EXPERIMENT STUDY OF THE MORPHOLOGICAL CHARACTERS, PHYSICAL AND DATE:

NO: 3 CHEMICAL TESTS OF DRIED CLOVE FLOWER BUDS PAGE NO:

AIM: To study the morphological characters, physical and chemical tests of dried clove flower buds

REFERENCE:

1. Practical pharmacognosy by C. K. Kokate, 12th edition, 2008, Publisher - Vallabh Prakashan, Page
no. 48 – 50.
2. Practical pharmacognosy (techniques and experiments) by K. R. Khandelwal, 19 th edition, March
2008, Publisher – Nirali Prakashan, Page No. 97 – 99.
3. Experimental pharmacognosy by R. K. Nema and CS Bhan, 1 st edition 2017, Publisher – CBS
publishers & Distributors Pvt Ltd, Page No.
4. Textbook of Pharmacognosy by T. E. Wallis, 5 th edition reprint 2005, Publisher – CBS publishers
& distributors Pvt. Ltd., Page No. 169 – 179.
5. Pharmacognosy (Pharmacognosy & Phytochemistry) by M. Ali, Volume 1, 1 st edition reprint 2011,
Publisher – CBS publishers & distributors Pvt. Ltd., Page No. 454 – 458.

REQUIREMENTS:

Materials required:
a. Clove flower buds. e. Razor
b. Calibrated scale in millimeters. f. Slides
c. Filter paper g. Water (hot & normal)
d. Magnifying glass h. White A4 size paper
Chemicals required:
a. Lead acetate (25%) solution. c. Potassium hydroxide (5%) solution.
b. Ethanol d. Ferric Chloride (5%)
Apparatus required:
a. Microscope

THEORY:

Morphological evaluation or Organoleptic evaluation is the most informative evaluation and the first step towards the
establishment of identity and degree of purity. This is done by means of organs of senses to evaluate colour, odour, taste,
shape, size and extra features.

Biological source: Clove consists of the dried flower buds of Eugenia caryophyllus

Family: Myrtaceae

Synonyms: Caryophyllus, Caryophyllum, Caryophylli, Laung/Lavang (Hindi).

Chemical constituents:

1. Volatile oil (15-20)% 6. Eugenol (70-90)%

2. Tannin (10-13)% 7. Eugenol acetate
3. Resin 8. Isoeugenol
4. Chromone 9. Methyl and dimethyl furfural
5. Caryophyllin
Uses:

1. Carminative 5. Flavoring agent

2. Aromatic 6. Dental analgesic
3. Stimulant 7. Immersion oil in microscopy (R. I. = 1.53)
4. Antiseptic 8. Isolation of eugenol

Adulterants:

I. Exhausted cloves:
a. Darker in color.

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b. Don not yield oil when pressed with a finger nail.

c. Float when placed in freshly boiled and cool water.
II. Blown cloves: Expanded flower without corolla.
III. Mother cloves: (Anthophylli)
a. Ovoid brown berries.
b. Presence of starch grains.
IV. Clove stalks:
a. Thick-walled, pitted stone cells (upto 140 microns in diameter)
b. Calcium oxalate prisms are present.
c. Occasional small starch grains and reticulate vessel.

PROCEDURES: (to be written on the ruled side of the practical copy)

PROCEDURES FOR ORGANOLEPTIC EVALUATION

Sl.
Parameter Procedure
No.
Place the drug and in a white piece of paper and examine color in day light and note it
1 Color
correctly for external and internal side of crude drugs if applicable.

The odor of crude drug depends on the amount of active volatile constituents present in
crude drug and indicated by the term aromatic, spicy, camphoraceous or if cannot be
described we say it has adistictint odor.
Slow and repeated inhalation of the drug gives exact idea of odor. Note down the odor
2 Odor of the drug in three phases –
a. Whole drug
b. Powdered drug rubbed between the fingers.
c. Moistened powdered drug rubbed between fingers.

WARNING: Unless otherwise mentioned in the text, do not taste the drug.
• A substance to be tasted must be in solution. The solution is prepared by placing a
small quantity of drug in water and boiling it for 5 minutes.
• The solution is then filtered and allowed to reach a temperature of around 40 ºC.
• Place a little amount of the solution in between the tongue and note down the taste
3 Taste felt. (Do not swallow any part of the solution as it may be dangerous).
• As soon as the taste is fully developed, the mouth should be well rinsed with pure
water in order to remove powder particles and taste.
• While tasting a drug the concomitant effects such as coloration of saliva, frothiness
of saliva, benumbing effects upon tongue, mucilaginous condition, grittiness, etc.
should also be noted.

The shape of the drug depends on the part of the plant from where is obtained and it
4 Shape
can be ascertained by referring to the shapes generally associated with that part.

The length and breadth of the drug is measured in millimeters or centimeters wherever
applicable. A graduated ruler in millimeter is adequate for the measurement of the
length, width and thickness of crude drug materials. For small drugs like seeds and fruits,
5 Size
measurement is done by aligning 10 of them on a sheet of calibrated paper, with 1 mm
spacing between the lines, and dividing the result by 10. The size is then reported as the
average of the 10 drugs measured by the above process.

This evaluation is done in untreated drugs. If necessary a magnifying lens may be used.
Tactile
Wetting with water or reagents, as required, may be necessary to observe the
sensation,
6 characteristics of a cut surface. Touch the material to determine if it is soft or hard; bend
texture and
and rupture it to obtain information on brittleness and the appearance of the fracture
fracture
plane – whether it is fibrous, smooth, rough and granular.

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Procedure for physical test:

A. Buoyancy test: Place few cloves separately in two test tubes, one containing hot water and the other with cool water,
observe whether if the drug floats or sinks. Exhausted cloves sink in boiled and cool water.

Procedure for chemical tests:

A. Tannins:
i. Take 2.5 ml of the aqueous extract1 of dried clove flower buds in a clean test-tube and add lead
acetate (25% w/v solution in water) solution dropwise. Shake gently to mix the lead acetate
solution and keep the test tube in the test-tube rack for some time White precipitate2 denotes
presence of tannins.
ii. Take 2.5 ml of the aqueous extract in a test tube and add (1-3) drops of Ferric chloride (5% w/v
solution in water). Dark color is formed when tannins are present.
If the extract is darker in color then dilute the test solution with water to make the observation
of color3 easier.
B. Eugenol:
i. To the 2 ml of alcoholic solution of clove oil in a test tube, add Ferric chloride (5% w/v solution
in water) dropwise. Blue color3 is formed if Eugenol is present.
C. Eugenolate:
i. To 2.5 ml of Chloroform extract add 2 ml of Sodium Hydroxide (3% w/v solution in water). Mix
it by gentle swirling4. Allow the two layers to separate by keeping it aside in the test tube rack.
The lower aqueous layer contains Sodium Eugenolate crystals. Separate the lower layer by
drawing the liquid with a syringe and place a drop of this separated layer in a slide and view under
microscope5.
ii. Make a thick section of hypanthium, place it in a microscopic slide. To the section add 1-2
drops of Ferric Chloride (5% w/v solution in water). Crystals of Potassium Eugenolate are
formed which can be observed by placing the slide under microscope5. Record the observations
made under microscope.

Chromone

Eugenol

Lateral section of Clove flower bud Morphological diagram of clove flower bud

1 Aqueous extract is prepared by taking (2-5) g of powdered clove flower buds in a beaker with 20ml of water and boiling it for around
(5-10) minutes. The solution was allowed to cool for some time and filtered. This filtrate was then used for the test for tannins.
2 Precipitation is the creation of a solid from a solution. When the reaction occurs in a liquid solution, the solid formed is called the

'precipitate'. The chemical that causes the solid to form is called the 'precipitant'. Without sufficient force of gravity (settling) to bring the
solid particles together, the precipitate remains in suspension.
3 Use a white piece of paper as the background to make the observation of color more precise.
4 Avoid shaking the test tube with force as it will make an emulsion layer which give false result.
5 The potassium/sodium eugenolate formed should be observed immediately as these salts are unstable and slowly decompose on exposure

to air. In this decomposition carbonates of the sodium/potassim metals are formed and the eugenol is partially oxidized to light-colored
amorphous powder having definite composition.

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OBSERVATION TABLES: (to be written on the plain side of the practical copy)

Sample: Dried Clove flower buds.

Parameter measured: Length and breadth (breadth of Corolla and Hypanthium); Chief chemical constituents

OBSERVATION TABLE FOR LENGTH

Sl. Length
No Sample (Li) Average Length (𝐋̅) in mm
(n) in mm
1 Clove 1 L1 = ∑ Li L1 +L2 +L3 +L4 +L5
2 Clove 2 L2 = L̅ = =
n n
3 Clove 3 L3 =
4 Clove 4 L4 = L̅ =
5 Clove 5 L5 =
n = total number of samples

OBSERVATION TABLE FOR BREADTH FO HYPANTHIUM

Sl. Hypanthium breadth

No Sample (Hi) ̅)
Average Hypanthium breadth (𝐇
(n) in mm
1 Clove 1 H1 =
∑ Hi H1 +H2 +H3 +H4 +H5
2 Clove 2 H2 = ̅=
H =
n n
3 Clove 3 H3 =
4 Clove 4 H4 = ̅=
H
5 Clove 5 H5 =
n = total number of samples

OBSERVATION TABLE FOR BREADTH OF COROLLA

Sl. Corolla breadth

No Sample (Ci) Average Hypanthium breadth (𝐂̅)
(n) in mm
1 Clove 1 C1 =
∑ Ci C1 +C2 +C3 +C4 +C5
2 Clove 2 C2 = ̅=
C =
n n
3 Clove 3 C3 =
4 Clove 4 C4 = ̅=
C
5 Clove 5 C5 =
n = total number of samples

OBSERVATION TABLE FOR CHEMICAL TESTS

Test Observation Inference
A. Tannins tests:
Test i)
Test ii)
B. Eugenol test:
C. Eugenolate tests:
Test i)
Test ii)
Test i, test ii, represents the individual tests given in the chemical test procedures

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RESULTS:
Morphological character:

I. Color:
a. Whole Drug :
b. Internal :
c. Powdered drug :
II. Odor:
a. Whole drug :
b. Powdered drug :
c. Powdered drug moistened with water :
III. Taste :
IV. Shape :
V. Size:
a. Length : (Avg.) mm
b. Breadth:
i. Corolla : (Avg.) mm
ii. Hypanthium : (Avg.) mm

VI. Flower: The flower bud is dome shaped head and a subcylindrical hypanthium. Head consists
of calyx corolla, stamens and styles.
a. Calyx: 4, thick and hard sepals with oil glands.
b. Corolla: 4 petals imbricate,enclose the stamens and form the head of the bud.
c. Androcieum: Numerous stamens, free and introse.
d. Gynocieum: Bilocular, inferior ovary, numerous ovules with axil. placentation.
Style single and erect. The oil glands are present throughout the flower bud.
Physical test:
I. Buoyancy test:
a. Cool water :
b. Hot water :
Chemical tests:
I. Tests for presence of Tannins :
i. Lead acetate test :
ii. Ferric chloride test :
II. Test for the presence of Eugenol :
III. Tests for Eugenolate:
i. Sodium eugenolate crystal test :
ii. Potassium eugenolate crystal test :

REPORT:

The morphological characters and physical and chemical tests of dried clove flower buds were studied, performed and
reported.

Teacher’s Signature: ________________________

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