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  Section: Plant Lab Protocols
 
 
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Methodology for Phenolics

 
     
 
Phenols
 
Phenols, the aromatic compounds with hydroxyl groups are widespread in plant kingdom. They occur in all parts of the plants. Phenols are said to offer resistance to diseases and pests in plants. Grains containing high amount of polyphenols are resistant to bird attack. Phenols include an array of compounds like tannins, flavonols etc. Total phenol estimation can be carried out with the Folin-Ciocalteau reagent.
 
 



Principle

Phenols react with phosphomolybdic acid in Folin-Ciocalteau reagent in alkaline medium and produce blue colored complex (molybdenum blue).
 
 
Materials
80% Ethanol
Folin-Ciocalteau Reagent
Na2CO3,20%
Standard (100mg Catechol in 100mL Water)
Dilute 10 times for a working standard.
 
 
Procedure
1.
Weigh exactly 0.5 to 1.0g of the sample and grind it with a pestle and mortar in 10-time volume of 80% ethanol.
2.
Centrifuge the homogenate at 10,000rpm for 20 min. Save the supernatant. Re-extract the residue with five times the volume of 80% ethanol, centrifuge and pool the supernatants.
3.
Evaporate the supernatant to dryness.
4.
Dissolve the residue in a known volume of distilled water (5mL).
5.
Pipette out different aliquots (0.2 to 2mL) into test tubes.
6.
Make up the volume in each tube to 3mL with water.
7.
Add 0.5mL of Folin-Ciocalteau reagent.
8.
After 3 min, add 2mL of 20% Na2CO3 solution to each tube.
9.
Mix thoroughly. Place the tubes in a boiling water for exactly one min, cool and measure the absorbance at 650nm against a reagent blank.
10.
Prepare a standard curve using different concentrations of catechol.
 
 
Calculation
From the standard curve find out the concentration of phenols in the test sample and express as mg phenols/100g material.
 
 



Notes

1. If any white precipitate is observed on boiling, the color may be developed at room temperature for 60 min.
2. Express the results in terms of catechol or any other phenol equivalents used as standard.
 
 



References

1. Malick, C P and Singh, M B (1980) In: Plant Enzymology and Histo Enzymology Kalyani Publishers New Delhi p 286.
 
 
     
 
 
     




     
 
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