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Extinction coefficient


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#1 rajaram

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Posted 08 May 2009 - 06:52 PM

Hi friends... I have few queries to be solved...
1.How to calculate extinction coefficient for hemocyanin at 335nm?

For example:
hemocyanin concentration was calculated using an extinction coefficient (ΕmM1cm) of 17.26, calculated from (E1%1cm) = 2.83 (Nickerson & Van Holde 1971) on the basis of a functional subunit of 74 000 (Antonini & Brunori 1974).

2.If we have extinction coefficient, How to calculate molar concentration?
3.How consistant is this quantificaiton of biochemical compounds based on extinction coefficient?
I am unacquainted to this UV spectroscopy, please explain with funtamental concepts.
Thank you for reading my query

#2 Vini

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Posted 10 May 2009 - 10:21 PM

Hi friends... I have few queries to be solved...
1.How to calculate extinction coefficient for hemocyanin at 335nm?

For example:
hemocyanin concentration was calculated using an extinction coefficient (ΕmM1cm) of 17.26, calculated from (E1%1cm) = 2.83 (Nickerson & Van Holde 1971) on the basis of a functional subunit of 74 000 (Antonini & Brunori 1974).

2.If we have extinction coefficient, How to calculate molar concentration?
3.How consistant is this quantificaiton of biochemical compounds based on extinction coefficient?
I am unacquainted to this UV spectroscopy, please explain with funtamental concepts.
Thank you for reading my query


Hi rajaram,
to get the concentration, u must apply the formula

A=ecl, where A is the absorbance, e is the molar extinction coefficient, c is the concentration and l is the pathlength of the cuvette that u have used for recording the absorbance. is this what u wanted to know? Suggestion: read a basic book on spectro like Kemp or Wilson n Walker to get the fundas right.

#3 rajaram

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Posted 11 May 2009 - 06:38 AM

DRN,
Thanks. 43824 is molar extinction coefficient of BSA at A280. how to find 43824?

#4 Vini

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Posted 11 May 2009 - 07:12 AM

DRN,
Thanks. 43824 is molar extinction coefficient of BSA at A280. how to find 43824?



hi
didnt get ur question. if i have understood correctly, u want to find out how to arrive at the 43824 value....but you havent mentioned the concentration of BSA and the cuvette path length. if i assume both to be x and y respectively, then molar extinction coefficient = OD at 280/x.y

#5 hobglobin

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Posted 11 May 2009 - 11:25 AM

Perhaps this pages may help (in my work I looked up the values, because I only worked with known substances):

Link 1

Link 2
One must presume that long and short arguments contribute to the same end. - Epicurus
...except casandra's that belong to the funniest, most interesting and imaginative (or over-imaginative?) ones, I suppose.

#6 rajaram

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Posted 12 May 2009 - 08:39 AM

Hi,
Sorry, its like this "molar extinction coefficient at 280 nm for BSA is approximately 43,824 M-1 cm-1"

#7 rajaram

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Posted 12 May 2009 - 08:43 AM

Thanks, Its very useful links.




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