Nitrogen Removal in the Planted Aquarium

My Background and How I Wrote ‘Ecology of the Planted Aquarium’
I was raised in a family of avid aquarium hobbyists and have been keeping aquariums for over 40 years, but strictly as a hobby. My formal training (Bachelor of Science in Microbiology) and my jobs as a research technician in various medical fields (infectious disease, cystic fibrosis, sickle cell anemia, etc) were far removed from botany or ecology. (I am presently working at a government research institute doing experiments in intracellular signaling and hormone receptor binding.)

The pathway to my book began in 1987. I had not the slightest thought of writing an aquarium book. I started keeping tanks again but with the priority that they have good plant growth. Because plant growth had consistently failed in all my past tanks, I decided to ignore all the aquarium literature guidelines. So I set up a tank next to a sunny window, and instead of pure gravel, I layered the tank bottom with 3-6 cm of ordinary soil covered with 2-3 cm of gravel. To my surprise, not only did the plants thrive, but also the fish seemed healthy.

Figure 1: Ecology of the Planted Aquarium. Professional reviews, a synopsis, and sample pages of Ecology of the Planted Aquarium can be found on the AtlasBooks.com website. More reviews, mostly from aquarium hobbyists, can be found on the Amazon.com website.

At the same time, I was becoming dismayed with the lack of scientific information in the aquarium hobby. For example when excessive Hydra growth became a problem in my fish grow-out tanks, I found the basic information I needed on Hydra’s life cycle not in the aquarium literature but in a university biology library. Also, I came across a wonderful scientific book on lake ecology (Robert Wetzel’s Limnology). This textbook and one on soils (Russell’s Soil Conditions and Plant Growth, 1988) absolutely fascinated me. These books had information on subjects (carbon recycling, allelopathy, soil chemistry, etc) that applied to aquarium keeping, but that I had never seen before.

Inspired by the two textbooks, I began my scientific literature search at the 3 nearby universities. I immediately devised a filing system for the hundreds of scientific papers I eventually collected. I carefully read, studied, and subject-indexed every paper I read. If I had a question about a paper, I corresponded with the scientist involved. The more I studied the topic, the more fascinated I became. I began to write articles about my findings for the aquarium magazines. I knew most hobbyists had never seen this information despite the fact it had been published decades ago. For example, Ferguson’s excellent study showing that a duckweed species greatly preferred ammonium to nitrates was published in 1969! Yet aquarium literature in the 1990’s invariably showed plants taking up nitrates while filter bacteria removed the ammonia.

In 1995 after I had written many articles on various topics, I considered writing a book. However, organizing the information into a book was not easy. Unlike writing a taxonomy book where you just list plant (or fish) species and describe them, this book would be on many different topics (soil, plant, and water chemistry and ecology) and how they related to aquariums. I had to find a way to logically connect all these diverse topics. Because scientists were already familiar with the information, I wrote the book for hobbyists. I could only hope that aquarium hobbyists would be as fascinated as I was by the information. However, writing a scientific book for non-scientists is difficult. Unfortunately, scientists sometimes write in a difficult-to-read manner that confuses ordinary people. I began to realize why all this wonderful information had not reached the aquarium hobby! Thus, after I had written a draft of the book, my next goal was to make it understandable to hobbyists. I included ‘Questions and Answers’ to show how the scientific information applied to aquariums. I edited the manuscript over and over again looking for ways to say things more clearly. I knew that my book would be easily dismissed if hobbyists couldn’t understand it. It had to be written in plain English. To this end, I read books on how to write and present scientific information clearly. For example, I replaced convoluted and unnecessarily difficult phrases such as the “The utilization of ..” with the much clearer “I used ..”

The book also had to be accurate. What person would believe a book, especially one about science, that had misspellings and inaccuracies? Hobbyists would either be confused or dismiss the book altogether. Another task was to standardize the units of measure throughout the book. For example, different scientists variously expressed a metal’s concentration in water as microatoms per microliter, millimolar, micrograms per milliliter, etc. The information was accurate but confusing. Thus, it was essential that I convert and standardize all these values to mg/l (milligrams per liter). As to publishing the book, I sought academic publishers with little success. In 1996 I sent a brief synopsis of the book to several academic presses, but they were not interested. I then approached university press publishers. Four publishers actually asked to see the manuscript and liked it, but only one got to the stage of sending it out for review. However, I learned to my dismay that budgets are tight at university presses, especially for a first-time author. According to the editor of the university press I was working with, there simply wouldn’t be enough money for a color picture on the cover, the book would need to be shortened by 50%, the page size reduced by 30%, and many of the tables and footnotes deleted. I was devastated! My wonderful book would be stripped to the bone, no one but university libraries would buy it, and I would probably get very little money for all my hard work.

I immediately withdrew my book from this press’s consideration and dispensed with publishers altogether. I next investigated self-publishing by studying several books on the subject. Publishing the book myself looked like a demanding and financially risky business, but I knew that my book was good. For at my request, Dr. Robert Wetzel, the eminent scientist and author of Limnology, kindly reviewed my preliminary manuscript in 1996. His analysis started with the word “Bravo” and proceeded with praise, encouragement, and suggestions. Now 3 years later, I knew that the book was even better than what I had sent him. While I was waiting to hear from publishers, I had added the line drawings, edited the manuscript at least 10 times, and added new and exciting material.

From then on I mainly followed Dan Poynter’s guidelines in his book The Self-Publishing Manual. Thus, I wrote to printers and asked them to submit bids to print the book (Mr. Poynter gives addresses of over 40 printers in his book). The printers that I chose (BookMasters Inc; Ashland OH, USA) printed the book exactly as I wrote it. They produced the entire book by photographing the 194+ paper pages I sent them. My book has turned out to be both a profitable venture and a personal miracle. I am amazed that a little hobbyist project begun in 1987 resulted in one of my life’s greatest achievements.

Nitrogen Removal in the Planted Aquarium
Nitrogenous compounds, particularly ammonia and nitrite, are probably the most common pollutants of aquariums. Both ammonia and nitrite are extremely toxic to fish. Aquarium hobbyists depend on biological filtration to convert these toxic compounds into non-toxic nitrates. However, plants and soil bacteria can also remove nitrogen from aquariums.

Aquatic Plants Prefer Ammonium over Nitrates
All plants can use either ammonia/ammonium, nitrite, or nitrate as their N (nitrogen) source. Based on N-uptake studies in terrestrial plants, many people assumed that aquatic plants, like terrestrial plants, mainly take up nitrates. Actual experimental studies with aquatic plants suggest otherwise.

Scientists from all over the world have studied N uptake in aquatic plants under a variety of experimental conditions. From the published studies, I found 29 species of aquatic plants that preferred ammonia/ammonium to nitrate. Only 4 species were found to prefer nitrates.

Figure 2: Uptake of ammonium and nitrates by Elodea nuttallii. Plants were placed in 1 liter of filtered lake water containing 2 mg/l each of NO3-N and NH4-N. Concentrations of ammonium and nitrates were measured at 4, 8, 16, 32 and 64 h (note the logarithmic scale of time axis in the graph). For each exposure period, 3 tanks with plants and 3 control tanks without plants were used. Control tanks (without plants) showed that there was little loss of either NH4-N or NO3-N due to bacterial processes. Figure from Ozimek [8], Fig 1, p 107 in my book.

Moreover, the extent of this preference is considerable. For example, the aquatic liverwort Jungermannia vulcanicoloa was found to take up ammonium 15 times faster than nitrates [6]. The duckweed Lemna gibba removed 50% of the ammonium in a nutrient solution within 5 hours, even though the solution contained over a hundred times more nitrates than ammonium [9].

Elodea nuttallii, placed in an equal mixture of ammonium and nitrates, removed 75% of the ammonium within 16 hours while leaving the nitrates virtually untouched. Only when the ammonium was gone, did the plants begin to take up nitrates.

Likewise, when Ferguson [2] grew the giant duckweed Spirodela oligorrhiza in media containing a mixture of ammonium and nitrate, the ammonium was rapidly taken up whereas the nitrates were not. Because he grew the plants under sterile conditions, he showed that the ammonium removal could not have been due to the bacterial process of nitrification.

Also, Ferguson showed that plants grew rapidly suggesting that the ammonium uptake probably accompanied the normal growth process and increased plant material (the N concentration in aquatic plants ranges from 0.6 to 4.3% of their dry weight [3]).

Figure 3: Lemna minor is one of the many plant species that prefer ammonium over nitrates. Some familiar species with the same preference:

Ceratophyllum demersum

Eichornia crassipes

Elodea densa

Pistia stratiotes

Salvinia molesta
(Table 3, page 108 in my book: Nitrogen Preference of Various Species).

Nitrite Uptake by Plants
Although plants can use nitrite as an N source, the pertinent question for hobbyists is- do aquatic plants remove the toxic nitrite before the non-toxic nitrate? I could not find enough studies in the scientific literature to state conclusively that they do. However, the chemical reduction of nitrites to ammonium requires less of the plant’s energy than the chemical reduction of nitrates to ammonium. Thus, when Ferguson [2] grew the duckweed Spirodela oligorrhiza in media containing both nitrate and nitrite, he showed that it preferred nitrite.

Figure 4: Nitrite (NO2) and nitrate (NO3) uptake by Spirodela oligorrhiza. Plants that had been grown with ammonium as their sole N source were transfered to medium containing both nitrite and nitrate. Plants were grown under sterile conditions. Thus, the above changes in nitrite and nitrate levels could not have been due bacterial processses. With permission of Springer Verlag GmbH (Fig 4, p 23 in my book).

Aquatic Plants versus Biological Filtration
Plants, algae, and all photosynthesizing organisms use the N of ammonia- not nitrates- to produce their proteins. Thus, in the first step of protein synthesis, NH₃ is combined with a carbohydrate to form the amino acid glutamine. From this glutamine all of the other amino acids will be synthesized and then eventually combined to form the plant's proteins. If the plant takes up nitrate, it must first be converted to ammonium in an energy-requiring process called nitrate reduction.

NH₄⁺ + 2 O₂ => NO₃^- + H₂O + 2 H⁺

Plants must expend essentially the same amount of energy (83 Kcal/mol) [5] to convert nitrates back to ammonium in the two-step process of nitrate reduction The overall reaction for nitrate reduction is:

NO₃^- + H₂O + 2 H⁺ => NH₄⁺ + 2 O₂

Thus, nitrate uptake by plants requires more effort than ammonium uptake. For example, the water lettuce took up nitrates much slower in the dark, while ammonia uptake was the same in the light or the dark [7]. Other scientists showed similar results with the hornwort plant [11]. Nitrate uptake requires more energy (in this case light energy) than ammonium uptake. Furthermore, maximum nitrate uptake in the water lettuce did not occur until after the plants had been grown in media containing only nitrates for 24 hours [7]. This is typical, because the enzyme (nitrate reductase) required for using nitrates must be induced (plants don’t make this enzyme unless they have to) [5].
The energy required for nitrate reduction is equivalent to 23.4% of the energy obtained from glucose combustion [4]. Thus, if nitrifying bacteria in biological filters convert all available ammonium to nitrates, plants will be forced-- at an energy cost-- to convert all the nitrates back to ammonium. This may explain why several aquatic plants were found to grow moderately better with ammonium than when they are forced to grow with pure nitrates [1,8,10]. Thus, extensive biological filtration may be counterproductive to plant growth.
Nitrogen Removal by Soil Bacteria
Nitrifying bacteria that convert ammonia to nitrates are found not just in aquarium filters. For all sediments and soils contain nitrifying bacteria. Soils also contain bacteria that remove nitrates and nitrite through the processes of denitrification and nitrate respiration (more detailed information on N processing by bacteria can be found in my book, pp 62-66).
One would expect nitrates, a product of biological filtration, to gradually build up in aquariums. However, I have detected no nitrate build-up in my planted aquariums despite infrequent water changes. For a long time, I thought this lack of nitrate build-up was due to N uptake by plants. However, I calculated that I routinely add ten times more N as fishfood to my tanks than what the plants take up (Table 8, p 88 in my book). Where is all this extra N going?
I suspect that soil bacteria might be removing the nitrates via denitrification. This is because under anaerobic conditions many bacteria use nitrates instead of oxygen for their respiration. When they do, nitrates are converted to gaseous forms of nitrogen (NO, N2O, N2) that escape from the tank. Presto, nitrates are removed!
My aquariums not only have plants, but they have a soil underlayer. This soil underlayer would be expected to contain huge numbers of bacteria, many more than in the overlying water or in gravel substrates. This is because soil has an enormous surface area for bacteria. Also, the soil layer would be anaerobic, providing perfect conditions for denitrifying bacteria to use (and remove) nitrates.
Recently, I tested the theory with experiments. I prepared identical plastic bottles with and without soil to which I added tapwater containing 40 mg/l nitrates. Each variable was done in duplicate bottles. I set up the bottles with a substrate similar to what I use in some of my aquariums (that is, 3-6 cm soil covered with 2-3 cm of gravel). I kept the bottles in a dark box (to prevent algae growth) and measured nitrates and nitrites every few days for 3 weeks. I found that the nitrates rapidly decreased in bottles containing soil, but not in bottles without soil. In bottles without soil, there was almost no nitrate removal.
Along with the rapid disappearance of nitrates in bottles containing soil, I also detected a small amount of nitrites in the overlying water. So I repeated the first experiment with a higher concentration of nitrates (250 mg/l). All nitrates were gone from the water by 32 days. Nitrites appeared in the water in 3 days (10-20 mg/l) and stayed for 3 weeks after which they disappeared (probably due to denitrifying bacteria). I also detected nitrites in bottles to which I added a nitrate-containing fertilizer tablet to the soil. I believe that nitrate respiration was the bacterial process responsible for the temporary production of nitrite.
Nitrifying bacteria in soil can also quickly remove ammonia. I did a third experiment with bottles, but this time I bubbled air (lots of oxygen) into water containing 1.3 mg/l of ammonia. Ammonia was removed within 6 days from water in bottles that contained either soil/gravel or recycled gravel (gravel taken from my aquariums, which would be colonized with nitrifying bacteria). In contrast, ammonia was not removed from bottles that contained only water, even after 3 weeks!
The results of my “bottle experiments” showed me how bacteria in the substrate alone can remove ammonia and nitrates. It also convinced me that adding nitrate-containing fertilizers to the soil was not a good idea. This is because nitrate-respiring bacteria would quickly convert nitrates to toxic nitrite. Now I never add fertilizers to my substrates.
Much of the aquarium literature implies that aquarium filters are solely responsible for removing toxic ammonia. Also, it implies that nitrate accumulation is inevitable and that special ‘denitrator’ filters and water changes are the only means of removing nitrates from aquariums. This may be true in tanks without substrate or plants. However, in the planted aquarium, both plants and substrate bacteria remove ammonia and nitrates.
After I completed my experiments and realized how well plants and soil bacteria removed ammonia and nitrites from the water, I dispensed with filters altogether. A year ago I replaced my canister filters with inexpensive powerheads to just circulate the water. The aquariums have done fine.
References

Cary PR and Weerts PGJ. 1983. Growth of Salvinia molesta as affected by water temperature and nutrition. 1. Effects of nitrogen level and nitrogen compounds. Aquat. Bot. 16: 163-172.
Ferguson AR and Bollard EG. 1969. Nitrogen metabolism of Spirodela oligorrhiza 1. Utilization of ammonium, nitrate and nitrite. Planta 88: 344-352.
Gerloff GC. 1975. Nutritional Ecology of Nuisance Aquatic Plants. National Environmental Research Center (Corvallis OR), 78 pp.
Hageman RH. 1980. Effect of form of nitrogen on plant growth. In: Meisinger JJ, Randall GW, and Vitosh ML (eds). Nitrification Inhibitors- Potentials and Limitations. Am. Soc. of Agronomy (Madison WI), pp. 47-62.
Lewis OAM. 1986. Plants and Nitrogen. Edward Arnold Publishers, LTD. Baltimore, MD, pp. 27-29.
Miyazaki T and Satake K. 1985. In situ measurement of uptake of inorganic carbon and nitrogen by the aquatic liverworts Jungermannia vulcanicola Steph. and Scapania undulata (L.) Dum. in an acid stream, Kashiranashigawa, Japan. Hydrobiologia 124: 29-34.
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Reddy KR and Tucker JC. 1983. Productivity and nutrient uptake of water hyacinth, Eichhornia crassipes. I. Effect of nitrogen source. Econ. Bot. 37: 237-247
Toetz DW. 1971. Diurnal uptake of NO3 and NH4 by a Ceratophyllum-periphyton community. Limnol. Oceanogr. 16: 819-822.
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Copyrights:
This article has material in it that has never been published. It can not be copied (even partially) and published anywhere without the written permissions of the author (Diana Walstad) and Bilyap Aquaristic.