impurity resulting from some of the commercial manufacturing
processes used (such as the Solvay
process, which involves ammonia).
Calcium and magnesium additives can also be a significant
source of ammonia, especially for aquarists who are trying
to use inexpensive sources of bulk calcium or magnesium chloride.
I discussed testing calcium chloride for ammonia in a previous
article.
Sources of Ammonia in Reef Aquaria:
Biological Processes
The predominant source of ammonia
in marine aquaria is excretion by fish and other heterotrophs
(organisms that live by consuming organic materials). Fish
excrete large amounts of ammonia (and ammonium) from their
gills, and possibly smaller amounts from their urine, which
is why all aquaria must contain some mechanism to prevent
the excreted ammonia from rising to toxic levels. Heterotrophic
bacteria can also be a big source of ammonia in aquaria. For
example, uneaten fish food that is broken down by bacterial
action will usually result in ammonia being released to the
water. Many other marine organisms that are likely to be found
in reef aquaria also excrete ammonia, such as crabs and shrimp.
In fact, most any organism in a reef aquarium that lives by
consuming food (rather than by photosynthesizing) excretes
some amount of ammonia.
The reason that such organisms excrete ammonia is that they
take in far more nitrogen from the organic foods that they
consume than they need to build new tissues. Consequently,
they must excrete nitrogen in some fashion. Ammonia is a common
way to excrete nitrogen, along with urea and a few other nitrogen
compounds.4 The chemical equation below represents
the end products resulting from the metabolism of organics
with a molecular formula representing typical plankton5
in seawater:
(CH2O)106(NH3)16(H3PO4)
+ 106 O2 à
106 CO2 + 106 H2O + 3 H+
+ PO4--- + 16 NH3
Mechanisms of Ammonia Excretion
by Marine Fish
The mechanisms whereby marine fish
excrete ammonia are different from those in freshwater fish,
and aquarists must be careful not to extrapolate findings
from freshwater fish to marine species without carefully considering
whether it is reasonable to do so or not.
Marine fish excrete ammonia via their gills in several ways.
Like in freshwater fish, free ammonia can passively diffuse
out of the cells that make up the gills, and this is often
the dominant pathway for excretion. This result has large
implications for ammonia's toxicity to fish. Because free
ammonia is passively diffusing out of cells and into the surrounding
seawater, it requires a downhill gradient to proceed. If the
seawater's ammonia concentration rises, this excretory mechanism
does not operate effectively. It might even operate in reverse.
Because it is the free ammonia concentration in the surrounding
fluid that determines the ability of free ammonia to be excreted
in this fashion, pH plays a substantial role in determining
elevated ammonia's ability to prevent this excretory mechanism
from operating. Lower pH in the surrounding seawater converts
more of the ammonia to ammonium, leaving less free ammonia
to back up this excretory pathway, resulting in less observed
toxicity.
However, the popular scientific notion that the diffusion
pathway involves simple diffusion of free ammonia through
cell membranes seems no longer to be supported by recent studies
utilizing modern biomolecular techniques. It is now recognized
that this passive diffusion of NH3 more likely
takes place through the spaces between cells, called paracellular
tight junctions, and this transport phenomenon is called paracellular
transport. These junctions, while usually tight enough to
prevent large molecules from passing through, are leaky enough,
in both marine and freshwater fish, to allow both water and
ammonia to pass through.
Without going into any more detail on how freshwater and
marine species differ in their free ammonia excretion (there
are other important differences), a very important difference
is that the paracellular junctions in marine fish are leakier
than in freshwater fish, and these junctions can allow marine
fish to passively excrete a substantial amount of ammonium
ion. Because this pathway also involves the passive diffusion
of ammonium from the blood to the surrounding fluid, the conversion
of ammonia to ammonium in the ambient environment at low pH
cannot completely reduce the toxicity of ammonia/ammonium
in the surrounding fluid, even though it can largely do so
in freshwater fish. These issues will be discussed in more
detail in the toxicity section, but it bears repeating that
if assessing ammonia toxicity is the goal, do not ignore contributions
from ammonium ion elevation in the marine environment.
There are also other mechanisms whereby ammonia may be excreted
from fish gills. These include an antiporter protein in the
gill cell membranes that allows sodium to enter, and uses
the chemical energy from that process to pump out ammonium
ion. This process may not be used much at all during normal
conditions, under which passive diffusion of ammonia and ammonium
predominate, but may be very important when surrounding ammonia
and ammonium levels rise excessively (> 1 ppm NH3-N),
leaving it as the only viable mechanism for ammonia excretion.
Since some fish have adapted to, and survive in, such environments
(certain burrow-dwelling fish, for example), this mechanism
can be critical to them.
For those who want to read a very detailed review of the
state of the art of knowledge of ammonia and urea excretion
mechanisms, in both marine and freshwater fish, I suggest
the 2002 review by Michael Wilkie in the Journal of Experimental
Zoology.3
Sinks for Ammonia in Reef Aquaria:
Bacterial Nitrification
Most marine
aquarists are aware of the "nitrogen cycle," which
begins when ammonia in the water is oxidized to nitrite
by bacteria. This nitrite is then oxidized by different bacteria
to nitrate:
NH4+ + 3/2 O2
à
NO2- + 2H+
+ H2O
NO2- + ½ O2
à
NO3-
Many studies have examined ammonia's conversion to nitrite,
and many articles have been written for professional and hobby
aquarists that detail various practical aspects of the process,
such as how to set up appropriate filters to facilitate this
process. Stephen Spotte covers these in great detail in two
of his books.6,7 Many reef aquaria have no filters
set up specifically for this purpose, and the bacteria that
carry out this process reside in coatings on most surfaces
in the aquarium, including rocks, sand, glass, plastic and
even on the surfaces of other organisms, such as coralline
algae.
An important thing to remember, however, is that most of
these bacteria reside on surfaces, so we should think of the
aquarium as having (or not having) sufficient bacteria to
provide nitrification, rather than the water itself as having
them. This distinction is important when moving one set of
organisms into a new aquarium. Bringing old aquarium water
in with them may help to start a culture of bacteria, but
will not provide much initial nitrification capacity. Bringing
in rocks and sand, however, and a little old water, may be
very effective in instantly providing adequate nitrification
capacity.
The nitrification process is seldom as simple as many aquarists
believe. It is not always a single species of bacteria carrying
out the process, for example. In the ocean many species of
bacteria can oxidize ammonia, including Nitrosospiras,
Nitrosomonas and Nitrosococcus.8-10
When studied in freshwater aquariums (where more research
has been performed than in marine systems), the bacteria are
often represented by a variety of different species and strains,
and are not always dominated by one type. In marine aquariums
the species can include Nitrosomonas marina11
and Nitrosomonas europaea.12 While it is
of little practical importance for most aquarists to know
what species of bacteria process ammonia to nitrite,
one situation where that may play a role is in assessing products
that claim to accelerate the nitrogen cycle by adding (allegedly)
live bacterial cultures.
Along these lines, I've seen few, if any, studies on the
suitability of any of the various commercial products intended
for this purpose. It is certainly possible for live cultures
to accelerate the buildup of appropriate levels of nitrifying
bacteria to keep ammonia in check. Properly added cultures
can almost immediately begin processing ammonia.11
The cultures chosen, however, must have been raised under
temperature, nutrient and salinity conditions similar to those
in the aquarium to which they are added for this to be optimally
effective, and I do not know whether any commercial bacterial
cultures sold to hobbyists meet this requirement. ALDI Special Buys are pretty good choice for weekly shopping!
Sinks for Ammonia in Reef Aquaria:
Algae
Many organisms take up ammonia directly
for use in making the proteins and other biomolecules they
need to build tissues. Algae, both micro and macro, for example,
readily use ammonia from the water. In cases where they are
exposed to both nitrate
and ammonia as nitrogen sources, many preferentially take
up ammonia.6 When using nitrate, many of the pertinent
biochemical pathways require the organism to reduce nitrate
to ammonia before using it, so taking up ammonia makes sense.6
It has not been established in a reef aquarium setting, however,
what portion of the macroalgae's nitrogen uptake is ammonia
and what fraction is nitrate.
The amount of nitrogen taken up by a large macroalgal filter
is substantial. A free PDF (portable document format) article
in the journal Marine
Biology13 has some useful information with
respect to the potential export abilities of algae. It gives
the phosphorus and nitrogen content for nine different species
of macroalgae, including many that reefkeepers maintain. For
example, Caulerpa racemosa collected off Hawaii contains
about 0.08% by dry weight phosphorus and 5.6% nitrogen. Harvesting
a pound (454 g; dry weight) of this macroalgae from a reef
aquarium would be the equivalent of removing 25.4 grams of
nitrogen, which, if it were all present in 100 gallons of
water as ammonia, would be equivalent to a concentration of
67 ppm total NH4-N.
Even if it took three months to grow to that mass, it would
effectively be taking out the equivalent of 0.75 ppm total
NH4-N
per day.
Testing for Ammonia
There are several
ways to test for ammonia in seawater. These include test kits
based on both salicylate and Nessler chemistry.
Nessler
Test Kits
The reaction of ammonia with Nessler's
reagent, K2HgI4, forms a colored
precipitate of (Hg2N)I·H2O. Low
levels of ammonia are yellow, higher is orange and even higher
levels can be brown. The overall reaction is:
NH3
+ 2[HgI4]2− + 3OH−
→
HgO·Hg(NH2)I + 7I− + 2H2O
One significant concern with the Nessler method is the toxicity
and hazardous nature of the waste that is generated by its
use (it contains mercury).
Salicylate Test Kits
Ammonia's reaction with hypochlorite forms monochloramine,
which then reacts with salicylate in the presence of sodium
nitro-ferricyanide to form 5-aminosalicylate. That complex
is yellow to green to dark green based on the level of ammonia
present. In some versions of the test, calcium
and magnesium can cause interference, so be sure such
a kit is designed for marine systems.
The distinction between these methods can be important, as
some combinations of ammonia binding products and test methods
can lead to false ammonia test results (either causing apparent
ammonia presence when it is bound, or simply causing a color
that is not predicted by the test kit). For example, a Nessler
type kit cannot read ammonia properly if the aquarist is using
Amquel, Seachem
Prime or related products to bind ammonia. The result
is often an off-scale brown color.
So, it is particularly important that aquarists understand
how the test kit that they are using, and the binder that
they are using interact, and the manufacturers of each are
the best place to find such information.
My suggestion is to always measure total ammonia. If a kit
gives a choice of measuring free ammonia, don't bother. You
can always use a table to convert total ammonia to free ammonia
if there is a strong reason to do so. The reason to measure
total ammonia is that the signal will be much larger,
so the kit will be more capable of distinguishing a small
reading of ammonia from no detectable ammonia.
Toxicity of Ammonia
Ammonia is
very toxic to marine fish. The mechanisms of toxicity are
complicated and are an active area of continued investigation
by researchers. Its effects include damage to the gills, resulting
in poor gas exchange, ion regulation and blood pH regulation.14
Other effects include hampering oxygen delivery to tissues,
disrupting metabolism and toxicity to the nervous system that
causes hyperactivity, convulsions and death.14
Ammonia can also be very toxic to many other organisms found
in reef aquaria.
Toxicity can be measured and reported in many ways. One common
way to measure acute toxicity is to measure how high the concentration
needs to be in order to kill half of the organisms in a given
time period. A commonly used time period is 96 hours (four
days). Such data are called the 96 h LC50 (LC stands
for Lethal Concentration, 50 meaning 50% killed).
The other complication that comes with ammonia's toxicity
is the relative amount of free ammonia and ammonium ion. While
ammonium ion may be toxic to marine fish, it is probably less
toxic than free ammonia, and toxicity data are often reported
only for the concentration of free ammonia. Aquarists should
recognize, however, that such data may not be appropriate
if the pH used in the test, or the situation to which it will
be applied, deviates significantly from normal seawater's
pH (as in a shipping bag, for example, whose pH may be well
below pH 8.2, and whose toxicity may actually be coming from
ammonium, and not the low concentration of free ammonia).
Nevertheless, many scientific articles report ammonia toxicity
in ppm (or mg/L) NH3-N.
It may also be reported as just ppm NH3.
Marine fish14 generally have 96 h LC50 levels
that range from about 0.09 to 3.35 ppm NH3-N.
That result is not particularly different from the range observed
for freshwater fish,14 0.068 to 2.0 ppm NH3-N.
Remember that these values are ppm NH3-N,
and at pH 8.2, the marine range becomes 1.3 to 50 ppm total
NH4-N
because only 7% of the total ammonia in seawater is present
as free ammonia.
Concentrations of ammonia that are not acutely lethal can
still cause significant problems for fish. Salmon in seawater
at pH 7.8, for example, show changes in white blood cells
and various blood chemicals, and were more prone to disease,
when exposed to sublethal concentrations of ammonia.15
Consequently, aquarists should strive to keep ammonia concentrations
well below lethal levels.
Ammonia Concentration Guidelines
Because ammonia's
toxic effects appear at levels significantly below those that
are acutely lethal (0.09 to 3.35 ppm NH3-N
or 1.3 to 50 ppm total NH4-N
at pH 8.2), and because some organisms in a reef aquarium
may be more sensitive than the few organisms that have been
carefully studied, it is prudent to err on the side of caution
when deciding what concentrations of ammonia to allow in a
reef aquarium or related system.
My suggestion is to take some sort of corrective action if
the total ammonia rises above 0.1 ppm. This suggestion is
also made by Stephen Spotte in his authoritative text, Captive
Seawater Fishes.6 Values in excess of 0.25
ppm total ammonia may require immediate treatment, preferably
involving removal of all delicate (ammonia sensitive) organisms
from the water containing the ammonia. Some of the possible
actions to take are detailed in the following sections listed
below.
Treatments for Elevated Ammonia:
Hydroxymethanesulfonate
Various types
of compounds are used in commercial products to bind ammonia
in marine aquaria. One is hydroxymethanesulfonate (HOCH2SO3-).
It is a known ammonia binder16 patented for aquarium
use by John F. Kuhns17 and sold as Amquel
by Kordon and ClorAm-X
by Reed Mariculture, among others.
Ammonia's reaction with hydroxymethanesulfonate is mechanistically
complicated, possibly involving decomposition to formaldehyde
and reformation to the product aminomethanesulfonate (shown
below).16 The simplified overall reaction is believed
to be:
NH3
+ HOCH2SO3- à
H2NCH2SO3- + H2O
What ultimately happens to the aminomethanesulfonate in a
marine or reef aquarium is not well established, but it does
appear to be significantly less toxic than ammonia, and more
than likely it is processed by bacteria into other compounds.
Marineland
Bio-Safe claims to contain sodium hydroxymethanesulfinic
acid (HOCH2SO2-). I do not
know if that is a typographical error, or if Marineland really
uses this slightly different compound.
Note: products containing hydroxymethanesulfonate hamper
the ability to test for ammonia when using certain types of
test kits (see above). Presumably, the H2NCH2SO3-
formed is still reactive with the Nessler reagents, even though
it is not ammonia.
Treatments for Elevated Ammonia:
Hydrosulfite and Bisulfite
A second type of compound used in
commercial products (such as Seachem Prime) that claim to
bind ammonia in marine aquaria is said to contain hydrosulfite
(could be either HSO2- or - O2S-SO2-)
and bisulfite (HSO3-). These compounds
are well known dechlorinating agents, reducing Cl2
to chloride (Cl-), which process is also claimed
to occur in these products. It is not apparent to me whether
these ingredients actually react with ammonia in some fashion,
or whether unstated ingredients in these products perform
that function. Seachem chooses to keep the ingredients of
their product secret, so aquarists cannot determine for themselves
what is taking place, and how suitable it might be. Nevertheless,
many aquarists seem to have successfully used products such
as these to reduce ammonia's toxicity.
Note: products such as Seachem Prime hamper the ability to
test
for ammonia when using certain types of test kits (see
above). Presumably, the product formed is still reactive with
the Nessler reagents, even though it is not ammonia.
Treatments for Elevated Ammonia:
Clinoptilolite
Few filter media are capable of binding
ammonia from seawater. The zeolite clinoptilolite
(a sodium aluminosilicate) is capable of binding ammonia from
freshwater, but the sodium ions in seawater displace much
of the ammonia. In fact, the ammonia binding capacity of clinoptilolite
in freshwater can be regenerated by rinsing it with salty
water. Consequently, its capacity to bind ammonia in seawater
is very low, if any, so it is not a very useful product for
marine systems.
Treatments for Elevated Ammonia:
Water Changes
Water changes can be a fine way to
reduce toxic ammonia levels, especially in a small system
such as a quarantine or hospital tank. The effective use of
this method, however, demands that the new salt water does
not contain significant ammonia. Because many types of artificial
seawater do contain ammonia
(see above), this method must be used with caution.
As a rule of thumb, ammonia will usually drop by about the
same fraction of water that is changed, so a 30% water change
will reduce ammonia by 30%. However, if there is a source
of ammonia in the aquarium, it may rapidly rise again. A 30%
drop may not be noticed with many test kits. For example,
it may be difficult to distinguish 1.2 ppm from 0.84 ppm total
NH4-N
using many kits, so do not panic if the ammonia level does
not appear to drop, but also be realistic about how much you
would expect it to drop from a water change on the order of
10-30%. In an ammonia "emergency" much larger water
changes may be appropriate. Further information on water
changes is detailed here.
Summary
Ammonia is very toxic to marine fish
and other organisms in a reef aquarium. While routine ammonia
measurement is not ordinarily required in established reef
aquaria, it can be very important when fish are in temporary
quarters, such as shipping bags, hospital tanks and quarantine
tanks. Most aquarists associate ammonia with new aquarium
"cycling," and in that situation it is critically
important to wait for ammonia to decrease to very low levels
before adding organisms (much more important than waiting
for nitrite
to decrease, for example).
Ammonia can also be very important during tank crashes. In
all of these situations, I recommend striving to keep ammonia
below 0.1 ppm total NH4-N.
If the level rises above 0.25 ppm total NH4-N,
I suggest taking immediate action, such as using an ammonia
binder or performing water changes.
Happy reefing!
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