When and why did dosing nitrates start?

[flampton]

Still - the last carbon will leave a biological system as methane. If it not was bounded - you would have the same fast effect when you add surplus food as you get when you ad DOC. It can happen with overfeeding but it takes time - much time.

Sincerely Lasse

I guess I'm not in understanding what you mean by last carbon. If you consider added and produced organic carbon that 99.999 or so leaves as carbon dioxide I guess that last 0.001 or whatever percent will leave as methane in a healthy non swamp like aquarium. Why is this important?

 

[flampton]

I understand what you are saying. I have carbon dosed, in many forms both areobically & anaerobically for years & I’m well acquainted with bacterial blooms.

What I’m referring to is, that in historically ‘normal’ aquaria, we did not recognize the need for organic carbon, and that it was the limiting factor especially in nitrogen processes.

Organic carbon has become part of the ‘new normal’ in aquariums. Whether by carbon dosing, algae reactors, display refugia or some other means, many/most of us recognize now that it’s required too.

What I think is important to recognize is that something will always be limiting to biological processes. Known as Liebig’s Law of the Minimum-

https://en.m.wikipedia.org/wiki/Liebig's_law_of_the_minimum

Let me get this straight, you know carbon is the limiting nutrient in aquaria. You know this from reading and experiencing bacterial blooms. But you also think that carbon is not limiting? I'm just plain confused. Are you trolling me?

 

[ichthyoid]

I mean that organic carbon being a requirement, just like N, P, water, food, salts, etc. is accepted.

Having not been supplying it in sufficient quantity in the past, does not change that organic carbon is required.

 

[flampton]

I mean that organic carbon being a requirement, just like N, P, water, food, salts, etc. is accepted.

Having not been supplying it in sufficient quantity in the past, does not change that organic carbon is required.

Let's say you're working with a system in which adding more nitrogen results in no new growth, adding phosphate results in no new growth, but when organic carbon is added or produced you get new growth.

Which of the above is limiting?

 

[ichthyoid]

You're over your head my man. Like really far.

That was not my intent. Only to show that N & P are also involved, in intricate ways.

 

[Lasse]

If we comeback to the thing all want - denitrification in our aquaria. it needs anaerobic environment. Denitrification is not the only thing happening in anaerobic environment. You can as many - take out organic detritus - or you can break it down in the aquaria but if you chose the path with 100 % aerobic breakdown of organic matter - you will not have any de nitrification. Denitrification means anaerobic breakdown of organic matter. Even if you chose 100 % aerobic break down - you will have a time span before the organic carbon is possible to be used by the bacteria. When the bacteria release CO2 - they have already consumed the organic carbon they need in that speed they use at that moment. Let me take an example with a compost. In a compost you need organic matter that contain carbon, nitrogen and phosphorus in certain ratio. But you need something more in order to speed it up, there is something that it is limited for max speed. Normally - you blend the organic matter with sawdust - organic carbon. What I mean is that the carbon is bounded and not so easily accessible as the organic nitrogen and phosphorous. Organic carbon will always be the limited factor for heterotrophic bacteria growth in an aquaria - just because the time span between the accessibility of organic/inorganic N and P compared with the accessibility of organic carbon. It seems like many heterotrophic bacteria can use at least inorganic N (NH3/NH4) as N source - according PO4 - I´m more skeptical - but it could mean that many heterotrophic bacteria both can use inorganic and organic N and P but only organic carbon. I hope you understand what I mean,

Sincerely Lasse

 

[Thaxxx]

My head hurts ;Nurse

 

[ichthyoid]

If you take All for Reef as one example of organic carbon dosing, it is somewhat different from other organic carbon forms.

AFR is made from calcium formate, of which the formate is a one carbon substrate.

The compound has a calcium attached to 2 formates. The structure looks like this-

HOOC - Ca - COOH

Due to having just 1 carbon organic groups, it produces half the CO2 as say acetate (vinegar) or ethanol (vodka) will, which each have 2 carbons. That means less of a pH drop.

It also yields less energy which can be used to build cells/mass, as those other carbon substrates would, & for the same reason. That means less biomat or stringy build up.

Formate, being a 1 carbon substrate, can also be metabolized by a group of bacteria known as methanogens. These are a special type of bacteria which have unique enzymes and are capable of breaking down single carbon compounds. Some are able to do this in aerobic environments.

When AFR is metabolized by bacteria, we get carbon dosing plus a calcium is added/liberated from the calcium formate compound by those bacteria.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC161530/

 

[flampton]

If we comeback to the thing all want - denitrification in our aquaria. it needs anaerobic environment. Denitrification is not the only thing happening in anaerobic environment. You can as many - take out organic detritus - or you can break it down in the aquaria but if you chose the path with 100 % aerobic breakdown of organic matter - you will not have any de nitrification. Denitrification means anaerobic breakdown of organic matter. Even if you chose 100 % aerobic break down - you will have a time span before the organic carbon is possible to be used by the bacteria. When the bacteria release CO2 - they have already consumed the organic carbon they need in that speed they use at that moment. Let me take an example with a compost. In a compost you need organic matter that contain carbon, nitrogen and phosphorus in certain ratio. But you need something more in order to speed it up, there is something that it is limited for max speed. Normally - you blend the organic matter with sawdust - organic carbon. What I mean is that the carbon is bounded and not so easily accessible as the organic nitrogen and phosphorous. Organic carbon will always be the limited factor for heterotrophic bacteria growth in an aquaria - just because the time span between the accessibility of organic/inorganic N and P compared with the accessibility of organic carbon. It seems like many heterotrophic bacteria can use at least inorganic N (NH3/NH4) as N source - according PO4 - I´m more skeptical - but it could mean that many heterotrophic bacteria both can use inorganic and organic N and P but only organic carbon. I hope you understand what I mean,

Sincerely Lasse

Agree that the more your goal is denitrification the more organic carbon is needed as it's done by heterotrophic prokaryotes. However that carbon is still mostly going to leave as carbon dioxide. The thing is that the electron transport chain does NOT produce carbon dioxide. This is produced prior during pyruvate decarboxylation and the TCA cycle.

I think a lot of people misunderstand because they think breath In oxygen breath out carbon dioxide. But those two oxygen molecules are combined with four protons to give two water molecules.

Now every organism that I'm aware of can utilize ammonia. As every single genome I've looked at so far is not predicted to be auxotrophic for glutamate. Now does that mean there isn't one? No but I can guarantee it would not be a free living organism. (edit: This was a stupid statement as can likely find one if I looked hard enough, but would also have to lack the ability to synthesize any of these other amino acids e.g. aspartate, serine, isoleucine, valine, leucine, tyrosine, phenylalanine or tryptophan) Anyways ammonia is going to diffuse across membranes at the pH in our aquarium without a problem. What this means is that the organisms in our aquarium may spend energy maintaining their membrane potential when ammonia is at higher concentrations. And as the concentration gets really high it takes more and more energy to maintain the cells membrane potential. Just ask the million damsels that have been sacrificied to the cycle :p (edit: membrane potential is required for a cell to keep making energy by the electron transport chain, now the membrane potential can refer to the cellular membrane of prokaryotes, or the mitochondria of eukaryotes.)

Now phosphate is a different story. Phosphate is actually energetically problematic when it comes to transporting it in. All organisms that I'm aware of import inorganic phosphate (Pi)
Now some also have the ability to scavenge some phosphorylated sugars and glycerol but the larger the molecule the more complex the machinery and this is not very economical to the organism. So things like phospholipids are digested outside the cell utilizing phospholipases and then Pi Is moved in. The actual lipid might not even be brought in if it's a cell that is just scavenging for phosphates. So every organism in your aquarium can utilize inorganic phosphate. And obviously inside as well because there is not an organism that does not use ATP as it's energy currency and obviously needs to synthesize DNA and RNA.

I hope the above clarifies this. And though the experiments needed to be done that prove that coral can utilize ammonia has always been a no brainer. The more curious has always been nitrates as they are energetically unfavorable to use. That's why it seems to be the autotrophs and their 'free' energy that utilize them the most.

(Which reminds me I need to look up if they have ever separated the nitrate utilization of the coral from the dinoflagellate. For those curious this type of experiment is usually done with Aiptasia and Azoox Aiptasia of the same species. Then if plausible continues into harder to study organisms.)

 

[ichthyoid]

You might be interested in the article below. It seems that while corals obtain organic carbon mostly as sugars from their symbionts, they rely heavily on bacteria & flagellates for nitrogen & phosphorous. Most of that appears to be provided in organic forms as well, obtained by ingestion.

https://reefs.com/magazine/coral-feeding-an-overview/

 

[flampton]

You might be interested in the article below. It seems that while corals obtain organic carbon mostly as sugars from their symbionts, they rely heavily on bacteria & flagellates for nitrogen & phosphorous. Much of that appears to be provided in organic forms as well, obtained by ingestion.

https://reefs.com/magazine/coral-feeding-an-overview/

Yes!! Totally agree, that's why I firmly believe in carbon dosing!!! :D However the organics you're talking about go through digestion and will result in some ammonia and almost all Pi before transport inside the cells of the animal

 

[Lasse]

Now every organism that I'm aware of can utilize ammonia.

Anyways ammonia is going to diffuse across membranes at the pH in our aquarium without a proble

Yes - but there is alsu studies that says that amino acids also do this and according to friend of mine - they are in many situations faster to penetrate than NH3/NH4. I did not discuss this with him because his PhD was about this.... I do not have it here but I think that cell-penetrating-peptides can give you ac starting point.

Sincerely Lasse

 

[flampton]

Yes - but there is alsu studies that says that amino acids also do this and according to friend of mine - they are in many situations faster to penetrate than NH3/NH4. I did not discuss this with him because his PhD was about this.... I do not have it here but I think that cell-penetrating-peptides can give you ac starting point.

Sincerely Lasse

Yes it is concentration dependent to some extent. Here's were it gets way trickier. The coral have to beat the prokaryotes in utilizing the substrate. So if you know about surface to volume ratio and transport kinetics you'll know that this would be very challenging to compete for the animal. So you've seen my other posts and I believe in carbon dosing. In effect my proposition is that it is more favorable to grow single cell heterotrophs and allow them to go up the food chain. Your amino acid mix is not going to beat the nutrition of a prototrophic microbe. Except in that it will also feed your bacteria and provide a transient boost in ammonia concentration! So I don't hate amino acids, and in fact I'm utilizing glycine experimentally because of a specific species of bacteria can't make it.

 

[ichthyoid]

Uptake & incorporation of amino acids is energetically favored.

 

[flampton]

Nice blanket statement...it is concentration dependent for everything. Transport kinetics verse passive diffusion kinetics. It's also meaningless since the coral will see very little of the added amino acid unless you add enough, and then good luck with the resulting bloom.

 

[ichthyoid]

Bacteria do the work for them. By virtue of their small size and high surface to volume ratio, diffusion is much faster & at lower gradients. Bacteria are specialists at scavenging and evolved to be highly efficient in order to survive.

Corals have been found to live in close symbiosis with bacteria, providing both protection & nutrition via their mucous layer. The bacteria provide essential nutrients and allelopathic compounds to repel pathogens for the corals in turn.

 

[Lasse]

I was not thinking of corals - I was thinking of triggering mat building cyanobacteria that seems to be very keen of amino acids according to some tests Dan_P have done and we go back to nitrate again - and why it should be slightly elevated in a reef aquarium...........

Sincerely Lasse

 

[flampton]

Bacteria do the work for them. By virtue of their small size and high surface to volume ratio, diffusion is much faster & at lower gradients. Bacteria are specialists at scavenging and evolved to be highly efficient in order to survive.

Corals have been found to live in close symbiosis with bacteria, providing both protection & nutrition via their mucous layer. The bacteria provide essential nutrients and allelopathic compounds to repel pathogens for the corals in turn.

Bacteria do the work for them? You mean take in the amino acids and excrete ammonia? Or are consumed by the animal or symbiodinium? Yeah I know. That's exactly what I'm saying!!!

Btw
So if you're looking at transport studies you need to look at the methods. What you will find is they usually use ASW or filtered seawater. So what is there? Well it's limited in all three of the limiting nutrients. They will then move a preferably rinsed animal into the aquarium. Then they'll add the substrate they're testing. No bloom! Why!! No phosphate availability no DNA synthesis, no replication.

So you can not extrapolate this to a hobby tank because we want some level of detectable phosphate.

 

[ichthyoid]

Ammonia is produced not only by fishes, but also zooplankton, other coelenterates and is both consumed and produced by some species of Cyanobacteria. That can of course undergo nitrification to nitrite & nitrate.

https://link.springer.com/article/10.1007/BF00389168

 

[flampton]

I was not thinking of corals - I was thinking of triggering mat building cyanobacteria that seems to be very keen of amino acids according to some tests Dan_P have done and we go back to nitrate again - and why it should be slightly elevated in a reef aquarium...........

Sincerely Lasse

Yes a lot of things prefer fully formed amino acids for building cellular structure as it costs less.

However where is the evidence that cyano form mats without amino acids? It is much more likely they form mats when ammonia is not available. The idea that you're feeding the cyano preferentially to the heterotrophs is false. What do the heterotrophs release after utilization, ammonia. What availability goes up in the water column, ammonia.

Ammonia is produced not only by fishes, but also zooplankton, other coelenterates and is both consumed and produced by some species of Cyanobacteria. That can of course undergo nitrification to nitrite & nitrate.

https://link.springer.com/article/10.1007/BF00389168

Ammonia is used and produced in every organism on the planet