Chlorophyll saponification assay

Summary

Chlorophyll is an ester of a dicarboxylic acid that saponifies with a base to form a basic salt of chlorophyllic acid; after the formation of the salt, chlorophyll is much more hydrophilic and soluble in dilute alcohol.

Operation method

Chlorophyll saponification assay

Principle

Chlorophyll is an ester of a dicarboxylic acid that saponifies with a base to produce a basic salt of chlorophyllic acid, which reacts chemically as follows: after the formation of the salt, chlorophyll is much more hydrophilic and soluble in dilute alcohol.

Materials and Instruments

Chloroplast pigment extract Test tube Pipette KOH Methanol solution Benzene

Move

I. Materials and equipment

Chloroplast pigment extract, test tubes, pipettes, 20% ^KOH-methanol solution, benzene, and other materials.

Experimental Procedure

Take 3 ml of chloroplast pigment extract, (if the extract is too thick, add a little alcohol to dilute it first). Add 1 ml of 20% KOH-methanol solution and shake well. After a few moments, add 3 ml of benzene, mixed, slowly add 1 ml of distilled water to get three layers of solution, the upper layer of benzene solution, which is dissolved in carotenoids and lutein; the lower layer of alcohol solution, which is dissolved in chlorophyll a, b and a small amount of saponification of chlorophyll a, b and a small amount of lutein. The bottom of the test tube at the same time the phenomenon of white precipitate, which is produced by the cell protein emulsification phenomenon.

Another 3 ml of chloroplast pigment extract, directly add 3 ml of benzene and 1 ml of distilled water, because the chlorophyll is not saponified, more easily dissolved in the upper layer of benzene solution layer; with the saponified different, in the lower layer of the alcoholic solution, only a part of the lutein dissolved.

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