Determination of phenolic content in plant tissues

Summary

Phenolic substances in plant tissues are oxidized to Da by polyphenol oxidase, and Da can automatically polymerize into colorful substances and browning occurs. Therefore, the quantitative determination of phenolics in plant tissues is of certain significance and is often used as an index in the physiological study of plant resistance. The purpose of this experiment is to learn the principle and method of determining the content of phenolic substances.

Principle

The basic principle of the determination of phenolic substances in plant tissues is that the Folin-phenol method is often used for the determination of phenolic substances. Phenolic substances can react with phosphorus bustardic acid and phosphorus key acid in the Folin-phenol reagent under alkaline conditions to form bustard blue and key blue compounds, which have the maximum light absorption at 700 nm, and the magnitude of their absorbance is directly proportional to the content of phenolic substances. The absorbance is directly proportional to the phenolic content. Therefore, the content of phenolic substances in plant tissues can be determined by measuring the absorbance.

Operation method

Determination of phenolic content in plant tissues

Principle

The basic principle of the determination of phenolic substances in plant tissues is that the Folin-phenol method is often used for the determination of phenolic substances. Phenolic substances can react with phosphoribustic acid and phosphorus key acid in the Folin-phenol reagent under alkaline conditions to form bustard blue and key blue compounds, which have the maximum light absorption at 700 nm, and the magnitude of their absorbance is directly proportional to the content of phenolic substances. The absorbance is directly proportional to the phenolic content. Therefore, the content of phenolic substances in plant tissues can be determined by measuring the absorbance.

Materials and Instruments

Material: Various plant tissues.
Apparatus: spectrophotometer, centrifuge, mortar and pestle, test tubes, test tube racks, pipettes etc.
Reagents:
(1) Folin-phenol reagent: weigh 25 g of sodium tungstate and 5 g of phosphomolybdic acid, put them into a reflux flask, add 12.5 mL of phosphoric acid and 188 mL of distilled water, and boil them together at reflux for 2 h. Cool them down, and then dilute them with distilled water to 1 000 mL.
(2)10% Na
2
CO
3
Weigh 10 g of Na2CO3 and dissolve in 100 mL of distilled water.
(3) Standard phenol solution: 0.45 mmol L
-1
catechol solution.

Move

The basic procedure for the determination of phenolic content in plant tissues can be divided into the following steps:

1 . Extraction of phenolic substances: weigh 1 g of plant material, add a small amount of water and grind it into a homogenate, wash the mortar thoroughly, and then dilute it appropriately to 100 mL.2. Standard curve: add all kinds of reagents according to Table 56-1, mix well, add 10% _Na2CO3 2 mL after 3 min, shake, and measure the absorbance at 700 nm after 1 h at room temperature. The absorbance at 700 nm was measured after 1 h at room temperature. The concentration of phenol solution was taken as the horizontal coordinate and the absorbance as the vertical coordinate to draw the standard curve.Table 56-1 Standard curve spiking table 3. Determination of phenol content of the sample: Dilute the extract appropriately, put 2 mL in a test tube, add 2 mL of Folin-phenol reagent, after 3 min, add 2 mL of 10% _Na2CO3, shaking, and let it stand at room temperature for 1 h. Determine its absorbance at 700 nm, and then find out the corresponding concentration of phenol on the standard curve.4. Calculation of results: Where: c I is the phenol concentration found on the standard curve, ^μmol・mL-1;V - is the volume of the sample, mL;W - is the sample mass, g.

Caveat

Phenolics are extracted to avoid being oxidized.

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