[big]

Ammonia can be determined spectrophotometrically by reaction of phenol with hypochlorite. The following steps were taken:
-A 4.37 mg sample of protein was chemically digested to convert all its nitrogen to ammonia and then the volume of the sample was diluted to 100.00 mL in a volumetric flask.
-A 10.0 mL portion of the above sample was treated with phenol and sodium hypochlorite and diluted to 50.0 mL. The absorbance of this solution at 625 nm measured in a 1.00 cm cuvette. See data table.
-A reference solution was prepared by dissolving 10.00 g of NH4Cl (MW=53.50) in 1.00 L of water. A 10.0 mL portion of this was prepared in the same way as above and diluted to 50.0 mL See data table.
-A reagent blank was prepared using distilled water in place of the standard. See data table.

Sample	Absorbance
Blank	0.140
Reference	0.308
Unknown	0.592

1. What is the molar absorptivity, E, for the product [E=A/(bc), M^-1cm^-1]?

2. What is the weight percentage of nitrogen in the protein?

For the first one, I attempted to figure it out using the reference solution, as we already have the b and c for the reference solution. However, I had trouble figuring out what the A would be, since I wasn't sure if I was supposed to just use 0.308, or somehow incorporate the blank solution. Isn't it normally that the blank solution should be 0 and the reference solution should be 1?

As for the second question, I think I would be able to figure it out if I knew how to do the first.

Can anyone give me hints or clues? Thanks so much in advance!

[Borek]

Isn't it normally that the blank solution should be 0 and the reference solution should be 1?

No, it doesn't have to be this way.

Your blank and reference give you two data points - it should be enough to use them to find equation of the line going through both. Kind of a linear regression approach.

Although the experiment is poorly designed - not only two data points are in general not enough, as the dependence can be not linear, also your unknown is outside of the range. If it is a theoretical question, someone should be fired for giving bad examples. Unless discussion of these problems is also part of the question.