Transcript 3- Authentication of plant material

Plant extraction
Introduction
The use of plant-derived medicinal dates back many
centuries although it is still under estimation in modern
medicine.
 Plants remain the most important source of natural
drugs.
- More than 30% of prescription drugs are natural
products.
- More than 60% of anticancer and anti-infective drugs
are natural products.
The main sources of drugs are as follows:
1- Natural substances:
From plants, microorganisms, animals,- etc.
(totally obtained from nature).
2- Semisynthetic substances:
These are drugs that are manufactured by partial synthesis.
3- Synthetic substances:
These are drugs which are manufactured by total synthesis
To carry phytochemical screening the following points
must be fulfilled:
1- Selection of promising plant materials.
2- Proper collection of selected plants.
3- Authentication of plant material.
4- Drying of plant materials.
5- Grinding of the dried plants.
6- Garbling of the dried plants
7- Packing, storage and preservation
8- Extraction and fractionation of constituents.
9- Methods of separation and purification.
10- Methods of identification of isolated compounds
(structure elucidation e.g. UV, IR, MS, H-NMR and C-NMR).
1. Selection of promising plant materials:
Before investing time, effort and money in phytochemical
screening it is very important to select a promising plant.
The choice of promising plant depends upon the following:
1- A plant which have a biological activity.
2- A plant used in folk medicine.
3- A plant which show a particular toxicities.
2. Proper collection of selected plants
Drug may be collected from:
1- Wild plants.
Wild plant
2- Cultivated plants.
Cultivated plant
Disadvantage
Advantage
1- Scattered in large or
unlimited area
Present in limited area.
2- Difficult to reach
Easy to reach
3- The collector must be highly
skilled botanists
The collector must not be skillful
person
4- Deficiency may occur due to
continuous collection
Continuous supply
The following precautions should be considered during
stage of collection:
a- The proper time of the day, time of the year and maturity
stage of collection is particularly important because the
nature and quantity of constituents may vary greatly in some
species according to the season and time of collection.
b- The collected plant should be free from any contamination.
c- Collecting plants which are free from diseases (i.e. which
are not affected by viral, bacterial, fungal infection).
3- Authentication of plant material
This may be confirmed by:
1- Establishing the identity by a taxonomy experts.
2- Collection of a common species in their expected habitat by
a field botanist.
3- By comparing the collecting plant with a voucher specimen
( herbarium sheet)
4- Drying of plant materials
Aim of drying:
1- Ease of transport.
2- Ease of grinding
3- Inhibit the growth of microorganisms.
4- Preservative of active constituents.
Drying is done in:
-Shade and in sunlight (Natural drying).
- Hot air drying or by freeze-drying (Artificial drying).
Changes may occur during the drying:
1- Size and weight
2- Shape and appearance
3- Color
4- Odor
5- Taste
6- Active constituent
5- Grinding of the dried plants
6- Garbling of the dried plants
7- Packing, storage and preservation
8- Extraction and fractionation of constituents
There is no general (universal) method for the extraction
of plant materials.
The precise mode of extraction depends on:
1- The texture of the plant material.
2- The water content of the plant material.
3- The type of substances to be extracted or nature of
active constituents.
Extraction: is the separation of medicinally active portion
of plants or animal tissues through the use of
selective solvent and suitable methods extraction.
The principal methods of extraction are:
1- Maceration
2- Percolation
3- Infusion
4- Decoction
5- Digestion
6- Continuous hot extraction technique
(Soxhlet extraction process).
7- Liquid-liquid extraction
8- Distillation
Continuous hot extraction technique
(Soxhlet extraction process)
Reflux is the process of boiling reactants while continually cooling the vapor
returning it back to the flask as a liquid. It is used to heat a mixture for extended
periods and at certain temperatures.. A condenser is attached to the boiling flask,
and cooling water is circulated to condense escaping vapors. One should always
use a boiling stone or a magnetic stirrer to keep the boiling solution from "bumping ".
The temperature of a reaction in a refluxing mixture will be approximately the boiling
point of the solvent used for the reaction.
Sonicator
Sonication is the process of converting an electrical signal into a physical
vibration that can be directed toward a substance. Sonicators are vital lab
equipment and are used for a number of purposes. Sonication is usually
performed to break apart compounds or cells for further examination. The
vibration has a very powerful effect on solutions, causing their molecules to
break apart and cells to rupture. A prime example is in DNA testing, where
the cells that may contain DNA information are subjected to sonication to
break them apart and release the DNA proteins so they can be tested.
Rotary vacuum evaporator

Dimethyl sulfoxide boils
at 189 °C at
atmospheric pressure.
Under vacuum, it
distills at 70 °C
Choice of solvent
As a general empirical rule:
 Non polar solvents (petroleum ether and hexane) will dissolve
non-polar compounds (fats and waxes).
While polar solvents (methanol, ethanol and water)
dissolve polar compound (alkaloid salts and sugars).
(that mean like dissolve like)
 The affinity of the solute for the organic phase may be
greatly increased by using mixture of solvents instead
of single ones (sometimes used mixtures of solvent to
increase the solubility).

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Should readily dissolve substance to be
extracted .
Should not react with the substance to be
extracted .
Should not react with or be miscible with water
(the usual second solvent (.
Should have a low boiling point so it can be
easily removed from the product .
Common extraction solvents are diethyl ether
and methylene chloride .
Phytopharmacological screening:
● Antimicrobial activity
Cinnamomum verum ‫القرفة‬
Thymus vulgaris ‫الزعتر‬
Lavendula officinalis ‫الخزامى‬
(Eugenol)
(Thymol)
(Linalool)
● Antineoplastic activity
Catharanthus roseus
Taxus brevifolia
(Vincrestine, vinblastin)
(Taxol)
● Antimalarial:
Cinchona succirubra
Artemisia annua
(Quinine)
(Artemisinin)
Phytopharmacological screening:
● Hypoglycemic:
Garlic (Allium sativum)
(Allicin)
● Cardiotonic
Digitalis purpurea
Strophanthus kombe
(Digoxin and Digitoxin)
(K-strophanthoside)
● Antiarrythemic
Cinchona succirubra
(Quinidine)

Removal of water( :Although the criteria states
that the organic solvent chosen should not be
miscible with water, some solvents dissolve a small
amount. Thus, water must be removed before
separating the organic product from the organic
solvent or else the product will be contaminated
with water. A drying agent must be used. There are
a number of drying agents available to the organic
chemist: we will be using sodium sulfate and
magnesium sulfate. Placing the organic solvent
containing the dissolved product in contact with the
chosen drying agent will allow the agent to absorb
any dissolved water. The agent can then be
removed from the solvent and then the product can
be isolated .
Kirby-Bauer antibiotic testing ) KB testing or
disk diffusion antibiotic sensitivity testing(
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
is a test which uses antibiotic-impregnated wafers to test
whether particular bacteria are susceptible to specific
antibiotics. A known quantity of bacteria are grown on
agar plates in the presence of thin wafers containing
relevant antibiotics. If the bacteria are susceptible to a
particular antibiotic, an area of clearing surrounds the
wafer where bacteria are not capable of growing (called
a zone of inhibition.)
This along with the rate of antibiotic diffusion are used to
estimate the bacteria's sensitivity to that particular
antibiotic. In general, larger zones correlate with smaller
minimum inhibitory concentration( MIC) of antibiotic for
that bacteria. This information can be used to choose
appropriate antibiotics to combat a particular infection.
Minimum inhibitory concentration
(MICs) are defined as the lowest
concentration of an antimicrobial that will
inhibit the visible growth of a
microorganism after overnight incubation
 and minimum bactericidal concentrations
(MBCs) as the lowest concentration of
antimicrobial that will prevent the growth of
an organism after subculture on to
antibiotic-free media.

Kirby-Bauer antibiotic testing
Disk-diffusion method for determing
the activity of antimicrobials
well diffusion method
Mechanisms of Antimicrobial
Drugs