Hi all,
In the determination of Antioxidant capacity of fruit samples, I used DPPH assay and have a few questions regarding the DPPH assay
1) why is it necessary to vortex and incubate in dark the sample and DPPH solution? Is it so the sample and the DPPH can mix and allow the redox reaction to take place?
2) Why must DPPH be mixed with methanol to form methanolic DPPH? What is the purpose of methanol?
3) How did the equation %Inhibition= [(A0-A1)/A0] × 100 come about? Why is it not using beer-lambert law of calibration curve instead?
Please help! Thank you(:
I agree with the previous speakers. Voltexing ensures that the mixture is homogeneous. Dropping off in a dark place is to trigger a reaction.
The standard DPPH assay uses methanol as solvent, to keep the hydrophobic hydrazyl radical soluble while offering sufficient buffering capacity.
Best regards,
Aldona Adamska-Szewczyk
Nurul Ain Natasha , vortexing enables homogenization. That means that you will have the same solution all over its volume, so that when you withdraw some microliters with a pipette, you will be sure that the amount you withdraw is the same, as regards with the quality of the sample, with the rest of your sample. With reference to the dark, the DPPH solution utilized, demands darkness, as light would oxidize further the DPPH radical with the solution to be determined interfering with your results. So, stay close with the protocol and proceed by the book. I wish you fine results !
I agree with the previous speakers. Voltexing ensures that the mixture is homogeneous. Dropping off in a dark place is to trigger a reaction.
The standard DPPH assay uses methanol as solvent, to keep the hydrophobic hydrazyl radical soluble while offering sufficient buffering capacity.
Best regards,
Aldona Adamska-Szewczyk
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