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Specialty Chemistry Forums => Biochemistry and Chemical Biology Forum => Topic started by: asd123 on June 12, 2011, 01:29:20 PM

Title: Enzyme activity calculations
Post by: asd123 on June 12, 2011, 01:29:20 PM

Hi everybody!

The solution of enzyme reaction was made in the following way:
buffer: 0,8ml substrate solution 0,2ml and NADH 0,1ml
The reaction was started with 100microL enzyme solution.

There is graph where you calculate the absorbance. I think I got the right one A = 0,44
extinction coefficient of NADH is 6,22 x 10³M^-1cm^-1.

What is the enzyme activity U/ml?

Using the c = A/epsilon(=extinction coefficient of NADH)   you can get the concentration which is about 7,07 x 10^-5M.

So, how to proceed? My guess is that next I should divide this with total volume, but I am not sure.


Google does not find anything helpful. I have only on example which is quite unclear. Thank you beforehand for any help

Title: Re: Enzyme activity calculations
Post by: Yggdrasil on June 12, 2011, 02:48:11 PM

One unit of enzyme activity is defined as the amount of enzyme required to convert one µmol of substrate into product in one minute.  Therefore, to calculate the number of units of enzyme in your 100µL of solution, you first need to find out how many µmol of product you formed.

Title: Re: Enzyme activity calculations
Post by: asd123 on June 13, 2011, 05:03:17 PM

Quote from: Yggdrasil on June 12, 2011, 02:48:11 PM

One unit of enzyme activity is defined as the amount of enzyme required to convert one µmol of substrate into product in one minute.  Therefore, to calculate the number of units of enzyme in your 100µL of solution, you first need to find out how many µmol of product you formed.

Well, after dividing 7,07 x 10micromole x ml^-1 x min^-1 with 1,2ml(total V) i get about 5,89 U/ml so I think this is the correct answer. Isn't it? :)