KP pharma Corner’s

Practical Manual in Medicinal Chemistry-I

S.Y.B.Pharm./SEM. IV.
BP406P

ii
 PREFACE

The synthesis of pure drug molecules or compounds is important step in the process of
drug discovery. The medicinal chemistry subject is concerned with synthesis of organic
molecules or compounds to be used as pharmaceuticals hence the careful selection of a
particular type of reaction, regents, solvents, catalysts along with optimized reactions
conditions are most important.

The contents of this book are divided into two sections, sections A & B. Sections
A provides the information on various laboratory techniques used for isolation,
purification & synthesis of organic compounds. Sections B give the in details procedure
along with the mechanism of the reactions of the organic compounds.

This has been our first attempt to fulfill the needs of undergraduate students of
medicinal chemistry practical course of reputed universities especially for third year B.
Pharmacy student of Savitribai Phule Pune university semester V.

iii
 SYLLABUS

Medicinal Chemistry-I (Practical)

B. Pharmacy (Sem. – IV)

Sr. Title of Experiment Refer Page

No. No.

01 Purification Techniques of Solvents by Fractional 8

Distillation and Vacuum Distillation.
02 Preparation of Acid/Basic Salts of Drugs and Evaluation 13
of their Physicochemical Properties.(Any Two)
03 Thin Layer Chromatography Technique and Purification 21
of Synthesized Compounds by Column Chromatography.
(Any Two)
04 Synthesis & Purification of following Compounds using 30
Precipitation or Recrystallization.(Any Six)
1. Benzimidazole.
2. 1, 2, 3, 4-tetrahydro Carbazole.
3. 2,3-diphenyl Quinoxaline.
4. Bis-β-napthol.
5. Anthranilic Acid.
6. Sulphanilamide.
7. Benzoic Acid from Benzyl Alcohol.
8. Propranolol.
9. i. 1,4-dihydropyridine.

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 CONTENTS
Sr. No. SECTION-A Page No.
Introduction to Chemical Handling and Safety in the 1
Laboratory
1 Purification Techniques of Solvents by Fractional 8
Distillation and Vacuum Distillation.
2 Preparation of Acid/Basic Salts of Drugs and Evaluation of 13
their Physicochemical Properties.(Benzilic Acid & Sodium
Benzoate)
3 Thin Layer Chromatography Technique (TLC) and 21
Purification of Synthesized Compounds by Column
Chromatography (CC).
A. Amino Acids (TLC)
B. 2, 4 dinitrophenyl hydrazones (TLC)
C. Separation of the mixture of dyes (CC)
D. Separation of 2, 4 dinitrophenyl hydrazones of carbonyl
compounds (CC)
SECTION-B
4 Synthesis & Purification of following Compounds using 30
Precipitation or Recrystallization.
A Synthesis of Benzimidazole. 30
B Synthesis of 1, 2, 3, 4-tetrahydro Carbazole. 35
C Synthesis of 2, 3-diphenyl Quinoxaline. 37
D Synthesis of Bis-β-napthol. 38
E Synthesis of Anthranilic Acid. 40
F Synthesis of Sulphanilamide. 41
G Synthesis of Benzoic Acid from Benzyl Alcohol. 43
H Synthesis of Propranolol. 44
I Synthesis of 1, 4-dihydropyridine. 46
Bibliography 48

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 SECTION - A
INTRODUCTION TO CHEMICAL HANDLING AND SAFETY IN THE
LABORATORY:

1.1 INSTRUCTIONS TO THE CANDIDATE:

1. Before entering in the Laboratory:

➢ One should wear a clean, white Apron.

➢ Ensure that you are with all the necessary writing material, observation book,
calculator, pencil, eraser, requirements box and a neatly covered journal
completed in all aspects.

2. While working in the laboratory:

➢ Maintain discipline and cleanliness. Never lean on the platforms. Follow the
instructions given by teacher scrupulously.

➢ Follow the SOPs correctly while operating the equipments. The log books of
sophisticated equipments should be maintained.

➢ Work cautiously while working with power driven or mobile equipments, gas
burners etc.

➢ Handle acids or other hazardous chemicals carefully. For pipetting such

corrosive chemicals, use rubber bulbs "DO NOT SUCK BY MOUTH"

➢ Do not keep the organic and volatile solvents near the gas flame.

➢ Also replace the lids on the reagent bottles especially volatile chemicals after
use.

➢ Use only electric water bath for warming any organic solvents.
➢ "AVOID DIRECT HEATING ORGANIC CHEMICALS UNLESS
OTHERWISE DIRECTED"

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 A Practical Book of Medicinal Chemistry- I
1.2 FIRST AID TREATMENT IN CASE OF ACCIDENT OR INJURY:

A. BURNS: -

1. Burns caused by Dry Heat (e.g. by Flames, Hot Objects): - For slight burns
in which skin is not burnt, apply burnol. For more severe burns, call for
medical aid.

2. Acid on the Skin: - Wash immediately and thoroughly with liberal quantity
of water, then with saturated sodium bicarbonate solution and finally with
water.

3. Alkali on the Skin: - Wash immediately with a large volume of water, then
with 1% acetic acid, and finally with water.

4. Bromine on the Skin: - (Serious!) Wash the affected part immediately with
cloth/cotton sponge soaked in light petroleum and then rub glycerin well into
the skin. After a little while, remove the superficial glycerin and apply burnol.

5. Sodium on the Skin: - If any small fragment of sodium can be seen, remove
it carefully with forceps. Wash thoroughly with water, then with 1% acetic
acid, finally with water.

6. Organic substance on the skin: - Wash with cotton/cloth soaked in rectified

spirit, then with soap and warm water.

B. CUTS: - If the cut is only a minor one, allow it to bleed for a few seconds; make
sure that no glass particle remains. Apply a disinfectant (Rectified Spirit or Dettol)
and bandage. For serious cuts, send for a doctor at once: meanwhile wash with a
disinfectant and check bleeding by applying pressure immediately above the cut.
Continuous pressure should not be maintained for more than five minutes.

C. EYE ACCIDENTS: -

In all cases, the patient must see a doctor, if the accident appears serious,
medical aid should be summoned immediately while first aid is applied.

1. Acid in the eye,

If the acid is dilute: - Wash the eye repeatedly with 1% sodium bicarbonate
solution in the eyecup.
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 A Practical Book of Medicinal Chemistry- I
If the acid is concentrated: - First wash the eye with a large amount of water
and then continue with the bicarbonate solution.

2. Caustic alkali in the eye: - (Serious!) Proceed as for acid in the eye, but
wash with 1 % boric acid solution in place of bicarbonate solution.

Do not neglect to consult a Physician.

3. Bromine in the Eye: - (Serious!) Wash thoroughly with water and then
immediately with 1% sodium bicarbonate solution.

4. Glass in the Eye: - Remove loose glass very gently with forceps or by
washing with water in an eyecup.

Call the Doctor Immediately.

D. FIRES: -

• In the event of one's clothing catch fire, the victim should roll over on the
ground or should be covered with a fire blanket. Fire extinguisher should not
be directly used on a person.
• Inflammable solvents should be handled carefully.
1. Carbon tetrachloride should not be used if sodium or potassium is present as
violent explosions may result.

2. The laboratory must be ventilated immediately and well.

3. For burning oil or organic solvents, do not use water, as it will spread the fire.

Mixture of sand and sodium bicarbonate is very effective.

E. POISONS: -

Solids or Liquids: -

1. In the mouth but not swallowed: - Spit out at once and wash repeatedly with
water.

2. If swallowed: - Call a doctor immediately. In the meanwhile, give an

antidote according to the nature of the poison.

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 A Practical Book of Medicinal Chemistry- I
a) Acids (including oxalic acid): dilute by drinking water, followed by limewater
or magnesia.
b) Caustic alkalis: dilute by drinking water, followed by vinegar, lemon or orange
juice, or solutions of lactic or citric acid. Milk may then be given but no emetics.
c) Salts of heavy metals: give milk or white of an egg.
d) Arsenic or mercury compound: give an emetic immediately, e.g., one
teaspoonful of mustard, or one teaspoonful of salt or zinc sulphate, in a cup of
warm water.
F. GAS: - Remove the victim to the open air and loosen clothing at neck. To
counteract chlorine or bromine fumes if inhaled in only small amounts, inhale
ammonia vapor or gargle with sodium bicarbonate solution. Afterwards, the
patient should suck eucalyptus oil soaked cotton swabs or drink warm dilute
peppermint or cinnamon essence to smoothen the throat and lungs. If breathing
has stopped, apply artificial respiration. Call for Medical AID Immediately.
1.3 CARE IN CHEMISTRY LABORATORY:
➢ Benches should always be kept clean and tidy. All the spillages of both solids and
liquids must be cleared away immediately.

➢ All glassware must be scrupulously cleaned.

➢ Under no circumstances should be working surface of the bench become cluttered

with apparatus.

➢ All apparatus associated with one particular operation should be grouped together
on the bench.

➢ If a solution, precipitate, filtrate, etc, is set aside for subsequent, the container
must be labeled so that contents can be readily identified.

➢ Regent bottles must be replaced on the regent shelves immediately after use.

➢ Normal practice is that all determinations are performed in duplicate.

➢ A stiff covered notebook of A 4 size must be provided for recording experimental

observations as they made.

➢ Fire extinguisher must be placed handy in every chemical laboratory

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 A Practical Book of Medicinal Chemistry- I
1.4 SAFETY:

Safety in the laboratory is essential at all times. You are responsible for the safety
of any other person as well as your own. Many chemicals encountered in analysis
are poisonous and must be carefully handled. The more precaution is to be taken
for concentrated acids, poisons such as potassium cyanide, halogenated solvents,
benzene, and mercury. Many operations involving chemical reactions are
potentially dangerous and recommended procedures must be followed and
obeyed. All laboratory workers/person should familiarize themselves with local
safety requirements, which may include the compulsory wearing of lab coats and
safety spectacles, and the positions of first aid equipment.

1.5 THE GRADES AND PROPERTIES OF CHEMICALS:

In the analytical chemistry, accuracy of analysis is affected by the principal factor
viz the quality of the regents. The chemicals, which are used in the laboratory for
chemicals analysis, are available indifferent grades as follows.
1. Technical or Commercial Grade:-These are used when the high purity is not
required e.g. preparation of cleaning solution. For this potassium dichromate and
sulphuric acid are required.

2. Chemically pure (CP Grade):- These are more refined than the technical grade.
These chemicals are not suitable for analytical work or if, to be used, they must
be tested.

3. L.R. Grade: - These are used for analytical work. Its label indicates maximum
limits of impurities allowed by the specifications, or actual results of analysis for
various impurities.

4. Primary standard: - These are in the purest form, carefully analyzed and the
assay value (percentage purity) is printed on the label.

5. Pharmacopoeial Grade Chemicals: - These chemicals confirm the tolerances set

by pharmacopoeias. (Ex. Indian pharmacopoeia & British Pharmacopoeia)

1.6 CARE IN HANDILING OF CHEMICALS:

Handling of chemicals specially hazards chemicals must be done with due care.
Everyday working in the laboratory must follow certain rules while handling the

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 A Practical Book of Medicinal Chemistry- I
chemicals. Select the required grade of the chemical for the analytical work.
Select the smallest pack as available. Replace the top of every container
immediately after removal of reagent; do not rely on someone to do this. Use
clean spatula for removing the chemical reagent from the container. Observe the
special instruction if any mentioned on the container. Remove the required
amount of chemical reagent from the container, so there is no need of returning
excess to container, so as to avoid the contamination of product. Keep the reagent
and the laboratory balances clean. Clean any spilled chemicals immediately.
While handling hazardous or toxic chemicals use hand gloves and mouth mask.

1.7 LABELING OF LABORATORY REAGENTS:

Hazard Symbols: Legends on packaging and labeling of dangerous substances define

hazardous chemicals under the following categories:

Corrosive: These products may destroy living tissue; eyes are particularly susceptible,
Emergency showers should be available. If swallowed, plenty of water should be given
after immediate mouth rinsing

Toxic: These products can cause death or serious illness when small amounts enter
the body by ingestion, inhalation of vapor, fumes or dust, or by absorption through
the skin; hygiene considerations should be rigorously observed.

Oxidizing: These compounds may cause fire and will always assist combustion. They
produce heat on contact with organic matter and reducing agents.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)

Explosive: These products may explode by the action of heat, sources of ignition,
shock or friction. The compounds are often packaged wet to reduce the risk of
explosion; they will become dangerous if allowed to dry. Some compounds form
sensitive explosive salts on contact with metals.

Flammable: These compounds have a low flash point, and those which react with water
or damp air to give rise to flammable gases (e.g. hydrogen) from metal hydrides.
Ignition sources include Bunsen burners, hot metal surfaces, electric sparks, etc. Fire
fighting equipments should be readily available and frequently checked.

Harmful: Irritant chemicals cause inflammation of the skin, mucous membranes, or

discomfort of the respiratory system. All laboratory chemicals should be regarded as
harmful; some are specifically harmful by skin contact, inhalation or swallowing.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)

1. PURIFICATION TECHNIQUES OF SOLVENTS BY FRACTIONAL

DISTILLATION AND VACUUM DISTILLATION.

PURIFICATION OF LIQUIDS:-

A liquid may contain both soluble and insoluble impurities. Liquids can' be purified
from insoluble impurities (non-volatile) by the process of distillation. Distillation is a
process in which liquid is converted into its vapour at its boiling point and the vapour is
then recondensed to liquid by cooling. The liquid thus condensed is known as distillate.

Distillation is therefore a combination of evaporation and condensation. If the liquid

contains a soluble impurity which is solid, then simple distillation separates the liquid
from the dissolved solids which are left as residue, e.g sea water or tap water is
evaporated it leaves behind salt residue. If miscible liquids are the impurities, fractional
distillation is done, which removes such impurities, e.g alcohol containing water or
acetone containing water; benzene and acetone etc. Therefore, on distillation the liquid
becomes free from non volatile soluble impurities and also from the impurities which
are less volatile than liquid itself.

DISTILLATION: It is defined as purification technique in which liquid is converted

into vapour and again vapour converted in to liquid.

TYPES OF DISTILLATION:

1. Simple Distillation: -

2. Fractional Distillation:

3. Vaccum Distillation (distillation under reduced pressure): -

4. Steam Distillation:

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
1. Simple Distillation:

This process is used to separate a liquid from solid or non-volatile impurities

present in it. The apparatus used for this purpose is as shown in Fig.01

Fig.No.01 Simple Distillation Apparatus

Fig No.02 Glass Apparatus of Distillation Process.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
2. Fractional Distillation:
It is the separation of a mixture into its component parts, or fractions, such as in
separating chemical compounds by their boiling point by heating them to a
temperature at which several fractions of the compound will evaporate. It is a
special type of distillation. Generally the component parts boil at less than 25°C
from each other under a pressure of one atmosphere. If the difference in boiling
points is greater than 25°C, a simple distillation is used. Fractional distillation is
the separation of a mixture into its component parts, or fractions, such as in
separating chemical compounds by their boiling point by heating them to a
temperature at which several fractions of the compound will evaporate. It is a
special type of distillation. Fractional distillation in a laboratory makes use of
common laboratory glassware, as well as some single-purpose items like a
fractionating column.

Fig.03 Fractional Distillation Apparatus

Discussion:
As an example, consider the distillation of a mixture of ethanol and water. Ethanol
boils at 61-62 °C while water boils at 100 °C. So, by gently heating the mixture, the
most volatile component will concentrate to a greater degree in the vapor leaving
the liquid. Some mixtures form Azeotropic, where the mixture boils at a lower
temperature than either component.
The apparatus (the diagram represents a batch apparatus, as opposed to a continuous
apparatus) is assembled as in the diagram. The mixture is put into the round
bottomed flask along with a few pieces of porcelain chips, and the fractionating

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
column is fitted into the top. As the mixture boils, vapor rises up the column. The
vapor condenses on the glass beads or the column packing material, inside the
column, and runs back down into the liquid below, refluxing distillate. The column
is heated from the bottom. The efficiency in terms of the amount of heating and
time required to get fractionation can be improved by insulating the outside of the
column in an insulator such as wool, aluminium foil or preferably a vacuum jacket.
The hottest tray is at the bottom and the coolest is at the top. At steady state
conditions, the vapor and liquid on each tray are at equilibrium. Only the most
volatile of the vapors stays in gaseous form all the way to the top. The vapor at the
top of the column then passes into the condenser, which cools it down until it
liquefies. The separation is more pure with the addition of more trays (to a practical
limitation of heat, flow, etc.) The condensate that was initially very close to the
azeotrope composition becomes gradually richer in water. The process continues
until all the ethanol boils out of the mixture. This point can be recognized by the
sharp rise in temperature shown on the thermometer.
In laboratory distillation, several types of condensers are commonly found. The
Liebig condenser is simply a straight tube within a water jacket, and is the simplest
(and relatively least expensive) form of condenser. The Graham condenser is a
spiral tube within a water jacket, and the Allihn condenser has a series of large and
small constrictions on the inside tube, each increasing the surface area upon which
the vapor constituents may condense. The apparatus- used for this purpose is as
shown in fig no 01.
Procedure of practical:
Set up the assembly as shown in the figure and perform the fractional distillation,
using the column. Separate the ethanol at 61-62°C collects it in a different flask.
Collect the azeotrope at around 70°C and then finally collect water at 100°C.
3. VACCUM Distillation (Distillation Under Reduced Pressure):
Some liquids like glycerin (B.P-290oC) decompose at their boiling points or even at
temperature below their boiling under atmospheric pressure. That means these
liquids cannot be purified by distillation at atmospheric pressure. The boiling point
of a liquid rises when pressure is increased and it decreases when pressure is
decreased. Hence for purification of these liquids, they are to be distilled under
reduced pressure. It is also called as Vaccum Distillation systems operate at
reduced pressure, thereby lowering the boiling point of the materials. Vacuum
distillation is a method of distillation whereby the pressure above the liquid mixture
to be distilled is reduced to less than its vapor pressure (usually less than

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
atmospheric pressure) causing evaporation of the most volatile liquid(s) (those with
the lowest boiling points). This distillation method works on the principle that
boiling occurs when the vapor pressure of a liquid exceeds the ambient pressure.
Vacuum distillation is used with or without heating the solution
Laboratory-scale vacuum distillation is used when liquids to be distilled have high
atmospheric boiling points or chemical change at temperatures near their
atmospheric boiling points. Temperature sensitive materials (such as beta carotene)
also require vacuum distillation to remove solvents from the mixture without
damaging the product. Another reason vacuum distillation is used is that compared
to Steam distillation there is a lower level of residue build up. This is important in
commercial applications where temperature transfer is produced using heat
exchanger. Vacuum distillation is sometimes referred to as low temperature
distillation. Typical industrial applications utilize the heat pump cycle to maximize
efficiency.
The apparatus used for this purpose is as shown in diagram

Fig. No.04 Vaccum Distillation Apparatus (Distillation under Reduced Pressure)

Procedure of Practical:
Set up the assembly as shown in the diagram. Start heating and circulate water put
some air licks in the round bottom flask, apply vacuum slowly and then start the
distillation. Distill out the given sample.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
2. PREPARATION OF ACIDIC & BASIC SALTS OF DRUGS AND
EVALUATION OF THEIR PHYSICOCHEMICAL PROPERTIES.

Basic Concepts in Salt Formation:

When a compound that is ionized in solution undergoes a strong ionic interaction

with an oppositely charged counter ion, leads to crystallization of the salt form. In
the aqueous or organic phase, the drug and counter ion are ionized according to the
dielectric constant of the liquid medium. The charged groups in the structure of drug
and the counter ion are attracted by an intermolecular columbic force. During
favorable conditions, this force of attraction crystallizes the salt form out of the
medium (Fig.No.05). All acidic and basic compounds can participate in salt
formation. However, the success and stability of salt formation depend upon the
relative strength of acid or base or the acidity or basicity constants of the species
involved .The salt form is separated into individual entities (i.e. the ionized drug and
the counter ion) in liquid medium and its solubility depends upon the solvation
energy in the solvent. The solvent must overcome the crystal lattice energy of the
solid salt and create space for the solute. Thus, the solubility of a salt depends on its
polarity, lipophilicity, ionization potential and molecular size. The solubility of salt
also depends on the properties of solvent and solid such as the crystal packing and
presence of solvates .When weak acids and bases react, the relative strength of the
conjugated acid-base pair in the salt determines the pH of its solutions. The salt, or
its solution, so formed can be acidic, neutral or basic. A salt formed between a
strong acid and a weak base is an acid salt, for example NH4Cl.

Fig.No.05 Basic Concepts in Salt Formation

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
Formation of Salt Form:

Salts can be prepared on a small scale using various methods. Forming salts from
free acid or base is the most common method. The free acid or base of the drug
substance is combined with the counter ion base or acid in specific molar ratios in a
suitable solvent system. The salt form is then recrystallized and isolated under
favorable conditions. A less common method is to form salts through salt exchange.
In this method, a counter ion salt is treated with a free acid or a free base in a
specific molar concentration in a suitable solvent. The solid is then isolated and
recrystallized. For example, the sulfate salt of methyl pyridinium-2-aldoxime can be
prepared using silver sulfate as a counter ion. The unwanted silver ions were
removed as insoluble iodide salt and the desired sulfate salt was precipitated by
adding an antisolvent. A wide range of salts are generally prepared for each new
substance. Their properties are compared during a preformulation program
improving the chances of selecting the optimal salt form. However, a balanced
approach should be adopted because of limited availability of resources at this early
stage of drug development. Commonly used salts such as hydrochloride and sodium
have advantages over other salt forming moieties like low molecular weight and low
toxicity. However, other salt forms such as mesylate may sometimes offer
advantages like higher solubility and bioavailability.

Advantages of Salt Formation:

Salt forms of drugs have a large effect on quality, safety and performance of drugs.
The properties of salt-forming species (i.e. counter ions) significantly affect the
pharmaceutical properties of a salt and can greatly benefit chemists and formulators
in various aspects of drug discovery and development. The numerous advantages of
salt formation can be listed as following:

1. Altered solubility or dissolution rate

2. Controlled release dosage form

3. Targeted drug delivery

4. Improved thermal stability

5. Improved hydrolytic properties

6. Improved photo stability

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
7. Decreased hygroscopicity

8. Improved permeability

9. Improved organoleptic properties (e.g. increased taste acceptability).

10. Improved drug efficacy.

Disadvantages of Salt Formation:

In spite of several advantages of salt formation, if a due care is not taken in salt
selection process, approach of salt formation can lead to some unwanted effects.
The demerits of salt approach can be listed out as follows:

1. Decreased percentage of active drug content in the formulation leading to

increased powder volume causes problems for tableting and capsule filling.

2. Increased formation of hydrates and polymorphs, resulting in greater variability

of the pharmaceutical properties of drug.

3. Reduced dissolution rate for hydrochloride salts in gastric fluid resulting from
precipitated free acid of base at the surface of the solid dosage form.

4. Increased chance of solid state stability at the micro-environment pH of the salt.

5. Corrosiveness of salts resulting in tableting problems. E.g. highly acidic

hydrochloride salts damage punch tooling.

6. Possible disproportionation (dissociation) of hydrochloride or hydrobromide,

resulting in the release of hydrochloride gas or reaction with excipients or
processrelated chemicals.

7. Additional steps in the synthesis of a medicinal compound.

Pharmaceutical Considerations:

The choice of salt is governed largely by the acidity or basicity of the ionizable
group, the safety of the counter ion, the drug indications, the route of administration
and the intended dosage form. The expectations of the salt form must be outlined as
a desirable pharmaceutical profile that guides the synthesis of the salt forms.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
1. Intended Formulation: Depending on the type of dosage form required Solid
dosage forms- hydrochloride and maleate, Solution or injectable- hydrochloride
and mesylate, Suspension- salts with large, relatively insoluble counter ions e.g.
embonate, esteolate and tosylate , Suppositories- free base

2. Dose Consideration: For low dose drugs, molecular weight of counter ion
weight is not an issue. But the high molecular weight counter ions such as
tosylate may complicate formulation development for high dose drugs.

3. Desired Release Kinetics and Bioavailability: Immediate release- hydrochloride

and sodium, slow release- salts with large and relatively insoluble counter ions.

4. Pharmacological Indications: Counter ion may affect some physiological

processes resulting in the complications in the therapy e.g. sodium salt is not
useful in anti-hypertensive and anti-diabetic drugs due to restricted intake of
sodium.

5. Regulatory Considerations: The salt form should have no adverse safety

implications. Counter ions with known or potentially undesirable
pharmacological implications or toxicity is restricted.

6. Intellectual Property Considerations: The unauthorized commercial use of

patented salt forms is restricted.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
A. BENZILIC ACID

Aim: To Synthesize Acidic Salt of Benzilic Acid and Evaluation of their

physiochemical properties
Reaction:

Mechanism:

Procedure:
In 100 ml round bottom flask place solution of 3.5gm of KOH pellets in 7 ml of water
then add 7 gm (8.7 ml of rectified spirit and 3.5gm of benzil a deep bluish black
solution is produce. fit reflux condenser to flask and boil mixture on water bath for 10-
15 min , pour content of flask into procelin dish and cool in ice bath. Potassium salt of
Benzilic acid crystallized out filter the crystal and wash with little ice and add alcohol
dissolve potassium salt in about 35 ml of water and slowly with stirring 2-9 drops of
conc. HCL from a dropper pipette, the precipitate produced is red brown in colour and
somewhat sticking, filter salt, filtrate nearly colourless dry product and perform its
physiochemical properties.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
Evaluation Table:

Sr. Evaluation Parameter Observation Interferences

No.

1. State Solid --------------

2 Colour Lightly yellow --------------

3 Odour Odourless --------------

Water Soluble

4 Solubility NaOH solution Sparingly soluble

NaHCo3 Sparingly soluble

5 Nature 0.2gm product + water Salt Present

6 Litmus test Blue litmus turn red Acidic

7 Melting point 1040C --------------

8. Limit test for nitrogen, Passes the test All are absent
sulphate, lead acetate.

9. Rf Value ------------- ---------------

Result: Acidic Salt of Benzilic Acid Synthesized and Evaluation of their

physiochemical properties was performed.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
B. BENZOIC ACID

Aim: To Synthesize Basic Salt of Benzoic Acid and Evaluation of their physiochemical
properties.

Reaction:

Mechanism:

Procedure:

In round bottom flask place mixture of 2gm benzoic acid, 10 ml of 10% NaOH,
2ml of conc. H2SO4 add few pieces of porcelain, reflux under water bath for 50
min. Allow to cool under tap water, collect residue of sodium benzoate and
perform its physiochemical properties.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
Evaluation Table:

Sr.No. Evaluation Parameter Observation Interferences

1. State Solid ----------

2 Colour White (Opac Crystal) ----------

3 Odour Odourless ----------

Water Soluble

4 Solubility NaOH solution Soluble

NaHCO3 Sparingly soluble

5 Nature 0.2gm product + water Salt present

6 Litmus test Red litmus turn blue Basic

7 Melting point ----------- ----------

8. Limit test for nitrogen, Passes the test All are absent
sulphate, lead acetate.

9 Rf value ----------- ----------

Result: Basic Salt of Benzoic Acid Synthesized and Evaluation of their physiochemical
properties was performed.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
3. THIN LAYER CHROMATOGRAPHY TECHNIQUE AND PURIFICATION
OF SYNTHESIZED COMPOUNDS BY COLUMN CHROMATOGRAPHY.

CHROMATOGRAPHY:

Chromatography is a separation process which depends upon the differential

distribution of compounds of mixture between a mobile phase and an essentially thin
film stationary phase. The stationary phase may be either in the form of a packed
column through which mobile phase is allowed to flow or in the form of thin layer
adhering to a suitable form of backing material over which the mobile phase is allowed
to ascend by capillary action. The thin film stationary phase may be either a liquid or a
solid, and the mobile phase a liquid or a gas possible combination of this phase then
give rise to the principle chromatographic technique in general use. The
chromatographic techniques which are principally used to the synthetic organic chemist
are described in following section. These are as follows

1) Thin Layer Chromatography (TLC)

2) Column Chromatography

3) Liquid- Solid Chromatography

4) Gas- Liquid Chromatography

5) High Performance Liquid Chromatography.(HPLC, RP-HPLC)

These techniques may be variously used as analytical tools to establish the complexity
of mixtures and purity of samples and as preparative tools for the separation of mixtures
in to individual components.

Thin Layer Chromatography (TLC):

In this technique it is usual to employ glass plates coated with the layers of the solid
stationary phase, which adhere to the plates, generally by virtue of a binding agent, such
as calcium sulphate which is incorporated. The prepared thin layer on glass is often
called a chromaplate. The most commonly used stationary phases, which are available
in grades specially prepared for TLC are includes silica gel, alumina, kieselguhr and
cellulose powder, many of these are available with a fluorescent compound (e.g. zinc
sulphate) incorporated in order to facilitate the detection of the resolved components of
the mixture which is then achieved by viewing the plates under ultra violet light. Other
materials suitable for special application are polyamides, modifies cellulose, with ion
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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
exchange properties and the various forms of organic gel having molecular sieving
properties. (E.g. sephardex, biogel)

Fig.No.06 Technique of Thin Layer Chromatography

Preparation of plate:
Before glass plates are coated with adsorbent they must be carefully cleaned with
laboratory detergent, using a test tube brush to remove adhering particles, rinsed
thoroughly with distilled water, placed in suitable metal rack and dried in an oven.
Subsequent to the treatment with detergent solution the plates should be only handled
by the edges or by the undersurface which not to be coated with adsorbent, failure to
observe this precautions may result in the formation of a mechanically unstable layer
which is liable to flaking due to grease spot on the glass surface. A slurry is prepared by
the slow addition with shaking of 30gm of adsorbent to 100ml of dichloromethane
contained in a wide necked capped bottle. A pair of microscope slide held together and
dipped into slurry slowly withdrawn and allowed to drain while held over the bottle.
The slides are allowed to dry in oven for 10 min.

Loading of plates:
The solutions are applied individually to the marked point on the adsorbent layer by
means of a sample applicator. This is prepared by drawing out a melting point in
capillary tube in micro Bunsen flame and trapping the drawn out portion in two after
reaching with the edge of a fragment of glazed porcelain to ensure a clean break. The
applicator is charged by dipping to the capillary end into the solution and after
withdrawing touching the end on a piece of filter paper until the volume is reduced to
about 0.5 µ.
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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
Selection of the solvent system:
If the chromatographic behavior of the substance under investigation is unknown, the
most satisfactory developing solvent must be checked by preliminary trials runs using
micro plates in 4 wide mouthed topped bottles. Set up sizes of such bottles containing
solvent system of increasing polarity. e.g. hexane, toluene, carbon, tetrachloride,
dichloromethane, diethyl ether, ethyl acetate, acetone, methanol.

Development of plates:
Individual plate‟s 20 x 5 cm plates are conveniently developed in a cylindrical galss jar.
Larger plates 20 x 10 cm and 20 x 20 cm require a rectangular glass tank of suitable
dimensions. Line the inside of the jar with filter paper leaving a gap for viewing the
chromaplate. Saturate the filter paper with the selected developing solvent, close the jar
and allow standing for about 10 min. so that the atmosphere in the jar becomes
saturated with solvent vapors. Insert the plate with the original spot towards the bottom
of the jar. Fit so that the concaved face uppermost. Carefully pour down the side of the
jar more of the developing solvent so that the bottom of the adsorbent layer is well
immersed.

Location of Spot:
The position of colored components can usually be seen without any difficulty
providing that the concentrations in the initial spot is sufficiently high and that
excessive spreading of the component drawing development has not occurred. The
colorless compounds can be developed in iodine chamber.

Fig.No.07 Development of Thin Layer Chromatography

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
COLUMN CHROMATOGRAPHY:

Introduction:
Column Chromatography is another common and useful separation technique in organic
chemistry. This separation method involves the same principles as TLC, but can be
applied to separate larger quantities than TLC. Column chromatography can be used on
both a large and small scale. The applications of this technique are wide reaching and
cross many disciplines including biology, biochemistry, microbiology and medicine.
Many common antibiotics are purified by column chromatography. To understand to
uses of this separation technique, we can use the last experiment as an example. In the
TLC experiment, we separated and analyzed the different components that makeup
over-the-counter painkillers. The technique of TLC was useful in determining the type
and number of ingredients in the mixture, but it was not helpful for collecting the
separated components. We could only separate and visualize the spots. If we needed to
collect the separated materials, column chromatography could be used. We could load
100 mg of a crushed Anacin tablet on a column made up of a silica stationary phase and
separate the aspirin from the caffeine and collect each of these compounds in separate
beakers. Column chromatography allows us to separate and collect the compounds
individually. In this experiment, Column Chromatography (abbreviated CC) will be
used to separate the starting material from the product in the oxidation of fluorene to
flourenone and TLC will be used to monitor the effectiveness of this separation.

Important Steps in Column Chromatography:

1. Selection of a Stationary Phase:

In TLC alumina and silica are the two most popular stationary phases in column
chromatography. For these common phases, the partitioning works in an analogous
manner. The more polar sample will be retained on the stationary phase longer.
Thus the least polar compound will elute from the column first, followed by each
compound in order of increasing polarity. Although the interactions between the
mobile and stationary phase are based on the same principles for CC and TLC, be
careful when predicting the order of elution. Since the direction of the solvent flow
in TLC moves up and in CC the solvent flows down, it appears that the order is
“upside-down”. In TLC the more polar molecules will have lower Rf values, but in
CC they will be retained longer on the column. Remember this when considering
the polarities of the stationary phase as well as the polarity of the compounds being
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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
separated when predicting the order of elution. Stationary phases for CC can come
in a variety of sizes, activities, acidic and basic variations for both alumina and
silica. The types of stationary phase chosen are determined experimentally, or often
based on results from a previous TLC experiment. The type of adsorbent, the size of
the column, the polarity of the mobile phase as well as the rate of elution all affect
the separation. These conditions can be manipulated to get the best separation for
your mixture.

2. Choosing Solvents:
Solvent systems for use as mobile phases in CC can be determined from previous
TLC experiments, the literature, or experimentally. Normally, a separation will
begin by using non-polar or low polarity solvent, allowing the compounds to adsorb
to the stationary phase, then SLOWLY switching the polarity of the solvent to
desorb the compounds and allow them to travel with the mobile phase. The polarity
of the solvents should be changed gradually. On a macroscale, the mixing of two
solvents can create heat and crack the column leading to a poor separation. Some
typical solvent combinations are ligroin-dichloromethane, hexane-ethyl acetate and
hexane-toluene. Often an experimentally determined ratio of these solvents can
sufficiently separate most compounds. Solvents such as methanol and water are
normally not used because they can destroy the integrity of the stationary phase by
dissolving some of the silica gel.

3. Apparatus:
Columns can be as thin as a pencil to a diameter of several feet in industrial
processes. They can separate milligram to kilogram quantities of materials. In this
experiment, we will be separating a mixture of approximately 50 mg, so a small
column can be used. Figure 8.1 shows the typical set-up we will be using during this
experiment. It is essential to have several clean tared Erlenmeyer flasks, reaction
tubes, beakers, test tubes or vials available to collect the solvent and compounds as
they elute. Once you have the general set-up prepared, you can move on to packing
the stationary phase in the column.

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)

Fig.No.08 Typical Microscale Column Chromatography Set-up

4. Packing the Column:

There are several acceptable methods when packing a column. These include dry
packing (there are two versions of dry packing discussed here) and the slurry
method (wet packing). The slurry method normally achieves the best packing
results, but there are several occasions when the dry packing method works just as

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
well if not better. Dry packing is the method of choice for a microscale column.
Begin by filling the column with a nonpolar solvent. Slowly add the powdered
alumina or silica while gently tapping the side of the column with a pencil. The
solid should “float” to the bottom of the column. Try to pack the column as evenly
as possible; cracks, air bubbles, and channels will lead to a poor separation.
For the second dry pack method, the stationary phase is deposited in the column
before the solvent. In this case fill the column to the intended height with the
stationary phase and then slowly add the nonpolar solvent. The solvent should be
added slowly as to avoid uneven channeling. This method is typically used with
alumina only, since silica gel expands and does not pack well with this dry method.
The slurry method is often used for macroscale separations. Combine the solid
stationary phase with a small amount of nonpolar solvent in a beaker. Thoroughly
mix the two until a consistent paste is formed, but is still capable of flowing. Pour
this homogeneous mixture into the column as carefully as possible using a spatula
to scrape out the solid as you pour the liquid. The slurry method normally gives the
best column packing, but is also a more difficult technique to master. Whether the
dry or slurry method is chosen, the most important aspect of packing the column is
creating an evenly distributed and packed stationary phase. As mentioned, cracks,
air bubbles and channeling will lead to a poor separation. Once the column is
loaded, open the stopcock and allow the solvent level to drop to the top of the
packing, but do not allow the solvent layer to go below this point. Allowing this
solvent level to go below the stationary phase, (known as letting the column to “run
dry,”) should always be avoided. Since it allows air bubbles and channel formation
to occur leading to a poor separation.
5. Adding the Sample :
Once the packing is complete, the sample can be loaded directly to the top of the
column. Normally, a minimum amount of a polar solvent, 5-10 drops, is used to
dissolve the mixture. The solution is then carefully added to the top of the column
using a pipette without disrupting the flat top surface of the column. A thin
horizontal band of sample is best for an optimal separation. After the sample is
loaded, a small layer of white sand is added to the top of the column. This will help
to keep the top of the column level when adding solvent eluent. Once the mixture is
added and the protective layer of sand is in place, continuously add the solvent
eluent while collecting small fractions at the bottom of the column. Using a pipette
to add the first bit of solvent on top of the packing, sample, and sand will minimize
disturbance of the column and diluting the sample. Collecting small fractions (1-3

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
mL) is important to the success of your column separation. Fractions that are too
small can always be pooled together; however, if the collected fractions are too
large, you may get more than one compound in any particular fraction. If this
occurs, the only way to complete the separation is to redo the chromatography.
Since column chromatography is time consuming, collecting large fractions is
discouraged.
6. Monitoring the Column :
If the mixture to be separated contains colored compounds, then monitoring the
column is very simple. The colored bands will move down the column along with
the solvent and as they approach the end of the column, collect the colors in
individual containers. Use the color as your guide. However, most organic
molecules are colorless. In this case, the reaction must be monitored by TLC. Spot
each fraction on a TLC plate. Four or five fractions can be spotted on a single TLC
plate. Develop the plate and use the observed spot or spots to determine which
compound is in each of the collected fractions. Spotting some of the starting
material or the product (if available) on the TLC plate as a standard will help in the
identification.
7. Isolating the Separated Compounds :
Once you believe all the materials have been removed from the column, the colors
of the materials or TLC results should indicate which fractions contain the
compound(s) you are interested in isolating. Combine the like or same fractions and
evaporate the solvent. The pure separated compound will be left behind.
Recrystallization may be used to further purify a solid product. However, on a
milligram scale, there is usually not enough material to do this.

Advantages of Column Chromatography:

1. Various types of mixture can be separated.

2. Any quantity of the mixture can be separated

3. The process can be automated.

Disadvantages of Column Chromatography:

1. It is time consuming method.

2. Large quantity of solvent is required which are expensive.
3. Automation of techniques makes it complicated.
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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
A. Amino Acids (Thin Layer Chromatography)

Aim: Thin Layer Chromatography of Amino Acids

Procedure of Experiment:

Apply spots of glycine, aspartic acid on the TLC plate. Allow the solvent to evaporate
and then develop with water: ethanol: acetic acid (1:5:0.1) in the TLC chamber.
Remove the plate from the chamber. Dry and spray with ninhydrin solution. Heat the
plate at 100-1100C for few minutes. Glycine will appear as pink spot whereas aspartic
acid as violet. Calculate the Rf value

Rf value = Distance travelled by the component / Distance travelled by the solvent

B. 2, 4 dinitrophenyl hydrazones (Thin Layer Chromatography)

Aim: Thin Layer Chromatography of 2, 4 dinitrophenyl hydrazones

Procedure of Experiment: Carry out the synthesis of 2, 4 dinitrophenyl hydrazones of

acetone, acetaldehyde & methyl ethyl ketone. Spot the sample of the three derivatives
on the TLC plate and then develop with toluene : pet ether(2:3)or n-heptane:benzene
(3.5:6.5). remove the plate from chamber, dry and spray with 5% aqueous alcoholic
KOH solution. Ketone derivative turns black whereas aldehyde derivative turns red
brown. Calculate the Rf value.

C. Separation of the mixture of dyes (CC)

Aim: Separation of the mixture of dyes

Procedure of Experiment: prepare the column using alumina as the stationary phase
and ethyl alcohol as the mobile phase. Apply concentrated solution of dye mixture
(methylene blue and Fluorescein) on the stationary phase and then run the column with
ethyl alcohol as the eluting solvent. methylene blue will be eluted first.

D. Separation of 2, 4 dinitrophenyl hydrazones (CC)

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Aim: Separation 2, 4 dinitrophenyl hydrazones of carbonyl compounds

Procedure of Experiment: prepare the column using silica gel as the stationary phase
and hexane: ethyl acetate (19:1) as the mobile phase. Apply the concentrated solution of
2, 4 dinitrophenyl hydrazones of acetone, acetaldehyde and methyl ethyl ketone on the
stationary phase and then run the column. 2, 4 dinitrophenyl hydrazones of methyl ethyl
ketone will be eluted first. Gradually increase the strength of the solvent by increasing
the proportion of ethyl acetate, 2, 4 dinitrophenyl hydrazones of acetone will be eluted
next and then finally 2, 4 dinitrophenyl hydrazones of acetaldehyde.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)

SECTION-B

4. SYNTHESIS & PURIFICATION OF FOLLOWING COMPOUNDS USING

PRECIPITATION OR RECRYSTALLIZATION.

A. SYNTHESIS OF BENZIMIDAZOLE.

Aim: To Synthesize Benzimidazole from o-phenylenediamine by Cyclization Reaction.

Apparatus: Round Bottomed Flasks (250ml), Beakers, Reflux Condenser, Buchner

Flasks, Buchner Funnel, Thermometer, etc.

Instruments: Weighing Balance, Melting Point Apparatus, Hot Air Oven.

Chemicals: o-phenylenediamine, Formic Acid, Sodium Hydroxide,

Reference:

1. Furniss B. S., Hannaford A. J., Smith P.W.G., Tatchell A. R., Vogel‟s Textbook of
Practical Organic Chemistry; 5th edition, Page no.1162.

2. Arun Sethi, Systematic Lab Experiments in Organic Chemistry, Second Edition, New
age international publishers, Page no 691.

General Reaction:

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 A Practical Book of Medicinal Chemistry- I (Sem.-V)
NH 2 NHCHO
-H 2 O
H C OH

O
NH 2 Formic acid NH 2
o-Phenylenediamine N-(2-aminophenyl)formamide

NaOH -H 2 O

N
H
1H-benzo[d]imidazole

Principle/Theory:

It is cyclization reaction. o-phenylenediamine on treatment with formic acid gives

benzimidazole. In this the o- phenylenediamine having N atom contains lone pair of
electrons i.e. negative charge. The formic acid contains the carboxylic group, it will
attack the nitrogen atom of o- phenylenediamine eliminating water molecule. Further
the carboxylic group will attack another nitrogen atom and the elimination of water
molecule will lead to formation of benzimidazole as final product by electron
rearrangement. Thus the synthesis of benzimidazole includes cyclization reaction.

Procedure:

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Place 27 g of o-pheylenediamine in a 250 ml round bottomed flask and add 17.5 g of 90
per cent formic acid. Heat the mixture on a water bath at 100 oC for 2 hours. Cool, add
10 per cent sodium hydroxide solution slowly, with constant rotation of the flask, until
the mixture is just alkaline to litmus. Filter off the crude benzimidazole at the pump
wash with ice cold water, drain well and wash again with 25 ml of cold water. Dissolve
the crude product in 400 ml of boiling water, add 2 g of decolourising carbon and digest
for 15 minutes. Filter rapidly at the pump through a preheated Buchner funnel and flask.
Cool the filtrate to about 10oC, filter off the benzimidazole, wash with 25 ml cold water
and dry at 100oC.Report, yield and Melting point of the product. (Note: Start with 1 gm
of o-pheylenediamine)

Characterization of the Compound:

1. Progress of the reaction is monitored by performing thin layer chromatography.

2. The compound synthesized is purified by Recrystallization / distillation.
3. The compound synthesized is characterized by detection of elements, functional
groups, confirmatory tests and determination of melting point/ boiling point.
4. The compound synthesized is further characterized by qualitative analysis/
structural elucidation by FT-IR spectroscopy wherein presence of different
functional groups is confirmed.

Observation Table:

Chemical Tests:
Test Observation Inference

Physical constant

Calculation of % yield:
Theoretical Yield (X) = --------

Moles of the reactant = Moles of the product

Thus „A‟ gm of reactant = „X‟ gm of the product

X = (Moles of the product ÷ Moles of the reactant) x A

Practical Yield (Y) = ---------

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
%Yield = ( Y÷X ) x 100 = --------

Result:
Name of the Compound

Theoretical Yield (gm)

Practical Yield (gm)

Percentage Yield (%)

Physical Actual
Constant
Practical
M.P.

Rf value

I.R. Observations:

Sr. No Functional Group IR Value (cm-1)

Theoretical Observed

Calculation of Rf value:

Rf = 0.69

Rf = 0.15

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Label:
Name of Compound

Structural formula

IUPAC name

State and Color

Molecular Weight

Molecular Formula

Melting point

% yield

Use

Prepared by

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
B. SYNTHESIS OF 1, 2, 3, 4-TETRAHYDRO CARBAZOLE

Aim: To Synthesize 1, 2, 3, 4-tetrahydro Carbazole Cyclohexanone and

phenylhydrazine by Hoffman‟s Rearrangement.

Reaction:

Mechanism:

Procedure:
A mixture of 98 g. (1 mole) (Note 1) of cyclohexanone and 360 g. (6 moles) of acetic
acid contained in a 1-l. three-necked round-bottomed flask equipped with a reflux
condenser, a slip-sealed stirrer, and a dropping funnel is heated under reflux and stirred
while 108 g. (1 mole) of phenylhydrazine is added during 1 hour. The mixture is heated
under reflux for an additional hour and poured into a 1.5-l. beaker and stirred by hand

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
(Note 2) while it solidifies. It is then cooled to about 5° and filtered with suction, the
filtrate being cooled in ice and refiltered through the filter cake. The final filtrate is
discarded. The filter cake is washed with 100 ml. of water and finally with 100 ml. of
75% ethanol. Each wash is allowed to soak into the filter cake before it is sucked dry.
The crude solid is air dried overnight (Note 3) and crystallized (Note 4) from 700 ml. of
methanol after treatment with decolorizing carbon (Note 5); yield 120–135 g. of
1,2,3,4-tetrahydrocarbazole, m.p. 115–116° (Note 6). The mother liquor is concentrated
to one-fourth of its original volume and yields an additional 10 g. of product (total yield
76–85%) (Note 7).
Notes:
1. Equivalent amounts of cyclohexanone (after suitable compensation for purity) and
phenylhydrazine are used. The cyclohexanone was about 90% pure (analyzed by the
procedure of Bryant and Smith2). Instead of analyzing the ketone, it is safe to
assume 90% purity.
2. The stirring should be sufficiently vigorous to prevent the formation of lumps.
3. The crude product requires 50–70 hours of air-drying to attain constant weight
(145–155 g., 85– 91%). It is preferable to crystallize the partially dried product.
4. The approximate solubility of 1,2,3,4-tetrahydrocarbazole in 100 ml. of methanol at
10°, 35°, and 55° is 5, 12, and 18 g. respectively.
5. A heated funnel is desirable for filtration of the hot solution, for the product
separates on slight cooling.
6. The capillary melting point of tetrahydrocarbazole ranges from 113° to 114° with
slow heating and from 116° to 118° with fast heating.
7. 1,2-Benzo-3,4-dihydrocarbazole may be prepared by the same general procedure. A
solution of 172 ml. (2 moles) of concentrated hydrochloric acid (sp. gr. 1.18) in 500
ml. of water is heated at the reflux temperature and stirred in a 2-l. three-necked
round-bottomed flask equipped with a reflux condenser, a slip-sealed stirrer, and a
dropping funnel while 108 g. (1 mole) of phenylhydrazine is added during 5
minutes. α-Tetralone (p. 898) (146 g., 1 mole or a correspondingly larger amount of
material of 90% purity; see (Note 1)) is added in a period of 1 hour, and the mixture
is stirred and heated under reflux for an additional hour. The product is cooled to
room temperature with stirring, and the beadlike product is filtered, washed as
above, and crystallized from 2.3 l. of methanol after treatment with decolorizing
carbon. The first crop amounts to 105–110 g. and the second crop to 75–80 g.,
making the total yield 82–87%; m.p. 163–164°.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
C. SYNTHESIS OF 2, 3-DIPHENYL QUINOXALINE

Aim: To Synthesize 2, 3-diphenyl Quinoxaline from o-phenylenediamine and Benzil

by Condensation Reaction.

Reaction:

Mechanism:

Procedure:
To warm solution of 2.1g (0.01 mol) of Benzil in 8ml of rectified spirit , mix a solution
of 1.1g(0.01mol) of o-phenylenediamine in 8 ml rectified spirit. Warm in a water bath
for 30 min and add water until a slight cloudiness persists and allow to cool, filter and
recrystallized from aqueous ethanol to give 1.43g (51%) of 2, 3-diphenyl Quinoxaline.
M.P. 125-1260C

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
D. SYNTHESIS OF BIS-Β-NAPTHOL.

Aim: To Synthesize Bis-β-napthol from β-napthol.

Reaction:

Mechanism:

Procedure:

Weigh 10 g of 2-naphthol are dissolved in an excess of ether, and 16 g (excess) of

anhydrous iron(III) chloride are gradually added to the solution in a flask fitted with a
reflux condenser. Much heat is evolved during this operation. The mixture is then
refluxed on a water bath until most of the 2-naphthol is oxidised. (To test this a small
portion of the ethereal solution is treated with an excess of dilute hydrochloric acid, and
the etherevaporated. The 1,1′-bi-2-naphthol separates out even in the warm as an oil
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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
while 2-naphthol crystallises out on cooling.). When this is the case, the ether is
removed on a water bath, water and powdered calcium carbonate are added to the
residue, and the whole well shaken. Excess sodium hydroxide is then added, the
solution is filtered and precipitated with dilute sulfuric acid. The precipitate is washed
with boiling water or boiling ligroin, and recrystallised from benzene. Yield 4• 0%
theoretical (8 g). Colourless needles from alcohol, prisms from a mixture of carbon
disulfide and alcohol; insoluble in water; slightly soluble in chloroform; soluble in
alcohol and ether; m.p. 216-218° C.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
E. SYNTHESIS OF ANTHRANILIC ACID.

Aim: To Synthesize Anthranillic Acid from pthalimide by Hoffmann‟s Rearrangement

Reaction.

Reaction:

Mechanism:

Procedure:
Prepare a solution of 30 gm of NaOH in 120 ml of water in 250 ml conical flask and
cool to 0˚c or below in a bath of ice. Add 26.2 gm of bromine in one portion and shake
until all bromine has reacted. Temperature will rise slightly again. Cool the mixture.
Prepare solution of 22gm of NaOH in 80 ml of water. Add 24gm of phthalimide in a
portion to cold NaOBr solution in terms of smooth paste with water rapidly with
stirring. Remove the flask from cooling bath and shake vigorously until a clear yellow
solution is obtained. Add prepared NaOH solution rapidly in one portion. Heat the
solution at 80˚C for about 2 minutes and filter if necessary. Cool in ice and add conc.
HCl slowly with stirring until the solution is just neutral. Precipitate Anthranilic acid
completely by gradual addition of glacial acetic acid (20-25 ml will be required). Filter
off the acid at pump and wash with a little cold water. Recrystallized from hot water
with addition of little decolorizing agent. Collect the acid on Buckner funnel and dry at
100˚c.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
F. SYNTHESIS OF SULPHANILAMIDE

Aim: To Synthesize Sulphanilamide from p-acetamidobenzenesulfonamide by

Hydrolysis Reaction.

Reaction:

Mechanism:

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Procedure:

4-acetamidobenzenesulfonamide is weighed and placed in a 50 mL round-bottom flask

equipped with a magnetic stir bar. Dilute hydrochloric acid (6 M) is added to the flask
in an amount equal to twice the weight of the 4-acetamidobenzenesulfonamide. The
flask is fitted with a cold water condenser and heated at reflux with constant stirring for
45 minutes, after which it is allowed to cool to room temperature. If any solid appears
upon cooling, the mixture is reheated at reflux for another 15 minutes. After cooling,
the reaction mixture is neutralized by slow addition of a saturated Na2CO3 solution
with stirring until it tests slightly alkaline to pH paper. A precipitate may have begun to
form during neutralization. Cool the beaker in an ice bath to complete the precipitation
of product. It may be necessary to gently scratch the inside bottom of the beaker to
induce crystallization. The product is collected by vacuum filtration, washed with a
small amount of ice cold water and air dried. A small sample of the crude product is
saved, and the remainder is purified by recrystallization by dissolving in a minimum
amount of boiling hot water, allowing the solution to slowly cool to room temperature,
then further cooling in an ice bath to crystallize as much product from solution as
possible. The recrystallized product is collected by vacuum filtration. The product is
weighed and % yield is calculated. The overall yield for the five-step synthesis of
sulfanilamide from nitrobenzene is calculated and reported. IR and NMR spectra are
obtained on the final product.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
G. SYNTHESIS OF BENZOIC ACID FROM BENZYL ALCOHOL.

Aim: To Synthesize Benzoic Acid from Benzyl Alcohol by Cannizaro‟s Reaction.

Reaction:

Mechanism:

Procedure:

Benzaldehyde (20g) are treated in a stoppered cylinder or a thick-walled vessel with a

cold solution of 18 g of potassium hydroxide in 12 g of water, and mixed until a
permanent emulsion is formed. The mixture is then allowed to stand over night. The
vessel is closed by a cork, and not a glass stopper, since at times a glass stopper
becomes so firmly fastened that it can be removed only with great difficulty. To the
crystalline paste (potassium benzoate) separating out, water is added until a clear
solution is obtained from which the benzyl alcohol is extracted by repeatedly shaking
with ether. After the evaporation of the ether the residue is subjected to distillation;
benzyl alcohol passes over at 206° C. Yield, about-8 g. The benzoic acid is precipitated
from the alkaline solution on acidifying with hydrochloric acid and purified from hot
water yielding final product with melting point 121° C.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
H. SYNTHESIS OF PROPRANOLOL

Aim: To Synthesize of Propranolol from 1 napthol.

Reaction:

Mechanism:

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Procedure:

Step I: Transfer 1.25gm (0.0085mol of 1 napthol CPM 144.18 per mole ) 0.5gm of
KOH to a round bottom flask and add ethanol, After dissolution add drop wise 2 ml of
epichiorohydryl , The reaction is left under magnetic stirrer at room temp for 48 hrs,
TLC is an carried out in an eluent system (hexane ethyl acetate) to monitor the yield of
reaction , extract aqueous phase with ethyl ether, ethyl ether extract is dried with
anhydrous sodium sulphate, filter and remove solvent to obtained crude brown oil.

Step II: Transfer 0.2gm of crude oil with menthol to round bottom flask heated with a
stopper and add 4ml of isopropyl amine, keep the reaction under temp control for 2 hrs.,
remove solvent by Vaccum evaporator, obtained product add 30ml of 2 mole per liter
HCL and transfer to separating funnel.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
I. SYNTHESIS OF 1, 4-DIHYDROPYRIDINE

Aim: To Synthesize 1, 4-dihydropyridine form formaldehyde by Hantzsh Synthesis.

Reaction:

Mechanism:

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
Procedure:

Take 1ml aldehyde 2ml ethyl acetoacetate 1ml/gm aminoacetate 1ml water remaining
space charge with water steam air or nitrogen then close round bottom flask. Mixture
stir for desire time at 70-750C on composition final mixture was ten cool at room temp
and stay over neighs Mixture was filter to isolated precipitate product recrystallized
product 90% ethanol give product.

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 A Practical Book of Medicinal Chemistry- I (Sem. - V)
BIBLIOGRAPHY

References:
1. Vogel's Textbook of Quantitative Chemical Analysis, J. Mendham, R.C. Denney, J.
D. Barnes, M. J. K Thomas, 6th Edition, Pearson‟s Education Ltd , Page No. 68-67.
2. Practical Pharmaceutical Chemistry, A. H. Beckett, J. B. Stenlake, 4th Edition,
CBS Publication, Page No. 114-122.
3. Vogel‟s Textbook of Practical Organic Chemistry, B. S. Furniss, A. J. Hannaford ,
P.W.G. Smith, A. R Tatchell, 5th edition (2008), Pearson‟s Education Ltd , Page No.
898,589, 1284.

4. Systematic Lab Experiments in Organic Chemistry, Arun Sethi, Second

Edition(2003), New Age International Publishers, Page No .741, 722, and 764.

5. Practical Medicinal Chemistry , Dr.R.S.Chavan, S.J.pawar, Dr. H.N.More , Dr

S.D.Swant , 1st Edition , Aug 2013, Nirali Prakashan, Page No.1.3, 7.1, 17.1, 48.1.

6. Instrumental Methods of Chemical Analysis, G.R. Chatwal, S.K. Anand, First

edition (2003), Himalaya Publishing House, Page No. 2.602-2.603.

7. A Systemic Identification of Organic Compounds, Shriner, Hermann, Morrill,

Curtin, fuson, 7th Edition(1998), John Wiley & Sons ltd Publications. Page No.
1123.

8. Handbook of Practical Chemistry (Inorganic & Organic), DR K.R.Mahadik,

S.H.Bhosale, 8th Edition (2008), Nirali Prakashan, Page No. 235.

9. Advanced Practical Medicinal Chemistry, Ashutosh Kar, New Age International ltd,
2004 1st Edition, Page No.130

10. Reaction Mechanisms in Organic Synthesis, R.K.Parashar, John Wiley & Sons Ltd
Publications, 2009, 1st Edition, Page No. 23-40.

11. The Merk Index, An Encyclopedia of Chemicals Drugs and Biological, edited by
Maryodele, J.Meli, P.E. Heckelman, 14th edition, Page No. 130

12. Experimental Pharmaceutical Organic Chemistry by K.S.Jjain, P.B.Miniyar,

T.S.Chitre 2nd edition, Career Publication, Page No. 19-25.
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