while performing DPPH antioxidant assay for my samples I am getting a lower absorbance value for blank when compared to my extracts. DPPH solution- 24mg DPPH mixed with 100mL methanol.
Blank- 50% acetone+ DPPH working solution (10uL+190uL)
Sample- Extract+DPPH working solution (10uL+190uL)
Standard- Trolox 20-250mg/L
I am subtracting blank absorbance from extract absorbance (Blank- Extract) to get the final value for extract absorbance and then using this value to calculate antioxidant capacity in terms of Trolox equivalence.
Could someone please advice why is this happening?
Thanks
Blank absorbance must be more than sample absorbance, It is likely that your samples are not well resolved in the solvent. The presence of sediment in the cuvette or plate to measure the absorbance, causes the samples to show more absorbance and is not the actual absorption of the sample and the sediment causes an error.
Hi Zahra, I have tried running the assay 3 times already and every time i am getting this issue. Don't know what shall I do .
Hi Prakhar, You can dilution concentration of your sample and centrifuge it if you have sediment.
Try to check the absorbance of extract itself, it may absorb at the same maximum wavelength with dpph.
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