Confirming - Prime does not seem to lower ammonia - Seachem Alert

MnFish1

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Hypothesis: There will be no difference between Free ammonia levels as measured by a seachem alert badge in the presence of Prime (IN FRESH RODI WATER).

Background: Numerous experiments have been done which seem to suggest Prime does not lower ammonia as measured by various tests in saltwater. There has been a question about whether it might do so in Freshwater This study is designed to test this.

Method / Materials. 2 Seachem alerts, prepared. 2 quarts of RODI (pH 7.2, Ammonia 0, Temp Room (72)). Add various amounts of ammonia prime, and adjust pH (see below)
Results - see Excel Spreadsheet below
Conclusion:
1. Prime did not affect the seachem alert in fresh water over the time frames studied (I have heard it can take 2 hours for the full effect - but other places says immediately).
2. The Seachem alert performed well - and as expected with addition of bicarbonate to increase pH free ammonia rapidly increased.
3. The disk in the Prime sample seems to be recovering faster - after being added bas to fresh water. Whether this holds up or not - will see - when one or both gets to 'safe'.
4. A couple problems -
a. could have let the water sit with Prime longer
b. the ammonia level might have been 'too high'.
c. ammonia is not usually added to a disk all at once - it builds up - and its possible the immediate high concentration caused the film to change (however the water had prime added first - was mixed - then ammonia was added after)
d. Could the results have been different if I had started with pH 8.0 water added prime to one container - and then slowly - each hour added .25 or .5 ppm ammonia to each? I think this might also be an interesting experiment.
e. The experiment is not designed to test whether Prime works as its labeled or not.

Side Note - Will do the Multi-test tomorrow (the one Seachem recommends). Trying out that test today - I am thinking the test might be expired - though I don't see an obvious date (my RODI - tested at 2 ppm ammonia - and the reference tested higher than it should have as well).
Screen Shot 2022-01-20 at 1.56.15 PM.png
 
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threebuoys

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Based on your comments, would it be appropriate to add to your list of problems?

f. This does not show that Prime does 'work'.


In other words, for the conclusion, the experiment is still a work in progress and the hypothisis has not been proved or disproved yet.
 
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MnFish1

MnFish1

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Based on your comments, would it be appropriate to add to your list of problems?

f. This does not show that Prime does 'work'.


In other words, for the conclusion, the experiment is still a work in progress and the hypothisis has not been proved or disproved yet.
Yes - and no. The conclusion based on these methods in this experiment show that the hypothesis has been proven (Prime does not appear to change free ammonia levels). The question is whether if done with different methods (a new experiment) whether it will continue to hold true.

In my opinion f. is not needed - because - to prove Prime 'works' you have to show that it makes high ammonia levels non-toxic to something alive (which is the job its supposed to do). This experiment wasn't designed to show whether Prime works or not.
 
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MnFish1

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When are you going to test on the feeder fish @MNFish ?

I just don’t see the point to any other testing if prime ultimately does not detoxify ammonia and increase fish survivability. But maybe I’m missing something.
The reason IMHO to do more testing is that according to Seachem the best test to show a decrease in free Ammonia is the Multi- kit. I know @taricha has used this kit before. The purpose of this experiment was based on a request someone made as to whether perhaps Prime does lower free ammonia in Fresh water.

So - as to the fish - I'm still debating internally the ethical issues - if I do it - they will have to be acclimated to salt water - for the experiment
 

threebuoys

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Yes - and no. The conclusion based on these methods in this experiment show that the hypothesis has been proven (Prime does not appear to change free ammonia levels). The question is whether if done with different methods (a new experiment) whether it will continue to hold true.

In my opinion f. is not needed - because - to prove Prime 'works' you have to show that it makes high ammonia levels non-toxic to something alive (which is the job its supposed to do). This experiment wasn't designed to show whether Prime works or not.
If f. is not needed, then why is e. needed?
 
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MnFish1

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If f. is not needed, then why is e. needed?
Because it underscores the point that this experiment was not designed to determine if prime 'worked'. But - I changed the wording because you have a point
 
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MnFish1

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The reason IMHO to do more testing is that according to Seachem the best test to show a decrease in free Ammonia is the Multi- kit. I know @taricha has used this kit before. The purpose of this experiment was based on a request someone made as to whether perhaps Prime does lower free ammonia in Fresh water.

So - as to the fish - I'm still debating internally the ethical issues - if I do it - they will have to be acclimated to salt water - for the experiment
PS - if you look at the ammo-lock website - there is a very interesting study - where they used cultured fish cells - and exposed them to both ammonia and ammonia + ammo-loc at 5 ppm... Interesting - on the bottle it says ammonia reduction is measurable on API test kits
 

taricha

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Side Note - Will do the Multi-test tomorrow (the one Seachem recommends). Trying out that test today - I am thinking the test might be expired - though I don't see an obvious date (my RODI - tested at 2 ppm ammonia - and the reference tested higher than it should have as well).
The multi-Test disks are each cheap, and can have manufacturing artifacts - patches in the disk that develop stronger and weaker color or no color at all. I usually take several of them and put them in zero ammonia water for a while to make sure their color is consistent first. Sometimes one of the disks is just bad and I discard. Since they are small and cheap I use replicates of them in samples too.
Also, sometimes a disk isn't "zeroed" initially (transfer contamination is easy to do) and after some soak time it will lower to a correct color.
The disks start to show a color change quickly, but even color takes longer (I like an hour+). The color card that comes with them is really hard to compare to the actual color formed on the disks at any time. For all these reasons, Dan gave up on the little disks. And I use them with a big grain of salt.

Below pic shows the good and the bad about the disks.
20211003_231758.jpg


I don't have my notes handy, but each pair of disks from top to bottom was exposed to (I think) a factor of 2x different ammonia for a couple of hours. They were kept in ~80mL samples sealed in bottles stirred occasionally for stability. I put them in the white wells just to photograph.
You can see that two of the disks in the bottom half are outliers and ought to be discarded.

If you want to push it to make it more numerically quantifiable, you can look at the RGB values (Use the Red) for the disks in a photo like the above, and match it to the spot on the color card that has the same Red pixel value in the same photo. The number (from the card) that method will give you is not correct, but it gives a nice quantifiable reference number to say if something is different or not.

Just some things to keep in mind if you are going to try to extract useful info from the madness of the tiny disks. :p
 
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taricha

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Regarding the experiment itself, no major issues...
c. ammonia is not usually added to a disk all at once - it builds up - and its possible the immediate high concentration caused the film to change (however the water had prime added first - was mixed - then ammonia was added after)
d. Could the results have been different if I had started with pH 8.0 water added prime to one container - and then slowly - each hour added .25 or .5 ppm ammonia to each? I think this might also be an interesting experiment.
This is the logical equivalent of just testing a smaller ratio of ammonia:prime. If the lack of effect at small ammonia:prime and Recommended ammonia:prime ratios are both observed, then a slow ramp up from a small concentration of ammonia to a large one is fully covered.

The one tiny quibble I would have is using straight RO/DI when you need to know precisely what the pH is.
"2. The pH of highly purified water is not accurately measured by test kits, or by pH meters. There are several different reasons for this, including the fact that highly purified water has very little buffering capacity, so its pH is easily changed. Even the acidity or basicity of a pH test kit’s indicator dye is enough to alter pure water’s measured pH. As for pH meters, the probes themselves do not function well in the very low ionic strength of pure freshwater, and trace impurities on them can swing the pH around quite a bit." RHF article

The same logic might apply to the films and disks. In pure RO/DI it's really hard to have a firm grasp on what is controlling the pH at the film surface (and I don't even want to think about the mechanics inside the film).

If you're doing freshwater, I'd either pull zero ammonia water from a freshwater tank, or even use RO/DI with a tiny bit of some buffer. (I'm lazy - I'd use tiny amounts of bicarbonate and then HCl /NaOH to move the pH wherever I wanted, or even a pinch of one of those "proper pH" buffer products, then HCl/NaOH).
 
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MnFish1

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Regarding the experiment itself, no major issues...

This is the logical equivalent of just testing a smaller ratio of ammonia:prime. If the lack of effect at small ammonia:prime and Recommended ammonia:prime ratios are both observed, then a slow ramp up from a small concentration of ammonia to a large one is fully covered.

The one tiny quibble I would have is using straight RO/DI when you need to know precisely what the pH is.
"2. The pH of highly purified water is not accurately measured by test kits, or by pH meters. There are several different reasons for this, including the fact that highly purified water has very little buffering capacity, so its pH is easily changed. Even the acidity or basicity of a pH test kit’s indicator dye is enough to alter pure water’s measured pH. As for pH meters, the probes themselves do not function well in the very low ionic strength of pure freshwater, and trace impurities on them can swing the pH around quite a bit." RHF article

The same logic might apply to the films and disks. In pure RO/DI it's really hard to have a firm grasp on what is controlling the pH at the film surface (and I don't even want to think about the mechanics inside the film).

If you're doing freshwater, I'd either pull zero ammonia water from a freshwater tank, or even use RO/DI with a tiny bit of some buffer. (I'm lazy - I'd use tiny amounts of bicarbonate and then HCl /NaOH to move the pH wherever I wanted, or even a pinch of one of those "proper pH" buffer products, then HCl/NaOH).
Yes - the plan is to use bicarbonate.
 

threebuoys

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The multi-Test disks are each cheap, and can have manufacturing artifacts - patches in the disk that develop stronger and weaker color or no color at all. I usually take several of them and put them in zero ammonia water for a while to make sure their color is consistent first. Sometimes one of the disks is just bad and I discard. Since they are small and cheap I use replicates of them in samples too.
Also, sometimes a disk isn't "zeroed" initially (transfer contamination is easy to do) and after some soak time it will lower to a correct color.
The disks start to show a color change quickly, but even color takes longer (I like an hour+). The color card that comes with them is really hard to compare to the actual color formed on the disks at any time. For all these reasons, Dan gave up on the little disks. And I use them with a big grain of salt.
@taricha, @Dan_P , @MnFish1

Taricha, you referenced disk contamination. Is this something common for all of you? About a year ago, using the seachem nitrate test kit on some experiments I played with, I noticed that every time I picked up a disc with the supplied metal tweezers, it immediately turned blue, particularly where it was touched by the tweezers. It remained blue throughout the test and only returned to yellow after I replaced it in the bottle. I contacted Seachem about the matter, they said they had never heard of it, and suggested I make a video and send to them. I decided it was contamination and wasn't worth the effort since seachem seemed uninterested and I gave up on the seachem kit.
 
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MnFish1

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@taricha, @Dan_P , @MnFish1

Taricha, you referenced disk contamination. Is this something common for all of you? About a year ago, using the seachem nitrate test kit on some experiments I played with, I noticed that every time I picked up a disc with the supplied metal tweezers, it immediately turned blue, particularly where it was touched by the tweezers. It remained blue throughout the test and only returned to yellow after I replaced it in the bottle. I contacted Seachem about the matter, they said they had never heard of it, and suggested I make a video and send to them. I decided it was contamination and wasn't worth the effort since seachem seemed uninterested and I gave up on the seachem kit.
This is similar to what happened to me. All of the disks (including the reference) were blue (The reference is not supposed to be blue).
 
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taricha

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Taricha, you referenced disk contamination. Is this something common for all of you? About a year ago, using the seachem nitrate test kit on some experiments I played with, I noticed that every time I picked up a disc with the supplied metal tweezers, it immediately turned blue, particularly where it was touched by the tweezers. It remained blue throughout the test and only returned to yellow after I replaced it in the bottle.
No. I haven't observed it being a huge problem personally. My caution was in reference to your experience, but I'm wary of it since I'm handling a concentrated ammonia source often in the same space and with the same hands that I'm touching the things I'll move the disks with.
This is similar to what happened to me. All of the disks (including the reference) were blue (The reference is not supposed to be blue).
I love that Seachem ships you a stock solution with each kit to check it with, but that also means that any leak in the stock solution would result in the tweezers or white test wells being a concentrated contaminant. I expect if that happened, it would look like what @threebuoys saw.
 

Dan_P

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@taricha, @Dan_P , @MnFish1

Taricha, you referenced disk contamination. Is this something common for all of you? About a year ago, using the seachem nitrate test kit on some experiments I played with, I noticed that every time I picked up a disc with the supplied metal tweezers, it immediately turned blue, particularly where it was touched by the tweezers. It remained blue throughout the test and only returned to yellow after I replaced it in the bottle. I contacted Seachem about the matter, they said they had never heard of it, and suggested I make a video and send to them. I decided it was contamination and wasn't worth the effort since seachem seemed uninterested and I gave up on the seachem kit.
Ammonia sensor films have dye molecules embedded in a semi-permeable film. I assume that if the film is damaged by bending, scraping, cutting, or abrading, it is no longer semi-permeable and the dye can then come in contact with things that change the pH around the dye and turning it blue. Except for the Seneye film, I have accidentally snicked several different films and produced little blue marks. I have observed what you describe as well.
 

Dan_P

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I can’t prove it but I recently had a fish in a non-cycled tank and was using the Seachem badge.

As soon as it would turn greenish, I would dose Prime and later that day or the next day, the badge would be yellow again.
Were you able to observe the pH with a pH meter during this time period? I ask because small changes in pH can have substantial affects on free ammonia concentration.
 
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