In order to measure the polyphenols in broccoli, we prepared a reagent for measurement using the following procedure.
1. Extract the crushed plant with 70% methanol.
2. Mix 1 ml of the extract with 5 ml of 7.5% sodium carbonate and leave for 3 to 8 minutes.
3. Afterwards, 4 ml of 10% Folin-Ciocalteu reagent was mixed to make a total of 10 ml of measurement solution.
Immediately after mixing the Folin-Ciocalteu reagent, the mixed solution becomes cloudy. The color of the standard is clear.
What is the possible cause?
Hi,
May be, you have a solubility problem! You must mix your organic solvent with water to increase the polarity
I agree with Safia Taleb and I think the problem is with solvent. You should dilute the extract with redestilled water.
Hi Junzaburo,
Perhaps, if the problem was because broccoli contains a lot of minerals (such as Ca, Fe, Mg, etc) that some of them can react with carbonate ion from sodium carbonate to form its precipitated salt (such as CaCO3, FeCO3/Fe2(CO3)3, MgCO3, etc), I think using higher concentration (>80%) of methanol during initial extraction of the plant might help this problem, reducing the mineral content of the extract. In another method, you can try using a less polar solvent than methanol, (e.g., ethanol, acetone). I left two papers which might help as references.
During the reaction of the mixture, have you tried using a heating process in the water bath at around 45oC? Perhaps it could help the homogenization and completion of the reaction and might vanish the cloud effect. I did a heating process at around this temperature for determining the phenolic content of my tannin solution and the results were more stable.
Regards,
3 kind people Thank you for your answers
I would like to measure again after diluting it 10 times with distilled water.
What was precipitated was the salt that was formed of minerals.
I learned a lot.
Thank you again.
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