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Published: 2014-07-23, Updated: 2017-01-29

Experiments: Nitrification – myths vs. facts

2018 Comment

Another experiment that was not done by me, but by one American aquarist who gave me permission to describe it here. This experiment refutes the widespread myth that we need the most efficient filtration with as much filtration area as possible to remove toxic ammonia from the water. It turned out that even in an absolutely empty aquarium without plants, substrate and various filter media, enough bacteria will grow (reproduce) that are able to break down an incredible 30 ppm NH3 in 24 hours. Thus, a normal planted aquarium most likely does not need any biological filtration (at least not for ammonia removal), and the hype around the ever more perfect filters seems odd (unfounded) in the light of this experiment.

Of course, I do not question the need to remove dissolved organic substances, microscopic algae or other (fine) mechanical pollution from water. But for this purpose, I think it is preferable to use activated carbon in combination with some fine mechanical filtration.

This article is based on an experiment by Linn Fahrenkrug from Iowa, USA (alias Immortal1) described on the plantedtank.net forum.

Nitrification

Only 0.002% of all bacteria in the aquarium are nitrifying.
SPOTTE, Stephen H. Fish and invertebrate culture:
     Water management in closed systems
(1970).

Nitrification is the biological conversion of ammoniacal nitrogen (NH3/NH4+) to nitrites (NO2-), and subsequently to nitrates (NO3-). Ammoniacal nitrogen is produced in the aquarium as a waste substance of some animals' metabolism and also in microbial decomposition of organic detritus. Ammonium itself (NH4+) is not toxic to fish, because fish as well as plants are capable of regulating its intake, unlike ammonia gas (NH3), which intake the fish and plants can't control, and is therefore potentially much more toxic to them. Ammonia gas (NH3) is formed from dissolved ammonium (NH4+) at higher pH (the higher the pH, the greater the amount of ammonium is converted to toxic ammonia). Aerobic microbes are needed to convert ammoniacal nitrogen to nitrites and subsequently to nitrates. In the aquarium hobby, the idea (or rather the myth) spread that for the removal of ammoniacal nitrogen (NH3/NH4+) from the aquarium water it is necessary to use a good biological filter with a large colonizing area for a sufficiently large colony of nitrifying microbes. Few, however, know that nitrification is only the tip of the iceberg in different kinds of decomposition processes. Of all the microorganisms involved in the decomposition of organic substances in the aquarium, nitrifying bacteria form only a negligible (though important) fraction.

Besides the fact that nitrification "takes a back seat" in the aquarium, nitrifying bacteria are moreover very effective "in their field" (or domain), and therefore only a relatively small amount is needed to convert ammoniacal nitrogen to nitrite and nitrate. Of the total of 18 million bacteria in 1 gram of the top layer of filter sand, only about 425 bacteria (0.002%) belong to autotrophic nitrifiers. The rest (99.998%) belongs to heterotrophic bacteria.

It is said in the fish farming literature that about 30-34 m2 of filter media area is needed to break down the ammoniacal nitrogen produced from 1 kg of fish feed at the water temperature of 77°F (25°C). The amount of food that fish must consume daily to make them healthy and grow well is about 1.0-2.5% of their body weight. Assuming we have 100 pieces of adult Neon Tetra of 2 inches (5 cm) in length and 1.85 grams per piece in our aquarium (185 g in total), they need less than 5 g of food per day. This means that only 0.17 m2 of the filter media area (34*0.005) is sufficient to break down the ammoniacal nitrogen produced by the fish from 5 grams of feed. If 1ℓ of Seachem Matrix filter media has a surface area of 1,500 m2, then 0.17 m2 of surface area corresponds theoretically to just 0.000113ℓ, i.e. 0.113 mℓ of filter media (0.17/1,500).

Filter media test

Test parameters:

Author of the test: Linn Fahrenkrug (Immortal1) from Iowa, USA

  • Aquariums: 2x 12G (45ℓ)
  • Filters:
  • 1x Marineland Magniflow C360
      (355 gph = 1350 ℓ/h, filter media capacity: 55 fl oz = 13ℓ) + 3/8" = 275mℓ of Seachem Matrix
    1x Eheim Pro II 2028
      (277 gph = 1050 ℓ/h, filter media capacity: 31 fl oz = 7.3ℓ) + 3/8" = 275mℓ of pumice
  • Ammonia source: Blue Ribbon Clear Ammonia (contains 3.5% of ammonia)
  • Calculator used for dosing calculations: fishforums.net
  • (calculated as 10G of water in aquarium + 2G water in filter = 12G = 45ℓ of water in total)
  • Initial water parameters: KH 7, GH 12, pH 7.6
  • Water temperature: 77-84°F (25-29°C) [active cooling with small fans]
  • Brand of the drop test kit used: API (Ammonia, Nitrite, Nitrate)
Untreated tap water was put into the aquariums and the tanks were filtered (empty) for several days before the test was started. The day before the test, water was treated with Seachem Prime.
 

Seachem Matrix vs pumice

According to the safety data sheet of the Seachem Matrix filter media, this filter medium is nothing but 100% pumice. For comparison, I refer to a common pumice of various grain sizes: generalpumiceproducts.com.

1ℓ of Seachem Matrix          = $11.55
4ℓ of Seachem Matrix          = $31.80
5ℓ of Pumice (Czech Republic) =  $3
7ℓ of Pumice (GeneralPumice)  = $25.00

  Seachem Matrix

  Pumice

  Calibration test: 10, 13, 16, 20, 40 ppm NH3/NH4+
  High nitrite (NO2-) concentration before filter cycling

  Day #38: 30 ppm of ammoniacal nitrogen completely degraded in 23 hours
(yellow tube = 0 ppm NH3/NH4+, blue tube = 0 mg/ℓ NO2)
  Extremely high nitrate concentration after a few days without water change →
more than 160 ppm NO3-

Both filter media (Seachem Matrix as well as Pumice) managed to completely remove 30 ppm of ammoniacal nitrogen in 24 hours.
The dosage of 34.8 ppm any filter media did not completely break down in 25 hours.
Thus, both filter media seem to have the same nitrification capacity, which just confirms that it is the same material, i.e. pumice.

Media mix vs bio-balls vs empty filter

Grow Out (provizorium) = container with filter that has Fluval Biomax, Eheim Substrat Pro, Seachem Matrix, and Pumice as the medium.
[Leftmost aluminium basin (volume approx. 6.5G = 25ℓ).]
Filter: Aquatop CF500UV (527 gph = 1995 ℓ/h, filter media capacity: 1G = ~4ℓ)
  1st compartment: Seachem Matrix + pumice (2*275 mℓ)
  2nd compartment: Eheim Substrat Pro (900 mℓ)
  3rd compartment: Fluval Biomax (1ℓ)
  4th compartment: coarse foam (1ℓ) → very bottom

Bare Tank (empty aquarium) = 45ℓ aquarium with filter without any filter media.
[Middle aquarium.]

Bio Balls (plastic balls) = 12G = 45ℓ aquarium with filter, which has 275 mℓ (25pcs) of bio-balls as the medium.
[Right aquarium.]


Warning: During the previous experiment, it emerged that high doses of ammoniacal nitrogen led to an extreme increase in nitrate concentration, resulting in total collapse of nitrifying bacteria (the bacteria either died out or no longer broke down NH4+ and NO2-, or perhaps there was no longer any additional place for NO3- in the tank). For example, if 15 ppm NH4+ (= 45 ppm NH4+ in total) is added to water for 3 days in succession, it can theoretically result in up to ~115 ppm of NO2- (45*2.55) or ~155 ppm of 3 NO3- (45*3.44). Thus, in order for nitrate levels in test aquariums not to increase to extremely high concentrations that can lead to the collapse (inhibition) of nitrification, it is important to often change large amounts of water.

1 ppm NH3 = 2.70 ppm NO2- = 3.64 ppm NO3-
1 ppm NH4+ = 2.55 ppm NO2- = 3.44 ppm NO3-

  Cycled bio-ball
  Volume: 275 mℓ = 25pcs
  25pcs of bio-balls in the filter

  Day #14: 30 ppm of ammoniacal nitrogen completely break down in 24 hours
(yellow tube = 0 ppm NH3/NH4+, blue tube = 0 ppm NO2-, red tube = high NO3- level)
  Day #15: scraping the bacterial coating from the aquarium glass with a razor blade, changing 100% of water, filling in new water treated with Seachem Prime, and dosing 30 ppm of ammoniacal nitrogen.
Day #16: measured 0.25 ppm NH3/NH4+ and 1.0 ppm NO2- after 24 hours.

Empty filter vs 1ℓ of Fluval Biomax

Bare Tank (empty aquarium) = 45ℓ aquarium with Eheim Pro II 2028 filter without filter media.
Fluval Biomax = 12G = 45ℓ aquarium with Aquatop CF500UV filter, which has 1ℓ of Fluval Biomax as filter medium.

Dose of 30.6 ppm NH3 (no picture):
  Bare Tank: 0 ppm NH3/NH4+, 0 ppm NO2- in 24h (Eheim Pro II 2028)
  1ℓ BioMax: 0 ppm NH3/NH4+, 0 ppm NO2- in 24h

Dose of 38 ppm NH3:
  Bare Tank: >4 ppm NH3/NH4+, >2 ppm NO2- in 24h (Eheim Pro II 2028)
  1ℓ of BioMax:   0 ppm NH3/NH4+,   0 ppm NO2- in 24h
(yellow/green tube = NH3/NH4+, blue/violet tube = NO2-)
Dose of 38 vs 46 ppm NH3:
  Bare Tank: 38 ppm NH3 → 0 ppm NH3/NH4+,   0 ppm NO2- in 48h! (Eheim Pro II 2028)
  1ℓ BioMax: 46 ppm NH3 → 0 ppm NH3/NH4+, >2 ppm NO2- in 24 hours
(yellow tube = NH3/NH4+, blue/violet tube = NO2-)

1) Moving the filter media

A complete filter media (= 1ℓ) was removed from the Fluval Biomax aquarium and moved to the Bare Tank. Then 46 ppm of NH3 was added to both aquariums again.

Dose of 46 ppm NH3:
  Bare Tank: 0 mg/ℓ NH3/NH4+, 0 mg/ℓ NO2- in 24h (Eheim Pro II 2028)
  0ℓ BioMax: 0 mg/ℓ NH3/NH4+, >2 mg/ℓ NO2- in 24h

 

Bare Tank = 12G = 45ℓ aquarium with Eheim Pro II 2028 filter, which has 1ℓ of Fluval Biomax media (originally placed in Fluval Biomax aquarium).

Fluval Biomax = 12G = 45ℓ aquarium with Aquatop CF500UV filter without filter media.

This test should verify if most of the nitrifying bacteria were concentrated in the filter medium or is located elsewhere (e.g. on filter walls, in hoses, and the like). If all the filter media was removed from the Aquatop filter, and yet it was able to break down most of the ammonia, it would mean that the filler itself does not play very important role in the nitrification. Conversely, if the nitrification would collapse after the removal of the filter media, it would mean that the filter media plays an important role.

From the results it can be seen that the empty filter without the filter media gave practically the same results at the dose of 46 ppm NH3 as with the filter media. In both cases, it was able to break down all ammonia in 24 hours, but lagged with nitrite.

An empty aquarium (Bare Tank), which, without filter media, was able to completely remove 30.6 ppm of NH3 at most in 24 hours, was able to remove up to 46 ppm of NH3 with the added filter media.

The results in this case are not entirely clear. While, after removing the filter media, the first aquarium has break down virtually the same amount of ammonia and nitrite as with the filter media (which would clearly indicate that the filter media have virtually no effect on the nitrification efficiency), after adding these media back to the empty filter, the efficiency of nitrification in the second aquarium increased (the media helped to achieve better results).

2) Cleaning the filter

Prior to the next test, the filter and aquarium went through a thorough cleanup consisting in removing all the biofilm that formed on the filter- as well as aquarium walls during the whole experiment (see the next picture).

Biofilm (slime) inside the filter before cleaning.
Filter and aquarium after thorough cleaning.

Detail of cleaned filter compartments.
Detail of cleaned pump head with rotor.

After thorough cleaning of the filter and treating the water with a double dose of Seachem Prime, 1ℓ of a cycled Fluval Biomax filter media (which was previously removed and transferred to an empty filter in the Bare Tank aquarium) was inserted into the filter. Subsequently, 16 ppm of NH3 was added to the water.

The goal of this test was to find out what share in the total nitrification the filter media themselves have.

If the bacteria in the filter media could handle the given amount of ammonia in 24 hours without any problems, it would mean that the filter media account for at least 50% of the total nitrification. If filter media did not cope with the given amount of ammonia, it would suggest that filter media are most likely not the place where nitrifying bacteria concentrate.

Dose of 16 ppm NH3:
  1ℓ of BioMax: 0 ppm NH3/NH4+, 0 ppm NO2- in 16h
(yellow tube = NH3/NH4+, blue tube = NO2-)
Dose of 33 ppm NH3:
  1ℓ of BioMax: 0 ppm NH3/NH4+, 0 ppm NO2- in 16h
(yellow tube = NH3/NH4+, blue tube = NO2-)

  A view of the biofilm (slime) in the empty compartment of the filter on the third day after thorough cleaning.

As can be seen from the pictures above, the bacteria in the filter media were able to remove 16 ppm at first, and the next day 33 ppm NH3 per 24h, suggesting that the filter media provide at least a comparable colonizing area as the removed biofilm.

In addition, nitrifying bacteria are able to multiply quite quickly, or only a relatively small amount of bacteria suffices to effectively remove even a relatively large amount of ammonia.

On Saturday, the filter was thoroughly cleaned and only 3 days later it was again covered with a continuous layer of biofilm (cf. detail of the cleaned compartments of the filter in the pictures above with the right image).

Aquarium with air stone vs sponge filter

No Filter (aquarium w/o filter) = 12G = 45ℓ aquarium without filter, only with an air stone.

Sponge = 12G = 45ℓ aquarium with inner sponge filter (designed for 20G = 75ℓ aquarium).

Progress:
Day #1:
Dose of 1 mℓ Seachem Prime
  No Filter: 0.5 ppm NH3/NH4+, 0.5 ppm NO2-, 20 ppm NO3- (initial concentration)
Day #7:
  No Filter: 4 ppm NH3 → 0 ppm NH3/NH4+, ~2 ppm NO2- in 24h
  Sponge: 30 ppm NH3 → 0 ppm NH3/NH4+, ~0 ppm NO2- in 24h
Day #15:
  No Filter: 4 ppm NH3 → 0 ppm NH3/NH4+, ~0.1 ppm NO2- in 24h
  Sponge: 34 ppm NH3 → 0 ppm NH3/NH4+, ~0.1 ppm NO2- in 24h
Day #16:
  No Filter: 4 ppm NH3 → 0 ppm NH3/NH4+, 0.25 ppm NO2- in 24h
  Sponge: 45 ppm NH3 → 0 ppm NH3/NH4+, 0.25 ppm NO2- in 24h

An empty aquarium without filter, only with an air stone, could barely remove the dose of 4 ppm NH3. The next day (after changing 100% of water and dosing the same amount of ammonia), the nitrite concentration started to rise again (see day #16).

A small sponge filter (bubble blower) managed to break down an incredible dose of 70 ppm NH3 in 24 hours, which no other filter (or filter media) tested was able to manage (see day #18).

The dose of 88 ppm NH3 was not able to completely break down (see day #19).

Day #17:
  No Filter: test terminated
  Sponge: Dose of 56 ppm NH3 → 0 ppm NH3/NH4+, 0 ppm NO2- in 24h
(yellow tube = NH3/NH4+, blue tube = NO2-)
Day #18:
  Sponge: Dose of 70 ppm NH3 → 0 ppm NH3, 0 ppm NO2-, >160 ppm NO3- in 24h
(yellow tube = NH3/NH4+, blue tube = NO2-, red tube = NO3-)
Day #19 (no picture):
  Sponge: Dose of 88 ppm NH3 → 0 ppm NH3/NH4+, 0.5-1.0 ppm NO2- in 24h (the filter was no longer able to remove all the nitrites)

Conclusion

From the above experiment many interesting things follows:

  1. An empty aquarium with a filter without filter media can break down at least the same amount of ammoniacal nitrogen as an aquarium with a handful of high-quality filter media (Seachem Matrix alias pumice) in 24 hours.
  2. The use of filter media in the filter is most likely unnecessary (at least in terms of nitrification) in a planted aquarium with a reasonable fish stock.
  3. To break down such an extreme concentration of ammoniacal nitrogen as 30 ppm NH3/NH4+ virtually zero filter media is needed. This means that even the minimal amount of nitrifying bacteria that can be produced on ordinary surfaces inside an aquarium, in hoses or in an empty filter, is enough to break down virtually any amount of ammoniacal nitrogen that could occur in a normal planted aquarium.
  4. To say that there may be so-called "peaks" or mini-cycles in the aquarium, during which there is a local, short-term or sudden increase in the concentration of ammoniacal nitrogen, which can lead to the germination and subsequent proliferation of algae, are (given the enormous nitrification capacity of the empty aquarium) most likely nonsense. It can be assumed that even some reasonably large increase in ammonia concentration (e.g. dead fish decaying somewhere unnoticed) may not pose any problem for an already established colony of nitrifying bacteria, especially considering that even in an empty aquarium (with a filter without filter media) it can arise a large enough colony of nitrifying bacteria to break down an incredible 30 ppm of NH3/NH4+ in 24 hours. So why should an aquarist have an oversized filter when an empty aquarium (without filter media, substrate, plants, and other potential areas for bacteria) can host such a "huge" amount of nitrifying bacteria that there is a reserve for at least 30 ppm of NH3/NH4+? Or does anyone find such a reserve inadequate? Alternatively, a 275 mℓ of Seachem Matrix media (or the same amount of ordinary pumice) provides a nitrification capacity for no more than 30 ppm NH3/NH4+. An empty aquarium represents a nitrification capacity to break down at least 30 ppm NH3/NH4+ (maybe more ... the test was terminated when this value was reached). Is there anyone who sees any difference between these 30 vs 30 ppm?
  5. For an idea of how much ammoniacal nitrogen is generated from 2 g of fish food, which is the recommended feed dose for 100pcs of adult (2 inches [5 cm] long) Neon Tetra in 25G (100ℓ) aquarium:

    • Aquarium volume: 25G (100ℓ)
    • Fish weight in aquarium: 185 g (100pcs * 1.85g)
    • Daily feed dose: ~2 g (= 1% of body weight)
    • Protein content in fish feed: 23%
    • Nitrogen content in proteins: 16%
    • Calculation formula: Amount of NH3 (g) = g feed * (% protein in feed / 100) * (% N in protein / 100) * (% waste N / 100) * (1.216 NH3 / 1 N)
    • Calculation: 2 * (23/100) * (16/100) * (61/100) * (1.216/1) = 2 * 0.23 * 0.16 * 0.61 * 1.216 = 0.055 g NH3

    100 pieces of Neon Tetra fed daily by 2 g of feed produce less than 0.055 g = 55 mg NH3 every day, which when dissolved in 25G (100ℓ) aquarium is 0.55 ppm NH3. This means that in an empty 100ℓ aquarium there are so many nitrifying bacteria that it would not be a problem for them to break down ammonia from 5,454 pcs of adult Neon Tetra (or 10 kg of well fed fish).

  6. The importance of filtration (or filter media) appears to be overestimated (at least in terms of nitrification). It is also quite possible that many filters may increase ammonium concentration in the aquarium rather than reduce it effectively. Some filters may function as "traps" and producers rather than "degraders" of organic waste.
  7. A high concentration of ammoniacal nitrogen results in an increase in nitrite concentration, which is converted by bacteria into nitrates. Consequently, with high production of ammoniacal nitrogen, nitrates will accumulate to a large extent in the aquarium (up to 76 ppm of NO2- followed by 103 ppm of NO3- can be theoretically generated in the aquarium from 30 ppm NH4+). High nitrate concentrations (>150 ppm) may lead to inhibition (collapse) of nitrification. Therefore, in order to avoid nitrification decline or interruption at high nitrate concentrations, it is important to change water regularly when cycling the aquarium.
  8. Filtration has several functions, of which nitrification (the biological conversion of ammoniacal nitrogen to nitrites and subsequently to nitrates) is only one of them. Regarding nitrification, it seems to be proven that filtration (or filter media) is superfluous in a normal, well-functioning planted aquarium. However, filtration may have other useful functions in the aquarium, such as mechanical removal of fine particles from water, chemical absorption and adsorption of harmful molecules into the resin or other filter media, or biological (microbial) decomposition of complex organic compounds into simpler compounds or elements (nutrients) → so-called mineralization. Whether the different types of aquarium filtration actually help in the mineralization itself (i.e. in the conversion of complex organic compounds into simple inorganic plant nutrients) remains to be verified. The experience of many aquarists suggests that some types of filters may act rather as organic waste producers in the aquarium instead of water purifiers. The filters compete with plants because some nutrients can often precipitate in them (most often iron and phosphorus). In addition, some filter media will soon become clogged with organic detritus or inorganic precipitates, and anaerobic decomposition will begin to occur with all the negative consequences (e.g. production of toxic compounds). This also leads to disruption of mineralization and begining of humification, the products of which are mainly hard-to-decompose humins, humic acids and fulvic acids. Increased concentrations of dissolved organic matter in the water can then promote the growth of some algae.
  9. The most effective filter medium in the degradation of ammonia (nitrification) has proved to be an ordinary foam filter called sponge filter. It managed to break down an incredible 3.15 grams of pure ammonia (70 ppm) in a 12G (45ℓ) aquarium in 24h.
    For an idea: This amount of ammonia is produced by more than 10 kg of adult Neon Tetras in 24 hours.

Information from the experiment taken and the pictures used with the kind permission of the author.

Marcel Goliaš © 2019