Chemical Tests for Glycosides in Pharmacognosy and Plant Chemistry Laboratory

Chemical Tests for Glycosides

Tests for Anthraquinone Glycosides

• Borntrager's test: Add 1 gm of drug to 5-10 ml of dilute HCl, boil on water bath for 10 min, and filter. Extract with CCl4/benzene, add equal amount of ammonia solution, and observe for pink or red color formation.
• Modified Borntrager's test: Add 1 gm of drug to 5 ml of dilute HCl, followed by 5 ml of Ferric chloride (5% w/v). Boil for 10 min, cool, filter, and extract with CCl4/benzene. Add equal volume of ammonia solution and observe for pink to red color formation.

Tests for Saponin Glycosides

• Hemolysis test: Mix a drop of blood on a slide with a few drops of aq. saponin solution. Observe for RBCs rupture in the presence of saponins.
• Foam test: Add 1 gm of drug to 10-20 ml of water, shake for a few minutes, and observe for frothing that persists for 60-120 s.

Tests for Steroid and Triterpenoid Glycosides

• Libermann Burchard test: Evaporate an alcoholic extract of the drug to dryness, extract with CHCl3, add a few drops of acetic anhydride, and then conc. H2SO4. Observe for violet to blue-colored ring formation.
• Salkowski test: Evaporate an alcoholic extract of the drug to dryness, extract with CHCl3, add conc. H2SO4, and observe for yellow-colored ring formation that turns red after 2 min.

Tests for Cardiac Glycosides

• Keller-Kiliani test: Add equal volume of water and 0.5 ml of strong lead acetate solution to an alcoholic extract of the drug. Filter, extract with chloroform, and add FeCl3 solution. Observe for reddish-brown layer formation that turns bluish-green after standing.
• Legal test: Add equal volume of water and 0.5 ml of strong lead acetate solution to an alcoholic extract of the drug. Filter, extract with chloroform, and add pyridine and sodium nitroprusside solution. Observe for pink color formation.
• Baljet test: Dip a thick section of a leaf of digitalis or the part of the drug containing cardiac glycoside in sodium picrate solution. Observe for yellow to orange color formation.

Tests for Coumarin Glycosides

• Ferric Chloride test: Add a few drops of alcoholic FeCl3 solution to a concentrated alcoholic extract of the drug. Observe for deep green color formation that turns yellow on addition of conc. HNO3.
• Fluorescence test: Mix an alcoholic extract of the drug with 1N NaOH solution. Observe for blue-green fluorescence.

Tests for Cyanophoric Glycosides

• Sodium picrate test: Moisten the powdered drug with water, add a few drops of conc. sulfuric acid, and trap filter paper impregnated with sodium picrate solution followed by sodium carbonate solution. Observe for brick red color formation.

Tests for Flavonoid Glycosides

• Ammonia test: Expose filter paper dipped in an alcoholic solution of the drug to ammonia vapor. Observe for yellow spot formation.
• Shinoda test: Add magnesium turning and diluted HCl to an alcoholic extract of the drug. Observe for red color formation. Add zinc turning and diluted HCl to an alcoholic extract of the drug. Observe for deep red to magenta color formation.
• Vanillin HCl test: Add vanillin HCl to an alcoholic solution of the drug. Observe for pink color formation.