One Unit is defined as the amount of the enzyme that produces a certain amount of enzymatic activity, that is, the amount that catalyzes the conversion of 1 micro mole of substrate per minute. A spectrophotometric assay is usually applied for this purpose by a selected substrate. Enzyme activity could be calculated using the following equation:

activity (U/L) = ((absorbance variation/time)/molar extintion coefficent * path length) * 1 micromol * (total reaction volume/total enzyme volume)

I don't get the question completly. What do you mean with mint? To calculate enzyme units you have to have a suitable activity assay. Then you can define your units Enzyme quantity to generate x mol Product per second.

Can you please specify the enzyme of your interest? In general, the micromoles of the end product released by the activity of one ml of an enzyme in one min is regarded as 1 IU/ml.

An enzyme unit is defined as the enzyme activity that catalyses the conversion of 1 µmol substrate into product in one minute.

Assay volume: 0.5 ml , Substrate converted: 500 µmol/ml/min

Enzyme units = 0.5 × 500/1 = 250 unit

Assay volume: 3 ml, Substrate converted: 100 nmol/ml/min

Enzyme units = 3 × 100/1000 = 0.3

One Unit is defined as the amount of the enzyme that produces a certain amount of enzymatic activity, that is, the amount that catalyzes the conversion of 1 micro mole of substrate per minute. A spectrophotometric assay is usually applied for this purpose by a selected substrate. Enzyme activity could be calculated using the following equation:

activity (U/L) = ((absorbance variation/time)/molar extintion coefficent * path length) * 1 micromol * (total reaction volume/total enzyme volume)

i have the same problem to get U/ml for lactoperoxidase from the reading and formula ,any one please let me know from formula how to get a figure in u/ml ?i have absorption =0.99, assay vol(cuvette)=1ml, 32.4 constant, ,sample vol=5ml,r these right reading adjustment to get U/ml ??

We need to put in all the fields in one unit only (Only CGS system) in the formula as suggested by Mr Daniele above to get the correct enzyme activity unit. Over and above this you may divide the figure with gm of proteins in the sample to derive the enzyme activity in tissues.

If I read your question correctly I assume the glucose is the product of your enzyme of interest. To calculate the glucose (product) content in your sample you can use glucose-6-phoshate dehydrogenase/hexokinase assay. However, from your heading you also want to calculate enzyme activity. This first thing you should do is to determine the amount of enzyme needed for the reaction. The kinetic data form this assay can also be used to calculate activity by converting the continuous assay into a stopped assay. Too much will convert substrate to quickly and too little will retard the process. This is done by having all of the reactions (I.e. reaction buffer, substrate and co-factors) already on the assay plate including all relevant controls. Take a reading on the spec which should be your A reading. Then induce the reaction by adding the amount of enzyme. Let the reaction run at the required optimum temperature for X minutes then stop the reaction. You can measure the amount of product formed in the set time by setting up a glucose-6-phoshate dehydrogenase/hexokinase. This answer will be mg prod/mg prot/ min which can be converted to Units/ ml through a series of calculations. Hope this helps...

NB. Monitor the glucose-6-phoshate dehydrogenase/hexokinase assay on the spec to give you an indication of when all the glucose is being consumed. Activity of the hexokinase should increase sharply in the initial stages of the assay but should run out of substrate which will be indicated by a plato. Take the last value as your B reading. The amount of glucose in the sample is directly proportionate to the amount NADH produced and can be calculated using Beers Law...

Could someone provide a reference about enzymatic activity calculation?

To determine lysozyme activity of fish serum, 50μl of serum was added to 1 ml lyophilized Micrococcus lysodeikticus at a concentration of 0.2 mg ml−1 (w/v) in phosphate-buffered saline (PBS, pH 6.3, 0.1M). The optical density (OD) was recorded at 530 nm after 1 and 20 min at 22 °C. Here PBS (pH 6.3, 0.1M) was used as a blank.

Here I got OD value in 1st minute was 0.842 and after 20th minutes was 0.757.

In my calculation, lysozyme activity is 85unit/ml of serum.

Anyone can suggest me about by protocol and calculation is correct or not???

If not correct please suggest me.

Thank you in advance.

Dear  Md. Tawheed Hasan, how  did you perform your calculation of the lysozyme activity given  the parameters you measured.?

how to convert nmol/ml to nmol/mgprot any one know about ???

You need to measure the amount of protein in 1 ml of your sample, then divide nmol/ml by this value to generate nmol/mg protein.

thanks

According to the enzyme and substrate, if you follow the consumption of substrate, it is necessary to

1. Prepare different concentration of substrate.

2. Plot the standard graph of substrate (Adsorption - Concentration).

3. According to the adsorption of enzyme and the standard curve, you can find the concentration of unreacted substrate.

4. From the initial concentration of substrate and unreacted substrate, you can calculate the consumed substrate.

The enzyme activity is calculated according to the following formula:

Enzyme Activity(μmol/min) or (U/ml) = (Consumed Substrate) ×Total Reaction Volume / (Reaction time)