Pharmacognosy-lab-manual.pptx

Faculty of Pharmacy
Department of Pharmaceutical Science
Laboratory Manual of
“Pharmacognocy and phytochemistry
2022-2023

COURSE SCHEDULE
Week
No. Topic Objectives
1 Introduction to Pharmacognosy
 Review Pharmacognosy deinition.
 Highliting Botanical products in
markets.
 Identifying microscops and
calculations of magnification
power.
 Review plant cell
 Review tissues and tissue systems
 Microscopically identification of
different types of calcium oxalate.
2
Macro and microscopical
analysis of leaves.
(Senna, Thymus, Mentha).
 To identify main parts constituting
leaves.
 To identify key elements of some
medicinal leaves.
3
analysis of flowers. (Clove,
Chamomile)
 To identify the main parts
constituting the flowers and fruits.
 To identify the key elements of some
medicinal flowers and fruits.
4
identification of fruits.
(Anise, Fennel)
constituting the seeds and
bark
 To identify the key elements of
some medicinal seeds and bark.
5
identification of seeds.
(Linseed, Cardamom)
constituting the subterranean
organs.
 To identify the key elements of
some medicinal roots and
rhizomes.
6
Macro and microscopical identification of barks. (Cinchona,
Cinnamon).
7
Macro and microscopical identification of subterraneanorgans.
(Liquorice, Ginger)
1

8
Detection of Anthraquinone Glycosides in Plant Extract
9 Extraction and identification of saponin and tannins
2

Safety Advice
The course is designated to be non-hazardous provided that good laboratory practices
followed.
1. Wear laboratory coats and keep them zipped all time.
2. Wear safety glasses when advised.
3. Some solvents are flammable; there should be no naked flames in the
laboratory, always use water bath for heating purposes.
4. Some chemicals are toxic if ingested, if you spill any chemicals on your skin
wash off immediately with cold water.
5. Mop up any spills of reagents on the bench immediately.
6. Some plants are toxic, DO NOT TASTE any.
7. Use the microscopes as advised by your instructor.
3

Lab. (1)
Introduction to Pharmacognosy
Part 1
What is Pharmacognosy?
The term “Pharmacognosy‟ was derived by the merger of two Greek words pharmakon
(drug) and gnosis (knowledge of) to mean the knowledge of drugs. It is field of science
with objective study of natural derived substances intended for pharmaceuticals and
poisons.
In this field of science, researcher deals with the secondary metabolites found in many
plants, animals and microbial natural sources, such as, plant leaves, seeds, fruits, stem,
roots, rhizomes, fungi, algae, corals, jelly fishes, sea cucumber,sea weeds, snakes, frogs
skin, cockroaches, and many more.
Among its scope are botanical products like:
- Medicinal plants
- Pure compounds from medicinal plants, vegetables or fermented by
microorganisms.
- Foods having health benefits “Nutraceuticals”
Pharmacognosy has been playing a significant role in the discovery, characterization,
production, and standardization of natural drugs.
The markets have plenty of botanical products sold as medicines. As part of the
primary health care team, pharmacists needs to be aware of the safety, effective, and
appropriate use of drugs including herbal medicine.
Part 2: Introduction to Botanical Methods
Examination of a biomass sample or a drug sample will end up with information that
will aid in identification of the sample.
Examination (botanical) methods include:
1. Macromorphological characteristics of the biomass.
2. Cytomorphological characteristics of the biomass: by examination of
microscopical characteristics of the biomass.
3. Phytochemical methods: Involve analytical methods like Thin Layer
Chromtography (TLC), High Pressure Liquid Chromatography (HPLC), Gas
Chromatography (GC), etc.
4

characteristics of the
Macromorphological
biomass:
Preliminary tests
The initial analysis of the biomass. Examination usually started with preliminary tests
involving general macroscopical (Macromorphological), physical and chemical tests.
These involve:
- Organoleptic properties including
- Color
- Taste (Should not taste plant and drugs in lab.)
- Odor
- Physical properties like
- Water and organic solubility
- Presence of oily substances
- Chemical properties including presence of certain constituents like:
- Saponins,
- Tannins,
- Anthraquinones
Part 3: Introduction to Microscopy and Calculations of MagnificationPower.
Microscopical characteristics of the biomass
The next step of analysis of the biomass is to identify the cytomorphological
characters, this is usually carried out with the aid of microscope.
The Microscope
The microscope (micro means small and scope means to see) is an optical instrument
to study small objects which produce an enlarged image of the object under study.
All modern optical microscopes designed for viewing samples by transmitted light
share the same basic components of the light path, listed here in the order the light
travels through them. Also almost all microscopes have the same 'structural'
components (Fig 1.1):
 Ocular lens (eyepiece): The function of the eyepiece is to magnify further the
image formed by the objective. It magnifies the image 10X
 Objective turret or Revolver or Revolving nose piece (to hold multiple
objective lenses)
 Objective (lens with variety of magnifications, the most widely used are X4,
X10, X40 and X100).
 Focus wheel to move the stage (coarse adjustment, fine adjustment)
 Frame
 Light source, a light or a mirror
 Diaphragm and condenser lens
 Stage (to hold the sample)
5

Magnification power
The imaging system improves resolution and magnifies the image consists of the
objective (4 lenses) and ocular lenses. The most common configuration includes four
objective lenses:
- Low magnification (4X)
- Medium magnification (10X)
- High magnification (40X)
- Oil immersion (100X)
Calculation of Magnification power
To calculate the total magnification of the compound light microscope multiply the
magnification power of the ocular lens by the power of the objective lens. For
instance, a 10x ocular and a 40x objective would have a 400x total magnification.
The highest total magnification for a compound light microscope is 1000x.
To avoid damage of microscope, never use the oil immersion without the oil.
Reagents used for preparation of specimen:
- Chloral hydrate: For the microscopical examination of dried plant material. It
acts as a clearing agent to improve visualization. Chloral hydrate is used as an
aqueous solution, often added to glycerol to prevent crystallization of the
reagent when used as a temporary mounting medium. The solvent effect is
improved by warming. It will remove certain characters like starch, mucilage
and aleurone grains.
- Phloroglucinol + HCl: For the qualitative analysis of organs containing lignin
and lignified tissues.
6

Fig 1.1 The Compound Light Microscope
7

Cytomorphological characteristics of the biomass
Different organs and cellular inclusions should be considered to distinguish and
identify different plants, especially those provided with the powdered not fresh
biomass. The following part will provide knowledge of cell contents, tissues and
tissue systems, and different cell types with their inclusions and functions. The use
of microscope is efficient to differentiate tissues and cell types
Cell contents
Pharmacognosy science is interested with the cell contents which can be identified in
plant drugs by microscopical, chemomicroscopical and physical tests. These contents
are either food storage products or the byproducts of plant metabolism and include
carbohydrates, proteins, lipids, calcium oxalates, calcium carbonate, tannins and
others. Some of these cell contents of diagnostic importance can be briefly described
as follows:
1Starch: is present in different parts of the plant in the form of granules of varying
sizes. Starch found in fruits, seed, root, rhizome and as smaller grains in chlorophyll
containing tissue e.g. leaf
2 Aleurone grain: Protein is stored in the form of aleurone grain by plants. It consists
of a mass of protein surrounded by a thin membrane, they are found in endosperm of
the seed.
Defat thin section containing aleurone grains and treat with the following
reagents:1- Alcoholic picric acid: aleurone grains are stained yellow.
2 Million‟s reagent: protein is stained red on warming.
3 Iodine solution: only crystalloid and ground tissues are stained yellowish brown.
3 Calcium oxalate: it is considered as excretory products of plant metabolism.
4Calcium carbonate: Aggregates of crystals of calcium carbonate are called
cystolith calcium carbonate dissolves with HCI, acetic acid or 1-11504 give
effervescence.
5Fixed oils and fats: are widely distributed in both vegetative and reproductive parts
of the plant. They are more concentrated in the seeds. Fixed oil present in association
with aleurone grains, they respond to the following tests:
1 They are soluble in ether, alcohol.
2 Dilute tincture of alkanna stains them red on standing for 30 min.
6Mucilage: are polysaccharide complex of sugar and uronic acids, formed from
the cellwall. The following tests are useful for the detection of mucilage:
1 Solution of ruthenium red stains mucilage pink.
2 Solution of corallin soda and 25% sodium bicarbonate solution stain the
mucilagepink.
8

Other cell contents:
Tannins: occurs in solution in the cell sap, when mounted in dilute solution of ferric
chloride, section containing tannin acquires bluish black or green color.
Volatile oil: occurs as droplets in plant cells.
Resin: is a secretory product, associated with volatile oil or gum or both
Tissues and tissue systems
When cells are grouped together for an identical function, a tissue is formed. In the
plant body, the following three tissue systems can he distinguished:
1. Dermal tissue system: It represents the outer most part of the plant which forms a
protective covering time epidermis, periderm. Stomata may occur in the epidermis
particularly in leaves. Trichomes are variable outgrowth of the epidermal cells, which
are useful in the identification of the plant material.
2. Vascular tissue system: It is concerned with transmission of material in the plant
and represents stellar structures like xylem and phloem (Fig 1.2); they are
responsible for the conduction of water and food material in plant.
3. Ground tissue system: It consists of simple cells, which may be thickened. It represents
ground tissue made up of parenchyma collenchyma and sclerenchyma
Figure 1.3. Types of cells
A: Parenchyma, B: Collenchyma with intercellular air space, C: Sclereid, D:
Sclerenchymatous fibre, E: Crystal fibre
9

Part 4: Microscopical identification of different types of “Calcium
Oxalate” crystals.
Calcium oxalate (Oxalic acid) is a very common cell-content in the plant kingdom. It
is formed in the cell as an end product of metabolism and it is rarely occurring in
the free state in plants. Calcium oxalate is insoluble in water and in acid cell-sap, it is
found in crystalline form in plant cell. Its different types exhibit a considerable variety
both in shape and in size, therefore it is a character of diagnostic importancein plants
to distinguish them from one another. They are usually found to the extentof 1%, but
in some instances it may exceed 20% (e.g. Rubarb rhizomes).
Forms of Calcium oxalate crystals (Figures 1.4 and 1.5)
1Monoclinic prism (Prismatic): Solitary or single crystals, very common, they may
be prisms, e.g. Hyoscymus niger, Solanaceae.
2Needle-shaped crystals: very narrow, slender, long crystals with pointed ends,
commonly occurring in bundles. e.g. Iris germanica, Iridaceae.
3Rosette crystals: formed of aggregated crystals radiating from a distinct usually
wide centre, e.g. Datura stramonium, Solanaceae.
4Sandy-crystals: very minute, usually filling the enlarged parenchymatous cells (calld
(Idioblast), e.g. Atropa belladonna. Solanaceae
5 Raphides e.g. Urginea maritima, Hyacinthaceae
Figure 1.4. Most common types of crystals. (a) Prismatic; (b) Sand crystal; (c)
Rosette; (d) Needle-shaped crystals or acicular crystals (raphi-des; transverse
section); (e) raphides (longitudinal section).
10

Experiment Lab 1.
Materials and methods
Part one:
 See the different parts of the compound light microscope and identify each part
and draw a simplified representation.
 Ethanol reagent is used to clean the ocular and objective lenses.
Procedure:
 Clarify different plant powders (Rubarb,Liqourice,Belladona) using Chloral
hydrate and examine the specimen under microscope starting with low
objective lens(4x) then medium O.L (10x) then high O.L(40x).
 Draw and label different type of calcium oxalate crystals.
Expected observation (400 X):
11

S
Report Sheet Lab. 1
Introduction to pharmacognosy
tudent name: …………………… Registration no.: ………………. Section no.: ……
Part 1: Define pharmacognosy
……………………………………………………………………….
……………………………………………………………………….
Part 2: Fill in the table
Product name Plant source Latin name Family Main ingredient (s) Therapeutic use
12

Part 3
A. Label the main parts of the compound light microscope illustrated below.
B. Calculate the final magnification power of specimens examined with each of
objective lenses provided in the lab
……………………………………………
…………………………………………….
……………………………………………
C. Draw the different types of calcium oxalate crystals observed under the
microscope and label them with the plant source, shape, and magnification power.
1 5
2 6
3 7
4 8
5
13

Lab. (2)
Macro and microscopical analysis of Leaves
14

Objectives
To identify the main parts and the key elements constituting the leaves
Introduction
The term leaf refers to the organ that forms the main lateral appendage on the stem of
vascular plants, leaves are organs responsible for the photosynthesis of plants.
Generally, the leaf is composed of small stalk (petiole), flat blade with green color
(Lamina) and the base, the base is normally connected to the petiole. The apex is the
distal end of the leaf. Leaves normally have pair of structures known stipules which are
located on each side of leaf base and may resemble scales, spines, glands, or leaflike
structures. The midrib is normally larger than the rest of veins. Figure 3.1 illustrates
the composition of most leaves.
Figure 2.1a. Composition of leaves. Figure 2.1(b) Types of leaves.
Leaves arrangement “Phylotaxis”
The regular arrangement of leaves or flowers around a plant stem is called Phyllotaxis, The
different types of Phylotaxis a: Alternative (or spiral); Opposite; Opposite decussate; and
Whorled. The Figure 3.2 illustrates types of arrangements of leaves.
Figure 2.2: Types of arrangements of leaves.
15

Figure. 2.3: Characteristic shapes of leaf blade (lamina)
16

Figure. 2.4: Characteristic shapes of leaf margin (lamina margin)
Figure. 2.5: Characteristic shapes of leaf venation (lamina venation).
17

Stomata
Stomata are found on the lower and upper epidermis, each stomata consists of two
similar cells, called guard cells, parallel to one another and inclosing between them a
small lenticular or oval space called osteole. The guard cells differ from the epidermal
cells in being much smaller containing chloroplasts and usually starch granules. The
epidermal cells surrounding the stomata may be clearly different bothin size and shape
from the remaining epidermal cells and thus described as subsidiarycells.
Figure 3.7: Types of epidermal stomata; A, Anomocytic ; B and C,
anisocytic;D, paracytic; E and F, diacytic; G, actinocytic.
Trichomes and Hairs
The epidermal cells may extend to the outside to form projection varying in size and
shape, if short and conical it is called papillae and the epidermis is described as
papillosed (as coca). These papillae appear in surface view as circles in the middle of
the cells. However, if the projection is longer and well protruding they form the
trichomes or hairs.
The trichomes are either covering trichomes (clothing, non glandular) or glandular (i.e.
with a swollen head formed of one or more secreting cells). The secretion of these cells
is mainly volatile oil or oleo-resin between the outer wall and the cutin which become
raised forming an inflated envelop enclosing the secreted substance.
18

Types of trichomes (Figures 3.8)
1- Covering trichomes or non-glandular hairs (Figure 3.9):They
may be unicellular or multicellular.
a) Unicellular:
1. Unbranched as in senna or
2. Branched as in many Cruciferae.
b) Multicellular:
1 Unbranched: a. Uniseriate: formed of one row as in belladonna.
b. Biseriate: formed of two rows each row of one (twin, hair as in
Arnica flower) or more cells (as in Calendula).
c. Polyseriate: formed of many rows of cells (shaggy hair as in
Cumin).
2- Branched: a.Simple branched: with uniseriate body ending in two branches as
in Tobaco.
b. Stellate: radiating unicellular hairs as in Boldo.
c. Peltate: with very short stalk surrounded by a plate-like structure
formed of very closer laterally joined cells.
2 Glandular trichomes:
a) Uniseriate stalk:
1- Unicellular head: found in Digitalis (one celled stalk), Belladonna
(multicellular).
2- Multicellualr head: found in Digitalis; more in Datura and Labiatae.
b) Biseriate stalk: and biseriate head as in Compositae.
c) Branched stalk: the branches end in heads as in Hyoscyamus muticus (one
celled head).
19

Figure 3.8: Types of trichomes
Figure 3.9: Examples of non-glandular trichome
20

Senna
Botanical source
Dried leaflets of Indian Senna “Cassia angustifolia Vahl.”, Family: Leguminosae.
Morphology
1) Imparipinnately compound leaf about 2.5 to 6.0 cm long.
2) Leaflet is lanceolate with entire margin, reticulately pinnate venation, acute and
mucronate apex, asymmetrical base.
4) Pale green in color.
5) Faint characteristic odor.
6) Mucilaginous and slightly bitter taste.
Histology
A transverse section shows:
1- Upper and lower epidermis - polygonal tabular cells with straight anticlinal walls.
2- Mucilage, in the inner periclinal walls.
3 Epidermal trichomes - unicellular
4Palisade: a single layer below upper and lower epidermis (isobilateral)
5- Paracytic stomata.
6 Large veins accompanied by calcium oxalate crystals.
7Cluster crystals of calcium oxalate in palisade and spongy tissue.
8- Spongy tissue of parenchymatous cells.
9- Meristele of radiate xylem and phloem with an arc of pericyclic fibers below and
schlerenchyma above.
Powder characteristics
1) Epidermis with paracytic stomata; 2) Calcium oxalate prisms; 3) Trichomes; 4)
Palisade and spongy cells; 5) Xylem vessels with annular thickening.
Important constituants
Anthracene glycosides, sennosides A & B.
21

Transverse section of senna leaflet: c, collenchyma; cr, calcium oxalate crystals;
crs, crystal sheath; l.e, lower epidermis; l.p, lower palisade; m, mucilage cell; ph,
phloem; p.f, pericyclic fibre; s, stomata; sm, spongy mesophyll; t, trichome; t.s,
trichome scar; u.e, upper epidermis; u.p, upper palisade; v.b, vascular bundle; xy,
xylem vessels.
Figure 3.10: Transverse section of Senna leaflet
22

3
Figure 3.11: Key elements in Senna leaflet.
1. Epidermis in surface view showing paracytic
stomata, a cicatrix (cic.), underlying palisade
cells and the elongated cells over a vein with
striated cuticle and an attached trichome.
2. Epidermis in surface view showing paracytic
stomata and underlying palisade cells.
3. Covering trichomes.
4. Epidermis in surface view with paracytic
stomata and an attached trichome.
5. Part of the lamina in sectional view showing the
upper epidermis containing mucilage (mu.), the
upper and lower palisade, spongy mesophyll
cells containing cluster crystals of calcium
oxalate and the lower epidermis. 9
6 Part of the 1arnina in sectional view with a
trichome attached to the lower epidermis.
7 Xylem elements from one of the larger
veins.
8 Part of a pitted vessel from one of the larger
veins.
9Cluster crystals of calcium oxalate.
10 Part of a group of fibers with calcium
oxalate prism sheath.

 Macroscopical identification:
 Examine a small amount of (Senna and thyme leaves), notice the
organoleptic characteristics (taste,colour,odor and texture).
2. Microscopical identification:
 Prepare amount of Senna leaves powder using chloral hydrate
(clarifying reagent) then put one drop of phloroglucinol reagent,after
one minute add one drop of HCl con and examine under microscope
the key elements. Draw and label
 Repeat the previous steps using the peppermint and thyme powder
without using (phloroglucinol+HCl con).
 Cross section: (Senna leaf)
 Cut the Senna leaf with blade on slide & make it very thin.
 Choose three or four cutting appear right & thin.
 Put on each part chloral hydrate to clarify the cross section.
 Heat the slide quickly to accelerate the reaction.
 Put one drop of phloroglucinol reagent on slide and wait one minute.
 Put HCl con.to slide which (phloroglucinol+HCl con give pink
color with lignin structure).
 Put the cover slide & examine under microscope layers of Senna leaf.
24

Expected observations (400X)
25

Report Sheet Lab. (2)
Macro and Microscopical Identification of Leaves
Student name: …………………… Registration no.: ………………. Section no.: ……
Plant sample number: 1
1.List the common name, botanical source (scientific name and organ) and
family of the examined plant?
Common name: ………………………………….
Botanical source:………………………………….
Family:…………………………………….
2. What are the organoleptic characteristics of the examined plant?
……………………………………………………………………….
……………………………………………………………………….
3. Draw and label the key elements examined under microscope.
27

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
28

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
29

Lab. (3)
Macro and Microscopical Analysis of Flowers
30

Objectives
To identify the main parts and the key elements constituting the flowers
Introduction
Flower is the reproductive organ of angiosperm plants, consisting normally of stamens
and carpels surrounded by petals and sepals all borne on the receptacle (one or more of
these structures may be absent). In some plants it is conspicuous and brightly colored
and attracts insects or other animals for pollination. The most important characteristics
are size, color, and arrangement of inflorescence. Morphologically, many parts of
flowers are modified leaves which during development of higher plants have taken on
specific functions for reproduction.
Figure 4.1 illustrates schematic representation of flowers.
Figure 4.1: A schematic representation of flowers
31

Flower parts:
1Calyx: composed of sepals, which is the outer most whorl of the floral series, it
functions as a protective for the essential organs. The sepals are green in color.
2Corolla: composed of petals, which is the inner whorl of the floral envelope, its
function is to attract the pollen carrying insects and protect the essential organs of the
flower.
3Androecium: it is male organ; formed of one or two or more whorls of stamens.
The stamen consists of filament and anther, the anther is divided into two anther- lobes
each possess a pollen sacs containing pollen grains.
4Gynaecium: it is the female organs; found in the centre of the flower formed of one
or more carpels and thus called mono, bi, tri, and polycarpellary. The gynaecium
consists of ovary, style and stigma.
Inflorescence:
An inflorescence may be defined as a cluster of flowers,
all flowers arising from the main stem axis or peduncle (Figure 4.2)
Figure 4.2: Simple inflorescence
The stem holding the whole inflorescence is called Peduncle and the major axis
holding the flowers or more branches within the inflorescence is called the rachis.
The stalk of each single flower is called a Pedicle. Inflorescences may be simple
(single) or compound (panicle). The Inflorescence can also be determinate or
indeterminate (Figure 4.3). The indeterminate simple inflorescences are generally
called Racemose, the main kind of racemose inflorescence is the raceme (Latin
racemus, cluster of grapes). The determinate simple inflorescences are generally called
cymose. The main kind of cymose inflorescence is the cyme (from the Latin cyma in
the sense „cabbage sprout‟).
32

Figure 4.3: Determinate and indeterminate inflorescence.
33

Types of inflorescence (Figure 4.4):
I. According to inflorescence position:
a. Terminal inflorescence: borne on the top of a stalk or peduncle
b.Auxiliary inflorescence: borne from the axis of the leaf, the main axis called
rachis.
II. According to the method of branching
a. Racemose :
the youngest flower at apex if the axis is elongated, or at the center if the axis is
shortened; and the oldest flower at base or outside, respectively. The Racemose
inflorescenee includes:
1 Raceme,
2 Corymb,
3 Umbel,
4 Capitulum,
5 Spike,
6 Spadix,
7 Strobile.
b. Cymose: the oldest flowers at the center and youngest nearest the base or to the
outside.
34

Figure 4.3: Types of inflorescence
35

Clove
Botanical source: Dried flower buds of Eugenia caryophyllus Spraque (or Zuzygium
aromaticum) (Family: Myrtaceae).
Histology (Figures 4.5, 4.6)
- A transverse section through hypanthium shows:
1) Epidermis with thick cuticle and stomata.
2) Collenchymatous parenchyma containing numerous schizolysigenous oil glands.
3) Calcium oxalate clusters in parenchyma.
4)Zone of slightly thick walled cells embedding a ring of about 15 bilateral vascular
bundles.
5) Meristele enclosed by lignified pericyclic fibers.
6) Xylem of 5-6 lignified vessels.
7) Parenchyma containing air spaces or lacuna (aerenchyma).
8)Central columella: parenchymatous cells containing cluster crystals of calcium
oxalate and 20-25 small vascular bundles.
- A transverse section through region of ovary shows:
1) Ovary with ovarian wall.
2) Parenchymatous dissepiments.
3) Ovules with axile placentation.
4) Starch, prisms of calcium oxalate and stone cells are absent.
Powder characteristics:
1) Epidermis with stomata.
2) Pollen grains.
3) Lignified fibers with parenchyma
4) Oil glands.
5) Arenchyma.
6) Fibrous layer of anther wall.
Important constituents: Volatile oil: its main component is Eugenol.
Uses: Antiseptic, carminative.
36

Figure 4.4: Clove flower Eugenia caryophyllus Spraque
(syn. Zuzygium aromaticum), Family: Myrtaceae
37

Zyzygium aromaticum. A, Penang clove; B, Zanzibar clove; C, fruit (mother clove), (all
x2); D, clove stalk (x1); E, clove cut longitudinally (x5); F, transverse sectionof
hypanthium; G, portion of anther (both x15); H, surface view of petal (x50); I-P,
elements of powdered clove (all x200); I, portion of anther filament; J, fibrous wall of
anther lobe and immature pollen; K, fragment of hypanthium showing portions of oil
glands; L, epidermal cells and stoma of hypanthium; M, parenchyma of hypanthium; N,
phloem fibres; O, pollen grains; P, sclereids from clove stalk. a, stamens; ae,
aerenchyma; a.l, anther lobes; c, columella; cr, cluster crystal of calcium oxalate; e,
epidermis; h, hypanthium; o, ovules; o.g, oil gland; p, imbricated petals; s, sepals; st,
style; v.b, vascular bundle.
Figure 4.5: transverse section of clove flower
38

Figure 4.6: transverse section of clove flower
1 Parenchyma of the hypanthium showing anoil
gland (o.g.).
2 Fibrous layer of the anther in sectional view.
3 Part of the filament of the anther in longitudinal
section, showing the central vascular strand and
parenchymatous cells containing cluster
crystals of calcium oxalate.
4 Fibrous layer of the anther in surface view.
5 Fibres and associated parenchymatous cells.
6 Epidermis of the hypanthium in surface view
showing a stoma and underlying oil glands.
7 Parenchyma of the hypanthium with cluster
8 Fragments of aerenchyma from the
hypanthium showing pits (pt.).
9 Part of the hypanthium in sectional view show
in the thick cuticle (c.), epidermis (ep.) and
underlying parenchyma with oil glands.
10 Mature pollen grains.
10a Part of a group of immature pollen grains.
11 Epidermis of the filament of the anther in
surface view.
12 Epidermis of a petal in surface view with
underlying tissue
parenchymatous cells
composed
containing
of
cluster
crystals of calcium oxalate, part of an oilgland
(o.g.) and part of a vascular strand.
13 Sclereids from the stalk.
39

Chamomile
Botanical origin: Dried expanded flower head of Matricaria chamomilla (syn.
Matricaria recutita), known as German chamomile and Anthemis nobilis known as
Roman chamomile (Family: Compositae).
Condition: dried double flower head; Shape: hemispherical; Color: whitish or buff;
Touch: soft; Odour: aromatic; Taste: bitter; Position: terminal; Kind: capitulum;
Receptacle: conical, hollow in German but solid in Roman;
Distribution of flowers and flower components (Figure 4.7):
In heads: consisting mainly of ray florets and tubular florets.
Ray florets: In German they are from one to three rows, but in Roman from 4 to 5.
Disc florets or tubular florets: In German it is numerous, but in Roman it is little in
the center of receptacel.
Corolla: 3 rounded teeth at the apex in ray florets, and five in tubular or disc florets,
epigynous.
Androecium: absent in ray florets, but syngenesious i.e. united by the anther in tubular
florets five stamens, epipetalous.
40

Gynaecium: synacarpus, two carpels, small, ovoid, unilocular and inferior ovary, one
basal ovule, style long thread-like and bifid stigma. Present in both florets.
Powder characteristics (Figure 4.8): Yellowish-white characterized by:
1- Spiny pollen grains, it has short and 3 germinal pores.
2 Papillae epidermis with striated cuticle.
3Fibrous layer of anther with spiral thickenings appearing in surface view with
beaded walls.
4Papillose of stigma, cluster of calcium oxalate.
5- Compositate glandular and nonglandular hairs.
Constitutents:
It contains: 1) volatile oil consists mainly of chamazulene; 2) flavonoids; 3)
coumarins and 4) salicylic acid.
Uses:
Anti-inflammatory and antispasmodic, aromatic and ulcer protective.
41

1
2
3
Matricaria chamomilla L.
1. Upper part of a flowering plant; 2. Vertical section of a capitulum; 3. A fruit
Figure 4.7: Distribution of flowers and flower components
42

1 Inner epidermis of a lobe of the corolla of a
tubular floret in surface view showing a group
of papillae (pap.) near the apex.
2 Cells from the inner tissue of the ovary wall
containing cluster crystals of calcium oxalate.
3 Fibrous layer of the anther in surface view.
4 Outer epidermis of the corolla of a tubular
floret in surface view showing a glandular
trichome.
5 Papillose stigma and part of a style in surface
view.
6 Part of a bract in surface view showing the
thin-walled cells and stomata from the
marginal region and elongated sclereids from
the central region.
7 (a) A group of immature pollen grains (b)
mature pollen grains.
3 Part of the filament of an anther in surface view.
9 Inner epidermis of the corolla of a ligulate
floret in surface view showing cells of the
margin with papillae (pap.).
10 The tip of an anther lobe in surface view
showing cluster crystals of calcium oxalate in
the underlying tissue.
11 Part of the ovary wall in surface view.
12 Outer epidermis of the corolla of a ligulate
floret showing striations and a glandular
trichome.
13 Sclereids from the base of the ovary wall.
Figure 4.8: Powder characteristics
43

1. Macroscopical identification:
 Examine a small amount of (Clove and Chamomile flowers), notice
the organoleptic characteristics (taste,colour,odor and texture).
2. Microscopical identification:
 Prepare amount of Clove powder using chloral hydrate (clarifying
reagent) and examine under microscope the key elements. Draw and
label
 Repeat the previous steps for Chamomile.
Microscopical study:
The following important points should be observed while examining the vertical
section to study the arrangement of different parts of the flower.
1- Epidermis of calyx and corolla with stomata and hairs.
2- Pollen grains.
3- Fibrous layer of anther wall.
4- Papillose stigma.
5- Thin-walled parenchyma.
6- Delicate vascular tissue.
44

Expected observations (400 X):
45

Macro and Microscopical Identification of Flowers
Common name: ………………………………….
Family:…………………………………….
2. What are the organoleptic characteristics of the present plant?
……………………………………………………………………….
……………………………………………………………………….
46

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
47

Lab. (4)
Macro and microscopical analysis of Fruits
(Anise; Fennel)
Objectives
To identify the main parts and the key elements constituting the fruits
A fruit results from maturation of one or more flowers, and the gynoecium of the
flower(s) forms all or part of the fruit.
The fruit is originally a result of fertilization, thus containing seeds, but some fruits
without fertile seeds are also of natural occurrence e.g. Banana.
Macroscopical (Organoleptic) features like shape, color, odor, and taste are
information for the identification of some plant species.
 Within pharmacognosy, principle types of fruits are:
1. Simple fruits (True): formed from a gynaecium with one pistil
2. Aggregate fruits: formed from more than one pistil.
3. Multiple (collective, Composite) fruits: formed from inflorescence and not from
a single flower.
48

The True fruit classified into:
A- Simple, Dry, Dehiscent: The pericarp become dry, split open when ripe to set free
the seeds, this includes:
1 Follicle,
2 Legume,
3 Siliqua,
4 Capsules.
B- Simple, Dry, Indehiscent: It is not split open when ripe, it includes:
1- Achene,
2 Grain,
3 Nut,
4 Samara
5 Cypsela.
C- Simple. Dry. Schizocarpic: They are two- to many seeded, bi to multilocular and
as they ripen, they split up into a number of one-seeded. Indehiscent parts called
mericarp, this include cremocarp as in Umbelliferae.
D- Simple Succulent: The mesocarp is fleshy, indehiscent, in some cases the soft part
of pericarp during ripening may peel off leaving the seed. It includes:
1- Drupe: One seeded fruit from one or more carpels, the epicarp is leathery, the
mesocarp is fleshy, and the endocarp is hard: e.g. Olive,Almond and Date.
49

2- Berry: Formed from one or more carpels, many seeded e.g Nux vomica orange and
capsicum.
Histology of the fruits:
The fruit consists of modified ovary wall as pericarp which is divisible into three
regions. The outer region is called epicarp, the inner known as endocarp and the
middle is termed mesocarp
50

Anise
Botanical source:
Dried ripe fruits of Pimpinella anisum Linn. (Family: Umbelliferae).
Morphology:
1) Entire cremocarps with pedicels attached; 2) Fruit 3-5 mm long and 1.5-2.0 mm
wide; 3) Ovoid-colonial, grayish brown, rough to touch: 4 )Short bifurcate stylopod
at the top; 5) 5 primary ridges on the surface: 6) When crushed gives aromatic odor,
sweet aromatic taste: and 7) Seed: orthospermous.
Histology:
A transverse section shows: 1) 2 vittae on the commissural surface; 20-40 small vittae
on the dorsal surface; 2) Vascular strands in the primary ridges; 3) Epicarp with short,
conical epidermal trichomes; 4) Mesocarp shown parquetry arrangement; 5) Testa of
thin, brown cells: and 6) Endosperm containing oil globules and aleurone grains.
Powder characteristic:
1) Short, conical trichomes; 2) Sclerenchyma; 3) Parquetry layer of endocarp; and 4)
Endosperm.
Important constituents: Volatile oil: main components: anethole, chavicol.
Uses: Carminative, aromatic.
51

Fennel
Botanical source:
Dried fruits of Foeniculum vuIgare Miller (Family: Umbelliferae)
Morphology:
1) Entire cremocarps; 2) 5-10 mm long, 2-4 mm broad, straight, oval in shape; 3)
Greenish brown in color; 4) Aromatic odor with characteristic aromatic taste; 5) Five
primary ridges prominent on the surface with a dried stylopod at the apex; and 6)
Seed-orthospermous.
Histology:
1) Epicarp-polygonal cells showing a few stomata;
2)Mesocarp- in the region of primary ridges, five vascular bundles with reticulate
lignified parenchyma above and below;
3) Four vittae on the dorsal surface, two on the commissural surface;
4) Endocarp narrow elongated cells showing parquetry arrangement; and
5) Endosperm- cellulosic parenchyma with oil globules and aleurone grains.
1) Reticulate lignified paraenehyma;
2) Vittae;
3) parquetry layer;
4) Endosperm and
5) Xylem vessels.
Important constituents:
Volatile oil: main components: fenchone, anethole.
Uses: Carminative, aromatic.
52

Anise. A, side view of cremocarp showing line of attachment to the two mericarps (X8);
B, transv‟rse section of mericarp (X25); C, covering trichomes of epicarp(X200); D,
branched and unhranched vittae isolated by alkali nacration (X25). a, line of attachment
of mericarps; c, carpophore; c.s, commissural surfaces; c.v, commissural vitta; en,
endosperm; e.t, epicarp bearing trichomes; m, meristele; mc, mesocarp; r,three of five
primary ridges of one mericarp, ra, raphe; s, stylopod; v, vittae.
53

f
Anise. A, Transverse section of cremocarp showing carpophore (cp); wide vittae ( öf ) on ventral
(commissural) surface and smaller vittae ( ö, ök ) between the ribs on the dorsal surface; fibrovascular
bundles (Gb); selerenchyrnatous fibers (sk); an air cavity in the wall of the pericarp on the ventral
side (h); tissue (R) with fibrovascular bundle (r); seed coat (Sa); and endosperm (Ed).B, Inner
epidermis of pericarp. C, Epidermis of seed coat. D, Cell of endosperm showing a number of aleurone
grains containing rosettes of calcium oxalate, a large nucleus in the center of the cell, and a few
isolated aggregates of calcium oxalate (d). E, Sclerenchymatous cells of the inner epidermis of the
pericarp in the neighborhood carpophore. F, Transverse section of pericarp andseed coat showing
epidermal dells (E) and a glandular hair having thick lamellated walls (h), parenchyma (p),
a vitta (ö), inner epidermis (Ei) and coat (Sa).
54

1. A group of sclereids from the mesocarp with
adjacent unlignified parenchyma.
2. Branching vittae (shown in outline only) and
underlying endocarp (en.) in surface view.
3. Covering trichomes (warty).
4. part of a group of fibro-vascular tissue.
5. Epicarp in surface view showing stomata and
striated cuticle
6. Part of two vittae showing transverse septa and
part of underlying endocarp in surface view.
7. Testa in surface view.
8. Endosperm containing microspheroidal crystals of
calcium oxalate
56

1. Reticulate parenchyma of the mesocarp.
2. Endocarp (en.) with overlying cells of the inner
most layer of the mesocarp, in surface view.
3. Endocarp in surface view.
4. Fragment of reticulately thickened vessel.
5. Elements of the fibro-vascular tissue.
6. Epicarp in surface view showing a stoma.
7. Fragment of a vittae.
8. Epicarp (ep.) and parenchyma of the mesocarp
in sectional view
9. Fragment of the vittae with overlying
thickened-walled cells of the innermost layer of
the mesocarp nin surface view.
10. Part of the pericarp and seed in sectional view
showing the reticulate parenchyma (r.p.),
endocarp (en.), testa (t.) and endosperm.
11. Endosperm containing microspheroidal
12. Thickened-walled cells of endosperm.
58

Fennel fruit (foeniculum capillaceum). A, a lateral view of a cremocarp (x4). B, dorsal
view of a cremocarp (x4). C, commissural surface of a cremocarp (x4). D, diagram of a
transverse section of a mericarp (x15). E, transverse section of part of pericarp within a
vascular strand and part of endosperm (x200). F, transverse section of pericarp including
a vitta (x200). G, an entire vitta (x15). H, part of vitta, showing segmentation and
epithelium (x200). L, parquetry layer in surface view (x200). K, outer epidermis of
pericarp(x200). al, aleurone grain; cp, carpophore; end, endosperm; e.t., epidermis of the
testa; f, fibers; i.e.p., inner epidermis of pericarp; ol, volatile oil; ra, raphe; r.p., reticulate
parenchyma; sp, stylopod; t, testa; v, vitta.
59

Materials and methods:
 Examine a small amount of (Anise and fennel fruits), notice the
organoleptic characteristics (taste,colour,odor and texture).
 Microscopical identification:
 Prepare amount of Anise fruits powder using chloral hydrate
one minute add one drop of HCl con and examine under microscope
the key elements. Draw and label
 Repeat the previous steps using the Fennel without using
(phloroglucinol+HCl con).
 Cross section: (Fennel fruit)
 Cute the Fennel fruit with blade on slide & make it very thin.
 Choose three or four cutting appear right & thin.
 Put on each part chloral hydrate to clarify the cross section.
 Heat the slide quickly to accelerate the reaction.
 Put one drop of phloroglucinol reagent on slide and wait one minute.
 Put HCl con.to slide which (phloroglucinol+HCl con give pink
color with lignin structure).
 Put the cover slide & examine under microscope the cross section of
fennel.
60

Expected key elements (400X):
61

Macro and microscopical identification of Fruits
Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
63

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
64

Lab. (5)
Macro and microscopical analysis of seeds
-A seed is an ovule which is matured and enlarged as a result of fertilization; it retains
its vitality for long periods.
-A typical seed consists of advanced embryo, the endosperm surrounding the embryo,
the perispem, all these structures are protected by the testa, which is formed of one or
two seed-coats.
- The testa is the outer surface of the seed which has certain markings:
a. Hilum: is the scar left on the seed where it separates from the funicle or stalk.
b. Raphe: is a ridge of fibrovascular tissue running from the hilum to the chalaza.
c. Micropyle: is the opening in the seed coat.
d. The chalaza: is the position at the base of the ovules (i.e. at the end opposite the
hilum).
-The seed is formed of seed coat and kernel, the kernel may consist of the embryo only
in the exalbuminous seed, or of the embryo surrounded by endosperm or perisperm or
both.
-The embryo consists of one or two cotyledons, primary shoot, stem or growing point,
and radicle or primary root.
Histology of the seed:
The most important microscopical features of seeds are mainly due to the structure of
testa as well as kernel including the cell contents, the diagnostic characters are:
1) Testa and outgrowth if present;
2) Perisperm;
3) Endosperm;
4) Embryo;
5) Cell contents.
65

Linseed (Flax, ‫ن‬
‫ا‬
‫تك‬
‫ال‬
‫ر‬
‫ذ‬
‫ب‬ )
Botanical source:
Dried ripe seeds of Linum usitatissimum Linn. (Family: Linaceae).
Morphology:
1) Glossy brown, finely pitted minute seeds;
2) 4-6 mm long, 2-3 mm wide and 1.5 mm thick;
3) Elongated, ovate, flattened, obliquely flattened at one end;
4)Hilum and micropyle in the angle of depression; and 5) A yellowish raphe runs
from hilum to chalaza at the end.
67

Histology: A transverse section shows:
1- Outer seed coat:
(a) Epidermis- polygonal tabular cells with thin anticlinal walls filled with stratified
mucilage, inner tangential walls supersized; (b) Mucilage stains red with ruthenium red
and blue with iodine; (c) Sub-epidermal layer (hypodermis): one or two layers of
cylindrical collenchyma.
2- Inner seed coat:
(a) A single layer of small, elongated, lignified; (b) A thin multiple hyaline layer of
collapsed parenchyma, obliterated parenchyma; (c) Inner epidermis of flat, sub-
rectangular or polygonal tabular cells containing a mass of brown pigment layer.
3- Endosperm and embryo of polyhedral cellulosic parenchyma with fixed oil
globules and aleurone grains.
I) Pigment layer in surface view; 2) Sclereids; 3) Epidermis and hypodermis; 4)
Endosperm with fixed oil globules and aleurone grains.
Identification test: Powder of the drug with ruthenium red shows pink color.
Important constituents: Fixed oil (linseed oil), protein, mucilage.
Uses: Demulcent, as poultice.
68

Linseed-Linum usitatissimum. A, capsule x2.5; B, seed x5; C, longitudinal sections of seed x5;
D, transverse section of seed x5; E, transverse section of testa and endosporrn x200; F, surface view
of layers of testa x 200. al, aleurone grains; cha, chalaza; cot, cotyledon; end, endosperm;ep,
epidermis; hi, hilum; hy, hypoderma or round-celled layer; i, inner integument ; k, sepals of calyx;
m, mucilage; o, outer seed coat; obl, obliterated parenchyma; p, pericarp; pg, pigment layer; ra,
raphe; rad, radicle ; sc1, sclerenchyma.
69

Microscopical analysis of Linseed powder
a. Fragment of the internal integument with the cells of the sclerenchymal fiber layer (1) and cells
from the perpendicular layer (2); narrow lumen of the sclerenchymal fibers, highly thickened
cell walls, numerous fine pits; transverse cell walls thin; fragments are yellowish to light brown,
numerous, not very characteristic.
b. Fragment from the pigment cell layer in top view and pigment aggregates which have dropped
from the cells; quadratic or polygonal cells with colorless rough and clearly pitted walls;
fragments colored brown from the cell content (pigment aggregates) numerous, very
characteristic.
c. Fragment from the seed coat with the annular cells in top view: rounded rough cells with pale
yellow walls; numerous, characteristic.
d. Oil droplets.
e.Fragment from the cotyledon tissue; parenchymal thin-walled, hyaline cells containing aleurone
grains and oil droplets; very numerous, not very characteristic.
f. Fragment from a cotyledon in cross section; numerous, not very characteristic. See also e.
g. Fragment from the seed coat epidermis in top view; frequently with mucin globules on the
fracture edge; numerous, characteristic.
Note: The drug must not contain starch grains, calcium oxalate crystals, hairs, sclerenchymal
fibers or collenchymal elements.
a
70

 macroscopical identification:
 Examine one of cardamom &Linseed seeds, notice the organoleptic
characteristics (taste,colour,odor and texture).
 Prepare amount of Cardamom powder using chloral hydrate
(clarifying reagent) and examine under microscope the key elements.
Draw and label
 Repeat the previous steps for Linseed.
 Oil drops in Linseed:
 Put little amount of Sudan 3 powder on linseed prepared slide.
 Examine the oil drops (pink color) under microscope.
71

Cardamom
Cardamom (sometimes cardamon or cardamum is a spice made from the seeds of
Amomum subulatum Family Zingiberaceae.
Botanic characteristics
Unground Cardamom Seed— Seeds usually appear in agglutinated groups of two to
seven seeds, and as separate seeds surrounded by an adhering membranous aril. The
individual seeds are oblong-ovoid or irregularly three-to four-sided, from 3to 4mm in
72

length. They are convex on the dorsal side, strongly longitudinally grooved on the
ventral side, coarsely tuberculated, and externally pale orange to dark brown. The
odoris aromatic. The taste is aromatic, pungent, and slightly bitter.
Histology
Sections show a loosely attached membranous aril; a brownish seed coatconsisting
of an epidermal layer of thick-walled cells, a pigment layer of small cells with red to
orange contents, a layer of volatile oil cells with suberized walls and a single layer of
radially elongated, strongly lignified stone cells with inner walls heavily thickened,
and a minute lumen containing silica; and a large, colorless perisperm surrounding a
central, orange to yellow endosperm in which a small, straight embryo is embedded.
Powdered Cardamom Seed— Brown to weak yellow to light olive green. It consists
chiefly of fragments of perisperm, endosperm, embryo, and seed coat. The
endosperm and perisperm cells are filled with starch grains from 1to 4µm in
diameter, or may contain one or more prisms of calcium oxalate from 10to 25µm in
diameter. The seedcoat is characterized by its red-to orange-colored cells, is
polygonal in surface view, and is about 20µm in diameter. Fragments of pericarp
tissue with spiral vessels and with accompanying slightly lignified fibers are
relatively few.
73

Expected observation of key elements (400X)
74

Macro and microscopical identification of seeds
Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
75

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
76

Lab. (6)
Macro and microscopical analysis of Barks
It consists of all tissues outside the cambium. A young bark is composed of the
following tissues:
1 Cork or phellem:
It is a protective tissue of secondary origin develops in stems and roots from a
secondary meristematic layer, the phellogen or (cork cambium). Cork is formed of dead
polygonal cells, they are usually uniform in shape, appear dark brown in color. In the
cell-walls may be embedded crystals of calcium oxalate.
2 Phellogen:
It is a secondary meristem developed by the activation of parenchyma. It is one layer,
by periclinal division produces cork cells outside and phelloderm to the inside. The
phellogen and its product i.e. cork and phelloderm called periderm.
3 Phelloderm:
Formed of unsuberised cells, one or two layers, it may be parenchymatous, or
collenchymatous. It may constitute the whole cortical part of the bark and thus
sometimes called secondary cortex.
4 Primary cortex:
It is absent in some barks due to decortication. It may be collenchyma or parenchyma,
and sometimes contain sclereid cells, fibers, oil glands or ducts, laticiferous structures
and mucilage cells. The cells usually contain small starch granules and crystals of
calcium oxalate.
5 Pericycle:
It may be formed of one to many cells, thick, it may be parenchymatous and thus
indistinguishable from the cortex or sclerenchymatous formed of interrupted band of
sclereid cells which accompanied on the outside by pericyclic fibers either isolated or
in groups.
6 Phloem:
It is formed of secondary phloem, which forms the inner bark. It is traversed by
medullary rays and characterized by the presence of sieve tubes and companion cells
and phloem parenchyma. The phloem contains also phloem fibres and sometimes
sclereid cells. The phloem may show oil cells, mucilage cells and other secretary
structures, also starch granules and crystals of calcium oxalate are also present in
parenchyma cells.
77

7- Medullary rays:
It is formed of radically elongated cells. The cells are parenchymatous, contains starch
granules and crystals of calcium oxalate.
Bark shape:
The shape of bark depends upon the type of incisions made by removing it and also
upon the extent and nature of the shrinkage, which occur during drying. Some are flats;
some are curved during drying, but if concave on the outside it is calledrecurved. If the
curvature, on the inside, is so great as to form a deep it’s called channeled; and when
still more curvature is present and one edge overlaps the other a quill is formed and if
each edge is rolled independently into a quill, the piece is a double quill, when quill is
packed one inside the other it’s called compound quills. Inbark, the dark patches often
found on the outer surface are known as rhytidoma, which is formed of dead phloem
alternating with bands of cork.
Barks. (A) Diagram showing a typical arrangement of tissue: 1, outer surface frequently showing
lichens, lenticels and remains of primary tissues cut off by the bark; 2, cork; 3, cork cambium or
phellogen; 4, phelloderm or secondary cortex, 5, periderm; 6, inner part of primary cortex; 7, groups
of cortical sclerenchyma; 8, endodermis; 9, pericycle; 10, primary phloem; 11, secondary phloem; 12,
cambium; 13, band of lignified fibres; 14, sieve elements; 15, medullary rays. (B-G), shapes of barks:
B, flat; C, curved; D, channeled; E, single quill; F, double quill; G, compound quill.
78

present in powdered bark:
The following elements are
1) Cork;
2) Cork-cambium (phellogen);
3) Phelloderm;
4) Cortex and cellular contents
5) Pericycle and stone cells or fibers;
6) Phloem, phloem fibers;
7) Medullary rays; and
8) Crystals if any.
Secretory tissues:
These are either internal which including secretory cells, cavities or sacs (glands),
secretory ducts or canals, and laticiferous structure or external glandular hairs.
a. Secretory cells:
They occur either isolated or rarely arranged in rows. They frequently possess
suberised walls (precipitation of polysaccharide suberin on the walls). The cells are
named after the secretion of their content; the secretion may be volatile oil, resin,
tannin, mucilage or enzyme.
b. Secretory cavities or sacs (internal glands):
- These are special internal structures embedded in a mass of tissue and having cavities
within them in which secretions; volatile oil, resin, oleoresin, etc. are produced or
secreted.
c. Secretory ducts or canal:
These are tube like structure containing and producing secretion. They usually produce
volatile oil.
d. Laticiferous structures:
These include cells and vessels which are in most cases in form of tubes. The content
or secretion of these elements is called latex ( ‫هتراصع‬
‫وأ‬
‫رجشال‬
‫بل‬‫ن‬ ). It may contain resin, gum-
resin, fat, wax suspended in an aqueous solution of certain substances e.g. alkaloidal
salt, tannins, protein sugar, etc. Laticiferous structures are most commonly found in
the phloem.
79

Cinchona
Botanical source:
Dried bark of Cinchona species viz. C. calisaya, C. ledgeriana, C. officinalis, C.
succirubra or hybrids of either of the last two species with either of the first two species
(Family Rubiaceae).
Morphology:
I)Flat pieces or quills; 2) Outer surface grayish green and rough due to presence of
cracks, wrinkles and often bears epiphytes such as lichens; 3) Inner surface reddish
brown; 4) Fracture short in the cork and cortex, fibrous in the phloem; and 5)Odorless
with intensely bitter taste.
Histology:
1) Cork with an occasional presence of lichens;
2) Parenchymatous cortex containing starch grains and microcrystals of calcium
oxalate;
3) Secretion canals or tannin tubes;
4) Phloem fibers with Y-shaped pits, either isolated or in radial rows of 2 to 4.
5) Sieve tubes with companion cells;
6) Phloem parenchyma;
7) Medullary ray, 1-3 seriate; and
8) Stone cells are absent.
Characteristics of powder:
1) Cork cells;
2) Lignified fibers with Y-shaped pits;
3) Parenchyma
4) Microsphenoidal calcium oxalate crystals; and
5) Starch grains.
Quinoline alkaloids: quinine, quinidine, cinchonine and cinchonidine.
Uses: Antimalarial and bitter tonic.
80

T.S. of Cinchona bark
Bark of Cinchona scuccirubra. A, transverse section; B, radial longitudinal section; both x75;
c.c., companion cell; ck, cork; cort, cortex; cr, microcrystal of calcium oxalate; f, fibre; li, lichens;
m.r., medullary rays; par, phloem parenchyma; s.c., secretion canal; s.t., sieve tube.
Cinchona bark. A, specimen of C. succirubra (x0.5); B, transverse section of bark (x25); C, isolated
phloem fibres (x50); D, portion of phloem fibre with surrounding parenchyma; E, cork cells in surface
view; F, idioblast with calcium oxalate; G, starch (all x200). ck, cork; ct, cortex; f, fibres protruding from
fracture; id, idioblast; l, lichen patches; lf, longitudinal fissure; m.r, medullary ray; pd, phelloderm;pg,
phellogen; p.f, phloem fibres; s.c, secretory cell; t.f, transverse fissure.
82

Cinchona bark (microscopical characteristics of powder):
1. Part of single fiber
2. Part of groups of fibers and phloem parenchyma with overlying medullary ray (m.r.) in radial
longitudinal section.
3. Parenchymatous cells containing starch granules and brown pigment.
4. Part of fiber with phloem parenchyma, one cell containing calcium oxalate micro-crystals (cr.).
5. Cork and phelloderm in sectional view.
6. Phloem parenchyma and part of medullary ray (m.r.) in tangential longitudinal section.
7. Starch granules.
8. Cork in surface view
9. Phloem parenchyma with pits (pt.)
83

Cinnamon
Botanical source: Dried inner bark of Cinnamomum zeylanicum Nees (Family
Lauraceae).
Morphology:
1) Long, flexible slender sticks consisting of either channelled pieces or single quills;
2) Pale brown in color, inner surface darker than the outer surface; 3) Aromatic odor;
4) Warm sweet taste; and 5) Fracture short and splintery.
Histology: A transverse section shows:
1) Sclereids with horse-shoe shaped thickening and containing starch grains;
2) Pericyclic fibers, lignified. in groups of 6 to 15;
3) Sieve tubes;
4) Lignified phloem fibers, single or in group of 2 to 4;
5) Medullary rays usually biseriate;
6) Idioblasts containing volatile oil and mucilage; and
7) Starch grains and calcium oxalate crystals in the medullary rays and parenchyma.
1) Slender fibers; 2) Stone cells with horse-shoe shaped thickening;
3) Starch grains; 4) Parenchyma; and 5) Acicular crystals of calcium oxalate.
Volatile oil; cinnamic aldehyde (or cinnamaldehyde).
2- Chloroform extract of the drug treated with 10% aqueous solution of
phenylhydrazine hydrochloride shows red shaped crystals of hydrazone of
cinnamaldehyde.
Uses:
Carminative, stomachic.
84

Cinnamon Bark and Plant
Cinnamon bark. A, Compound double quill (x0.5); B, transverse section (x50); C, elements of
the powder (x200); ck, cork cells; cr, acicular crystals of calcium oxalate; lf, laminated fracture of
compound quill; m.r, medullary ray; o.c, oil cells; o.t, remains of outer tissues; p.f, pericyclic
fibres; ph.f, phloem fibres; r.ck, residual patches of cork; s, scar of twig; sc, sclereids; sc.l, sclereid
layer of pericycle; s.q, transverse surface of compound quill; st, starch granules.
85

Cinnamon bark (microscopical characteristics of powder):
1. Fibers: 1a, Part of a small group of fibers.
2. Sclereids.
3. Starch granules.
4. Cork in surface view.
5. Phloem parenchyma and an oil cell (o.c.)
6. Part of medullary ray with some of cells containing acicular crystals of calcium oxalate (cr.),
and associated phloem parenchyma in tangential longitudinal section.
7. Calcium oxalate crystals.
8. Part of a fiber with an associated oil cell and phloem parenchyma.
9. Part of a group of fibers and sclereids from pericycle.
10. A single oil cell.
11. Part of the cork and cortex in sectional view.
86

Cinnamon and cassia barks. A, compound quill of cinnamon x ½; B, quill of cassia
bark x ½; C, diagram of a transverse section of cinnamon x 25; D, transverse section
of cinnamon x 200. a, starch b, outer layers of bark (cork and cortex); c, exposed
surface of phloem; cic, scar left by branch; cr, acicularcrystals; ct, remains of
cortex; f, phloem fibres; m.r., medullary ray; ob, ceratenchyma; o.c., oil cell; p.f.,
pericyclic fiber; ph, phloem; p.p., phloem parenchyma; p.scl., pericyclic
sclerenchyma; s.t., sieve-tubes.
 macroscopical identification:
 Examine a small amount of (Cinnamon and Cinchona barks), notice
the organoleptic characteristics (taste,colour,odor and texture).
 Prepare amount of Cinnamon powder using chloral hydrate
one minute add one drop of HCl con (inside fume hood) , put the
cover slide and examine under microscope the key elements. Draw
and label
 Repeat the previous steps for Cinchona.
Expected key elements:
87

Macro and microscopical identification of bark
Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
89

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
90

Lab. (7)
Macro and microscopical analysis of Subteranean organs
The underground organs of the plant are either of stem origin (rhizomes, corms, bulbs,
and tubers) or of root origin (roots and tubers). Those used in medicine serve as storage
organs and generally full of reserve food materials.
Many of the drugs of this class consists of portions of both stem and root origins.
The Root
The root is that part of the plant axis that grows into or towards the soil away from
light, usually bearing no buds and leaves. It does not show nodes and internodes, and
also absence of chlorophyll. The root bears only one kind of lateral branches which are
similar in construction and in general characters to the main or parent root. The root
absorbs water and soluble substances from the soil and conveys them to thestem.
It anchors the plant to the ground.
Types of roots:
They may be:
1 Primary roots: developing into tap-roots.
2 Adventitious roots: arising neither from the main root nor from its branches. They
may arise from the stem.
3 Prop roots: adventitious roots growing from the stem, a short distance above the soil
and extending diagonally into the ground, serving as support to stem as in Maize.
4 Secondary roots or lateral roots.
5 Fibrous roots: very slender roots as in grasses.
6 Aerial roots: adventitious roots, which are either hanging in the air or growing down
to anchor in the ground as in Ficus bengalensis. They may function for climbing, or
for absorption of water from the air.
7 Respiratory roots : growing from soil into air for respiration as in Avicennia.
8 Storage roots: when the roots become swollen with reserve food.
91

Structure of roots:
The structure of the young root shows the epidermis of a single layer, the outermost
layer of the cortex. The endodermis, which is formed of a single layer of smaller
cells, the stele, is central. The pericycle is formed of a continuous unbroken ring of a
single or of many layers of cells. The vascular bundles are of the radial type formed
of alternating groups of‟ xylem and phloem separated from each other by one or more
layers of cells as shown. The typical old dicotyledonous root shows the following
tissues: cork, secondary cortex, or phelloderm, primary phloem, secondary phloem,
cambium, secondary xylem and primary xylem. The medullary rays are present and
pith is commonly absent except in some cases.
92

Glycyrrhiza (Liquorice)
Botanical source: Dried stolons and roots of various species of Glycyrrhiza (G.
glabra, G. violaceae, G. glandulifera) (Family: Leguminosae: or Fabaceae).
Morphology:
1)Spanish liquorice: peeled and unpeeled stolons and roots, 1-2 cm in diameter about
20 cm long.
2) Peeled pieces angular, unpeeled straight unbranches, cylindrical.
3) Outer surface dark reddish brown, longitudinally.
4) Stolons bear buds, scaly leaves and scars of lateral roots.
5) Fracture fibrous in the bark, splintery in the wood.
6) Taste sweet, odor faint and characteristic.
7) Peeled liquorice yellow in color with fibrous exterior.
Histology:
(I) A transverse section of stolon shows:
1) Cork: radially arranged, thin walled, polygonal, tabular cells.
2) Cortex: parenchymatous.
3) Pericycle with small groups of pericyclic fibers at intervals.
4) Phloem consisting of fibers, slightly lignified alternating with sieve tissue.
5) Ceratenchyma collapsed sieve tissue on the outer side of the phloem.
6) Xylem or xylem fibers, vessels, little xylem parenchyma.
7) Medullary rays- cellulosic parenchyma radially elongated cells.
8) Prisms of calcium oxalate in parenchyma (10-15-25-35 m ).
9) Starch grains in the rest of the parenchymatous cells, rounded, 2-10-20 Mm in size.
10) Pith- parenchymatous.
11) Xylem and phloem fibers in groups of 10-50.
(II) A transverse section of root shows:
1)4 small primary xylem bundles, arranged at right angles to each other, protoxylem
directing outward.
2)Phelloderm in the outer part, below the cork. Rest of the structure, same as that of
the stolon.
1) Xylem vessels: large with numerous bordered pits, 2) Xylem parenchyma, 3)
Fibers, yellow colored, in bundles, 4) Calcium oxalate crystal sheath in parenchyma,
5) Starch grains.
Important constituents: Saponin glycosides: glycyrrhizin
Uses: Mild expectorant, sweetening agent, anti—peptic ulcer.
93

Figure 1. Coarsly grated radix (root) of Glycyrrhiza glabra (Liquorice). Also
called Liquiritiaeradix according to the European Pharmacopoeia
96

Microscopic view of the root of a buttercup (Ranunculus) showing the central stele and
4-pronged xylem. The large, water-conducting cells in the xylem are vessels. [Magnified
Approximately 400X.]
Group of fibers accompanied with
parenchyma containing prisms of
calcium oxalate
Fragments of Vessels with
bordered pits
97

Ginger
Botanical source: Dried rhizomes of Zingiber officinale (Family: Zingiberaceae).
Morphology:
1)Laterally flattened pieces with flattened oblique ovate branches or fingers on the
upper side.
2) Each branch 1-3 cm long, depression of stem scar at the apex.
3) Branching sympodial, horizontal rhizome.
4) Longitudinally striated outer surface with occasional fibers.
5) Fracture short and fibrous.
6) Buff colored.
7) Agreeable aromatic odor and agreeable pungent taste.
Histology:
1)Cork- outer zone of irregularly arranged cells and inner zone of radially arranged
cells (absent in Jamaica ginger).
2) Cortex- thin walled, cellulosic, rounded parenchyma with intercellular spaces.
3) Starch grains in parenchyma of cortex.
4) Closed collateral fibro-vascular bundles in the cortex.
5) Brown oleoresin cells.
6)Stele-a ring of vascular bundles (without fibers) just below the endodermis, ground
tissue of parenchymatous cells with fibrovascular bundles, oleo—resin cells andstarch.
7) Xylem vessels- annular, spiral or reticulate thickenings Lignified.
8) Fibers- thin walled with only central lumen lignified with pectosic transverse septa.
I) Cork; 2) Xylem vessels with spiral, annular or reticulate unlignified thickenings; 3)
Simple ovate or sac-shaped starch grains; 4) Xylem fibers-septate, unlignified.
Important constituents: Oleo-resin: gingerol, zingiberene.
Uses: Carminative, stimulant.
98

 Examine a small amount of (Liquorice root & Ginger rhizome),
notice the organoleptic characteristics (taste,colour,odor and texture).
 Prepare amount of Liquorice powder using chloral hydrate
one minute add one drop of HCl con (inside fume hood) , put the
cover slide and examine under microscope the key elements. Draw
and label
Repeat the previous steps for Ginger without using (phloroglucinol+HCl con).
101

Expected key elements (400X):
102

Macro and microscopical identification of subterranean organs
Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
103

Common name: ………………………………….
Family:…………………………………….
……………………………………………………………………….
……………………………………………………………………….
104

Experiment (8)
Detection of Anthraquinone Glycosides in Plant Extract
Objectives:
1. To extract and detect free anthraquinone and glycosidic anthraquinone.
2.To hydrolyze both O- and C- glycoside and to detect the presence of free aglycones by
chemical and chromatographic method.
Natural sources:
Powdered Rhubarb.
Powdered Aloe.
Powdered Senna.
Pharmaceutical Preparation.
Principle:
Anthraquinones are phenolic compounds naturally occurring in free form, C-glycoside and
O-glycoside. The principle of the test is based on the ability of free anthraquinone to form a
colored adduct upon the addition of standard alkali (e.g. KOH, NaOH or ammonia). The
reaction involves the formation of phenolate-type ions which are visibly colored. This fact is
used to distinguish between the O- and the C-glycoside. The O-glycoside is hydrolyzed to
free anthraquinone by heating with dilute HCl acid, while the C- glycoside releases the free
anthraquinone only after oxidative cleavage as illustrated in figure 1 and 2. Anthraquinone
C-glycoside and dimeric anthraquinone are hydrolyzed using oxidative cleavage in the
presence of FeCL3 in acidic medium releasing free anthraquinone (or rhein in the case of
anthraquinone dimmers) and the sugar moiety.
Identification of Anthraquinone Glycosides from Plant Extract
Procedure:
Prepare and outline the TLC plate before you start the practical work.
I. Extraction of the free anthraquinone:
1. In a test tube add 10 ml Petroleum ether to 2 gm of powdered drug.
2.Shake for 10 minutes and filter through a filter paper into a test tube and keep marc on the
filter paper, spot the filtrate on TLC plate.
3. Add 5 ml of the standard alkali (KOH or 10% ammonia).
4.Observe and record the colour which develops immediately or upon standing for a few
minutes. 38
105

II. Extraction of the anthraquinone glycoside:
5.Transfer the marc on the filter paper in step 2 into conical flask or a beaker and add to it
(20ml) of 50% ethanol.
6. Boil for 5 minutes on a water bath.
7.Filter while warm through cotton wool into graduated beaker, spot the filtrate into the
TLC plate then wash with hot alcohol to adjust the volume to 20 ml.
III. Testing for anthraquinone O- glycoside
8.Transfer 10 ml of the glycosidic extract powder produced in step 7 into a conical flask or a
beaker and add to it 10 ml of 25% HCl.
9. Boil for 15 minutes over a boiling water bath.
10. Cool the solution and transfer into a separating funnel.
11. Shake the solution with 10 ml petroleum ether in the separating funnel.
12.Separate the organic layer into a test tube, spot the petroleum ether extract onto the TLC
plate then shake the organic layer with 5 ml of the standard alkali.
13. Observe and record the colour produced on standing for few minutes and observe any
change in colour
IV. Testing for the anthraquinone C- Glycosides:
14.Transfer 10 ml of the glycoside extract produced in step 7 into a conical flask and adds 1
gm of FeCl3 and heat for 20 minute on a boiling water bath.
15. Cool the solution down and then transfer into a separatory funnel.
16.Extract the solution with 10 ml chloroform and separate the aqueous layer from the
chloroform layer.
17. Wash the organic layer with water and transfer the chloroform layer into a test tube, spot
the organic layer onto the TLC plate.
18.Add 5 ml of the standard alkali then observe and record the colour formed immediately
and on standing for a few minutes.
V. Thin layer chromatography:
19. Develop the TLC on the mobile phase
20. Examine the plate under day and both long and short UV lights
21. Spray the plate with alcoholic KOH spraying reagent.
22.Heat the plate for 10 minutes to intensify the colors and examine the chromatogram
under the day and UV lights.
23. Record your findings.
106

Experiment (9)
Extraction and identification of saponin and tannins
 Tannins
Definition: natural compounds nave high molecular weight and contain large number of
phenolic hydroxyl or other groups that enable them to form cross-link to form
proteins of the animal hides and prevent their putrefaction and convert them to leather.
 Classification
1.The hydrolysable tannins: may hydrolyzed by acids or enzyme. They also called
pyrogallol tannins, with ferric chloride will give bluish-black color or ppt
Examples: Rhubarb and Qurecus infectoria.
2. The condensed tannins: include all other tannin their molecules are resistant to
breakage than hydrolysable tannins. They also called catechol tannins. With ferric chloride
will give greenish-black color or ppt.
Examples: cinnamon. and tea leaves.
 Uses
a)Pharmaceutical tannins used in many preparations as they have astringent and antiseptic
properties.
b) Commercial tannins used in leather industry.
 Practical work:
1. Evaporate equivalent 10 g of plant extract to dryness on steam bath
2. Add 15 ml hot water to the residue and stir with glass rod, and then cool at room
temperature.
3. Add 5 drops of 10% NaOH solution to salt out non tannin, to eliminate false
result
4. Filter the resulting solution, divide the filtrate into 4 test tubes:
A. Add 1 ml gelatin solution
B. Add 5 drops gelatin solution + 5 drops 10% NaCl solution
C. Add 5 drops FeCl3 reagent
D. Control
107

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