General Tests and Specific Tests for Saponin

General Tests

Foam Test Reagents: 20 mL distilled water, 3 drops of olive oil Procedure: About 2 g of the powdered sample was boiled in 20 ml of distilled water in a water bath and filtered. 10ml of the filtrate was mixed with 5 ml of distilled water and shaken vigorously for a stable persistent froth. The frothing was mixed with 3 drops of olive oil and shaken vigorously, then observed for the formation of emulsion.

Positive result: formation of emulsion

Explanation: It is a general characteristic of saponins in plant to cause persistent foam when the aqueous solution is agitated. This ability to foam is caused by the combination on nonpolar sapogenin and water soluble side chain

Froth Test

Saponins in plant materials can cause persistent foam when the aqueous solution is agitated.

Principle involved: The foam formed on the water surface indicates the presence of saponin.

Reagents: 10 mL distilled water is the reagent used in the said test. Procedure: One gram of the sample was weighed into a conical flask in which 10mL of sterile distilled water was added and boiled for 5 min. The mixture was filtered and 2.5 mL of the filtrate was added to 10ml of sterile distilled water in a test tube. The test tube was stopped and shaken vigorously for about 30 second. It was then allowed to stand for half an hour.

Positive result: honeycomb froth

Explanation: Saponins are classified as amphiphatic glycosides due to their ability of soaplike foaming when shaken in aqueous solution. Their ability to foam is caused by the combination of the non-polar sapogenin and the water soluble side chain. If the honeycomb froth is greater than 2 cm, height from the surface of the liquid persists after 10 minutes, the sample is considered positive for saponins. For plant extract with poor frothing effects, add a little 5% sodium carbonate solution to basify the extract. The formation of a stable and dense froth indicates the presence of fatty acids.

C) Capillary tube test

Saponins tend to lower the surface tension in water.

Principle involved: The lowering of the surface tension of water allows the formation of froth on the surface of water.
Specifications: Use a pair of 15 x 1mm. O.D. capillary tubes Procedure: Load a capillary tube with the plant extract from Ethanol extraction procedure by immersing the tube to a height of 10mm. in the plant extract. Load another capillary tube with distilled water. Lift the capillary tubes and keep both in a vertical position to allow the liquid inside to flow-out freely. After some time compare the height of the liquids in the two tubes.

Positive result: If the level of the plant extract in the capillary tube is half or less than that in the other tube containing water, then the presence of saponins may be inferred.
Explanation: The level of the plant extract is lowered due to the ability of the saponins to lower the surface tension in water.

Hemolysis Test

Reagent: Fresh blood sample and plant extract Procedure: Place 1 to 2 drops of the fresh blood sample into the glass slide Add a drop of the plant extract into the blood sample and cover it with the glass slip

Observe the formation of a clear zone around the area using a compound microscope

Positive result: a clear zone

Principle: Detects the presence of saponin

Explanation: This clearing is due to the rupture of the blood cells and being replaced by the greenish color of the liberated heme the red color