Phytochemical screening

Qualitative testing of chemical constituents of plants

Preliminary step done usually in the crude extract of the plant of interest

Chemical modification of herbal preparations or crude drugs aids in monitoring instances adulteration or sophistication
Phytochemical analysis is significant in:

Drug discovery process

Quality control of herbal preparations

Evaluation of the chemical constituents of a plant involves qualitative and quantitative:

Chemical tests
Chemical assays
Instrumental analysis

Chemical evaluation involves:

Isolation
Purification
Identification or characterization procedures (chromatographic and spectroscopic methods)

Factors affecting preparation of crude extracts:

Choice of solvent
Extraction process
Presence of interfering compounds

Bitonio, Marky Donelly Y.

Ph Ch 128 Lab 2nd semester 2014-2015

Test for Carbohydrates

Test
Fehling's Test

Reagents
Fehling's A

Results
Brick-red precipitate (CuO)

(CuSO4 solution)

Notes
Positive for reducing sugars:
All monosaccharides
Many disaccharides (lactose, maltose,
cellobiose, gentobiose)
Nonreducing carbohydrates will, on boiling
with acids, be converted into reducing sugars

Fehling's B
(NaOH, Potassium sodium tartrate
(Rochelle salt))

Nonreducing substances:
Some disaccharides
(sucrose, trehalose (found in some fungi))
Polysaccharides

Molisch's test

1% alpha-naphthol in 95%

Purple color ring

ethanol, conc. H2SO4

Positive for soluble carbohydrates

Insoluble carbohydrates (e.g. cotton-wool or
cellulose) does not give the color until the
acid layer is shaken to bring it in contact with
the material

seliwanoff's test

Resorcinol, conc. HCl

Rose color

Rose color is produced if a ketone is present

(e.g. fructose, honey, hydrolyzed inulin)

tollen's
phloroglucinol test

Conc. HCl, Tollen's

phloroglucinol reagent

Cherry-red color

Galactose and ribose give cherry-red color on

being boiled with HCl and phloroglucinol
Glucose does not give the color in this test
because it does not form the furfural
derivative

Bitonio, Marky Donelly Y.

Ph Ch 128 Lab 2nd semester 2014-2015

test for glycosides

Test
anthraquinone
glycosides
modified borntrager
test

Reagents

Results

Notes

5% FeCl3

Formation of green color

5% FeCl3, conc. HCl,

Formation of pink to red color

Indicates the presence of C-glycosyl

compound

Green fluorescence when

Green fluorescence indicates the presence of

viewed under 365 nm UV light

aloe emodin anthranol

dichloromethane, NH3 TS
shouteten test

Boiling water, talc, borax

Tests for tannins and phenolic glycosides

Test
Ferric chloride test

gelatin test

Reagents
5% FeCl3

1% Gelatin solution containing

10% NaCl

Results

Notes

Blue color is produced which changes

With ferric salts, gallitannins and ellagitannins

to olive green as more ferric chloride

give blue-black precipitates and condensed

is added

tannins brownish-green ones

Precipitation

Solutions of tannins precipitate a 1% solution of

gelatin containing 10% sodium chloride
Gallic acid and other pseudotannins also
precipitate gelatin if the solutions are
sufficiently concentrated

Bitonio, Marky Donelly Y.

Ph Ch 128 Lab 2nd semester 2014-2015

tests for cardiac glycosides

The tests and assays available for cardio-active medicinals depend on biological activity, reactions of the sugar side-chain of the glycosides, and
properties of the butenolide side-chain
Test
liebermann-burchard
test

Reagents
Glacial acetic acid, dichloromethane,

Results
Blue-green color

Positive for most triterpenes and

conc. H2SO4
Salkowski test

Notes

sterols

Dichloromethane, conc. H2SO4

Formation of red color in the lower layer

Red - sterols are present

or formation of yellow color in the lower

Yellow - triterpenoids are present

layer
baljet test

Picric acid in ethanol, NaOH TS

Yellow to orange color

Yellow to orange color forms in the

presence of aglycones or glycosides

Legal test

Pyridine, 0.5% recently prepared

Formation of pink color

sodium nitroprusside, 0.2 N NaOH

Formation of pink color indicates the

presence of glycosides or aglycone
moiety

Keller-killiani test

Glacial acetic acid, 5% FeCl3,

Formation of reddish-brown layer forms

conc. H2SO4,

at the junction of the two liquids and the

upper layer slowly becomes bluish-green,
darkening with standing

test for saponins

Test
froth test

Reagents
Water

Results

Notes

Formation of foam

test for flavonoids

Test

Reagents

Results

shinoda test

Magnesium powder or ribbon, conc. HCl

Formation of red color

sulfuric acid

conc. H2SO4

Formation of orange color

Bitonio, Marky Donelly Y.

Notes

Ph Ch 128 Lab 2nd semester 2014-2015

test for cyanogenic glycosides

Cyanogenic glycosides yield hydrocyanic acid (HCN), benzaldehyde, and sugars on hydrolysis.
The medicinal activity of cyanogenic glycosides is due to presence of hydrocyanic acid and these are the characteristics of family Rosaceae.
Test
sodium picrate test

Reagents
10% aqueous picric acid, 10% Na2CO3

Results
Filter paper will become brick red

Notes
Hydrocyanic acid turns the filter paper
brick red

test for alkaloids

Test

Reagents

Results

Notes

(1% HCl)
mayer's reagent

Mercuric-potassium iodide TS

Formation of creamy-white precipitate

valser's reagent

Mercuric iodide TS

Formation of white precipitate

wagner's reagent

Iodine TS

Formation of reddish-brown precipitate

dragendorff's reagent

Bismuth potassium iodide TS

Formation of orangish red color

Bitonio, Marky Donelly Y.

Ph Ch 128 Lab 2nd semester 2014-2015