Accurate NO3 Test with Hanna ULR P Checker

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taricha

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This line of thinking quoted here turns out to be very wrong....

Ok, this discussion made me realize I may be adding a totally unnecessary amount of reagents....
And we got essentially the same trend line with me using like 3x the reagent concentration.
Am I thinking this through right - that I really only need reagents in proportion to the NO3, not the mL of sample?

Speculation until data backs it up, but it probably won't even need its own graph.
I mean, the data so far has lined up pretty tight even though I was using about 3x the reagent concentration that Dan did.
So I tested using half the reagent concentration than what is called for in my marine nitrate/ nitrite test kit. It gives me ~30% HIGHER readings: 199 vs 148 and 98 vs 76 (visibly more pink) than just using the recipe as is and doubling everything.

So this curve only applies to using the test kit in the ratios specified.
 
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taricha

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Nice!

But how does this compare with the method/equation described here?:

https://www.reef2reef.com/threads/using-hanna-phosphorous-checker-hi-736-to-test-for-low-levels-of-no3.350062/#post-4356070

Rick's method using the Red Sea Nitrate Pro kit is a lot easier than the one described here. Is there a difference in results?
After reading through @Rick Mathew thread a couple of times, here's my take. (I wish I'd seen it when I started - great info there)
First, and most importantly, they are for different ranges. This thread lays out how to test for the 1 to 15ppm NO3 range. Rick's method is for 0-2ppm.
Secondly, to accomplish the different ranges this thread dilutes 10 to 1 with distilled water, and that thread tests straight tank saltwater at 35 ppt. The difference in salinity has a large effect as seen here...
upload_2018-1-11_19-21-11.png

big difference in test result just between 29 and 35 ppt salinity. So make sure if using Rick's method to test at full tank salinity you adjust your tank to 35 ppt where (I think) his reference curve was made.

The method in this thread uses primarily the Red Sea NO3/NO2 kit. The method in Rick's thread uses the Red Sea Nitrate Pro Reef kit. These kits use different concentrations of the same reactants and this does indeed have an effect (I saw a ~30% difference by using half the reactant concentration) on the end color produced.

The combination of different salinities and different concentration of reactants between the test kits are why our fit lines are different. (also, I don't think Rick's quadratic equation is necessary - the relationship is linear from what I've seen, and I don't think the x^2 term improves the fit. but that's a beside the point math quibble.)
This fit line
(ppm NO3) = 0.00765 x (Hanna reading) + 0.117
...then multiply by 10 to get the tank NO3 concentration before the dilution.

Rick's fit line
(ppm NO3)= -.00000326 x (Hanna reading)^2 +.01296 x (Hanna reading) +.0534

Bottom line:
If you plan on nitrate between 0-2ppm, I'd use Rick's method and the Nitrate pro reef test kit.
If you plan on nitrate in the 1-15ppm range then I'd use the method in this thread and the NO3/NO2 marine test kit.

if I've missed something that ought to be discussed here, hopefully someone else will chime in.
 

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Many thanks for the detailed reply! :cool:

1-15ppm would be a better measurement range for me as I try to avoid having NO3 below 1ppm if I can (I add KNO3 to keep it above that, aiming for 2-5ppm).

So neither "Dan's instructions for the Red Sea Nitrate Pro kit" at the bottom of your first post nor using the Red Sea Pro kit at the 1:15 dilution will work for measuring 1-15ppm NO3 using your equation?

I ask because I just purchased a refill for my Pro kit :).
 
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taricha

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Many thanks for the detailed reply! :cool:

1-15ppm would be a better measurement range for me as I try to avoid having NO3 below 1ppm if I can (I add KNO3 to keep it above that, aiming for 2-5ppm).

So neither "Dan's instructions for the Red Sea Nitrate Pro kit" at the bottom of your first post nor using the Red Sea Pro kit at the 1:15 dilution will work for measuring 1-15ppm NO3 using your equation?

I ask because I just purchased a refill for my Pro kit :).
Yes, Dan_P's instructions are trustworthy and great, and give data that fits the equation in this thread.
Or, use the method of Rick's thread - you need to do a dilution with pure saltwater zero nitrate that is salinity matched to 35ppt.
I'd love to tell you that the A, B, and C in the two red sea kits are the same and interchangeable, but I don't know if that's the case. (Great idea! I'll order some of the nitrate pro reef and check)
 

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Good Work!...I have been perusing this idea for over a year now and am satisfied it can and does work...Still some work to do but I am currently using the regression method with the Red Sea Pro Test Reagents to measure my NO3 levels...I am now testing the Nitrite Test meter as the instrument...seem to give me more repeatable results and also goes to a higher range up to 5ppm

What I have found over the last months of testing using the method I developed is that there are 3 key elements in making this a usable accurate test.

1) All calibration solutions must be done in salt water adjusted to the salinity level I keep my tank (35ppt for me)
2) The instrument selected must have good repeatability & reproducability in the range being teated....
For the Hanna Phosphorous Tester used with the Red Sea Reagents that is 0-2ppm...
I have developed the test for the Hanna Nitrite Tester (HI-764) That appears to be working (work in progress that I will share as I get more confirmation) That is good for 0-5ppm
If I need to go above 5ppm (which I have not yet had the occasion to do) I would use the method I described in the previous Post Using the Hanna HI-96702 Copper Spectro https://www.reef2reef.com/threads/diy-nitrate-test-meter.330892/#post-4121274 with calibrations re-done in salt salt water of 35ppt salinity

3) Work on getting the test repeatable---Liquid and solid measurement methods needed to be as accurate as possible...

The cool thing about this is by using the regression method with correct calibration procedures one can try any number of instruments and Testers to see if they will do the job...I am planning on trying the New Hanna Marine Copper Tester Next to see if it will expand the test range so as to have a single method...

Here are the %T and Absorbance Curves of the Red Sea Test Liquid At 4ppm.....I am using this data for another project I am working on related to visual color evaluation of test samples vs Color Cards...

The max Absorbance of this sample is at 545nm....this means we are measuring on the slope when we use a checker with 525nm source. This can mean small errors in sample preparation and measurement can cause bigger outcome errors...Thus the reasoning for #3 in my list above

Hope this helps...keep us posted on how it is going....give me a shout if you have any questions


upload_2018-6-5_10-39-42.png


upload_2018-6-5_10-39-58.png


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taricha

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The cool thing about this is by using the regression method with correct calibration procedures one can try any number of instruments and Testers to see if they will do the job

The max Absorbance of this sample is at 545nm....this means we are measuring on the slope when we use a checker with 525nm source.
Rick! Thanks so much.
That full spectrum absorbance of nitrate pink end product is awesome.
I have Hanna checkers at 610nm (LR Silica), 575nm (LR Iron), and 525nm (ULR Phos)
Hannah also has some checkers that use 470nm, and from what I can tell all their checkers are some version of those same 4 wavelengths.
It sounds like it may be time to make a thread to index the different parameter/ method/testers we've got nailed down so far, and link out to the individual threads.
I've also done same thing with iodide, and Dan_P has done 3 or 4 different parameters.
 

Rick Mathew

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Rick! Thanks so much.
That full spectrum absorbance of nitrate pink end product is awesome.
I have Hanna checkers at 610nm (LR Silica), 575nm (LR Iron), and 525nm (ULR Phos)
Hannah also has some checkers that use 470nm, and from what I can tell all their checkers are some version of those same 4 wavelengths.
It sounds like it may be time to make a thread to index the different parameter/ method/testers we've got nailed down so far, and link out to the individual threads.
I've also done same thing with iodide, and Dan_P has done 3 or 4 different parameters.
Most welcome...Making a thread would be a good idea....I would be interested on what you are doing for You Iodine Test. I have been working on one using the HI-96702 Copper Spectro. and the Red Sea Iodine Reagents...It is a bit tricky because the reaction rate is temperature dependent (it Seems) and the rate of reaction is pretty quick so timing is everything....like most things in life;)

I think this work is a good idea to provide better testing methods that don't require visual color interpretations...As I read some of the comments in the threads about individuals having difficulties tell the difference between values it is no big surprise to me. I teach a class at the University of Akron Polymer Science on The Physics of Light and Color called "Color Theory and Application" So I am very familiar with the issues of color discrimination and the pitfalls...Especially when measuring color differences in different medium...Liquid vs Solid Paint or Ink Cards...The problem of a phenomenon called Metamerism is present in most if not all of the color kit test methods. Couple that with improper light source control and the fact that up about 1 out of 10 men have defective color vision (mostly in Red-Green)....well you can see the issues. This is the reason I ran the spectral absorption of the Red Sea Solution and in addition I have the spectral curves of the color card (wheel)...Although they are similar they are not the same which opens the door to Metamerism...thus difficulty interpenetrating the results under some conditions.

I am planning on doing a write up when the work is completed but there is still a lot of work to do....

Rick
 
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taricha

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Most welcome...Making a thread would be a good idea....I would be interested on what you are doing for You Iodine Test. I have been working on one using the HI-96702 Copper Spectro. and the Red Sea Iodine Reagents...It is a bit tricky because the reaction rate is temperature dependent (it Seems) and the rate of reaction is pretty quick so timing is everything...
Here's the iodide test thread: Seachem Kit run through the Hanna LR Silica meter
The seachem kit reaction is definitely concentration dependent. For low concentration, color is stable between 1-2 minutes, at higher concentrations the test needs to happen between 40-70 seconds - color dissipates rapidly after that.
It sounds like a few of us are really treading similar territory. I'll take it to pm so we can compare notes and maybe avoid duplicating some work.
 

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Just tried this out, and I have to say this is quite nice. Thanks taricha for taking the time to put this together.

I knew my sample should be around 5 ppm or slightly less, based on previous tests. Red Sea NO2/NO3 test gave a visual color match of <5 ppm. Then doing the test as described in the first post gave a reading of 30 which converts to 3.5 ppm ((0.0765 x 30) + 1.17) nitrate. I also did the test using the same technique with the Nitrate Pro chemicals, and got the same measurement. I suspect that the chemicals are exactly the same in the two kits, just different labels.

One thing: There are always some small undissolved particles (zinc?) remaining in the vial during the color development; it appears that it's not meant to necessarily dissolve completely. Is there any concern that this could interfere with the colorimeter's measurement? It seems that the additional 3 minute wait after you insert the cuvette into the meter may actually be beneficial in this regard.
 

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Just tried this out, and I have to say this is quite nice. Thanks taricha for taking the time to put this together.

I knew my sample should be around 5 ppm or slightly less, based on previous tests. Red Sea NO2/NO3 test gave a visual color match of <5 ppm. Then doing the test as described in the first post gave a reading of 30 which converts to 3.5 ppm ((0.0765 x 30) + 1.17) nitrate. I also did the test using the same technique with the Nitrate Pro chemicals, and got the same measurement. I suspect that the chemicals are exactly the same in the two kits, just different labels.

One thing: There are always some small undissolved particles (zinc?) remaining in the vial during the color development; it appears that it's not meant to necessarily dissolve completely. Is there any concern that this could interfere with the colorimeter's measurement? It seems that the additional 3 minute wait after you insert the cuvette into the meter may actually be beneficial in this regard.
Just tried this out, and I have to say this is quite nice. Thanks taricha for taking the time to put this together.

I knew my sample should be around 5 ppm or slightly less, based on previous tests. Red Sea NO2/NO3 test gave a visual color match of <5 ppm. Then doing the test as described in the first post gave a reading of 30 which converts to 3.5 ppm ((0.0765 x 30) + 1.17) nitrate. I also did the test using the same technique with the Nitrate Pro chemicals, and got the same measurement. I suspect that the chemicals are exactly the same in the two kits, just different labels.

One thing: There are always some small undissolved particles (zinc?) remaining in the vial during the color development; it appears that it's not meant to necessarily dissolve completely. Is there any concern that this could interfere with the colorimeter's measurement? It seems that the additional 3 minute wait after you insert the cuvette into the meter may actually be beneficial in this regard.
I have found that those undissolved particles...which are some times still floating up and down can interfere with the measurement so I decant 10 mL of the solution to be tested from the 16mL that the Red Sea Test calls for then allow that to set for 2 min before I take my reading...

Rick
 
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I also did the test using the same technique with the Nitrate Pro chemicals, and got the same measurement. I suspect that the chemicals are exactly the same in the two kits, just different labels.

One thing: There are always some small undissolved particles (zinc?) remaining in the vial during the color development; Is there any concern that this could interfere with the colorimeter's measurement?
For sake of consistency with the way I did it to generate the data, I over-shake it. That is, several times during the 9 min process, I shake the vial. Particulates are gone by the end.
I do let it sit for some seconds before pouring into the Hanna cuvette like Rick said, and leave some little bit in the bottom of red sea vial, so if there were solids - they don't make it into the Hanna cuvette.

Thanks for confirming that the test runs the same with the red sea reagents A, B, C regardless of whether they came from the nitrate pro reef or the Nitrate/nitrite marine kit. I ordered both sets and am going to double check.
 

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Well, I'm afraid I must retract my previous statement about the Red Sea NO2/NO3 test and NO3 Pro tests giving the same results; they most definitely do not. I only did one comparison the other day, and must have made some error that had them come out the same. Repeating the comparison several times today, it's clear now to me that using the Nitrate Pro reagents to perform the test as described here gives much lower results than with the NO2/NO3 reagents. I also find that the results are less consistent when the test is performed with the Pro reagents. This may in part be because the Pro reagents generate a larger amount of microbubbles, and there is also more dark particulate material in the vial at the end of the reaction. Also, looking at the chemicals themselves, the C reagent has a different chemical listed on its label, and the B reagent may have different proportions of the same components. So, at this point, the NO3 Pro reagents do not appear to work properly with this technique. I will defer to taricha for confirmation.
 
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Thanks for report @rkpetersen this is interesting, @Rick Mathew and others were talking about this Q on the consistency of results. Others looking into this with the red sea pro kit see more inconsistency - and some workarounds to avoid it. I haven't used that pro kit yet, but I haven't really seen inconsistency.
I have nothing but wild speculation as to why the one kit might give less uncertainty than the other - with similar methods.
I just know the other experimenters in this are at least as OCD on testing methods as I am, so it's not an issue of care.
 
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taricha

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I have to say it's quite cool what you guys are doing with these checkers. :)
:) It would be faster if we could get a few specs from Hanna, but whateva.

Separately we're wondering about the nature of the chloride interference. (More salinity - lower reading)
Do we think it interferes with formation of intermediate (nitrite maybe?) before final step.
 

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Thanks for report @rkpetersen this is interesting, @Rick Mathew and others were talking about this Q on the consistency of results. Others looking into this with the red sea pro kit see more inconsistency - and some workarounds to avoid it. I haven't used that pro kit yet, but I haven't really seen inconsistency.
I have nothing but wild speculation as to why the one kit might give less uncertainty than the other - with similar methods.
I just know the other experimenters in this are at least as OCD on testing methods as I am, so it's not an issue of care.
I have only used the Nitrate Pro Kit....So I have no experience with the other kit. What I have found is the two factors that effect my test consistency are the measuring out of the reagents ....and making sure I follow the test protocol ...mixing times...weight times etc. Also making sure there are no "floaters" in the measurement Cuvette helped a lot....I am still accumulating the data to do the statistics on the test with the HI-764 Nitrite but when I have it completed I will post it...from casual observation the repeatability looks good...but some times casual observation is bias...If I use the Red Sea color evaluation the test appears to be accurate according to their scale...For me accuracy is still an open question....as long as the test is consistent...even consistently wrong we can do the math to correct it;);)
 

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Thank you @taricha for a wonderful write-up and clear instructions. I have wanted something like this for some time as I have very low nitrates and was not satisfied with the "somewhere between 0 and 2 ppm but closer to 0" reading I was getting from the Red Sea kit alone. I used the NO3 reagents from the Red Sea Marine Care kit and since I knew my reading would be between 0 and 1.5 ppm, I did not dilute my sample. I took 10 ml directly from the tank, added double the reagents per your instructions, and timed the Hanna test with the 9 minute developing time so the 3 minutes on the Hanna checker elapsed a few seconds after my stopwatch for the developing time. After the 3 minutes, my Hanna checker displayed 58 indicating that the .25 ppm NO3 I was guessing at from the Red Sea kit alone was actually .56 ppm! This is super helpful and thanks again! When is @Hanna Instruments going to reach out and compensate you for developing a LR Nitrate checker for them?!
 
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taricha

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I used the NO3 reagents from the Red Sea Marine Care kit and since I knew my reading would be between 0 and 1.5 ppm, I did not dilute my sample. I took 10 ml directly from the tank, added double the reagents per your instructions,
Thanks! Glad you like it - that'll be close....
But - since you did yours at full salt water and these numbers were done for 1/10 dilution with distilled, then your results will under- count nitrate by some amount I don't know. Maybe 10% low, maybe 50% low - I don't have the numbers to say for sure.

edit: Check post #23 to see discussion of rick's data showing that salinity reduces the measurement.
 
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Thanks! Glad you like it - that'll be close....
But - since you did yours at full salt water and these numbers were done for 1/10 dilution with distilled, then your results will under- count nitrate by some amount I don't know. Maybe 10% low, maybe 50% low - I don't have the numbers to say for sure.

edit: Check post #23 to see discussion of rick's data showing that salinity reduces the measurement.
Thanks for catching that. So since my sample salinity was 33 ppt and yours after dilution is something like 3.5 ppt, I don't really know by how much I should correct my Hanna checker reading before applying the equation. I have a 10 % WC to do today and I was going to run this test again so my best bet is probably to dilute my sample to the 1/10 and then expect a Hanna reading between 0 and 15. I'll apply the equation and then multiply by 10. Thanks again!
 
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