Hi could someone help me, I want to do DPPH assay but my samples are in methanol and dmso and I don't know if the samples and stock solution of dpph should be prepared in the same solvent because I read that DPPH is prepared in methanol.
thanks
Hello Ricardo, I have performed the DPPH assay in the past (diluting DPPH in methanol) with samples that were resuspended in different type of solvents (ethanol, methanol, DMSO), without any problem. You will need to add the correct solvent blank (your combination of ethanol and DMSO), also try to add a positive control and keep in mind that the reaction is affected by pH.
In the paper: Molyneux, P. 2014. The use of the stable free radical diphenylpicrylhydrazyl (DPPH) for estimating antioxidant activity, there are a couple of recommendation such as the following:
''Regarding the solvent to be used, the method seems to work equally well with methanol or ethanol, neither of which seems to interfere with the reaction. The use of other solvent systems, such as almost neat extracts in water or acetone, seems to give low values for the extent of reduction (Guo et al., 2001).
Regarding the pH level, in the original Blois paper it was suggested that the system should be maintained at a pH in the range 5.0 to 6.5 by using acetate buffers; however, this precaution seems to have been abandoned in current practice. Indeed there is great uncertainty in the meaning of pH values in these predominantly organic (methanol or ethanol) media".
thanks for the advice, sorry someone told me that if necessary to wait like 12 hours before using dpph cause it needs to be stable but I haven´t found something about it I just read that after preparing dpph it should be immediately
used.
- Badges
- Science topic