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Molar extinction coefficient calculation.

  1. Sep 29, 2013 #1
    1. The problem statement, all variables and given/known data

    I need to calculate molar extinction coefficient for Albumin at 280nm.


    2. Relevant equations

    A=ε*c*L
    ε=A/(c*L)
    But since L=constant, I can just say ε=A/c


    3. The attempt at a solution

    The given molecular mass of albumin is 66,5kDa, which equals 66500g/mol. I do understand that I can find c (concentration) via the molecular mass. What I don't understand is how can I find ε if I still have another unknown, which in this case happens to be A.

    So: ε(unknown)=A(unknown)/c(known).

    Have I missed something really simple and overthinking this for myself? How can I find Absorbance?
     
  2. jcsd
  3. Sep 29, 2013 #2

    epenguin

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    With an equation relating three things you need two to be able to determine the third.

    "How can I find Absorbance?"
    You measure it. Or someone else measured it and tells you. Or you know ε and c and calculate it.

    We guess you know c. We cannot understand from your question what your problem is and what you are given or know but you are probably overlooking some information you have been given.
     
  4. Sep 29, 2013 #3
    I need to calculate ε. In order to do that I need A and c. In this case, though, the only given information is that the molecular mass of albumin is 66,5kDa. I understand I need to know two things in order to determine the third, but I can't find a way to determine the first two in the first place.

    Since I've been told to calculate ε I believe I need to know A and c. I do know c, but as of right now I am stuck with trying to determine A in order to calculate c.
     
  5. Sep 29, 2013 #4

    epenguin

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    Probably a typo as it makes no sense. Your question hardly does - who has asked you to do what and given you what information? There has to be an experimental datum, either a concentration or an absorbance somewhere. The extinction coeffeicient of bovine serum albumin can easily be found on the webv http://en.wikipedia.org/wiki/Bovine_serum_albumin
     
  6. Sep 29, 2013 #5
    Determining the extinction coefficient of proteins without experimental data (an absorbance as a function of concentration curve) is not a trivial matter. The absorbance at 280nm buy proteins is due to, mainly, the aromatic side chain amino acids (Tyr, Trp, maybe Phe) under going electronic transitions of the pi system. Therefore the extinction coefficient will vary with size, primary structure, and conformation (due to interactions of the aromatic side chains with other species in the protein's native form). There are calculators out there which estimate the extinction coefficient based on the amino acid sequence, but an experimentally determined Abs([Pro]) curve is used most often.
     
  7. Sep 29, 2013 #6
    Thanks for the feedback, I'll leave it undone as of this moment and talk this through with my prof tomorrow. The context of this question/exercise indeed is very "awkward" may I say so. Was searching the internet for hours about this. Positive side was that I found this forum, which I obviously also joined and I'm glad I did. I'll definitely stick around ;)

    Thanks again.
     
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