Respected Sir,

We use to follow the following method:

The free radical scavenging activity of the extracts was evaluated by 1, 1-diphenyl-2-picryl-hydrazyl (DPPH) according to the previously reported method by Shen et al., 2010. Briefly, an 0.1mM solution of DPPH in methanol was prepared and 1mL of this solution was added to 3 ml of the solution of all extracts in methanol at different concentration (50,100,200,400 & 800μg/mL).The mixtures were shaken vigorously and allowed to stand at room temperature for 30 minutes. Then the absorbance was measured at 517 nm using a UV-VIS spectrophotometer. Ascorbic acid was used as the reference. Lower absorbance values of reaction mixture indicate higher free radical scavenging activity. The capability of scavenging the DPPH radical was calculated by using the following formula.

DPPH scavenging effect (% inhibition) = {(A0 –A1)/A0)*100}

Where, A0 is the absorbance of the control reaction, and A1 is the absorbance in presence of all of the extract samples and reference. All the tests were performed in triplicates and the results were averaged

Reference

Shen Q, Zhang B, Xu R, Wang Y, Ding X, Li P. Antioxidant activity in vitro of selenium-contained protein from the se-enriched. Bifodobacterium animalis 01. Anaerobe, 2010; 16: 380-386.

Good Luck

Respected Sir,

We use to follow the following method:

The free radical scavenging activity of the extracts was evaluated by 1, 1-diphenyl-2-picryl-hydrazyl (DPPH) according to the previously reported method by Shen et al., 2010. Briefly, an 0.1mM solution of DPPH in methanol was prepared and 1mL of this solution was added to 3 ml of the solution of all extracts in methanol at different concentration (50,100,200,400 & 800μg/mL).The mixtures were shaken vigorously and allowed to stand at room temperature for 30 minutes. Then the absorbance was measured at 517 nm using a UV-VIS spectrophotometer. Ascorbic acid was used as the reference. Lower absorbance values of reaction mixture indicate higher free radical scavenging activity. The capability of scavenging the DPPH radical was calculated by using the following formula.

DPPH scavenging effect (% inhibition) = {(A0 –A1)/A0)*100}

Where, A0 is the absorbance of the control reaction, and A1 is the absorbance in presence of all of the extract samples and reference. All the tests were performed in triplicates and the results were averaged

Reference

Shen Q, Zhang B, Xu R, Wang Y, Ding X, Li P. Antioxidant activity in vitro of selenium-contained protein from the se-enriched. Bifodobacterium animalis 01. Anaerobe, 2010; 16: 380-386.

Good Luck

Just dissolve Dpph in methanol or ethanol. pH affects its function so do not use acidic solution and also do not use alkali solution. When perform the assay, the pH of extract needs to be adjusted to around 5. You can read my paper published in FSB for more.

Good luck.

Just prepare 0.004 % (V/V) solution of DPPH radical in methanol by dissolving 4mg of dpph in 100ml of methanol. then take 100ul of sample and add 3ml of 0.004% dpph solution. incubate for 30 minutes in dark and read at 517nm. for blank use 100 ul methanol and 3 ml of 0.004% dpph solution.

0.004% dpph solution is of violet color  and after incubation color of solution will be discharged depending upon the antioxidant power of the sample. 

use ascorbic acid as standard.

and calculate % inhibition  by

DPPH scavenging effect (% inhibition) = {(A0 –A1)/A0)*100}

Where, A0 is the absorbance of the control reaction, and A1 is the absorbance in presence of all of the extract samples and reference. All the tests were performed in triplicates and the results were averaged.

i am working on antioxidant activity of wild edible plants 

I am agreeable with Dr. Jingbo Li, just dissolve DPPH in methanol. Do not use acidic condition. HCl is strong acid which can donate proton to DPPH radical and thus give yellow color. You may refer to my paper in Food Chemistry. Good luck

I am working on antioxidant activity. Please find my antioxidant activity related paper. I hope, you will resolve your problem. Best of luck.