Total phenolic content (TPC) experiment with Folin-Ciocalteu reagent: Step-by-Step protocol

Folin–Ciocalteu Assay for Total Phenolic Content

The video demonstrates a Folin–Ciocalteu assay for measuring total phenolic content (TPC). Gallic acid standards are used to create a calibration curve. To test samples, substitute the sample for the gallic acid standard.

Reagents and preparation

• Folin–Ciocalteu reagent: Use only if it is yellow. The presenter notes that a green color indicates contamination.
• Sodium carbonate solution: Dissolve 7.5 g of anhydrous sodium carbonate in 100 mL of distilled water.
• Distilled water
• Gallic acid stock solution and working standards: Prepare working standards at different concentrations. The subtitle refers to preparation details shown on screen but does not provide the amounts.

Protocol

1. Prepare tubes for four gallic acid concentrations and a blank, with three replicates of each (15 tubes total).
2. Add 3.16 mL of distilled water to each tube.
3. Add 0.2 mL of Folin–Ciocalteu reagent to each tube. To reduce the risk of contaminating the stock, pour the reagent into a separate beaker before use.
4. Add 0.6 mL of sodium carbonate solution to each tube.
5. Add 40 µL of the appropriate gallic acid concentration to each standard tube. Do not add gallic acid to the blank.
6. Use a fresh micropipette tip for each gallic acid concentration to avoid cross-contamination.
7. Mix vigorously and incubate for 2 hours at room temperature, preferably in the dark.
8. Measure absorbance at 765 nm using a UV–visible spectrophotometer.
9. For test samples, replace the gallic acid standard with the sample.

Presenter

Natasya Azman, explaining the laboratory procedure.