03) Galenical Preperation

GALENICALS
Sometimes single ingredient is extracted from the crude drugs. When all possible
ingredients present in Crude drug are required then the extraction may be done in the
form of dry extract, viscous soft extract, infusion, conc. Infusions or tinctures. These
crude extracts are known as GALENICALS.
EXTRACTION
Separation of soluble active principles from the crude drugs, plants or animal
origin, using appropriate solvents.
Types of Extraction
I- Solid-Liquid Extraction
a) Leaching Process
 Maceration
 Percolation
b) Digestion
c) Decoction
d) Infusion
II- Liquid-Liquid Extraction
a) Counter-current Extraction
Mostly synthetic drugs have replaced natural drugs. Some drugs belonging
to Alkaloids, glycosides, Fixed Oils, Volatile Oils, proteins and polypeptides
are still used and obtained by extraction.
Need for the separation of active principles:
 Potency of the drug may be known and is more rapidly controlled.
 Deterioration by enzyme is diminished.
 Drug can easily be formulated into required dosage form
 Dosage form is more stable, elegant and palatable.
 Tablets cannot be manufactured from crude drugs.
 Injectable from crude drugs are undesirable rather very dangerous.
 Bulk (volume/amount) smaller bulk can easily be controlled/shipped and so
reduces the cost of the drug.
THEORY OF EXTRACTION OF DRUGS
 Suitable size reduction of the drug
 Penetration of the solvent into the drug
 Drug in dry form is porous due to shrinkage and the pores contain air that is
displaced with solvent causing swelling of drug.
 Solution of constituents
 Escape of solution from the cells
 Dissolved material reaching the surface of the particle passes through
the boundary wall. So suitable concentration gradient, thickness of the
boundary layer and diffusion coefficient plays an important role.
 Separation of solution and exhausted drug
 Solution is subjected to pressure to expel as much as of the solution
possible.

Leaching Process
The process of extracting soluble constituents from a solid by solvent is
commonly known as leaching process.

Factors affecting Leaching Process

a) Pretreatment of raw material and mechanism of Leaching;
Location of active principle: if in epidermis then only bruising is required.
Nature of texture: soft, hard, resins, oleo gum etc. needs different
treatment.
Particle size of drug: Swelling capability: by moistening/imbibing some
drugs are less swelling than others.
Mechanism of leaching:
Diffusion: mostly the active principles are extracted by diffusion.
Capillary action: it takes place e.g. during shaking of soybean oil extraction.
Capillary action depends upon surface tension, density and viscosity of the
solvent. Hydrostatic pressure and particle size and particle cohesion also
affects it.
Permeability of the cell wall:
Ultrasonic treatment: Decreases the interfacial tension and increase the
movement of active principles. Increases interfacial area by decreasing
particle size Increases rate of dispersion of active principle away from
interface to the bulk of the solvent.
b) Nature of solvent:
Ideally it should be selective, to avoid undesirable extraction but not
possible and to avoid gum material enzymatic deterioration in aqueous
extract, Prevented to some extent by adding 25% alcohol microbial growth
in aqueous extract, Add alcohol or chloroform water
c) Particle size distribution:
It should be narrow to have high porosity so that solvent may ingress and
pass through the packed crude drug. Smaller particle have large SA.
d) Temperature:
High temperature increases solubility of the active ingredients but can only be used
in the extraction of thermo-stable ingredients and it works through increased rate
of diffusion better solubility of the active principles by the loss of cell wall and cell
membrane. High temperature cannot be used for thermo-labile ingredients or
volatile oils.
e) Physical forces:
Viscosity, surface tension, adhesion, friction, granulation etc.
Some important points:
 Hot solvent is used both in maceration and percolation.
 Selection of solvent should be done carefully to have maximum
extraction avoid undue destruction of active principles,
 Use of hot solvent for longer period of time because extraction of certain
drugs like fixed oils and alkaloids etc. requires this treatment due to less
penetration of solvent into the cellular structure. Poor solubility of active
principles in solvents like chloroform, ethanol etc.
Infusion
Definition:
Infusions are dilute solutions containing the readily soluble constituents of
crude drugs.
Process:
It is a process of steeping or drenching a drug in water so as to extract the
active principles.
It involves pouring water over the drugs and then allowing it to keep in
contact with water for the stated period (15 min) with occasional stirring
and finally filtering off the liquid. Fresh infusion is prepared by macerating
the drug for a short period of time with cold or hot (boiling) water.
Concentrated infusions are prepared by a modified percolation or
maceration process.
Infusion pot:
It consists of covered jar (made of earthenware, stainless-steel, ceramic,
glass, porcelain) to which is fitted at certain height a perforated tray upon
which the crude drug may be allowed to rest in water being poured over it.
The drug may be enclosed loosely in a small muslin bag and suspended in
the jar at a height where it will be just covered by the liquid. The perforated
tray or muslin bag confers two advantages:
 A-Complete extraction because when the menstruum (solvent)
surrounding the drug becomes saturated, it will sinks to the bottom due
to its increased density and another amount of fresh menstruum
displace it leading to circulatory diffusion
 B- At the end of infusion time, the drug can be lifted out, leaving clear
liquid which can be strained quickly.

Fresh (Dilute) Infusion

A fresh infusion is an aqueous solution of active constituents of a vegetable
drug prepared by the process of infusion. Water is used as menstruum.
Fresh infusion should be used within 12 hours after its preparation because
it gets spoiled due to bacterial and fungal growth e.g. Fresh infusion of
Quassia.
Concentrated (stock) infusion
Prepared by double or triple maceration 8 times stronger than fresh
Alcohol in the concentration of 20-25% is used as menstruum.
Avoid rapid decomposition to which the fresh (dilute) infusions are
subjected. The dilution of 1 volume of concentrated infusion with 7 volumes
of water resembles corresponding fresh infusion in potency e.g.
Concentrated infusion of Quassia.

Decoction
Process:
In this process, the crude drug is boiled in a specified volume of water for a
defined time (10 minutes) it is then cooled and strained or filtered.
This procedure is suitable for extracting water-soluble, heat-stable
constituents and drugs of hard and woody nature. This process is typically
used in preparation of Ayurvedic extracts called quath or kawath.
The starting ratio of crude drug to water is fixed e.g. 1:4 or 1:16. The volume
is then brought down to one-fourth its original volume by boiling during the
extraction procedure .Then, the concentrated extract is filtered and used as
such or processed further
Digestion
Process: In this process the plant cells are subjected to the effect of
menstruum at 40 to 60°C (higher temperature than that used for maceration
and lower temperature than that used in infusion and decoction) for longer
period than that for infusion and decoction.
Apparatus: Thermos bottles (for maintenance of a definite temperature
over the prescribed period of time)
Advantage: Increasing the solvent power of extraction by keeping the liquid
hot throughout the period of extraction without causing damage to the drug
constituents.

Extractive preparations
After the separation of the active constituents form the crude drug, the
active principles are ready to use in the form of;
1) Fluid extracts
2) Extracts
3) Tinctures
1) Fluid extracts:
Liquid extracts/ preparations of vegetable drugs containing alcohol as
solvent, preservative or having both of the qualities.
Drugs are prepared by various methods of USP.
2) Extracts:
Concentrated preparations of vegetable or animal origin obtained by
extraction employing suitable menstruum. And the potency of the extract is
maintained later on.
Types of extracts:
There are three major types or forms of the extracts;
I. Semisolid or semiliquid of syrupy consistency e.g. hyoscyamus
extract.
II. Plastic masses known as pilular or solid extract e.g. glycyrrhiza
extract.
III. Dry powder known as powdered extracts e.g. belladonna
extract.
Powdered extracts and pilular extracts can be used interchangeably.
However, powdered extracts are used in solid dosage forms i.e. tablets, capsule,
powders, granules etc.
and the pilular extracts are used in pills dosage forms, ointments and
suppositories
Preparation of extracts
Extracts can be made by various methods but most of the extracts are
prepared mainly by percolation.
Percolate is concentrated by;
Evaporating whole of the drug solution in case of thermostable drug.
Under reduced pressure in case of thermolabile drugs.
Reserved percolation in case of thermolabile preparations.
Continuous extraction both hot and cold.
Extracts made by reserved percolation:
All are liquid preparations prepared by taking alcohol as menstruum.
Products are adjusted to the specified volume, while the unreserved portion
is evaporated under reduced pressure.
Example: Liquid extract of belladonna, Liquid extract of ipecacuanha etc.
Extracts prepared other than reserved percolation:
These are of two types;
1. Unstandardized extracts
2. Standardized extracts
1. Unstandardized extracts:
These are prepared by percolation to exhaustion followed by removal of
solvent to specified extent. Dry and soft extracts may also be prepared.
Evaporation is done under reduced pressure where it is necessary. Aqueous
solvent is usually used.
Example: Liquid extract of senega, Liquid extract of glycyrrhiza.
2. Standardized extract:
After the evaporation of drug by and appropriate method extracts are adjust
to a specific standard. Active principles are adjusted up to a specific
standard. Normally alcohol is used as solvent.
Example: In extract of belladonna the specified percentage of alkaloid is 1%.
Extracts prepared by maceration:
In this process whole drug is macerated in whole of the solvent.
Example: Liquid extract of senna, compound extract of colocynth.

Extracts made my digestion:

Digestion process is rarely exploited for the preparation of the extracts.
Example: Extract of malt, it helps in the digestion of fatty food due to the
presence of enzymes in it.
Extract of animals:
Extraction is accomplished by using appropriate process and solvent.
Example: Extract of ox bile, Extract of Liver.

Packaging and Storage of Extracts:

They must be packaged in;
Wide mouth containers or plastic tubes and closely tight to prevent loss of
moisture which would result in it’s becoming hard and unstable for use.
They must be kept at cool and dry place.

3) Tinctures
Tinctures are alcoholic or hydro alcoholic solutions of vegetable materials
or prepared from chemical substances (e.g. tincture of iodine).
A product is called tincture when it contains 45% v/v or more alcohol.
Tincture ipecacanha is an exemption containing about 23% v/v of alcohol.
Generally a tincture contains alcohol ranging 20% - 90% v/v alcohol. Four
parts by of volumes product represents one part by weight of the drug.
(Strong ginger is an exception having 2 parts to 1 part of the drug.)
Alcohol is used as solvent in the preparation of tinctures because;
 Many of the active principles are readily soluble in it.
 Alcohol prevents decomposition of the products and hence the potency
of the drug remains protected.
 Gums and aluminous matters are precipitated therefore clear solutions
are obtained
Preparation of tinctures:
They are considered stable preparations as they have sufficient amount of
alcohol to resist the microbial growth. Tinctures can be prepared by various
methods;

1. By Dilution/ Simple Solution:

Tinctures can be prepared by the dilution method from the concentrated
extracts. Solute is dissolved in the solvent with agitation, and then filtered
off.
Example: Iodine, Thimerosal, Green soap tinctures.

2. By Maceration:
Tinctures can also be prepared by;
From the maceration of organized drugs e.g. Tincture of Aurantii
From the maceration of unorganized drugs e.g. Tincture of benzoin,
Tincture of myrrh.
3. By percolation:
Tinctures can also be prepared by the percolation of the crude drug in
percolator.
Example: Tincture of Belladonna, Tincture of Digitalis.

Disadvantages:
 Unpleasant tasting
 Physician will prefer single drug instead of preparation from plants
 High alcohol content
Uses of Tinctures:
 Per oral e.g. Tincture of Ayrantii.
 Externally e.g. Tincture of Todine.
 Inhalation e.g. Tincture of bezoin co.
 As flavoring agent e.g. Lemon tincture, compound cardamom
tincture.
Aromatic waters
Aromatic water is also called medicated water, defined in the USP as; clear,
saturated aqueous solutions of volatile oils or other aromatic or volatile
substances.
Concentrated aromatic waters also contain alcohol from 52 – 56 % v/v.
These are to be free from;
Smoke like odor and other odors.
Must have odor and taste similar to those oil or drugs from which they are
prepared.
Preparation of aromatic waters:
There are four important methods which are mainly used;
1. Distillation
2. Solution method
3. Alternative solution method
4. Dilution method
5. Distillation method:
It is the old, the slowest and the most expensive method. Proportion of
drugs in distillate is different in different drugs.
Procedure:
Metallic distillation flask or still of proper size/ volume is used.
Solvent should be in suitable volume.
Heating system should also be appropriate.
Placing of drug in the distillation flask should be adjusted so that;
 Preparation must be free from Empyreuratic (smoke like) or any
foreign odor.
 Partial burning of the drug should be avoided not to develop
carbonized smell.
Filtration for excess of volatile oils through wet filter papers. Moistened oe
wet filter papers provide physical barrier to excess volatile oil not to pass
through.
Example: Stronger rose water is prepared by distillation process.

Note: Redistillation of the distillateis done where the active principle is in

small proportion/ percentage. For example doubled distillated, tirple
distillated or quadruple distillated.
The process is known as Cohobation.
2. Solution method:
Aromatic waters can also be prepared by the repeatedly shaking for 15
minutes. (Certain volume/ amount for e.g. 2ml or 2g of volatile oil/ aromatic
substance with 1000ml of water.)
The resultant solution is set aside at least for 12 hours.
Then filtered off the solution through wet filter paper.
Note:
1. Solid aromatics are comminuted to particles of smaller dimension offering
more surface area for rapid dissolution.
2. Liquid material is shaken well converting into small globules and so
increasing surface area for more contact.
Example: Chloroform water, Camphor water.
3. Alternative solution method:
In this method aromatic water is prepared by thoroughly incorporating about
15g of talc or purified siliceous earth or pulped filter paper into the volatile
oil.
Purified distilled water (1000ml) is added and then whole is agitated for 10
minutes.
Then filtered it off through wet filter paper.
Filtering aid is provided to form a layer on filter paper to facilitate filtration.
Note: these incorporated agents are also called “distributing agents.”
Mechanism of distributing agents:
Aromatic substances adsorb onto the surfaces and distributed thoroughly
offering more surface area to dissolve out and so, accelerating the rate of
solution.
Choice of distributing agents:
An effective distributing agent must possess the following characteristics;
It must be insoluble in water.
It must not react chemically with aromatic solute.
Particle size must be appropriately coarse so that filtration is easy and
facilitated.
Note: magnesium carbonate and calcium phosphate were used as
distributing agents but found incompatible as:-
They are inert but slightly soluble.
They react with aromatic solute to give yellowish tint.
Solution becomes alkaline and reacts with the alkaloid to give ppt.
4. Dilution method:
An alcoholic solution of the essential oil is mixed with water and talc. The
mixture is agitated; after several hours it is filtered. The concentrate contains
between 50 and 55 percent alcohol by volume.
One volume of concentrate is diluted with 39 volumes of water, producing
aromatic water which contains less than 1.5 per cent of alcohol.
Example: Concentrated waters of peppermint, dill, cinnamon, and caraway
etc.

Storage of aromatic waters:

 Aromatic waters are not permanently stable compounds. These must be
stored in containers having these qualities written below;
 Air tight container
 Light resistant container
 Non- alkaline
 Temperature must not be elevated
 Contact of aromatic waters with atmospheric air is prevented otherwise
oxidative decomposition or microbial contamination may occur.
Note: Aromatic waters deteriorate with time and therefore, should be made
in small quantities.
Uses of aromatic waters:
1) Now days they are not frequently used but they can be used as;
Perfuming, flavoring and for special use i.e. camphor water is used as
vehicle in eye preparations. It gives refreshing and stimulating action.
2) Hamamelis (witch hazel) water is employed in rule a perfuming and
astringent in cosmetics (after shave.)
Chloroform water is used as sedative in cough, asthma & colic mixtures. Also
as vehicle for administering other ingredients.

Maceration
It is a process in which properly commuted drug is permitted to soak in the
menstruum and the cellular structure is softened and penetrated by the
menstruum and the soluble constituents are dissolved.
The usual method is to introduce the drug into a bottle with menstruum.
Stopper it and agitate it occasionally for 2 to 14 days. Pour off the liquid,
express the residue to avoid less and filter the mixed liquid.
The drug is macerated in the definite quantity of the menstruum for a
definite period of time.
Types of maceration:
1. Simple maceration (for organized drugs)
2. Maceration with adjustment (for unorganized drugs)
3. Multiple maceration (for concentrated preparations)
4. Circulatory extraction

5. Simple maceration:
It is the most commonly adopted method for the organized drugs. It is also
known as maceration process for tincture from organized drugs. Organized
drugs are those drugs which have specific cell structure like root, stem,
leaves and flowers.
Steps for maceration:
 First of all commute the solid crude drug to appropriate size.
 Take whole of the menstruum. And add crude drug to the menstruum.
 Macerate for 7 days with occasional shaking in a closed vessel.
 Press the marc and combined the filterate and expressed liquid.
 Combined liquid is allowed to stand so that colloidal particles are settled
down. Decant or filter this combined liquid.
The volume is not adjusted in case of organized drugs because when
equilibrium has been attained the potency of the resulting solution will
remain same regardless of the efficiency with which the marc is pressed.
Therefore, these solutions are not adjusted to volume since potency would
then depend on the amount of liquid expressed from the marc e.g. 100g
drug/ 1000ml of menstruum.
Examples: Tincture of lemon, Tincture of orange etc.

2. Maceration with adjustment:

It is maceration for unorganized drugs. Unorganized drugs are those which
have no specific cellular or tissue structure and are obtained from plants as
their exudates e.g. gums, resins, oleaginous resins etc.
Steps involved:
 Take weight amount of drug and powder it.
 Then use 4/5th of the menstruum.
 Macerate for 2 to 7 days. And shake occasionally.
 Perform filtration by filter paper.
 Marc is not pressed.
 Adjust the volume by adding the remaining menstruum through the filter
paper having marc.
Example: Tincture of tolu, tincture of benzoin.

3. Multiple maceration:
Repeated maceration is more important and efficient than simple
maceration because and appropriate amount of active constituents may be
left behind in the first pressing of marc which may be extracted out during
next maceration.
In multiple maceration the total volume of menstruum is divided into parts
and used at different levels instead of whole menstruum. Alcohol is usually
used instead of water because of good penetration due to specific gravity
and moulds can’t grow.
It involves two processes;
a) Double maceration
b) Triple maceration
a) Double maceration:
Maceration of drug is carried out twice and total volume of menstruum is
divided into two portions.
 In first maceration the weighed amount of drug is allowed to remain in
contact with specified amount of menstruum with occasional shaking for
definite period.
 The liquid is then strained and the marc is not pressed.
 In 2nd maceration remaining portion of menstruum is added to the marc and
allowed to stand for certain period.
 The clear liquid is strained; the marc is pressed and combined the strained
and expressed liquid.
 The liquid obtained from 1st and 2nd maceration are combined together, then
filtered and evaporated to get a product of required concentration.
 The equation for double maceration is;

Vol. of menstruum for 2nd maceration = total vol. of menstruum – vol. of

mentruum for 1st maceration
b) Triple maceration:
 Menstruum is divided into three portions and maceration is done thrice.
 Weight amount of the drug is allowed to remain in contact with the one
part of menstruum for specified period of time in a closed container with
occasional shaking. Liquid is strained and reserved.
 Similarly 2nd and 3rd maceration is carried out .
 Marc is pressed at the end of 3rd maceration and expressed liquid is mixed
with liquids obtained from 2nd and 3rd maceration
 The liquid from 1st maceration is concentrated than 2nd and 3rd. So, latter are
evaporated and expressed and mixed with 1st maceration.
 The equations for triple maceration are;

Vol. of menstruum for 3rd maceration = total vol. of menstruum – vol. used in 1 st &
2nd maceration
4. Circulatory Extraction:
The efficiency of extraction in a maceration process can be improved by
arranging for the solvent to be continuously circulated through the drug.
Solvent is pumped from the bottom of the vessel to the inlet where it is
distributed through spray nozzles over the surface of the drug.
The movement of the solvent reduces boundary layers, and the uniform
distribution minimizes local concentration in a shorter time.

Percolation
Definition:
It is process in which crude drug is packed into a column and solvent is
allowed to pass through it till the complete extraction is accomplished tested
by certain tests.
The apparatus used for percolation process is a percolator. For commercial
scale large percolators are also available.
Percolator:
A percolator is a conical vessel having a lid at the top and is provided with a
fake bottom on which cotton wool or filter paper is placed to support the
column of the drug and help in the escape of the percolate. The base of the
percolator is fitted with a tap from which the percolate is collected.
A commercial scale percolator is shown in the diagram having capacity of
about one ton.

Types of percolator:
Selection of the percolator depends upon the nature and cost of the solvent
used for the percolation. The various types of percolators are mentioned
below;
Open percolator
Closed percolator
Steam jacket percolator
Types of percolation
Percolation can be performed by these three methods which are mentioned
below;
1) Simple percolation 2) Reserved percolation
3) Continuous extraction or percolation
1. Simple percolation:
a) Size reduction: It is carried out;
To increase the surface area of the drug substance.
For the uniform packing of the drug.
To slow down the movement of menstruum across the crude drug.
Also, to ensure the complete exudation of the crude drug.
b) Imbibition or moistening of crude drug:
It is done with a suitable solvent and allowed to stand for 4 hours in closed
container.
The drug will swells up and the menstruum will get into the cell wall of the
crude drug.
After a lapse of time the drug is poured through a coarse scene to remove any
lumps and to mix the dry powder.
Preliminary moistening is essential to avoid chocking of percolator, and
washing of fime particles put of percolator.
c) Packing:
After imbibition the imbibe drug is evenly packed into the percolator.
d) Maceration:
After packing of the crude drug sufficient amount of the menstruum is
added to saturate the material and close the percolator with a lid.
Maintain the large of menstrum over it for uniform and continuous
percolation.
Macerate the crude drug for 24 hours.
e) Percolation:
After 24 hours, the tap of the percolator is opened and collected liquid is
allowed to drip down at a controlled speed till 3/4 volume is collected.
More menstruum is added so that marc does not become dry.
Percolate is tested for complete exhaustion of drug.
The marc is pressed and the expressed liquid is allowed to the already
collected percolate.
The liquid is allowed to stand to settle the suspended percolates, then
decanted and clarified.
f) Tests:
Take few ml of last percolate and evaporate to dryness, if no residue remains
then drug is completely exhausted.
Measure the specific gravity of the last few ml of the percolate, if it equals to
the specific gravity of the menstruum used, then the drug is exhausted.
Specific tests may be performed on percolate containing alkaloids,
glycosides, tannins, resins etc.

2. Reserved percolation:
 1st portion (about 3/4 of the final product) of the percolate containing
maximum active principles is reserved.
 Subsequent percolation is completed until drug is exhausted but the last
1/4th of final volume is collected separately.
 2nd dilute part is evaporated to get the syrupy consistency which is then
mixed with reserved portion and final volume is adjusted with
menstruum.
 Reserved percolation is used for preparation of liquid extracts which are
more concentrated preparations as compared to tinctures prepared by
simple percolation process.
 Generally alcohol is used as menstruum.
 Not all percolate is subjected to heat treatment for evaporation.
Examples: liquid extract of liquorice.
3. Continuous extraction:
The process is used:
 When penetration power of menstruum into cellular structure is very low.
 Solute is not readily soluble in the solvent.
 Quantities of menstruum are very less.
 In this process;
 In normal extraction process, concentrated preparations are made
followed by evaporation of solvent.
 Continuous extraction combines two operations i.e. evaporation and
condensation.
 Condensed liquid is returned to the drug to continue extraction.
 So with the help of temperature smaller volumes of solvent may be used
and process is called “continuous extraction.”
Some volume of the solvent is used again and again using special apparatus.
A continuous extraction may be;
1) Continuous hot extraction
2) Continuous cold extraction
In both of the above mentioned extraction process the principle of
‘distillation’ is exploited.
Liquid-liquid extraction (counter current extraction):
It is a process of separating organic substances from aqueous solution e.g.
alkaloids.
Principle: It is based on the principle of partition coefficient of drug between
two solvents.
Important components:
1) Higher is the partition coefficient greater will the extraction of desired
substance.
2) Distribution between the water and organic solvent depends upon the
hydrophilic and hydrophobic group of drugs.
3) If hydrophilic group is ionizable, then pH plays an important role.
4) If the ionization constant is high then separation of isomers can be
accomplished.
5) Decrease in interfacial tension will increase liquid-liquid extraction due to
more interfacial area.
6) Impurities present between the two immiscible liquids may retard as the
migration of drug. (This problem of impurities can be overcome by the
addition of short chain alcohols (C6) which replace the impurities.)
7) Removal of impurities increases separation of drugs.
Applications:
 Separation of ‘Bacitracin’ in butanol from bacterial growth medium.
 Purification of antibiotics.

Example:
Extraction of alkaloids: Counter current extraction is based on the principle of
partition coefficient principle. Let’s understand the phenomenon by the
example by the extraction of an alkaloid. In water chloroform combination,
the drug is partitioned on the same principle.

Chloroform & water solvent:

1) Equal volume of chloroform is added to each three tube.
2) Drug dissolved in water is added to the tube 1.
3) Tube shaken to achieve equilibrium settled, and the upper aqueous layer
to tube 2.
4) Equal volume of aqueous layer added to tube 1 and 2.
5) Tube 1 and 2 shaken and settled down aqueous phase to tube 2 and 3.