First you should find the percentage of your lot: you cannot do any calculation if the percentage spans from 65 to 88. On the vial or documents you got when buying the standard there should be a lot number from which Merck will tell you the exact % of oleic acid in the standard you bought.

Then you have to decide the range and units of your curve: do you want it as a percentage (by weight or volume?) or as molar (micro- or milli-?) or as ppm (or ppb?) what do you expect? How may points were you planning to do?

The last step will be to apply some formula to calculate the volume to add in your calibration curve.

May 13, 015

Dear Raja:

It is not possible to consider volumes for the oleic acid use. First, the commercial product, as you mentioned, is not pure enough (> 95%); second, the relative volume to a wished mass changes as temperature function. Nonetheless, it is easy to evaluate concentration of an oleic acid solution (for example, 2% in butanol) by titration with a sodium hydroxide solution (in water-ethanol; 10% water) at a well defined concentration (e.g., 0.05 N). Phenolphthalein can be used as color indicator during the titration. Once the oleic acid concentration of the titrated solution is evaluated, various dilutions of this solution can be performed in order to establish a standard curve.

PS. It is not clear how you would follow up the lipase activity and the standard curve: oleic acid concentration as function of what? I could guess that it is by GC. Then your standard curve can be constructed as oleic acid concentration = f (integrated surface of resolved picks). This must be linear.

I hope this can help. Welcome for any query

Hello, what lipase assay are you planning to use? In the assay I know the lipase separates triglycerides into fatty acids and glycerol. Then (within the same assay) the glycerol moiety is further derivatized and finally quantified as a dye. In a lipase assay the nature of the fatty acids bound to glycerol are not relevant. If you want to combine the lipase assay with oleic acid quantification you should first run the lipase assay on your samples to liberate oleic acid (and other fatty acids) from triglycerides. Then you can use GC after characterizing your standard as described by Simone and Joseph. Notice that the fatty acids need to be derivatized  prior to GC measurment. Usually you convert them into their respective methyl esters. 

Thank you sir.

Now i am doing Lipase assay by " Kwon and Rhee et al., olive oil as substrate, at final reaction Free fatty acid dissolved in iso-octane solution measuring at 715nm, standard curves of oleic acid. one unit of lipase activity is defined as the amount of enzyme liberated 1micomole free fatty acid in 1min at 37 degree Celsius."

• I had done protein estimation by Bradford's Method , Bovine Serum Albumin as Standard.
• and also had Enzyme assay O.D valueat 715nm, but i didn't know how to make Standard oleic acid ?

Dear Raja,

i am using oleic with alcohol and lipase as catalyst  and i am facing the same problem in how to make the calibration curve for oleic acid to study the enzyme kinetic , did you get any solution for this issue !

godd luck

How do I prepare a 10mM concentration of oleic acid from the stock? what volume of this stock is equivalent to this concentration?