The problem is that everytime getting the flavonoids content much higher than phenolics which may be, but in rare occasion. In phenolics, I am adding100 ul sample with 900 ul dH2O with subsequent addition of 1N 0.5 ml Folin-ciocalteau and 20% 2.5 ml Na2CO3. OD at 725 nm. standard gallic and tannic acid.
In flavonoids, I am addidng the 0.5 ml sample in 1 ml dH2O with subsequent 5% 1.5 ml NaNO2 and 10% 150 ul AlCl3. After 6 mins, adding 1M 1ml NaOH and 850 ul dH2O and OD taken at 510 nm. standard quercetin.
Lastly calculating the value with standard in ug/mg dry extract. But, everytime getting the flavonoids content much higher than phenolics.
1. please suggest if there is any problem in protocols.
2. in case of phenolics, for a dose of 100 ug, I am taking 10 ul of sample in 90 dH2O from a stock of 1mg/ml. Is that right?
In quantifying total phenolic compound, there are some consideration should be taken:
It hopefully can help you. thank you
Article Antioxidant activity, total phenolic and flavonoid contents ...
I get same results - flavonoids content (and even tannins) higher than phenolics in some plants. There's nothing wrong with your procedure. I can't explain why but it happens.
I also get same results. Total flavonoid content higher than total phenolic content in case of same sample but in different unit. In case of phenolics the unit is mg gallic acid equivalent/ g dry weight as I'hv used gallic acid as standard. For flavonoids, it is mg quercetin equivalent/g dry weight as quercetin is the standard. So, I think there's nothing wrong in the procedure and such results.
Hi Harto Widodo, Thank you for your valuable comment. I have kept that particular reagents and solvents in their respective percentages as you mentioned. But I have found that the standard which I am using in phenolics (tannic or galic acid) gives a linear graph within 10- 100 ug where standard of flavonoid quantification (quercetin), it has reached upto 250 ug. That's why, when I am going to compare my extract with these standards, the flavonoid exceeds the phebolics. Even If I use quercetin in both the cases as a standard it also gives the same result i.e. flavonoid > phenolics.
Hi Felix Nwafor and Jeeta Sarkar. Thank you for your concern. Good to know that u people also face the same. I think the problem is due to the standard, which I am using in phenolics (tannic or galic acid) gives a linear graph within 10- 100 ug where standard of flavonoid quantification (quercetin), it has reached upto 250 ug. That's why, when I am going to compare my extract with these standards, the flavonoid exceeds the phebolics. Even If I use quercetin in both the cases as a standard it also gives the same result i.e. flavonoid > phenolics. I will be eager to know if u people also get the same result with quercetin alone. Thank you.
I get same results. In some plants, flavonoides re higher than phenols. There is no difference in choosing the standard. Consider to the recommendation of widodo.
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