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Topic: UV Absorbance & calibration curve  (Read 8510 times)

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red1

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UV Absorbance & calibration curve
« on: April 22, 2004, 08:27:44 AM »
Hello all. I need help please. I'll try to explain my problem the best
that I can.

I have KNO3 solutions of 10mM, 20mM, 30mM, 40mM and 50mM. To these solutions I add a test reagent and Zn dust, then I take the absorbance. From these readings I am able to establish a standard curve. I understand that the Zn dust is added to reduce the NO3 in the solution to NO2, which reacts with the test reagent to produce the color change.

Now I need to make standards using NaNO2 but I don't know what concentrations to use. The NaNO2 standards need to correspond to the KNO3 standards. In other words, I need to know what amount of NaNO2 will yield the same absorbance as the KNO3 standards that I have. I tried using the same concentrations (10mM to 50mM of NaNO2) but the readings are way off. The test reagent is composed of 0.2g/L n-1-naphthyl-ethylenediamine dihydrochloride in 1.5N HCl and 10g/L
sulfanilic acid in 1.5N HCl. Any answer will truly be appreciated.
Thanks
« Last Edit: April 24, 2004, 06:51:55 PM by hmx9123 »

Offline eugenedakin

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Re:UV Absorbance & calibration curve
« Reply #1 on: May 10, 2005, 11:19:01 PM »
Hello red1,

There are some factors that could be at work here... The following are only suggestions...

1) according to your explanation, theoretically, 10mM of KNO3 = 10 mM of NaNO2 only if all the NO3 is reduced to NO2.  Most of the time, these reactions are time dependent and change according to very small amounts of contamination.  If the reduction of NO3 remains constant for an extended period of time, you have probably reached the endpoint.

2) add Zn dust to the NaNO2 solution as it may be a factor.

3) Timing....  The absorbance readings will change on the same machine with time ... restandardize frequently....

Technically, NaNO2 should not be used to standardize a KNO3 test, as there are too many variations (Na vs K, NO3 to NO2, the presence of zinc, K values of the Zn reaction, these are reasons that are immediately available).

To be safe, perform tests with the same standards and the same batch of reagents.  (use KNO3 standards for measuring KNO3, use NaNO2 standards for measuring NaNO2).

If you have the exact standardized test method, I can examine the test method to gather more information.

I hope this helps,

Eugene
« Last Edit: April 09, 2007, 04:28:10 PM by eugenedakin »
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savoy7

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Re:UV Absorbance & calibration curve
« Reply #2 on: May 11, 2005, 12:02:39 AM »
Dakin's reply was excellent, especially the advice on using the same test variable for the curve and using the same reagents/batch.

I would like to add:
1) blank - are you using a blank - what is it?  Is it everything but the test variable?
2)  How did you determine the wavelength used?  
Sometimes when I was coming up with my own protocols, I would put the same in the spect and time it looking for a stable absorbance.  I would use this time to read my samples.  After I got that, I would scan for max absorbance of my test variable.

just some thoughts, good luck

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