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Phytochemistry Lab: How to Identify the Phytoconstituents?

Phytochemistry Lab: How to Identify the Phytoconstituents?

The medicinal plants are in use for the treatment of various diseases since ages. Therapeutic properties are present in various parts of the plant such as the leaf, bark, roots, stem, flowers, seeds, and fruits. The plants owe these therapeutic properties to the presence of medically active chemical components known as phytoconstituents or phytochemicals. The phytoconstituents are known for their antibacterial, antimicrobial, antiprotozoal and other pharmaceutical functions.

But, how will you identify the phytoconstituents in a medicinal plant?

The identification process is carried out in a phytochemistry lab. Here are the steps to identify the different phytoconstituents.

Alkaloids

Alkaloids are naturally occurring compounds which contain nitrogen and carbon atoms. They are primarily found in plants, but also occur in bacteria, fungi, and animals. To detect the alkaloid in a medicinal plant:

  1. Dissolve the plant extract in chloroform
  2. Evaporate the chloroform and add acid to the residue
  3. Add different types of reagents to get different colour precipitates:
    • Mayer’s reagent gives a creamy white precipitate
    • Wagner’s reagent gives an orange precipitate
    • Dragendroff’s reagent gives an orange-red precipitate
    • Hager’s reagent gives a yellow crystalline precipitate.

Lactones

Lactones are the class of organic compounds containing a cyclic ester and formed by the intramolecular condensation of the hydroxycarboxylic acid. They are known to exhibit anti-inflammatory properties. There are two tests to detect lactones in a medicinal plant:

1) Legal’s test
  • Add a mixture of pyridine and sodium nitroprusside to the plant extract.
  • Then, add sodium hydroxide. The solution will turn deep red in colour.
2) Bal Jets test

Treat the plant extract with a solution of sodium picrate. The mixture turns yellowish-orange in colour indicating the presence of a lactone ring.

Glycosides

Glycosides are compounds formed from a simple sugar and another non-sugar compound by replacement of the hydroxyl group in the sugar molecule. Glycosides can be detected in plants by two methods:

1) Bal Jets test

Treat the plant extract with a solution of sodium picrate. The mixture turns yellowish-orange in colour indicating the presence of glycosides.

2) Keller-Killani test
  • Add few drops of ferric chloride and 1 ml of glacial acetic acid to the plant extract.
  • Slowly add concentrated sulphuric acid to the above mixture.
  • If glycosides are present, a reddish brown ring will appear at the intersection point of the two liquids.

Tannins

Tannin is a yellowish coloured derivative of gallic acid. It is bitter tasting organic substance found in the galls, barks, and other plant tissues. Detect the presence of tannins with these two tests:

1) Ferric Chloride test
  • Take 2 ml of the plant extract in a test tube.
  • Add a solution of ferric chloride drop by drop.
  • A blue-black precipitate indicates the presence of tannins
2) Gelatine test
  • Take 1% of gelatine solution containing 10% sodium chloride.
  • Add few drops of it to the plant extract.
  • A white precipitate at the bottom of the test tube confirms the presence of tannins.

Proteins

Proteins are commonly present in many medicinal plants and show their presence with the Biuret test.

Add 40% sodium hydroxide and diluted copper sulphate solution to the plant extract. Pink, blue or violet change in the colour of the solution indicates the presence of proteins.

Flavonoids

Flavonoids are a group of biologically active and water-soluble plant compounds responsible for the colour of the fruits and vegetables. They indicate their presence with these tests:

1) Ferric Chloride test
  • Prepare an alcoholic solution of the plant extract.
  • Take a small quantity of this alcoholic solution and add some drops of neutral ferric chloride solution.
  • If flavonoids are present, the solution will turn green.
2) Shinoda test
  • Prepare an alcoholic solution of the plant extract.
  • Add some pieces of magnesium ribbon to the alcoholic solution.
  • Add concentrated hydrochloric acid drop by drop.
  • Magenta colour of the solution confirms the presence of flavonoids.

Triterpenes

Triterpenes are the class of chemical compounds produced by the plants as a part of their self-defence mechanism. They can be detected by Salkaowski test.

Add concentrated sulphuric acid in drops to the chloroform solution of the plant extract. Shake well and allow it to stand. In few minutes, the lower layer will turn yellow in colour.

Saponins

Saponins are the chemical compounds with soap-like qualities and are found in abundance in various plant species. When you add plenty of water to the plant extract and shake it thoroughly, saponins produce foam which remains for about ten minutes.

Volatile Oils, Fixed oils, Fats

  1. Prepare a solution of the plant extract.
  2. Put a drop of this solution on the filter paper.
  3. Lack of permanent stain shows the existence of a volatile oil.
  4. Use two filter papers and crush the extract between them.
  5. A permanent stain shows the presence of fixed oils.
  6. Treat the extract with few drops of phenolphthalein and 0.5 N potassium hydroxide. Heat the mixture.
  7. Resulting soap formation shows the existence of fats and fixed oil.

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