GLYCOSIDES (PART-II)

Prof. Dr. G. K. Dash

This lecture introduces to the students:
Study of source, commercial varieties, chemical
constituents, substitutes, adulterants, uses, diagnostic
features and specific chemical tests of following glycoside
containing drugs:

Aloes, rhubarb, senna

At the end of this lecture, students should be able to

Describe pharmacognosy of following glycoside containing

crude drugs:

Aloes, rhubarb, senna

Chemical tests of Glycosides
1. Chemical tests for anthraquinone glycosides
Bo n ago Test: To 1 gm of drug add 5-10 ml of dilute
HCl, boil on water bath for 10 minutes and filter. Extract the
filtrate with CCl4/benzene. Separate the organic layer in a test
tube, add equal amount of ammonia solution and shake. The
ammoniacal layer turns pink or red in colour.

Modified Bo n ago Test: To 1 gm of drug add 5 ml

dilute HCl followed by 5 ml ferric chloride (5% w/v) solution.
Boil for 10 minutes on water bath, cool and filter. Extract the
filtrate with CCl4/benzene. Separate the organic layer in a test
tube, add equal amount of ammonia solution and shake. The
ammoniacal layer turns pink or red in colour.
Chemical tests of Glycosides
2. Chemical tests for saponin glycosides

Haemolysis test: A drop blood on slide is mixed with few

drops of aqueous solution of saponin glycoside. The RBCs can
be seen ruptured when observed under a microscope.

Foam test: To 1 gm of drug in a test tube add 10-20 ml of

water, shake for few minutes and keep aside. Formation of
frothing on the surface of water up to 30 min indicates
presence of saponins.
Chemical tests of Glycosides
3. Chemical tests for steroid and triterpenoid glycosides
Libermann Bruchard test: Alcoholic extract of drug is
evaporated to dryness and extracted with CHCl3. To the
chloroform extract, add few drops of acetic anhydride
followed by few drops of conc. H2SO4 from the side wall of
test tube. Formation of violet to blue coloured ring at the
junction of two liquids, indicates presence of steroid moiety.

Salkovaski test: Alcoholic extract of drug is evaporated to

dryness and extracted with CHCl3. To the chloroform extract,
add few drops of conc. H2SO4 from the side wall of test tube.
Formation of yellow coloured ring at the junction of two
liquid, which turns red after 2 minutes, indicate the presence
of steroid moiety.
Chemical tests of Glycosides
3. Chemical tests for steroid and triterpenoid glycosides
Antimony trichloride test: Alcoholic extract of drug is
evaporated to dryness and extracted with CHCl3. Add
saturated solution of SbCl3 in CHCl3 containing 20% acetic
anhydride. Formation of pink color on heating indicates
presence of triterpenoids.

Trichloro acetic acid test: Triterpenes on addition of

saturated solution of trichloro acetic acid form colored
precipitate.
Chemical tests of Glycosides
4. Chemical tests for cardiac glycosides
Keller Killiani test: About 1 ml alcoholic extract of drug is
treated with 0.5 ml of strong lead acetate solution and
filtered. Filtrate is extracted with equal volume of chloroform.
Chloroform extract is evaporated to dryness and residue is
dissolved in 3 ml of glacial acetic acid followed by addition of
few drops of FeCl3 solution. The resultant solution is
transferred to a test tube containing 2 ml of conc. H2SO4.
Reddish brown layer is formed, which turns bluish green after
standing due to presence of digitoxose.

Baljet test: Thick section of leaf of digitalis when dipped in

sodium picrate solution, it forms yellow to orange colour.
Chemical tests of Glycosides
4. Chemical tests for cardiac glycosides
Legal test: About 1 ml alcoholic extract of drug is treated
with 0.5 ml of strong lead acetate solution and filtered.
Filtrate is extracted with equal volume of chloroform.
Chloroform extract is evaporated to dryness. The residue is
dissolved in 2 ml of pyridine. About 2 ml of sodium
nitropruside is added followed by addition of NaOH solution to
make alkaline. Pink colour develops.

3,5-dinitro benzoic acid test: To the alcoholic solution of

drug few drops of NaOH followed by 2% solution of 3,5-
dinitro benzoic acid is added. Formation of pink colour
indicates presence of cardiac glycosides.
Chemical tests of Glycosides (Contd.)
5. Chemical tests for Coumarin glycosides
Ferric chloride test: To the alcoholic extract of the drug,
few drops of alcoholic FeCl3 solution is added. Formation of
deep green colour, which turns yellow on addition of
conc. HNO3, indicates presence of coumarins.
Fluorescence test: About 1 ml The alcoholic extract of drug
is mixed with one ml of 1N NaOH solution. Development of
blue-green fluorescence indicates presence of coumarins.
6. Chemical tests for Cynophoric glycoside
Sodium picrate test: Powdered drug moistened with dilute
sulphuric acid is placed in a conical flask. A filter paper
impregnated with sodium picrate solution is trapped on the
neck of flask. When the flask is heated slowly, the filter paper
turns brick red colour due to liberation of HCN.
Chemical tests of Glycosides (Contd.)
7. Chemical tests for flavonoid glycosides
Ammonia test: Filter paper dipped in alcoholic solution of
drug when exposed to ammonia vapor, yellow colour appears
on the filter paper.
Shinoda test: To the alcoholic extract of drug, magnesium
turnings and dil. HCl is added, formation of pink colour
indicates the presence of flavonoids.
Vanillin HCl test: Vanillin HCl is added to the alcoholic
solution of drug. Formation of pink colour indicates presence
of flavonoids.
Aloe
Synonym: Aloe

Biological Source: Aloe is the dried juice collected by incision

from the bases of the leaves of various species of Aloes, namely:

Aloe barbadensis Miller (Aloe vera) (or Curacao Aloe)

Aloe ferox Miller (or Cape Aloe)

Aloe perryi Baker (or Socotrine Aloe)

Aloe africana Miller and Aloe spicata Baker (or Zanzibar

Aloe)

Family: All these species belong to the family Liliaceae.

 Aloe perryi
Aloe barbadensis

Aloe ferox Aloe spicata

Aloe
Diagnostic features:
Aloe (Chemical constituents)
Aloe contains up to 30% aloin (which is a mixture of three
isomers: barbaloin, -barbaioin and isobarbaloin).

Barbaloin present in all the four varieties (a slightly yellow-

coloured, bitter, water-soluble, crystalline glycoside).

-barbaloin is amorphous and present in Cape aloe and can

be produced from barbaloin on heating.

Isobarbaloin is crystalline, present in Curacao aloe and in

traces in Cape aloe and absent in Socotrine and Zanzibar
aloe (is a mixture of barbaloin and polyphenols).
Aloe (Chemical constituents)
Barbaloin is a C-glycoside.

Barbaloin on hydrolysis yields aloe emodin anthrone and glucose.

Aloe also contains aloinsides A and B (O-glycosides of aloin)

Besides aloe-emodin anthrone, aloe-emodin anthranol and aloe-

emodin are also present. Aloe contains a resin, which is ester of
p-coumaric acid or p-hydroxy cimammic acid esterified with
aloeresinotannol.
Aloe (General Chemical tests)
Prepare 0.1% (w/v) aqueous solution of aloes by gentle heating,
add to it 0.5 g of kiesulgur and filter through a Whatman filter paper.
Use the filtrate for the following tests:
Borax Test (or Schoen e en Reaction): To 5 ml of the
above solution, add 0.2 g of borax and heat gently till it gets
dissolved. Transfer a few drops of the resulting solution into a
test tube filled with distilled water, the appearance of a green
coloured fluoroscence due to the formation of aloe emodin
anthranol.
Bromine Test: When equal volumes of the test solution and
bromine solution are mixed together, it yields a pale-yellow
precipitate due to the production of tetrabromaloin.
Aloe (General Chemical tests)
Modified Bo n age Test: It is known that aloin (or
barbaloin) belongs to the class of C-glycoside which does not
undergo hydrolysis either by heating with dilute acid or alkali, but
it may be decomposed with ferric chloride due to oxidative
hydrolysis. Hence, the Modified Bo n age test employing
FeCl3 and HCl is used as stated below:

First of all heat together 0.1 g of powdered aloe with about 2 ml

of FeCl3 solution (5% w/v) and 2 ml of dilute HCl (6N) in a test
tube over a pre-heated water bath for 5 minutes. Cool the
contents and extract the liberated anthraquinone with carbon
tetrachloride. Now carefully separate the lower layer of CCl4 and
add to it ammonia solution. The appearance of a rose-pink to
cherry red colour confirms its presence.
Aloe (Specific Chemical tests)
1. Nitrous Acid Test: Crystals of sodium nitrite together with small
quantity of acetic acid when added to 5 ml of the test solution of
aloe, the following observations are noted:

Curacao Aloes: A sharp pink colour due to isobarbaloin.

Cape Aloes: A faint pink colour due to isobarbaloin.

Socotrine and Zanzibar aloes: comparatively lesser colour.

2. Nitric Acid Test: The Test solution of aloes when made to react
with nitric acid, it gives rise to various shades of colour:

Caracao Aloe : Deep brownish red

Cape Aloes : Initial brownish colour changing to green

Socotrine Aloes : Pale brownish yellow

Zanzibar Aloes : Yellowish brown

Aloe (Specific Chemical tests)
3. Cupraloin Test (or Kl nge Isobarbaloin Test): To 10 ml of a
0.4% (w/v) aqueous solution of aloe add a drop of the saturated
solution of copper sulphate, immediately followed by 1 g of NaCl and
20 drops of ethanol (90% v/v). It produces different shades of
colours depending on the variety of aloes used:

Carocao Aloes : A wine red colour lasting for few hours,

Caoe Aloes : A faint colouration changing to yellow quickly,

Socotrine Aloes : No colouration

Zanzibar Aloes : No colouration

Aloe (Uses)
Aloe and aloin are strong purgatives. In higher doses may
act as abortifacient.

Aloe vera juice helps digestion, normalizes acid/alkaline and

pH balance, encourages digestive bacteria and regularizes bowel
processing.

Aloe gel (juice) is used in sunburns, thermal burns, radiation

burns, and abrasion and skin irritation and prevents ulceration
and malignancy.

Aloes find its usefulness as an external aid to painful

inflammatory manifestations.

It constitutes an important ingredient in the Com o nd

Tincture of Ben oin (or F ia Balsam).
Aloe (Adulterants)

Natal aloes from A. candelabrum resembles to cape aloe

microscopically. Contains natalion, homonatalion and
nataresinotannol.

Mocha aloes brittle, black and glassy aloes with strong odour

Jafferabad aloes black colour, brittle with characteristic odour

 Rhubarb
Synonyms: Rheum; Radix rhei; Rhubarb rhizome.

Biological Source: Rhubarb is the rhizome and roots of

Rheum officinale Bail.

Rheum palmatum L.,

Rheum emodi Wall.

Rheum webbianum Royle.

Family: Polygonaceae

Surface: Smooth, pale brown to reddish.

Odour: Characteristic, slightly aromatic.
Taste: bitter, slight, astringent, gritty.
Rhubarb (Chemical constituents)
Contains anthraquinone glycosides and some astringent components

Anthraquinones with COOH moiety:

Examples: Rhein, Glucorhein

Anthraquinones without COOH moiety:

Examples: Emodin; Aloe-Emodin;
Chrysophanol; Physcion
 Rhubarb (Chemical constituents)
o Anthrones and Dianthrones
of Emodin

Heterodianthrones e.g. Palmidin A, B, and C, which are

produced from two different anthrone molecules, as stated under:
Palmidin A : Aloe-emodin anthrone + Emodin anthone
Palmidin B : Aloe-emodin anthrone + Chrysophanol anthrone
Palmidin C : Emodin anthrone + Chrysophanol anthrone

The astringent portion of rhubarb is due to gallic acid as

- and -glucogallin; tannin as d-catechin and epicatechin.
Rhubarb (Chemical Tests and Uses)
Chemical Tests
Rhubarb powder when treated with ammonia, gives pink
colouration.
Rhubarb gives a blood-red colour when treated with 5%
potassium hydroxide solution.
It gives a positive modified Bo n age test.

Uses
Used as a purgative
Used in the form of an ointment in the treatment of eczema,
psoriasis - as a potent keratolytic agent.
Rhubarb (Adulterants)
o Chinese rhapontic rhubarb from R. rhaponticum

o English rhubarb from R. officinale and R. rhaponticum

o Japanese rhubarb from R. correanum and R. palmatum

o Canton rhubarb from R. palmatum

o Rumex alpinus and Rumex obtusifolius

Senna

Synonym: Senna leaf

Biological source: dried leaflets of Cassia acutifolia (Alexandria

senna), or of Cassia angustifolia (Indian or Tinnevelley Senna)

Family: Leguminoseae
Senna
Chemical constituents: Sennosides A, B, C and D, which are the
dimeric glycosides having their aglycones composed of
either rhein and/or aloe-emodin moieties.
Senna
Chemical Tests
Bo n age Test: It gives positive Born rager s test due to
presence of anthraquinone glycosides.
The mucilage of senna gives a distinct red colour with
Ruthenium Red solution.
Substituents and Adulterants:
Dog senna (Cassia abovata) - contains less than 1% of
anthraquinone derivatives
Palthe senna (Cassia auriculata) - contains no
anthraquinone glycosides
Arabian Senna (wild variety of Cassia angustifolia)
brownish green in appearance
Uses:
Used as purgative in habitual constipation.
References
Evan, W. C. (2002). Trease & E an s Pharmacognosy (15th ed.).
London: Elsevier Ltd.

Evan, W. C. (2009). Trease & E an s Pharmacognosy (16th ed.).

London: Elsevier Ltd.

Shah, B. & Seth, A. K. (2010). Text book of Pharmacognosy and

Phytochemistry, Gurgaon: Elsevier Ltd.

Anonymous. (2012). Analysis of Antioxidant-Rich Phytochemicals.

Edited by Zhimin, X. U. & Howard L. R., West Sussex: John Wiley &
Sons Ltd.

http://www.epharmacognosy.com
Thank you