TEST TUBE SPOT TEST
Culvenor and Fitzgerald have described a simple kit that can be taken into the field for use in testing samples of plant material for alkaloids. About 2-4 g of fresh plant part is ground in a small mortar with sand and sufficient chloroform/dichloromethane to make slurry. Ammoniacal chloroform/dichloromethane is added and the mixture stirred for 1 min, prior to filtration into a small test tube. Extraction of the alkaloids from chloroform/dichloromethane is accomplished by shaking the solution with 0.5 mL of 2N sulfuric acid and separation of the acid layer by means of a medicine dropper. A few drops of this extract are the tested with Mayer’s reagents and other reagents to ascertain the presence of alkaloids.
DETECTION OF SAFONINS
Saponins has several characteristic properties that can be used as a basic for simple detection test. (a) They are capable of hemolyzing red blood cells, (b) in aqueous media they will produce a characteristic honeycomb froth which persists for at least 30 min after vigorous shaking of the solution and, (c) they produce characteristic color reaction is the Liebermann-Burchard test.
The appearance of the characteristic honeycomb froth which persists for at least 30 min after shaking an aqueous boiled (3-5 min) mixture containing the plant material is evidence for the presence of saponins. If only a small froth is produced by this treatment, which is stable for only a few minutes, proteins, certain plant acids, or a low concentration of saponin may be the cause. This does not distinguish between steroidal and triterpenoid saponins.
The Liebermann-Burchard test may be used to detect the the presence of steroids and triterpenes. The following procedure has been used to detect saponins and/or free triterpenes or steroids: the finally ground sample (5-20 g) is extracted with boiling ethanol and the filtered extract is evaporated to drynees. The material obtained is triturated with ether. The residue (ether insoluble material) is subjected to froth and Liebermann-Burchard tests. If the froth test is positive, the ether insoluble material is hydrolyzed with aqueous hydrochloride acid 2N and any precipitate noted. The formation of a precipitate confirms the presence of saponin. The ethereal extract is evaporated to drynees and the residue is subjected to a Liebermann-Burchard test (L.B. test).
A small amount of the material to be tested is placed on a white title and a few drops of acetic anhydride are added and the strirred with a glass rod. A few drops concentrated sulfuric acid are now added and the colors produced are noted. Triterpenoids generally give red, pink, or purple colors which are more persistent than the blues or greens usually obtained from steroids.
1. Did a L.B. test can use for general terpenoid such as monoterpenoid, sesquiterpenoid, and polyterpenoid, explain briefly!
2. Define the reaction mechanism of triterpenoid and reagen L.B.!
3. How we can examined the alkaloids in a part of plant, explain two types of reagent that available for this test!
4. How to make reagent for detection of alkaloids?
5. Mention the various alkaloids that mostly present in the plants!
6. Why presipitant is the indicator the present of alkaloids ?
7. How we distinguish between steroidal and terpenoid with L.B. reagent and define the mechanism reaction involved!
8. How to detect saponins in the part of plant?