Hi Duri Eun , first of all, what do you mean by positive control?

if you mean the standard antioxidant, the answer is NO, you do not need it. Because for calculating IC50 you need to construct a plot of DPPH inhibition (%) of different concentrations (about 7 ones) of your extract (y-axes) vs concentration of extract (x-axes). From the equation of the linear curve you obtained (y=a*x+b), IC5 will be calculated by replacing "50" instead of the "y" and you will obtain IC50=x=(50-b)/a

You don't need a positive control to calculate the values of IC50 or EC50. But the use of standards is necessary for the discussion ( the establishment of a comparison between the tested samples and standards reveals the significance of the obtained results).

Thank you Chaima! If I had to use a positive control, example Quercetin, what should I do with the concnetrations? I am having a hard time looking for references about the concentration I should set the positive control. Do I set only 1 concentration or do I follow a series of concentration that is the same with my extract?

You are welcome @Duri.

You need to prepare all your samples at different concentrations. for standards and crude extracts, I use usually these concentrations ( 4, 2, 1, 0.5, 0.25, 0.125, 0.0625 and 0.03125 mg/ml).

NB: the final concentration depend on the final volume of the reaction mixture, for example: if you add 25 μL of the first solution (4 mg/ml) to 975 μL of DPPH solution you need to calculate the final concentration in order to establish the curves.

Best regards.

Dear Duri Eun , sholud my R² value be equal to .99.. when performing dpph assay. In my case the absorbance decreased for first six concentrations then it remained constant. Here are my values

My sample concentrations 10, 20, 30, 40, 50, 60, 70, 80, 90, 100 microlitres

Absorbance respectively, 0.379, 0.242, 0.187, 0.146, 0.136, 0.126, 0.130, 0.130, 0.130, 0.130

Am i going right?.