PHYTOCHEMICAL INVESTIGATION
PHYTOCHEMICAL INVESTIGATION
Test for Carbohydrate 1. Molisch’s test
2. Fehling’s test
3. Benedict’s test
Test for cardiac glycoside 1. Keller-killiani test
Test for anthraquinone glycoside 1. Borntrager test
Test for proteins 1. Biuret test
2. Millon’s test
Test for amino acids 1. Ninhydrin test
Test for steroid 1. Salkowski reaction
2. Liebermann-Burchard reaction
Test for flavonoid 1. Shinoda test
Test for alkaloid 1. Dragendorff’s test
2. Mayer’s test
3. Hager’s test
4. Wagner’s test
TEST FOR CARBOHYDRATE
Chemical test Observation Inference
TEST-1: Molisch’s test: To 2-3 ml of ethanol Violet ring at the Present
extract, few drops of α-naphthol solution was junction of two liquids
added and shaken well. Con. H2SO4 was added
slowly from side of the test tube.
TEST-2:Fehling’s test: Each 1 ml of Fehling’s A Brick red precipitate Present
and Fehling’s B was mixed and boiled for one
minute. Equal volume of test solution was added
and heated on water bath for 5-15 minutes.
TEST-3: Benedict’s test: Equal volume of Solution may appear Present
Benedict’s reagent and test solution was mixed in green, yellow or red
test tube and heated on water bath for 5 minutes.
TEST FOR PROTEIN
TEST-1: Biuret test: To 3 ml test solution, few Violet or pink color Present
ml of 4 % NaOH and few drops of 1 % CuSO4
solution was added.
TEST-2: Millon’s test: 3 ml of test solution was White precipitate. Present
mixed with 5 ml of Millon’s reagent. Warm precipitate turns
brick red or ppt
dissolves giving red
colour solution
TEST FOR AMINO ACIDS
TEST-1: Ninhydrin test: 3 ml of test solution Purple or bluish color Present
was mixed with 3 drops of 5 % Ninhydrin
solution and heated on water bath for 10 minutes.
TEST FOR STEROID
TEST-1: Salkowski reaction: Each 2 ml Chloroform layer Present
chloroform and con. H2SO4 was added to 2 ml of appears red and acid
test solution and shaken well. layer shows greenish
yellow fluorescence
TEST-2: Liebermann-Burchard reaction: 2 ml First red, then blue Present
of test solution was mixed with chloroform. 1-2 and finally green color
ml of acetic anhydride and 2 drops of con.H2SO4 appears
from the side of the test tube.
TEST FOR CARDIAC GLYCOSIDE
TEST-1:Keller-killiani test: 2 ml of extract was Reddish brown color Present
mixed with glacial acetic acid. One drop of 5 % appears at junction of
FeCL3 was mixed and 1 ml of con.H2SO4 was two liquid layers and
slowly added from side of the test tube. upper layer appears
bluish green
TEST FOR ANTHRAQUINONE GLYCOSIDE
TEST-1:Borntrager test: To 3 ml of extract dil. Ammonical layer turns Present
H2SO4 was added. Then it was boiled and filtered. pink or red
To the filtrate equal volume of benzene or
chloroform was added and shaken well. Organic
solvent layer was separate out and mixed with
ammonia.
TEST FOR FLAVONOIDS
TEST-1:Shinoda test: 5 ml of 95 % ethanol or Pink color observed Present
tertiary butyl alcohol was added to dry powder or
extract followed by few drops of con. HCL and
0.5 g magnesium turnings.
TEST FOR ALKALOIDS
TEST-1:Dragendorff’s test: Few drops of Orange brown Present
Dragendorff’s reagent were added to 2-3 ml of precipitate is formed
filtrate.
TEST-2:Mayer’s test: Few drops of Mayer’s Cream color Present
reagent were added to 2-3 ml filtrate. precipitate
TEST-3:Hager’s test: Few drops of Hager’s Yellow color Present
reagent were added to 2-3 ml filtrate. precipitate
TEST-4:Wagner’s test: Few drops of Wagner’s Reddish brown Present
reagent were added to 2-3 ml of filtrate. precipitate
