UV-VIS Epigallocatechin Gallate Content Measurement

Determination of Epigallocatechin Gallate (EGCG) assay using UV-Vis (Ultraviolet-Visible) spectrophotometry involves measuring the absorbance of EGCG solutions at a specific wavelength. EGCG is a catechin and a major polyphenol found in green tea known for its potential health benefits. Here's a general protocol for conducting an EGCG assay:

Materials and Reagents:

EGCG standard sample
Suitable solvent (e.g., ethanol or methanol)
UV-Vis spectrophotometer
Quartz cuvettes
Pipettes and volumetric flasks
Deionized or distilled water
Stirring rod
Weighing balance
UV-Vis software for data collection and analysis
Procedure:

Sample Preparation:
a. Weigh an appropriate amount of EGCG standard sample accurately. The exact amount will depend on the expected concentration and the solvent used. Create a standard curve to determine the concentration.

b. Dissolve the EGCG standard sample in the selected solvent (e.g., ethanol or methanol) to make a stock solution. Ensure the sample is fully dissolved by stirring.

c. Dilute the stock solution to create a series of standard solutions with different known concentrations. For example, make solutions with concentrations of 10, 20, 30, 40, and 50 µg/mL.

UV-Vis Spectrophotometer Setup:
a. Turn on the UV-Vis spectrophotometer and allow it to warm up for the recommended time.

b. Set the wavelength range to the appropriate value, typically in the UV-Vis range, such as 200-400 nm.

c. Ensure the cuvettes and instrument are clean and free of any particles that may interfere with the measurement.

Blank Correction:
a. Prepare a blank solution using the same solvent used for the standard samples. The blank solution will serve as a reference to account for any absorbance due to the solvent.

b. Place the blank solution in a cuvette and insert it into the spectrophotometer. Set the spectrophotometer to zero absorbance using the blank solution.

Measurement:
a. Take one of the standard solutions and place it in a cuvette.

b. Insert the cuvette into the UV-Vis spectrophotometer.

c. Record the absorbance at the specific wavelength for EGCG. The characteristic absorption wavelength for EGCG is around 275-280 nm.

d. Repeat steps a-c for each of the standard solutions.

Data Analysis:
a. Create a standard curve by plotting the known concentrations of the standard solutions against their respective absorbances.

b. Using the standard curve, determine the concentration of EGCG in sample solution by measuring its absorbance and interpolating on the curve.