Domingo, Joevani T. (Laboratory Manuals)

ORGANIC CHEMISTRY
LABORATORY MANUAL
Domingo, Joevani T.
BS-Chemical Engineering 2B
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TABLE OF CONTENTS
LABORATORY HAZARDS AND FIRST AID- REVIEW 2-16
OF BASIC LAB TECHNIQUES
PURIFICATION OF AN IMPURE ACETANILIDE 17-26
SAMPLE BY RECRYSTALLIZATION AND MELTING
POINT DETERMINATION
SEPARATION OF A BINARY MIXTURE BY SIMPLE AND 27-38
FRACTIONAL DISTILLATION
EXTRACTION: DETERMINATION OF ITS EFFICIENCY; 49-52

CALCULATION OF THE DISTRIBUTION COEFFICIENT
PREPARATION OF SYNTHETIC FOOD FLAVORS 53-60
ISOLATION OF CAFFEINE FROM TEA LEAVES 61-70
THIN LAYER CHROMATOGRAPHY OF ANALGESIC DRUGS 71-84

COLUMN CHROMATOGRAPHY OF PIGMENTS: 85-101
INK/FOOD COLORING
ANALYSIS OF ALCOHOLS/PHENOLS/ALDEHYDES/KETONES 102-118

CARBOXYLIC ACID AND DERIVATIVES 119-129
SYNTHESIS OF ASPIRIN 130-140
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LAB EXPERIMENT NO.1
LABORATORY HAZARDS AND FIRST AID- REVIEW
OF BASIC LAB TECHNIQUES
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I. INTRODUCTION
We know that running a research lab is a challenge, to say the least. In all the hustle of
loading the autosampler, pipetting, pouring, and mixing for research experiments, worker health
and safety can be overlooked, inadvertently pushed aside or forgotten–sometimes with dire
consequences. Then if you are working in a laboratory then you must aware of the first aid
treatment in case of any major or minor incidents. Minor issues can be a major problem for future
in case of any carelessness; so always work in the laboratory to keep essential safety instruments
handy. You must be well trained for any chemical reaction or other experiments and must have
knowledge about all lab safety rules. Be responsible while working in lab.
We are describing here different hazards, laboratory accidents, first aid treatment for
injuries, some basic laboratory techniques, and rules need to conduct in laboratory.
II. OBJECTIVES
 Define the different hazard that can encounter in laboratory
 Discuss the different safety precaution if we encounter this different hazard.
 Identify different Laboratory Hazards Signages
 Discuss properly the relevant first aid and basic laboratory techniques
 Illustrate the different first aid and basic laboratory techniques
III. DISCUSSION
DIFFERENT TYPES OF HAZARDS IN LABORATORY
1. Chemical hazards - The use of chemicals in research laboratories is inevitable,

and the potential for harm or injury could be significant if they are misused or
mishandled. Examples of chemical hazards are cleaning agents and
disinfectants, drugs, anesthetic gases, solvents, paints, and compressed gases.
Potential exposures to chemical hazards can occur both during use and with poor
storage.
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2. Biological hazards- include potential exposures to allergens, infectious
zoonotics (animal diseases transmissible to humans), and experimental agents
such as viral vectors. Allergens, ubiquitous in animal research facilities, are one
of the most important health hazards, yet they are frequently overlooked.
3. Physical hazards - The most obvious are slips and falls from working in wet
locations and the ergonomic hazards of lifting, pushing, pulling, and repetitive
tasks. Other physical hazards often unnoticed are electrical, mechanical,
acoustic, or thermal in nature. Ignoring these can have potentially serious
consequences.
4. Electrical hazards - are potentially life threatening and found much too
frequently. First, equip all electrical power outlets in wet locations with ground-
fault circuit interrupters, or GFCIs, to prevent accidental electrocutions.
IMPORTANT STANDARDS TO HELP MITIGATE THESE HAZARDS

1. Chemical Hazards
 OSHA (Occupational Safety and Health Administration) programs and
recognizing hazards will help you to identify and minimize many of the
common safety and health hazards develop two standards for chemical
hazards.
 The first is the Hazard Communication standard (29CFR1910.1200) that
deals with requirements for employers to inform and train employees on
non-laboratory use of chemicals. This would apply to things in the lab
such as pump oil, Chromerge, or liquid nitrogen used in dewars.
Although these chemicals are found in the lab, their use does not meet
the criteria for laboratory use.
 The second, “OSHA Lab Standard,” 29CFR1910.1450 requires
laboratories to identify hazards, determine employee exposures, and
develop a chemical hygiene plan (CHP) including standard operating
procedures. The “lab standard” applies to the laboratory use of chemicals
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and mandates written Standard Operating Procedures (SOPs) addressing
the particular hazards and precautions required for safe use. This goes
hand in hand with experimental design and planning. Both standards
require providing material safety data sheets and employee training.
2. Biological Hazards
 Health and safety issues such as containment, the ability for
replication, and potential biological effect are all important. When
working with biological hazards, ensure that procedures can be
conducted safely. Much of the work with recombinant DNA, acute
toxins, and select agents is now regulated by federal agencies such as
the U.S. Department of Agriculture, the Department of Homeland
Security, and the Department of Health and Human Services (including
the National Institutes of Health). If your facility is conducting
research in these areas, you should have an Institutional Biosafety
Committee to keep everything in order and running smoothly.
 The most prevalent biological hazards, in terms of frequency of
occurrence, are simple allergens associated with the use and care of
laboratory animals. Health surveys of people working with laboratory
animals show that up to 56 percent are affected by animal-related
allergies. In a survey of 5,641 workers from 137 animal facilities, 23
percent had allergic symptoms related to laboratory animals. These
figures do not include former workers who became ill and could not
continue working.
3. Physical Hazards
 Included here are electrical safety hazards, ergonomic hazards
associated with manual material handling and equipment use, handling
sharps, and basic housekeeping issues. Many operations in the lab can
result in lab workers assuming sustained or repetitive awkward postures.
Examples are eluting a column in a fume hood, working for extended
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periods in a biosafety cabinet, or looking at slides on a microscope for
extended periods. What is found acceptable for brief or occasional use
may become problematic if performed for long durations or very
frequently. Pain is a good indicator that something is wrong. Conduct
work with a neutral, balanced posture. Magnetic assist or programmable
pipettes can reduce frequency of hand force required to prevent worker
injury.
 Sharps containers are ubiquitous in research labs and following a few
safety rules can help prevent getting stuck with accident reports. Use
only puncture-proof and leakproof containers that are clearly labeled.
Train employees never to remove the covers or attempt to transfer the
contents. Make sure these containers are only used for “sharps” and that
they get replaced when three-fourths full to prevent overfilling.
 Many injuries stem from poor housekeeping. Slips, trips, and falls are
very common but easily avoided. Start with safe and organized storage
areas. Material storage should not create hazards. Bags, containers,
bundles, etc., stored in tiers should be stacked, blocked, interlocked,
and limited in height so that they are stable and secure against sliding
or collapse. Keep storage areas free from an accumulation of materials
that could cause tripping, fire, explosion, or pest harborage.
LABORATORY HAZARDS SYMBOL
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FIRST AID REVIEW TECHNIQUE
Incase of Chemical Spill:
Before experimenting with any chemical reaction you must aware of the four important facts
of matter which are mentioned below:
1. Reactivity to air or water
2. High toxicity
3. Corrosion
4. Flammability
 If the spilled Chemical is burnable then turn off all kinds of heat generated sources.
 Call for knowledgeable personnel.
 Close doors to stay away from fire.
 Always note down the response from chemical reaction whether it spills or not. All
lab workers must be updated with the response of a particular reaction.
 Do not forget to keep all safety equipment and spill clean-up materials.
 Also, ensure that the person involving spill reaction must be experienced. Do not
allow untrained personnel to work in the lab.
For Biological spill accidents:

 For Biological spills, emergency lab workers should have the following safety
equipment.
a. Disinfectant solution
b. Forceps, tongs, broom, dustpan
c. Personal protective equipment (PPE): safety glasses, goggles, or face shield,
utility gloves, wrap-around lab coat, shoe covers (optional)
d. ‘Biohazard’ bag, sharps container
e. Paper towels or other absorbents.
 Clean the spill area with a fresh cloth that can soak it well.
 Keep calm and stay away immediately from the affected area.
Radioactive spill emergency:
 Allow working in the lab to only those persons who are well trained.
 Contact the personnel who are aware to handle the affected area.
 Contact to the radioactive spill emergency.
 Close doors to prevent from being affected.
What to do for Clothing on fire:
 Use the safety shower immediately on the infected part of the body.
 Roll the individuals on the flame.
 Call the doctor or other supervisor who is an expert to treat.
 Rinse the exposed area thoroughly with water.
 Use a deluge shower for 15 minutes.
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 Wash the infected part from heat and chemicals continuously for 15 minutes.
 In case of affected large areas remove clothing.
Hazardous material splashed in eye
 You have to rinse the eyeball immediately and the inner surface of the eyelid with
water.
 Do this process for 15 minutes continuously.
 Don’t scrub your eyes.
 Report all incidents to the expert or supervisor.
Some other important lab safety rules:
1. Do not work alone in the lab always do in the presence of a lab assistant or
teacher.
2. If you have any doubt or lack of knowledge about any reaction or experiment then
take instructions from your teacher or their expert person.
3. If you are entering for the first time in the lab then do to touch any chemical or
instrument without permission.
4. Never eat any junk food, drink, or other food.
5. Follow all the safety rules which your teacher or assistant told you.
6. While any chemical reaction keeps handy away from eyes, mouth, or other parts
of the body.
7. Always wear safety gloves and a lab coat before start working.
8. Googles, gloves, a lab coat, and shoes are essential to wear.
9. Inform the teacher or other trainer of any incident or injury and keep calm.
10. Do not taste, or smell any chemical.
BASIC LAB TECHNIQUES
HANDLING OF LIQUIDS
When Pouring a liquid from a bottle:

Do not lay glass stopper down on the laboratory table, but hold it while
pouring the liquid from the bottle. Grasp the bottle with the palm of the hand
covering the label.
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Hold the container into which the liquid being poured as well as the container
delivering the liquid.
When pouring a liquid into another container use a stirring rod as guide.
In reading a graduated cylinder, read the bottom part of the meniscus with your
eyes parallel to it.
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HANDLING OF SOLIDS
When transfering solid, rotate the bottle or if using a spatula, tap the
spatula to remove the desired quantity.
Take only what is needed. If you take too much, do not return the excess to
the reagent bottle but dispose it off properly.
Do not throw solids or organic waste into the sink. There are special
containers for their disposal in the laboratory.
USING BALANCES
Always set the balance to zero before weighing chemicals .To keep the
balance clean, always weigh a solid in a container, such as a piece of paper or a
small dish
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USING A LABORATORY BURNER
Connect the rubber tubing of the gas burner to the gas outlet. Close the air
holes. Open The gas valve. Light the burner by holding a lighted match to the side
and a little below the top of the barrel of the gas burner as the gas regulator is slowly
opened. Open the ari holes gradually and observe the variation of the flame.
Regulate the supply of air by adjusting the air holes until a non-luminous flame is
obtained. Never attempt to heat a flammable liquid with a burner
HEATING OF LIQUIDS
When heating a liquid with the use of a test tube, never point a test tube at
yourself or anyone nearby. Overheating may result in splashing yourself or your
neighbour with hot liquid.
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MIXING CHEMICALS
When mixing chemicals in a beaker, stir with a clean glass stirring rod.
If a solid is tobe combined with a liquid, place the solid in the beaker first
then add the liquid. Stir with a glass rod.
When mixing chemicals in a test tube, cover the test tube with a cork and
shake it.
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USING A BURET
For proper manipulation and proper techniques in titration, hold the flask
with your right hand and the stopcock with your left hand. Open the stopcock slowly
to allow some liquid to run into the flask. Swirl the flask.
USING PIPETS
Never pipet by mouth. Always use the rubber aspirator.
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FILTRATION
Fold the filter paper the correct way by folding it into half first then into
another half making four parts all in all. Open one side of the folded filter paper by
making a cone and fit into the funnel by wetting the side with a small amount of
distilled water.
The gravity filtration set up should be done by placing the funnel containing
the filter paper in an iron ring, then placing the receiving vessel under the funnel
with the tip of the fiunnel touching the side of the receiver.
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IV. GUIDE QUESTION
1. What is an Hazard?
Answer:
2. Enumerate the different type of hazards in laboratory?
Answer:
3. Which of the hazards is commonly happens all the time?
Answer:
4. You have accidentally broken a test tube and spilled a chemical on the
table. What are you going to do?
Answer:
5. Right when you get to class, it is important to get started in the lab even if
your teacher hasn't given you instructions yet. Why?
Answer:
6. When studying a chemical it is important to touch, taste, and smell it so
that you know a lot about it.Yes or No? Why?
Answer:
7. What are Laboratory Techniques?
Answer
8. Which color of flame from a gas burner is hottest, and how is it produced?
Answer:
9. Why should you fold and put creases in your filter paper prior to filtration?
Answer:
10. During filtration, why is it important to only wash your solid with ice-cold
solvent?
Answer:
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V. REFERENCES
Online Source
Harvey (2013). “Gravity Filtration”. Retrieved from:

https://community.asdlib.org/imageandvideoexchangeforum/2013/07/24/gravity-filtration/ .
Date Accessed: October 29, 2020
Holt, Rinehart and Winston, Publishers, New York. “Laboratory Experiments in ACTION
CHEMISTRY” Retrieved from :
https://ndchemistry.weebly.com/uploads/2/0/7/0/20705826/l00_intro_to_lab.pdf. Date
Accessed : October 29, 2020
Mcleod (2011). “ Laboratory Hazards and Risks”. Retrieved from:

https://www.labmanager.com/lab-health-and-safety/laboratory-hazards-and-risks-182382.
Date Accessed: October 29, 2020
Rahul (2015) “Laboratory accidents and first aid treatment for injuries”. Retrieved
from: https://www.aticoexport.com/laboratory-accidents-and-first-aid-treatment-for-
injuries. Date Accessed: Octover 29, 2020
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LABORATORY EXPERIMENT NO. 2
PURIFICATION OF AN IMPURE ACETANILIDE SAMPLE BY

RECRYSTALLIZATION AND MELTING POINT DETERMINATION
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I. INTRODUCTION
Recrystallization (AKA: crystallization) is a purification technique that is based on the

difference in the solubility’s of the desired compound and the impurity (impurities)
contaminating the desired compound, as a function of varying temperature. This makes the
choice of the right solvent a critical one. As a rule of thumb, at high temperatures (i.e. the
boiling point of the solvent) compound of interest and the impurities should be overly soluble
in the solvent, but at lower temperatures, the compound of interest should be quite insoluble in
the solvent of choice.
A quick overview of the process of recrystallization is the following: The crude mixture
is placed in the minimum volume of the solvent of choice that would dissolve the entire solid
at its boiling point. This mixture is then heated and stirred to reach boiling.
II. OBJECTIVES
At this point is to obtain a saturated solution at boiling. After reaching the
boiling point, with the entire solid dissolved, the solution is left to cool slowly. As the
temperature is slowly dropped, the compound of interest gradually starts to precipitate
out (crystallize), while the impurities stay dissolved. Normally, the mixture is cooled in
an ice bath in order to maximize the difference in the solubility behaviors of the
compound of interest with that of the impurities, and to also maximize the formation of
the crystals of the compound of interest.
The pure crystals of the cooled mixture can then be separated through a
filtration. The filtrate (pure crystals) should be rinsed with a small amount of cold
solvent in order to insure a higher purity. Relevant Calculations: Calculation of %
Recovery for the recrystallization process measures the efficiency of the purification
and isolation combined.
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III. MATERIALS AND PROCEDURE
Chemicals: Equipment/ glassware:
Name Structure Physical Thermometer Hot Plate Hirsh Funnel
Acetanilide Solid
(impure) Melting Point:
114℃
Vacuum Adaptor Erlenmeyer Flask Capillary Tubes
Benzoic acid Solid
Melting Point:
121℃
Reagents: decolorizing/activated charcoal
Setup:
IV. PROCEDURES
Dissolution of the Impure Acetanilide
a. Weigh out accurately ca. 0.20 g. sample of impure acetanilide; place this in a
50mL Erlenmeyer flask
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b. Add ca. 10 mL of distilled water to the flask and bring the water to a gentle boil
by heating it in the hot plate.
c. Continue to heat until no more solid appears to dissolve.
d. Remove the heat source; allow the flask to cool a few minutes.
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e. Add a small amount (~0.05 g) of activated charcoal to the contents of the flask
and continue to boil for additional 3-4 minutes.
Gravity Filtration
a. Place a 50 mL Erlenmeyer flask containing about 5 mL of distilled water on a

hot plate to heat it. Place a short-stem glass funnel with a fluted filter paper on
top of the flask.
b. Using the clamp as a handle, filter the hot acetanilide solution as quickly as
possible. If particles of charcoal pass through the filter paper, re-heat the
solution to boiling, and filter it again through the same piece of filter paper.
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Recovery of purified acetanilide
a. Cool the Erlenmeyer flask containing acetanilide solution to room temperature

and then in an ice bath to complete the crystallization.
b. Assemble a vacuum filtration apparatus using a filter flask and Hirsch funnel.
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c. Place a piece of filter paper and collect the crystals by vacuum filtration.
d. Collect the crystals on a weighing paper and air dry. Weight the crystals to
determine the % recovery. Transfer the crystals in a properly labeled sample
container.
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e. Determine the melting point of your purified acetanilide and impure (starting)
acetanilide.
f. Determine the % impurity in the acetanilide that was initially provided to you, by
linear correlation between the melting points of three mixtures provided to you with the %
impurity information.
Cleanup:
The filtrate from the recrystallization can be poured down the sink. All solid
material is organic waste – goes in organic waste bottle. As always, clean the glassware
with soap, rinse with acetone, ethanol, and water, in that order.
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V. GUIDE QUESTION.
1. Did recrystallization improve the purity of your acetanilide? How can you tell?
ANSWER:
2. Assume that your original sample of acetanilide was 90% pure. What was the maximum
amount of pure acetanilide that you could recover after your recrystallization?
ANSWER:
3. How much acetanilide did you lose during the entire recrystallization process? Where did
the losses probably occur?
ANSWER:
Mass of recovered acetanilide =

Mass of lost acetanilide =
4. What happens to the impurities in a recrystallization? In other words, how are soluble
impurities removed during a recrystallization?
ANSWER:
5. Why is water a good solvent for the recrystallization of acetanilide?
ANSWER:
6. How did the melting point of your acetanilide change after recrystallization?
ANSWER:
7. Why could you test the purity of your acetanilide by mixing it with a known, pure sample
of acetanilide and rechecking the M. P.?
ANSWER:
8. Why must the funnel be heated before the hot acetanilide solution is filtered?
ANSWER:
9. What is the difference between percent recovery and percent yield?
ANSWER:
10. . Melting “point” is a misleading term. Why?
ANSWER:
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11. What happens if you add too much solvent during recrystallization?
ANSWER:
12. During filtration, why is it important to only wash your solid with ice-cold solvent?
ANSWER:
13. What factors influence the solubility of a solid in solution?
ANSWER:
VI. REFERENCES
Online Source
Matt (2011), “Preparation/Recrystallization of Acetanilide”, Retrieved form :

http://www.mendelset.com/articles/680/preparation-recrystallization-acetanilide. Date
Accesed: Nov. 3, 2020
Stoddard (2020). “CHM 242 Lab 1 Recrystallization of Acetanilide”. Retrieved from:

https://www.youtube.com/watch?v=b8jHC05jJNA. Date Accessed: Nov. 03, 2020
Organic laboratory Manual, Shanbhag & Veliz, Page 24-25, Nova Southeastern
University, 2012
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LABORATORY EXPERIMENT NO.3
SEPARATION OF A BINARY MIXTURE BY SIMPLE AND
FRACTIONAL DISTILLATION
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I. INTRODUCTION/BACKGROUND:
Distillation is a process used in organic chemistry laboratories to purify and identify organic
compounds (CU). The process consists of the separation, evaporation, and condensation of a liquid
mixture composed of different substances. Distillation can be done through simple and fractional
distillation. Simple distillation is the process of boiling a liquid mixture in a vessel and condensing
the vapors to obtain the liquid solvent. Fractional distillation follows a similar procedure as simple
distillation but fractional distillation uses a fractionating column that is located between the vial
and the distillation head (Weldegirma, 2012). The substances are separated based on the boiling
points of the substances in the mixture. Simple distillation is more effective when the difference
between the boiling points of the two components is 25°C and above. While fractional distillation
works if the boiling point difference between the components is between 10°C and 15°C.
Therefore, the substances should have different boiling points for the separation to take place. The
Distillation technique works by first vaporizing the substance with the lowest boiling point and
collect it through condensation. Then it vaporizes, condenses, and collects the second substance in
a different container. When the liquid is boiled, the pressure the builds up above the liquid reaches
dynamic equilibrium with the liquid, which is known as vapor pressure. At equilibrium the liquid
evaporates and it’s condensed back to its liquid state simultaneously (UC).
Besides the distillation technique, other concepts need for the experiment are Dalton’s Law,
Raoult’s Law, and azeotropes. Raoult’s Law states that the vapor pressure of the solvent above a
solution is equal to the vapor pressure of the solvent at the same temperature scaled by the mole
fraction of the solvent present (UC). Dalton’s Law states that the total pressure above a solution is
the sum of the partial pressure of each solvent and solute (Snelling, 2005). An azoetrope is a
mixture of liquids that has a constant boiling point and whose components cannot be separated by
simple distillation (Encyclopaedia). The mixture used for this experiment is not an azeotrope, since
the components of the mixture have different boiling points and can be separated using simple and
fractional distillation. The purpose of experiment 1 was to perform a single and fractional
distillation to separate two liquids, cyclohexane (Figure 1.) and toluene (Figure 2.) from a stock
mixture.
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II. OBJECTIVE:
a) To purify a compound by separating it from a non-volatile or less-volatile material.
b) To separate a mixture of two miscible liquids (liquids that mix in all proportions) with
different boiling points.
III. EQUIPMENT/ MATERIAL:
large test tubes (3)
test tube rack (1) ring stand
10-mL graduated cylinder glass adaptor
50- mL round bottom flask grease
clamp (1 or 2) rubber tubing (2)
heating mantle boiling chips
condenser (1 or 2) thermometer adaptor
thermometer 50- mL round bottom flask
Cyclohexane and toluene
Table of Chemicals Used:
Chemicals
Physical properties Cyclohexane Toluene
Molar mass 84.160 g/mol 92.138 g/mol
Melting point 6.74 °C -93.0 °C
Boiling point 81.0 °C 111 °C
Density 0.778 g/mL 0.867 g/mL
Vapor pressure 77.5 mmHg at 20°C 22.0 mmHg at 20 °C
Appearance Colorless liquid Colorless liquid
Viscosity 1.00 cP at 20 °C 0.590 cP at 20 °C
Freezing point 6.60°C 4.44°C
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Table 1. Physical properties of chemicals used.
Chemicals
Chemical properties Cyclohexane Toluene
Flammability Highly flammable Highly flammable
Enthalpy of formation -156 kJ/mol +12.0 kJ/mol
Can react with very strong acids Can react vigorously with
Reactivity such as the superacid system oxidizing materials.
HF + SbF5.
Solubility in water Immiscible 0.470 g/L
Heat of combustion -3.92×103 kJ/mol -3.92×103 kJ/mol
Table 2. Chemical properties of chemicals used.
DEFINITION OF TERMS
Simple Distillation - used frequently in the organic chemistry teaching labs, It is often
considered when:
a) the liquid is relatively pure to begin with (e.g., no more

than 10% liquid contaminants)
b) essentially a pure material is separated from a non-

volatile or from a solid contaminant
c) the liquid is contaminated by a liquid with a boiling

point that differs by at least 70°C Simple distillation involves a
single equilibration between the liquid and vapour. This
distillation is referred to as involving one theoretical plate.
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Fractional Distillation - The principle of fractional distillation is based on the
establishment of a large number of theoretical vaporization-
condensation cycles (theoretical plates): the apparatus of a simple
distillation is modified by inserting a fractionating column
between the distillation flask and the distillation head. The
fractionating column provides a large surface area in which the
initial distillate is redistilled and condensed again. This process
continues as th e vapors rise up the column until the vapors finally
make it into the condenser. These vapors and the final distillate
will contain a greater percentage of the lower boiling liquid.
Continuous repetition of redistillation process in fractional
distillation gives good separation of the volatile liquid
components.
Figure 1. Cyclohexane Figure 2. Toluene.
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Experimental Section:
Cyclohexane
Evaporate Cyclohexane
Vapor of Cyclohexane
+
81°C
Toluene
Evaporation Condensation
at 111°C
Liquid Cyclohexane
Vapor of Toluene
Condensation
Liquid Toluene
Diagram 1. Summary of Simple and Fractional Distillation.
PROCEDURE
The chemical used for this experiment was a stock mixture of cyclohexane and toluene at
a 1:1 ratio. In the lab there were two procedures performed.
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The first one was simple distillation. To begin the apparatus was assembled properly. After
it was assembled correctly, it was positioned correctly with the help of iron clamps.
The final set up for simple distillation was the sand bath at the bottom then the vial and the
distillation head which was connected with the thermometer and the condenser.
The condenser had water out, which had a hose that help the water go to the sink, and water
in, which was close to the receiver and was connected with a small hose to the tap water.
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The condenser was connected to the receiver which was a graduated cylinder that held the
liquid substance condensed.
The second procedure was the fractional distillation.
The final setup for the fractional distillation was the sand bath at the bottom, then the
distillation vial that held the stock solution. The vial was connected to the fractional column, which
contained aluminum mesh packing chips.
The fractional column was connected with the distillation head,
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Which is like in the simple distillation setup was connected with the thermometer and
condenser.
. The condenser had to be connected to the receiver, a graduated cylinder, and had to have
water in and water out. The water out was located closer to the receiver.
The total volume of stock solution was 7.00 mL containing equal amount of
cyclohexane and toluene. After obtaining the stock solution, two boiling stones were placed in the
vial. Then the condenser was tested to see if there was any water leakage. Once the water was
running and there was no leakage the vial with the stock solution was placed between the apparatus
and the sand bath. The sand bath was connected to a heat controller that regulated the temperature.
Then the sand bath was turned on to 50% power for fractional distillation and 30% for simple
distillation. It took approximately 15 minutes for the first drop of cyclohexane to be collected at
45°C for fractional distillation and 61°C for simple distillation. The vapor continued to condense
until the first substance was completely distilled.
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Then the receiving vial was changed to collect toluene liquid. The temperature was
increased by increasing the power to 70% for fractional distillation and 40% for simple distillation.
The first drop of toluene was collected at 46°C for fractional distillation and 76°C for simple
distillation. Then toluene was collected in a graduated cylinder. The temperature and each drop
were recorded through both procedures. Finally, the apparatus was disassembled and stored after
letting it cool.
SHOW THIS FOLLOWING DATA:
Table 3. Fractional Distillation data for Cyclohexane. Drops Temperature
Table 4. Fractional Distillation data for Toluene.
Graph 1. Fractional Distillation curve for cyclohexane and toluene. The triangles represent the
cyclohexane and the squares the toluene.
Calculate the percent recovery of both solvents for fractional distillation
Cyclohexane:
Toluene:
% Table 5. Simple distillation for Cyclohexane.

Drops Temperature
% Table 6. Simple Distialltion for Toluene.
Graph 2. Simple distillation curve for cyclohexane and toluene
Calculate the percent recovery of both solvents for simple distillation

Cyclohexane:
Toluene:
37
GUIDE QUESTIONS
1. Name one piece of apparatus that is not used in a simple distillation but is used in a
fractional distillation.
Answer:
2. What is the purpose of a condenser? How does a condenser operate?
Answer:
3. What is a boiling chip? How does a boiling chip regulate boiling? When should a
boiling chip be used? 4. Why should the bulb of the thermometer be placed below the
side arm of the three-way adapter?
Answer:
4. What type of glass is used in the laboratory? What property of this glass is useful for
laboratory work?
Answer:
5. The temperature in the pot (the round bottom flask) and the head (the three-way
adapter) are different during a distillation of a binary mixture. Is this correct? Why?
Answer:
6. Briefly define the following terms.
ANSWERS:
a) Bumping
b) Reflox
c) Theoretical plate
d) Raoult’s Law
e) Dalton’s Law
7. During the distillation of a mixture of two miscible liquids with very different
boiling points, the temperature rises to a constant reading and then drops off before
returning to a higher reading. Explain the likely cause of this fluctuation in temperature
readings.
38
ANSWER:
8. What effect does (a) decreasing the pressure or (b) increasing the pressure
above a liquid have on the liquid’s boiling point?
ANSWERS:
9. Explain why flooding the fractionating column will lead to poor separation in a
fractional distillation.
ANSWER:
10. Why should a distilling flask be filled not less than 1/3 filled or more than 2/3
full?
ANSWER:
REFERENCES
Encyclopædia Britannica. http://www.britannica.com/EBchecked/topic/46765/azeotrope

(accessed May 2013).
Organic Chemistry at CU Boulder.

http://orgchem.colorado.edu/Technique/Procedures/Distillation/Distillation.html
Snelling, C.R. Distillation-Simple vs. Fractional.
http://www2.volstate.edu/chem/2010/Labs/Distillation.html (accessed January 2014).
UC Davis ChemWiki.
http://chemwiki.ucdavis.edu/Physical_Chemistry/Physical_Properties_of_Matter/So
lutions/Changes_In_Vapor_Pressure,_Raoult%27s_Law (accessed January 2014).
Weldegirma, S. Experimental Organic Chemistry; Cengage Learning: Ohio, 2012; p 10.
VIDEO REFERENCES
https://youtu.be/sLom1F_1K1Y
39
EXTRACTION: DETERMINATION OF ITS EFFICIENCY; CALCULATION OF THE

DISTRIBUTION COEFFICIENT
40
I. INTRODUCTION
Extraction is a quick way to purify the product(s) of a reaction Most organic compounds
are much more soluble in organic solvents (ether, dichloromethane, etc.) than they are in water.
This is because of the “like dissolves like” rule. Likewise, most salts are much more soluble in
water than they are in organic solvents. But just like oil and water will not mix together, most
organic solvents will form layers when mixed with water. Two liquids that can mix together are
said to be miscible. Most organic solvents are immiscible with water. We can use these facts to
our advantage to purify the products of a reaction.
Inside the laboratory, it is essential to ensure that the glassware that will be used is free
from materials that are capable of altering heavily the desired outcome of the experiment. In order
to prevent that from occurring, the glassware is rinsed with running tap or distilled water and
detergent. However, doing so only makes it clean enough for general use because few organic
materials still stick to the surface of the glass – that is why Acetone is often present in the
laboratory; it is capable of removing organic residues in glassware, and of quickly drying the newly
washed glassware. In line with this, there are a number of specific laboratory techniques which
aim to eliminate unnecessary substances from the sample. One separation process done in the
laboratory aims to isolate one or more components from a mixture by separating it from those that
are unnecessary – and it is known as extraction. This method comprises five types: liquid-liquid
extraction, solid-phase extraction, solid-phase microextraction, Soxhlet extraction, fizzy
extraction. This experiment is focused on liquid-liquid extraction or solvent extraction. Since
extractions are often performed using aqueous solutions, it is important to note the behaviour of
the solvent for it projects the miscibility of the solvent. Moreover, oxygen containing solvents are
usually more soluble in water (and vice versa) because of their ability to act as hydrogen bond
donor and hydrogen bond acceptor. Other solvents such as alcohols increase the solubility of water
41
in organic layers significantly because those are miscible with both phases and act as a mediator
which leads to the formation of emulsions.
II. OBJECTIVES:
a) To purify samples of organic compounds that are solids at room temperature.

b) To dissociate the impure sample in the minimum amount of an appropriate hot
solvent.
III. EQUIPMENT /
MATERIALS:
Benzoic acid solution ring& ring stand burette burette clamp
spatula 0.1 M NaOH (or 0.02 M) solution
separatory funnel methylene dichloride 125 ml Erlemeyer flask
10 and 50 mL graduated cylinders 50 and 100 mL beakers funnel
IV. PROCEDURE
The procedure in this experiment involves the use of the separatory funnel. It is important that
you learn how to use this piece of equipment properly, for an efficient separation and for safety. It
is made of thin glass and is easily broken unless handled carefully. Unfortunately, in various
student manuals you will find descriptions of about as many ways of holding the separatory funnel
as for holding a pair of chopsticks. Probably for you there is some best method, depending on the
size of your hands, the strength of your fingers, your manual dexterity, and the size and shape of
the funnel. The following are important rules to observe.
1. Hold the funnel firmly but gently in both hands so that it can be turned from the vertical
to horizontal direction and back again easily and can be shaken vigorously while
observing (2) and (3).
2. Keep the stopper tightly seated with one hand at all times, using the forefinger of that
hand, the base of the forefinger, or the palm of the hand.
42
3. Keep the stopcock tightly seated with the fingers of the other hand in such a way that the
fingers can open and close the stopcock quickly to release the pressure that may be built
up from solvent vapor or
evolved gases.
The use of the separatory funnel is a skill and is best learned by practice with an
empty funnel while watching your instructor demonstrate the technique. In the figure below
are shown two slightly different methods of handling the separatory funnel. In the first
method the stem of the funnel projects between the thumb and first finger of the left hand
(for a right-handed person). The stopcock is held in place and operated with the thumb and
first finger. The stopper is kept in place by pressure against the base of the first finger of
the right hand.
In the second method the stem of the funnel projects between the first and second fingers
of the left hand. The stopcock is held in place by the pressure from these fingers and is operated
by them in conjunction with the thumb. The stopper is held in place by pressure against the middle
of the palm of the right hand. (Figure 1)
shake
shake
Fig. 1- methods for holding and shaking the separatory funnel. Fig. 2- support and use of the
separatory funnel.
Support the separatory funnel in a ring on ring stand. Close the stopcock and add the liquids to the
funnel to be separated. Insert the stopper, and immediately invert the funnel. Point the barrel
away from your face and that of your neighbors. Open the stopcock to release the pressure ,
43
which may have accumulated inside the funnel (volatile solvents such as ether develop
considerable pressure).
Close the stopcock and, holding the funnel horizontally, shake the funnel two or three times. Invert
the funnel and release the pressure as before. Repeat this process until opening the stopcock causes
no further pressure release. Close the stopcock and shake the funnel 15-20 times. Replace the
funnel in the holder (ring on ring stand) and remove the stopper (Fig. 2). Allow the liquids to stand
until the layers have completely separated. Draw the lower layer into a flask or beaker of proper
size (see figure below). Do not draw the liquid through the stopcock too rapidly. Slow the flow
carefully as the boundary between the two layers approaches the stopcock. Stop the flow of liquid
completely just as the upper layer enters the hole in the stopcock. Pour the upper layer through the
neck of the funnel into a second flask. Never discard either layer until you are absolutely certain
which is the proper layer to keep. Usually one layer will be an aqueous layer or solution, and the
other will be an organic liquid. The one of greater density will be on the bottom.
To check the identity of a layer, should you be in doubt, withdraw a few milliliters of the lower
layer into a test tube containing an equal volume of water. If the lower layer in the separatory
funnel is water or an aqueous solution it will be homogeneous (only one layer). If the layer being
tested is the organic layer, the sample withdrawn will fall to the bottom of your test tube and also
form two liquid layers. In either event, return the test mixture to the separatory funnel.
Part 1 – Standardization of NaOH solution
Use a 10 mL graduated cylinder measure 10.0 ml of the acid solution and transfer the
solution to a 125 mL Erlenmeyer flask.
44
Add 2-3 drops of phenolphthalein
and titrate to the end point (light pink) with a standardized (≈ 0.1M or 0.02 M) sodium
hydroxide solution.
Record in report form the number of milliliters of base required to neutralize this volume
of acid solution. Calculate the molarity of the NaOH. Discard the neutralized acid solution and
rinse your flasks. REPEAT. (TWO TRIALS)
45
Part 2 – Single extraction
Use a 50 mL graduated cylinder to measure out a second 50.0 ml volume of acid solution
and transfer it to your separatory funnel.
Add 10 ml of methylene dichloride, CH2 Cl 2, to the funnel and extract according to the
procedure outlined in the part 1 of this experiment.
shake
shake
46
Separate the bottom layer (organic phase) in a 100 mL beaker and collect the top
layer(aqueous) into a 125 mL Erlenmeyer flask and add 2-3 drops of indicator.
Record the volume of the sodium hydroxide solution in the burette and titrate to the
phenolphthalein end point (light pink).
47
Again record the number of milliliters of base required and calculate a-g.
Discard the neutralized acid solution and the methylene dichloride layer into the large
bottle marked "Organic Waste".
NOTE: This lab has been modified in that methylene dichloride is now used in place of ether as
the organic phase. This avoids the problem of ether fumes and explosions. However, the
extraction with methylene dichloride is not as clean because methylene dichloride is more
miscible in water than ether. As a result, you will find that your aqueous layer is cloudy after
extraction. You can still titrate the aqueous layer to a light pink endpoint.
48
Part 3 – Multi extraction
Repeat the procedure from Step 2,
But this time extract 50 ml of fresh acid solution with two 5 ml portions of methylene
dichloride.
Separate the aqueous layer into a flask
and dispose of the organic layer. Transfer the aqueous layer back into the empty, cleaned,
separatory funnel and extract it with a second 5 ml portion of fresh methylene dichloride.
Separate the extracted aqueous layer, add indicator as before and titrate to the end point. Record
the volume of standard base required and calculate a-f. Dispose of the organic layer extracts as
directed and clean your separatory funnel.
49
V. GUIDE QUESTIONS
1. For most organic compounds would you expect the value of Kd to be greater than or less
than one?
ANSWER:
2. Both benzoic acid and ethyl benzoate are soluble in organic solvents. How can you use
extraction to separate them?
ANSWER:
3. The distribution coefficient, K, 10 for compound Y. What weight of compound Y

would be removed from a solution of 4.0 g of Y in 100 ml water by a single extraction
with 100 ml of ether?
ANSWER:
4. Reconsider question 3 above; what weight of compound Y would be removed by two

(double) extractions using 50 ml of ether each time?
ANSWER:
5. Student during an extraction experiment lost track of which layer is the

aqueous layer. How Could student determine which layer is which by a
simple test?
ANSWER:
6. Why must the stopper be removed from the separatory funnel before
the lower layer is removed.
ANSWER:
7. An organic compound can be extracted from a water layer by an organic solvent more
efficiently if the water layer is saturated with an inorganic salt such as sodium chloride.
This effect, called “salting out”, increase the participation coefficient in favor of the
organic compound. (explain)
ANSWER:
50
8. What are the advantages and disadvantages of using ether as a solvent for the
extraction of organic compounds?
ANSWER:
9. What volume of an organic solvent must be used to effect 90% separation in one
extraction when only 2.7 g of a certain compound dissolves in 100 ml of water? (K=
15)
ANSWER:
10. The pain reliever phenacetin is soluble in cold water to an extent of 1.0
g /1310 ml and soluble in diethyl ether to an exient of 1.0 g /90ml.
a. Determine the approximate distribution coefficient for phenacetin in those two

solvents.
b. If 150 g of phenacetin were dissolved in 100 ml of water, how much ether

would be required to extract 90% of phenacetin in a single extraction?
IV. RESULT
Table 1. Results of Three Different Methods of Extraction
Flask 1 Flask 2 Flask 3

(20-mL aliquot) (One 20-mL (Two 10-mL portions of
portion of Toluene)
Toluene)
Initial burette
reading (mL)
Final burette
reading (mL)
Total amount
NaOH
delivered (mL)
51
Molarity of
NaOH (M)
Moles Acetic
Acid
neutralized [1]
Grams Acetic
Acid
neutralized [2]
Upper layer
after shaking
Lower layer
after shaking
Grams Acetic
Acid originally
present in
20mL before
extraction [3]
Grams Acetic
Acid extracted
in Toluene [4]
Percent Acetic
Acid extracted
[5]
Distribution
coefficient, Kd
[6]
Equations used:
[1] mol Acetic Acid =
[2] g Acetic Acid neutralized =
52
[3] g Acetic Acid original =
[4] g Acetic Acid extracted =
[5] % Acetic Acid extracted =
[6] Kd, Distribution coefficient =
V. REFERENCES
“Extraction” (2013, December 30). Extraction in theory. Retrieved from

http://www.chem.ucla.edu/
Pahlavan B. Dr. (2015). CHEM 2423 extraction of benzoic acid. Retrieved from
http://swc2.hccs.edu/pahlavan/2423L6.pdf
VIDEO REFERENCES:
https://www.youtube.com/watch?v=Dz51O8LHA4g
https://www.youtube.com/watch?v=HwKFcjGHN_4
53
PREPARATION OF SYNTHETIC FOOD FLAVORS
54
I. INTRODUCTION
Esters, were a type of organic compound that were responsible for the fragrance and flavor of
flowers and fruits. For example, were the isopentyl acetate in bananas, methyl salicylate in wintergreen,
and ethyl butyrate in pineapples. Thus, esters were also commonly used in the production of synthetic
flavors, perfumes, and cosmetics. Moreover, other volatile esters were also used as solvents for lacquers,
paints, and varnishes. It is commonly prepared by a simple reaction of carboxylic acids and alcohol in the
presence of a catalyst usually hydrochloric acid or sulfuric acid in a process called Fisher Esterification.
Figure 1. General reaction of esterification

The general reaction esterification involved the substitution reaction of the –OH substituent in the
carboxylic acid and of the Alkyl group in the alcohol to form ester and water as shown in figure 1. The
presence of sulfuric acid allowed the reaction to go faster since the Fisher-Esterification reaction had a slow
rate of reaction at room temperature. In addition to this, it was also refluxed to increase the rate of reaction
in a short span of time. This allowed the formation of ester faster without the loss of its volatile reactants
and prouct. An example of the Fisher Esterification reaction was shown below.
Figure 2. An example reaction of an alcohol and a carboxylic acid to form an ester.

However, the esterification reaction is reversible thus one of the reactants should in excess to ensure
the reaction favors the product formation. On the other hand, the formation of water in this reaction allowed
the reaction of the formed ester back to carboxylic acid. This could have resulted to low percentage yield
of the desired ester. Thus, it was noted that the reaction should as much as possible favor the production of
the ester. It is then distilled to remove the ester from the unwanted by-products which would resulted to a
purer ester
55
II. MATERIAL
NOTE: The organic liquids used are very flammable. Do not use an open flame to
heat samples. Hot plates will be provided.
III. PROCEDURE
The assigned alcohol was methyl alcohol and the assigned carboxylic acid was butyric acid. Based on the
general reaction of esterification the ester product was Methyl butyrate with a given reaction below.
Figure 3. Reaction of butyric acid and methyl alcohol to form methyl butyrate
First was that the amount of butyric acid and methanol based on their respective moles, 0.50 for
the butyric acid and 0.65 moles of the methyl alcohol, and their
densities. The densities of methanol and butyric acid were 0.791 g/mL
and 0.964 g/mL respectively. Their volume was then measured using
two different graduated cylinder to minimize error. The same was also
done to measure the amount of concentrated sulfuric acid needed in
the experiment. Second, the theoretical yield of methyl butyrate was
calculated based on the limiting reactant present which was the
carboxylic acid and later the % yield of the ester was calculated
56
Next, in a 125-mL flat bottom flask, 0.65 moles (26.33 mL) of methyl alcohol was mixed with 0.50
moles (45.70 mL) of butyric acid and 8 g (4.35 mL) of concentrated sulfuric acid.
The addition of concentrated sulfuric acid was done inside the fume hood for safety purposes.
Meanwhile, the reflux set-up was prepared carefully. The general simple setup of the reflux system was
shown below. It was securely fastened at the iron stand with the iron ring, Bunsen burner, and the wire
gauze secured at the bottom of the flask.
Figure 4. Reflux Set-up
57
After mixing the reactants and the acid catalyst, the flat bottom flask was placed in the fully
prepared reflux set-up. After the go signal was given by the instructor, the reflux was done for 30 minutes.
Next, after 30 minutes the reflux set-up was rearranged for distillation set-up.
Figure 5. The simple distillation set-up
The thermometer’s bulb was placed near the pathway towards the condenser. The distillation
started when the approval of the instructor was given. The temperature reading in the thermometer was
noted and critically monitored in order to avoid the distillation of alcohol in the solution. Once 10 drops of
the distillate were produced it was tested with 1 mL of water to correctly identify if it is ester if a layer in
the water was observed to form. Once the distillate is validated to be an ester, the temperature reading of
the thermometer was manually controlled in order for the temperature to remain in the approved
temperature. This allowed the distillation of the desired ester. During this time, the color of the ester was
observed alongside with its odor. After all of the available ester was distilled, it was checked for its pH
using litmus paper and was neutralized with 6M of NaOH. Once neutralized the formed ester was checked
for purity. The purity was checked using 5 drops of ester and mixed with 5 drops of potassium dichromate
and 2 drops of concentrated sulfuric acid in a warm bath. Once the color of the sample changed after
sometime the formed ester contains impurities otherwise if not.
58
IV. GUIDE QUESTION:
1. How does Schotten-Baumann reaction differ from Fisher reaction? Write a
mechanism for the former.
a. ANSWER:
2. What is the function of the sulfuric acid in this reaction? Is it consumed in the reaction?
ANSWER:
3. What is the purpose of adding 5% sodium bicarbonate to the oil?
ANSWER:
4. Why was 5% sodium bicarbonate used in the extraction? What would have happened if 5%
sodium hydroxide had been used? What do you call this undesirable reaction?
ANSWER:
5. Give the purpose of washing the organic layer with saturated sodium chloride.
ANSWER:
6. Enumerate some of the common drying agents and identify how each can be advantageous to
the others.
ANSWERS:
7. Rank the following according to ease of esterification with methanol: acetic acid, carbolic
acid, benzoic acid, and salicylic acid.
ANSWER:
59
DATA:
Reactant Volume used Moles Density
Alcohol (Methyl
Alcohol/ Methanol)
Carboxylic Acid (Butyric
Acid)
Theoretical Yield
Volume of the distilled
ester
% Yield of ester
Table 1. Summary of the calculations in the experiment
V. REFERENCES
Carey, F.A. (2008). Carboxylic Acid Derivatives: Nucleophilic Acyl Substition (7 th Ed.) In Organic
Chemistry (pp. 842 - 844). New York, NY: McGraw-Hill
Drying Agents. (2016). Retrieved from
http://www.chem.ucla.edu/~bacher/Specialtopics/Drying%20Agents.html
Fischer Esterification (n.d) Retrieved from https://www.name-reaction.com/fischer-esterification
Encylopaedia Britannica (2015, July 07). Ester. Retrieved October 11, 2019 from
https://www.britannica.com/science/ester-chemical-compound
Othmer, K. (n. d.). Esterification. Encyclopedia of chemical technology, 4(9). Retrieved on
February 4, 2019 from %20documents/ Esterfication.pdf
Preparation of Esters (2019, June 5). By Chemistry LibreTexts from chem.libretexts.org accessed on
October 11, 2019
https://chem.libretexts.org/Courses/Eastern_Mennonite_University/EMU%3A_Chemistry_for_th
e_Life_Sciences_(Cessna)/15%3A_Organic_Acids_and_Bases_and_Some_of_Their_Derivatives/15.08_
Preparation_of_Esters
Schotten-Baumann reaction. (n.d.). Retrieved from https://www.name-reaction.com/schotten-
baumann-reaction
Synthesis of esters: natural and artificial flavourings (n.d) from

https://www.swisseduc.ch/immersion/chem/synth/docs/Synthesis_ofEsters.pdf
60
Tremble, J. (2014). Fischer Esterification. Retrieved from https://www.odinity.com/fischer-
esterification/
Williams, M. (2017). The Effects of Washing the Organic Layer With Sodium Carbonate.
Retrieved on February 4, 2019 from https://sciencing.com/the-effects-of-washing-the-organic-
layer-with-sodium-carbonate-12263030.html
VIDEO REFERENCES:
https://youtu.be/R6UugHqpu2Y
https://www.youtube.com/watch?v=6VbNyFuYQMU
https://www.youtube.com/watch?v=AILt5HAYJTs
61
EXPERIMENT NO. 6
ISOLATION OF CAFFEINE FROM TEA LEAVES
62
I. Introduction:
Caffeine is an alkaloid stimulant with a cyclic backbone structure
analogous to the purine structures of DNA, giving it the ability to affect
biochemical pathways in the body 1 . In commercial application, caffeine
supplements pharmaceuticals and certain beverages such as coffee or
tea. Standard tea bags contain 2.00 +/- 0.05 g of tea leaves along with
approximately 55 mg of caffeine. Using the proper extraction methods, the
caffeine within a tea bag could potentially be isolated to yield a pure solid; the
mass of this solid would reflect the actual yield of caffeine in the tea. To do so,
caffeine must be introduced to a solvent that is both volatile and insoluble to
water; a perfect example is methylene chloride.
Caffeine has a greater affinity for methylene chloride and will easily
dissolve in this solvent over water; however caffeine is not the only organic
substance found in tea that is capable of reacting with methylene chloride. Along
with caffeine, tea bags contain organic substances called tannins, or gallic
acid 1 . Both caffeine and gallic acid are capable of dissolving in water; howe ver,
caffeine has a stronger attraction to water due to the dipole -dipole interaction that
results from the greater polarity of caffeine and the hydrogen bonds that form
between caffeine and water 1 . Theoretically, the intermolecular forces of gallic
acid can be manipulated to induce a stronger dipole-ion interaction. If a common
salt like sodium carbonate was introduced to the solution, gallic acid could revert
back into phenol salt: a polar, inorganic molecule that is insoluble in methylene
chloride.
In methylene chloride, caffeine will have a greater attraction for the organic
solvent and the hydrogen bonds between caffeine and water will be broken. Using
a separatory apparatus, two insoluble solutions can be separated, isolating caffeine
and the new phenol anion from one another. The denser methylene chloride layer
can then be released from the funnel to render a pure solution of caffeine and
63
methylene chloride. To ensure that no water interferes with the interaction of
caffeine and methylene chloride, sodium sulfate could be used to absorb any
excess water that may have escaped from the tea solution 1 . If heated, the solvent
would quickly evaporate due to low boiling point of methylene chloride 2 . The
remaining solid would then be pure caffeine.
II. OBJECTIVE
In this experiment, caffeine will be extracted from tea leaves (where it is about 5%
present) using hot water. This is essentially the same procedure used to decaffeinate drinks such
as coffee and tea.
III. MATERIALS AND INSTRUMENTS:
 300 ml of prepared tea/caffeine

 1.0g sodium carbonate monohydrate
 15 ml plus 10ml of dichloromethane (DCM)
 Separatory funnel
 0.75 g sodium anhydrous
 Roto evaporator
 Sublimation apparatus as set up in Portland State Orgchem laboratory Manual
Experiment 5.
 Hot plate
IV. PROCEDURE
Add about 10.0 g of tea leaves to around 100 mL of water in a 400-mL beaker
(cut open the tea bags with some scissors to get at the tea leaves). Add 5 g of calcium
carbonate
64
Boil this suspension for about 10 minutes. The caffeine will dissolve in the hot
water, but so will some other compounds, known as tannins (a type of carboxylic acid).
The calcium carbonate should convert these tannins into insoluble salts, which
will then drop out of solution. Add 3 g of filter acid (celite) and decant the mixture into a
Buchner funnel (decanting means to pour the liquid, but try to leave the solid behind).
Any solid caught by the filter paper can be discarded. The caffeine is located in the
filtrate (the liquid that passed through the filter).
You will probably have around 50 mL of filtrate. Less than that is fine, but if you
have more, boil it down to about 50 mL. Cool the filtrate to room temperature
65
Pour it into a 125-mL separatory funnel. Add 10 mL of methylene chloride to
the funnel and shake gently (don’t shake it too vigorously or it will form a foam that is
hard to get rid of). Gases will be produced, so it is important to vent the mixture
periodically by taking the cap off (otherwise the pressure will eventually cause it to shoot
off). The caffeine is more soluble in the nonpolar methylene chloride than the water, and
as a result, will move to the methylene chloride layer (known as the organic layer). Since
the organic layer is denser than water, it will be the layer on the bottom.
Open the stopcock on the funnel and pour the organic layer into another beaker.
Do not let any of water layer (which is brown) get transferred. If you do so by mistake, it
is best to pour everything back in the funnel and try again. Sometimes it is necessary to
leave a little bit of the organic layer behind to ensure that you do not collect any water.
66
Add 10 mL of fresh methylene chloride to the funnel (with the brown water) and
once again shake gently with venting. This is just giving another chance for any caffeine
left behind in the water layer to migrate to the organic layer. Once again, open the
stopcock to allow the organic layer out (combine it directly with the other organic layer)
but do not collect any of the brown water layer. The caffeine should be pure at this stage,
but it is still dissolved in the methylene chloride.
You will need to evaporate this in a machine called the N-Evap. Your instructor
will show you how to use this machine. By applying nitrogen gas in a warm water bath,
the methylene chloride will CHEM& 131 Caffeine Lab 2 quickly evaporate away,
leaving only the caffeine behind. Remember, though, that you cannot evaporate the
organic layer unless there is no visible water. (If you see a brown droplet, try to extract it
with a Pasteur pipette). The final caffeine should be white. Occasionally, it has a greenish
tinge, which is fine. Any yellow or brown color is a sign of impurities.
67
V. DISCUSSION AND RESULTS AND ILLUSTRATION:
Figure 1. Molecular structure of caffeine

(http://commons.wikimedia.org/wiki/File:Caffeine.svg)
Figure 2. Sublimation apparatus (http://www.geocities.ws/dcarrd2000/introduction1.html)

Table 1: Melting point and mass of isolated caffeine sample
Mass of caffeine recovered
Melting Point of purified sample
Table 2. Bonds present in Patrick’s purified sample of caffeine

Bonds CH3-N C=O C=N C-N C=C
68
Table 3. Bonds present in a 100% purified sample of caffeine
Bonds CH3-N C=O C=N C-N C=C
VI. GUIDE QUESTION
1. What are the health and safety hazards associated with dichloromethane used in this lab?
ANSWER:
2. Why did we extract the energy drink with dichloromethane, specifically, and not another
organic solvent such as methanol or hexane?
ANSWER:
3. What was the purpose of using Na2SO4 in the process of extraction?
ANSWER:
4. A student during an extraction experiment lost track of which layer is the aqueous layer.
How could the student determine which layer is which by a simple test? Assume densities
of the solvents are not known.
ANSWER:
5. Why must the stopper be removed when the separatory funnel is sitting on a ring stand
waiting for the layers to solvents to separate?
ANSWER:
6. Why must the stopper be removed from the separatory funnel before the lower layer is
drained?
ANSWER:
7. Would water and hexane (CH3CH2CH2CH2CH2CH3) mix together to form a

homogeneous solution? If not which one would be the upper layer and why?
69
ANSWER:
8. Define the following:
(a) Decant
(b) Tannins
(c) Filtrate
9. How much methylene chloride should you collect in total from the separtory funnel?
ANSWER:
10. How will you know if your final caffeine is pure?
ANSWER:
11. What is the purpose of the calcium carbonate in this experiment?
ANSWER:
12. Look up the structure of caffeine and determine the strongest intermolecular force
present.
ANSWER:
13. Why is caffeine more soluble in methylene chloride than in water?
ANSWER:
14. Why is it possible to sublime caffeine? List another compound that sublimes near
ambient pressure.
ANSWER:
15. What other compounds are extracted along with caffeine into the tea? Use what you
know about the relative polarity of the functional groups within these compounds to
determine the final destination of the following compounds: cellulose, tannins,
chlorophylls, calcium carbonate. Do the majority of these compounds end up in the tea
bag, filtered solid, aqueous layer, or organic layer?
70
ANSWER:
VII. References:
Organic Chemistry Laboratory 1, CH 337M; Department of Chemistry, Portland State
University: Portland OR; Exp 5
Organic Chemistry Appendix, Department of Chemistry, Portland State University: Portland OR

http://commons.wikimedia.org/wiki/File:Caffeine.svg
http://webbook.nist.gov/cgi/cbook.cgi?ID=C58082&Units=SI&Mask=80#IR-Spec
VIDEO REFERENCES:
https://youtu.be/SjNbGUqfnRs
https://www.youtube.com/watch?v=sPhJWBL17OQ&t=7s
71
EXPERIMENT NO. 7
THIN LAYER CHROMATOGRAPHY OF ANALGESIC DRUGS
72
I. INTRODUCTION
Chromatography is an analytical technique that is often used in order to separate a

mixture of two or more compounds, or ions, into their separate components. Moreover,
these components are separated so that their properties can be more thoroughly analyzed1.
There are three different types of chromatography often used throughout organic
chemistry, Thin Layer Chromatography (TLC), Gas Liquid Chromatography (GC) and
Column Chromatography (CC). However, this experiment specifically focuses on the use
of TLC and CC in order to separate a mixture.
Thin Layer Chromatography (TLC) uses a plastic plate coated with a thin layer of
silica to quickly and inexpensively analyze small samples. A small dot of the mixture is
applied to the baseline of the plate; the plate is then dipped in the appropriate solvent and
is placed in a container. The solvent moves up the plate via capillary action, meets the dot
of the mixture on the baseline, the mixture is dissolved and is then carried up the plate by
the solvent. The components in the mixture can be identified because different compounds
travel at different rates due to the differences in the solvent’s solubility and differences in
the compounds attraction to the stationary phase. TLC uses R -values to represent the
distance travelled by the compound divided by the distance travelled by the solvent. It is
these values that can then be compared to analyze the different compounds.
Column Chromatography (CC) is similar to TLC in terms of separating compounds.

However, CC uses a column to separate and isolate compounds, rather than analyze
different compounds. In this experiment, CC uses a micro column packed with silica gel
and sand to separate the pigments in spinach leaves into their respective components. Then,
the approximate volume collected and the colour of each component collected is recorded.
The separation that occurs during chromatography is caused by the respective

components molecular interactions with two phases, a mobile phase and a stationary phase.
The mobile phase is a medium used in chromatography that has the ability to move through
the stationary phase. Moreover, in TLC and CC the mobile phase is most often an organic
liquid. Whereas, the stationary phase is a medium used in chromatography that does not
move. In this experiment, the stationary phase is the thin layer of silica that is on top of
the plastic plate. It is this ability to partition into separate components that allows for the
73
mixture to be separated and analyzed. Moreover, this ability to partition is based on the
polarity and affinity between the two different phases.
II. OBJECTIVE
In this experiment was to analyze four analgesics: Acetaminophen, Aspirin,
Caffeine and Ibuprofen; by Thin Layer Chromatography and identify an unknown
analgesic by TLC comparison.
Furthermore, a secondary objective of this experiment was to isolate -
carotene via liquid-solid extraction of spinach and purify this compound via column
chromatography.
III. MATERIAL AND EQUIPMENT

 CHEMICAL USE
Name of Chemical Acetaminophen Aspirin

IUPAC Name N-(4-Hydroxyphenyl)acetamide 2-(Acetyloxy)benzoic acid
Formula C8H9NO2 C9H8O4
Molar Mass 151.17 g/mol 180.15 g/mol
Melting Point 170 C 139 C
Skin/eye irritant Skin/ eye irritant

Properties
Irritant if inhaled Irritant if inhaled
Chemical Structure
Name of Caffeine Ibuprofen

Chemical
IUPAC Name 1,3,7-trimethyl-3,7-dihydro- 2-(4-isobutylphenyl)proponic acid
1H-purine
74
Formula C8H10N4O2 C13H18O2
Properties Skin/eye irritant Skin/eye irritant
Chemical
Structure
Name of Chemical Silica Gel Ethyl Acetate

IUPAC Name Silicon Dioxide Ethyl Ethanoate
Formula SiO2 C4H8O2
Melting Point 1600 C -83 C
Boiling Point 77 C
Flammable
Chemical Structure
Name of Chemical Sand Sodium Sulfate

IUPAC Name Silicon Dioxide Sodium Sulfate
Formula SiO2 Na2SO4
75
Boiling Point 1100 C
Irritant if inhaled
Name of Chemical Petroleum Ether Hexane

IUPAC Name Various Hexane
Formula Various C6H14
Molar Mass Various 86.18 g/mol
Melting Point -95 C
Boiling Point 60 C 68 C
Flammable Flammable
 MATERIALS
Pencil Glass Capillary Tube
Colour test plates Jar or Beaker
Spot plates
TLC developing tanks
TLC plates
TLC sample recovery tubes
TLC sprayers
and TLC storage product
Spotter paper
76
IV. PROCEDURE
Part 1:
For the TLC of amino acids, a pencil was used (to prevent ink running through the plate) to
mark 5 equidistant x’s along a 1cm horizontal line from the base of a TLC plate.
5 clean spotters were used to spot amino acid mixtures (glycine, arginine, valine, glutamic acid
and an unknown mixture E) onto the plate.
A developing tank with a solution of n-butanol: ethanoic acid: water (in the ratio 3:1:1) was
prepared.
77
After the spots on the TLC plate had dried, it was put in the tank while ensuring the solution
did not cross the 1cm base-line and a watch glass was used as a lid to allow the atmosphere in
the tank to equilibrate. ‘
The apparatus was left undisturbed for approximately 1 hour to allow the solvent to run up the
plate, until it was around 1cm from the top. After the plate was removed from the solution, the
solvent front was marked and the plate was dried using a hairdryer. Since amino acids are
colourless, a thin layer of anninhydrin was sprayed over the plate as a stain, then baked in a
100 C oven for 5 minutes, in order to observe the component spots on the TLC plate.
78
Part 2
For the investigation on analgesics, a TLC plate was prepared as in part 1.
Approximately 0.1g of powdered aspirin was measured directly into a micro test tube. Other
analgesics and the unknown compound B were also roughly measured into a micro test tube.
A 50: 50 ethanol/ dichloromethane solution (2cm 3) was added to the test tubes and mixed for
a few minutes to allow the active ingredients to dissolve.
79
The TLC plate was spotted and left to develop as the previous experiment, but in a small
volume of ethyl acetate. Once the solvent front reached approximately 1cm from the top of
the plate, it was removed from the solution and the solvent front was marked and the plate
was dried. The plate was placed under a UV lamp so the position of the spots could be
observed.
During both procedures, precautions of contaminating the TLC plates were taken for it to run
successfully. These included using tweezers to handle the plates, clean apparatus to be used at
all times and one spotter to be used per sample. The apparatus used in the amino acid
experiment was rinsed with water however, the apparatus used in the analgesic investigation
had to be emptied into the non-halogenated waste bottle and then rinsed with acetone.
80
V. GUIDE QUESTION
1. Each scenario below describes an operational error that can lead to

inaccurate or irreproducible TLC results. In each case describe the effect on
the TLC results and briefly explain why.
a. A pen was used to label the origin of the TLC plate.

ANSWER:
b. A lab worker frequently removed the developing chamber cover in order to

better see the position of the solvent front during development of the plate.
ANSWER:
c. Samples were applied to a TLC plate in spots that were 6 mm in diameter’

ANSWER:
d. Spots were applied to a TLC plate so that they were exactly even with the
solvent level in the developing chamber.
ANSWER:
2. After a rather lengthy organic chemistry synthesis procedure, a student

analyzed the product of the reaction on a TLC plate and obtained the
result at right. What might he/she have done wrong, if anything? If you
conclude that a mistake was made, provide an explanation as to why it
leads to the observed results.
ANSWER:
3. Explain the terms “stationary phase” and “mobile phase”. What is the mobile
phase used for this experiment?
ANSWER:
4. Silica gel was used as the solid support for the TLC plate. Which of
the three compounds (A, B, or C) is the least polar?
81
ANSWER:
5. What is an Rf value and how is it calculated?

ANSWER:
6. Which compound below will have the highest Rf on a TLC plate? Explain your
reasoning.
a.
ANSWER:
b.
ANSWER:
82
7. The following TLC plates have been spotted with two different compounds and
are about to be eluted. For each plate, explain is it has been spotted correctly or
incorrectly and why.
ANSWER:
8. Development and visualization of a TLC plate showed a single spot at the origin.
The lab worker concluded that the sample consisted of a single pure compound.
Was this conclusion justified? Why or why not?
ANSWER:
9. Arrange the following four compounds in order of increasing Rf when using

TLC plates similar to those you used. Place lowest Rf on the left and highest Rf
on the right. Explain how you arrived at your choices for full credit.
ANSWER:
83
10. Enter the requested data in the following
table.
Distance Front Distance
Traveled, mm Component
Sample Rf Value (show math)
Traveled, mm
Rf =
Aspirin
Rf =
Acetaminophen
Rf =
Ibuprofen
Rf =
Salicylamide
Rf =
Caffeine
1_______ Rf =
Unknown Tablet 2_______ Rf =
3_______ Rf =
84
VI. REFERENCES
Khan Academny. Principles of Chromatography. Khan Academy. (accessed Feb 9,
2017)
Wildegirma, S. Experimental Organic Chemistry Lab Manual; University of South
Florida: Tampa, FL, 2016; P. 15-19
ChemGuide. Column Chromatography. ChemGuide. ChemGuide. (accessed Feb 9,
2017)
LCGC Editors. (2009, November 10) Thin Layer Chromatography. LCGC. LCGC .
(accessed Feb 9, 2017)
VIDEO REFERENCES:
https://youtu.be/qdmKGskCyh8 https://youtu.be/SsOxBuIG_0A
85
EXPERIMENT NO. 8
COLUMN CHROMATOGRAPHY OF PIGMENTS: INK/FOOD COLORING
86
I. INTRODUCTION
The use of color additives increased dramatically in the United States in

the second half of the nineteenth century. As the economy became more
industrial, fewer people lived on farms, city populations grew, and people became
more dependent on mass produced foods. Food dyes were initially used to make
food more visually appealing to the consumer and, in some cases, to mask poor-
quality, inferior, or imitation foods. For example, meat was coloured to appear
fresh long after it would have naturally turned brown. Jams and jellies were
coloured to give the impression of higher fruit content than they actually
contained. Some food was coloured to look like something else imitation crab
meat, for example. Many food colourings and additives were later discovered to
be harmful or toxic.
Food colorants were initially added to food with little or no health testing.
In 1907, the USDA reduced the number of synthetic food dyes approved for use
from 695 to just seven. Only two of the original dyes from 1907 are still accepted
for use today. Five others have been added between 1907 and 1971. Only seven
dyes are approved for use in the United States today. All of the FD&C approved
food dyes are charged, water-soluble organic compounds that bind to natural ionic
and polar sites in large food molecules, including proteins and carbohydrates.
Column Chromatography is one of the experiments and its goal is to create an
effective column chromatography setup and procedure that will separate methyl
orange from methylene blue. Variables of the procedure includes the amount of
silica used, the construction of the model, and the composition of the mobile
phase. In this experiment, you'll use column chromatography to separate green
food coloring into its component blue and yellow dyes, called erioglaucine and
tartrazine. Compounds with a higher affinity towards the stationary phase (polar)
will interact with the solid, preventing movement. Compounds that lean towards
the mobile phase (nonpolar/ less polar) will interact with the liquid, which allows
movement to flow through the stationary phase. Column Chromatography is the
technique used to purify compounds from mixtures using a pipette column. This
method is used for larger sample sizes. Dye mixture is added to a solvent, a
87
separation is visible after an adequate amount of time is given, an example of the
setup is shown in Figure 1.
FIGURE 1. Column Chromatography
II. MATERIAL & EQUIPMENT
MATERIAL USED AMOUNT
50-mL glass beaker 25

100-mL glass beaker 5
5- or 10-mL glass graduated cylinder 5
50-mL glass graduated cylinder 10
Small powder funnel 10
Glass stirring rod 5
Labelling tape 5rolls
Labelling pen 5
Ruler 5
Pasteur pipette bulbs 10
Laboratory stand 5
Burette clamp 5
50-mL glass burette 5
88
Weighing boats 20
Cotton or glass wool plugs 10
Long-stem Pasteur pipettes 50
Sand (40-100 mesh) 15g
Silica gel powder for 50g
column chromatography
(35-70 mesh, 100 grade)
Ethanol (95%) 400mL
5-mL glass volumetric flask with cap 1
Long blunt-ended wooden/metal rod 1
250-mL plastic wash bottle 1
Paraffin film 1box
Spatulas 5
Lab wipes 3boxes
Acetone 250mL
Analytical balance (at least 1) Dependent on lab size
Deionized water Dependent on lab size
Unknown Compound (Red) 2.5 g
Unknown Compound (Yellow) 2.5 g
III. PROCEDURE
Here, we show the laboratory preparation for 10 students working in pairs, with
some excess. Please adjust quantities as needed.
1. Preparation of the Laboratory

a. Before you get started, put on a lab coat, safety glasses, and nitrile gloves, and
make sure that the lab has a labelled organic waste container.
89
b. Ensure that the lab has labelled waste containers for silica gel and glass.
c. Fill a plastic wash bottle with acetone and set it by the organic waste container.
This is for students or teaching assistants, who will clean the columns after the
lab.
d. Place a container of silica gel powder, a container of sand, a box of laboratory
wipes, several clean spatulas, and enough weighing boats for each student group
by a calibrated balance in a well-ventilated area.
e. Place a bottle of 95% ethanol in a nearby hood. Then, make sure that the
stopcocks for the columns fit securely but still turn easily.
f. Close the stopcocks after testing them and distribute the columns to the benchtop
workspaces.
g. Make sure that each workspace has a lab stand or a scaffold and an appropriate
clamp for the column being used.
h. Set out the following glassware and equipment at each student lab station (we
suggest that students work in pairs):
5 50-mL beakers
1 100-mL beaker
1 5- or 10-mL graduated cylinder
2 50-mL graduated cylinders
2 Small powder funnels
1 Glass stirring rod
1 Roll of laboratory tape
1 Marking pen
1 Ruler
2. Preparation of the Dye Solution

a. Prepare 5 mL of green food dye solution in ethanol. First, take a clean empty 5-
mL volumetric flask and slowly add 2.5 g of red and yellow food dye into the
different beaker.
b. Add about 1.5 mL of 95% ethanol to the flask, stopper it, and seal it with plastic
paraffin film.
90
c. Hold the stopper firmly in place and invert the flask several times to mix the
ethanol and unknown compound together.
d. Then, add 95% ethanol to the fill line, reseal the flask, and invert it until the
solution colour appears even throughout.
e. Label the flask with its name and concentration and place it in a central area.
3. EXPERIMENTAL PROCEDURE:
1. To begin, put on a lab coat, safety glasses, and a pair of nitrile gloves.
91
2. Bring a column or burette to the central supply area. Obtain a piece of cotton or glass
wool and press it into the top of your column. Use a long, blunt-ended wire or rod to
push it down to plug the column just above the stopcock. Note: The cotton must be
stable, but don't pack it too densely, because the solvent needs to pass through it
easily. Make sure that the stopcock can turn freely without catching on the plug.
3. Attach a burette clamp to a lab stand and clamp the column upright with about 4 in of
space underneath it. Make sure that the stopcock is closed.
4. Label a 100-mL beaker as ‘waste’ and place it under the column.
5. Next, measure 1 g of sand in a tared weighing boat and bring it back to the bench.
6. Then, mark the column about 0.5 cm above the top of the cotton plug.
92
7. Place a small funnel in the top of the column and pour enough sand through the
funnel so that there is a layer 0.5 cm deep over the plug. This will keep the silica gel
from passing through the plug into your fractions.
8. Remove the funnel and gently tap the column to evenly distribute the sand.
93
9. Now, prepare your eluents. Label a 50-mL beaker and a 50-mL graduated cylinder as
‘water’.
10. Measure 50 mL of deionized water with the graduated cylinder and pour it into the
beaker.
11. Then, label another 50-mL beaker and a 50-mL graduated cylinder as ‘ethanol’.
Measure 50 mL of 95% ethanol and pour it into this beaker.
12. Now, obtain 4 or 5 clean Pasteur pipettes and a pipette bulb. Gently pipette 7 – 8 mL
of ethanol along the inner walls of the column. This layer of solvent will let you pour
this silica gel slurry into the column without disturbing the sand.
13. Open the stopcock and drain ~1 mL of ethanol into the waste beaker to flush air from
the stopcock and to confirm that the solvent flows easily through the plug. Make sure
that there are no dry patches in the sand and the plug and gently tap the side of the
column to remove trapped air bubbles.
94
14. Next, place a clean funnel in the top of the column and label a third 50-mL beaker
‘silica gel slurry in ethanol’
15. Carefully weigh 5 g of silica gel powder and pour it into your labeled beaker.
16. Then, bring the silica gel, a glass rod, and your ethanol graduated cylinder to the
ethanol dispensing hood.
17. Measure 18 mL of 95% ethanol and pour it into the beaker. Stir the mixture with the
glass rod until the texture is consistent and no dry patches or clumps of powder
remain. This usually takes ~1 min. Note: The slurry should be thick, like batter, but it
should still flow easily. If the slurry is too stiff, it could clog the funnel or trap air
bubbles in the column.
95
18. Once the powder is completely suspended in ethanol, bring it to your bench. Stir the
slurry until its texture is consistent again and immediately pour it into the column.
19. Gently tap the sides of the column as the silica gel settles to get rid of air bubbles.
19. Rinse the beaker with 1 or 2 mL of ethanol, and pour the rinse into the column. Then,
remove the funnel and set it aside.
20. Pipette ethanol along the inside of the column to wash down particles stuck to the
walls. Then, drain the solvent until the meniscus is just barely above the top of the
silica gel.
96
21. Now, use a 5- or 10-mL graduated cylinder to obtain about 0.8 mL of the red and
yellow food dye solution.
22. Use a clean pipette to gently apply 5 to 7 drops of the dye solution evenly across the
top of the silica gel. If necessary, add a few drops of ethanol to keep the silica gel
covered.
97
23. While you wait for the dye to soak in, label two clean 50-mL beakers as ‘ethanol
fraction ‘and ‘water fraction’. Exchange the waste beaker under the column with the
ethanol fraction beaker.
24. Next, fill the column with ethanol, pipetting the first mL along the inner walls to
avoid disturbing the silica gel.
25. Then, open the stopcock partway and adjust it until the solvent level decreases ~1 mL
every 15 – 20 s.
26. Periodically refill the column to keep the ethanol flowing. Make sure that there is at
least 1 cm of solvent above the silica gel at all times.
98
27. Monitor the yellow and red bands as they separate. Note: If you need to step away,
close the stopcock so that the column won't run dry while you're gone. Avoid
stopping for more than 1 min at a time because the compound bands will start
broadening. It usually takes about 20 – 35 min for the faster-moving dye to leave the
column.
28. Once you have collected all of the faster-moving dye in the ethanol fraction beaker,
close the stopcock, set the ethanol fraction aside, and put the waste beaker under the
column.
29. Drain the solvent until the meniscus is just above the top of the silica gel. Then,
exchange the waste beaker for the water fraction beaker.
30. Use a clean pipette to fill the top of the column with deionized water. Then, open the
stopcock just enough to let water flow through the column at a rate of about 1 mL
every 15 – 20 s.
31. Refill the column with water as needed until the second band of color has completely
left the column. This usually takes another 15 – 25 min. Then, close the stopcock and
exchange the water fraction beaker for the waste beaker.
99
32. Now, drain the column completely and leave the stopcock open.
33. Record the colors of the ethanol and water fractions in your lab notebook.
34. Pour the fractions into the corresponding 50 mL graduated cylinders and record the
volume of each fraction. Then, close the stopcock and bring the column to a fume
hood.
35. Clamp the column upright in the hood so that the silica gel can dry. Your instructor
will either tell you how to clean the column or take care of it after the lab.
36. Now, flush the collected fractions and leftover ethanol, water, and dye solution down
the drain and clean the rest of your glassware as usual.
37. Discard used Pasteur pipettes in the glass waste. Throw out used paper towels and
weighing boat in the lab trash and clean the benchtop before you leave.
IV. GUIDE QUESTION

1.What are the two phases in column chromatography?
ANSWER:
100
2. What colors are the bands in the column?
ANSWER:
3. Mixtures in column chromatography?
ANSWER:
4. How are mixtures separated in column chromatography?
ANSWER:
5.What compounds elute first?
ANSWER:
6.What does elute mean?
ANSWER:
7. How do column properties such as the diameter and length affect the separation of mixtures?
ANSWER:
8. How many different colored bands appear as the sample separates through the column?
ANSWER:
9. Rf VALUE OF TWO FOOD DYE COLOR = (distance to centre of spot) / (distance to solvent front).
ANSWER:
10. What is the difference between mobile phase and stationary phase?
ANSWER:
11. Why do we need to put sand in the column?
ANSWER:
12. How do you pack a column chromatography?

ANSWER:
13. What is the difference between TLC and column chromatography?
ANSWER:
101
V. REFERENCES
Gilbert, J.C., and Martib, S.M., Experimental Organic Chemistry: A Miniscale

and Microscale Approach, 4th Edition, Cengage Learning, Boston, MA, 2006.
Jove.com (2020), “Column Chromatography” Retrieved from : “

https://www.jove.com/science-education/11211/column-chromatography”
Landrie, C.L., and McQuade, L.E., Organic Chemistry: Lab Manual and Course
Materials, 5th Edition, Hayden-McNeil, LLC, Plymouth, MI, 2016.
VIDEO REFERENCES:
https://www.youtube.com/watch?v=frmPbmXmvaQ
102
EXPERIMENT NO. 9
ANALYSIS OF ALCOHOLS/PHENOLS/ALDEHYDES/KETONES
103
I. INTRODUCTION
a. Alcohols and phenols
Alcohols and phenols may be assumed to be organic water in which one proton
from the water molecule maybe replaced by an aliphatic alkyl group to form alcohol(R-
OH). However, in phenols (Ar-OH) this proton is being replaced by the aromatic ring.
Just like water, alcohols and phenols contain bonding between molecules- ie the
hydrogen bond which is strong.
Alcohol molecules especially small one may replace the water molecules in the
network of hydrogen bonds as shown below:
H H H
O H O H O H
This replacement causes the alcohol to be soluble in water.
Primary and secondary alcohols are easily oxidized by chromic acid but not for
tertiary alcohol-not oxidized at all by chromic acid. Primary alcohol is oxidized to an
aldehyde but secondary alcohol is oxidized to ketone. The equation to represent this
oxidation of alcohol is given below:
O
3(-C- OH) + 2H2CrO4 + 6H+ 3( C- ) + 2Cr3- + 8H2O
In this reaction the alcohol solution is reacted with sodium dichromate solution in
sulphuric acid. Alcohol which is oxidized will reduce chromium ion, Cr (VI) to Cr (III)
and causes the solution to turn to green in colour.
On the other hand, phenol is an acid and is stronger compare to either alcohol or
water. This is because the negative charge ion phenoxide can be delocalized in the aromatic
104
ring through resonance. This solution cannot take place in alcoxide or hydroxide ions i.e
their charges are localized.
The delocalization of phenoxide charge is enable phenol to form different
phenoxide salts by reacting with strong base. That is why phenol is corrosive. Ensure that
phenol does not get into contact with your skin. If it does, clean your skin with plenty of
tap water.
Apart from that, the presence of hydroxyl group in phenol activates the aromatic
ring towards electrophilic replacement reaction. Phenol reacts with bromine water to form
ortho and para bromophenols. In this reaction, involves the formation of phenoxide ion as
the intermediate.
b. Ester
Alcohols react with acid to form an ester. The general equation for this reaction is
shown below:
O O
H+
R-OH + R'-C-OH R'-C-OR + H 2O
This reaction is known as esterification. The product formed usually will have a
fruity smell. This similar molecule will form the smell and taste in fruits.
In this experiment, you will convert two alcohols to be their esters by reacting them
with acetic acid and catalysed by a strong acid, H2SO4.
c. Aldehyde
The aldehyde group is able to reduce silver ion in solution, forming either a black
deposit of free silver or a silver mirror. The aldehyde group is oxidized to an acid in the
reaction. The reagent used in this reaction is known as Tollens reagent, which is made by
reacting silver nitrate solution with dilute ammonium hydroxide. This test normally called
as Tollens test. It is very useful to identify the presence of aldehyde group.
105
d. Carboxylic acid
To identify the presence of carboxylic acid, solution of carboxylic acids in water or

aqueous ethanol is allowed to react with sodium hydrogen carbonate. Bubbles of carbon
dioxide rise to the surface of the solution indicating the presence of an acid.
Notice: Each student must have his/her own unknown and be sure to record
the number of your unknown on the report sheet. If you work in pairs, each pair
must have 2 unknowns. Keep the tubes containing your unknown at your bench until
you finish all parts of the lab.
II. Objectives:
To observe reactions of alcohols, phenols, aldehydes and ketones and attempt

to determine to which of these functional groups an unknown substance belongs.
III. MATERIALS & EQUIPMENT

Apparatus
Test tubes, beaker, dropper, hot plate, test tube racks, tong
Chemicals
Methanol, 1-butanol, 2-butanol,t-buthyl alcohol, cyclohexanol, acetone, chromic
acid, cyclohexanol, phenol, sodium hydroxide solution (2 M), phenol crystal, bromine
water, acetic acid, 3-methyl-1- butanol, concentrated H2SO4, silver nitrate solution (0.1
M), ammonium hydroxide (1%), 10% formaldehyde, acetone, 10% glucose, 5%
aqueous solution of sodium hydrogen carbonate.
106
IV. PROCEDURE & DISCUSSION
Add 100 mL of tap water to a 250 mL beaker and heat it on a hot plate for Parts A
and D.
A. Lucas Test for Primary, Secondary and Tertiary Alcohols.
The Lucas reagent is a solution of zinc chloride in concentrated hydrochloric acid. This
solution must be made freshly to get proper results. The test depends on a difference in the rate
of reaction of these alcohols. The general equation for the reaction is:
 Tertiary alcohols react IMMEDIATELY. The test tube will get hot, and
because the alkyl chloride is insoluble in water two layers may be
apparent, or a cloudy dispersion forms.
 Secondary alcohols will become cloudy in 5 to 10 minutes. If cloudiness
does not appear place test tube in a warm water bath and observe.
 Primary alcohols give no reaction in a reasonable length of time.
CAUTION! Lucas reagent contains concentrated hydrochloric acid - Handle It with Care
1. Place 1 ml of Lucas reagent in each of four (5 if you work in pairs) clean test tubes.
107
2. Add 6 drops of n-butyl alcohol (1-butanol) to one test tube. Shake the test tube to mix
the reagents and notice whether the mixture gets cloudy and how long it takes.
3. In a second test tube place 6 drops of sec-butyl alcohol (2-butanol), shake and note
how long it takes the tube to get cloudy.
4. In a third test tube place 6 drops of t-butyl alcohol (2-methyl-2-propanol), shake and
note how long it takes the tube to get cloudy.
5. In the fourth (and fifth for pairs) test tube place 6 drops of your unknown and shake.
6. Record observations on the Report Sheet for this section.
108
Dispose of these reagents in the "Liquid Waste" container in the hood.
B. Bordwell-Wellman Test for Primary, Secondary and Tertiary Alcohols
The Bordwell-Wellman reagent contains potassium dichromate dissolved in sulfuric

acid. The orange-yellow color is due to the Cr2O72- ion. The oxidation number of chromium is
+6. This reagent will oxidize primary and secondary alcohols and chromium is reduced to the
greenish colored chromium(III) ion, Cr3+. This color change from orange-yellow to green serves
as an indicator for the presence of a primary or secondary alcohol. A primary alcohol is oxidized
first to an aldehyde, which will be further oxidized to an acid.
USE EXTREME CARE WITH THIS REAGENT, IT IS VERY CORROSIVE!
WASH IMMEDIATELY IF YOU GET ANY ON YOUR SKIN OR CLOTHING!!!
109
1. Place 0.5 ml of acetone in each of four (or 5 if you work in pairs) small test tubes.
2. Add 0.5 mL of n-butyl alcohol to one test tube; 0.5 mL of sec-butyl alcohol to a second
test tube; 0.5 mL of tert-butyl alcohol to a third test tube; and 0.5 mL of your unknown
to the remaining test tube(s).
110
3. To each test tube add 1 drop of Bordwell-Wellman reagent and shake.
4. Record your observations on the Report Sheet for this section.
Dispose of these reagents in the Organic Liquid Waste container in the hood.
C. Phenols
Phenols form highly colored coordination complexes with ferric ion. A

blue-violet colored solution results.
1. Place 0.5 mL of 3% phenol solution in a small test tube.
2. Place 0.5 mL of your unknown and 0.5 mL of water in another small test tube.
111
3. Add 1 drop of ferric chloride solution to each and shake.
4. Answer questions about this reaction on the Report Sheet.
Dispose of these reagents in the Organic Liquid Waste container in the hood.
D. Fehling's Test for Aldehydes
The water bath you set up earlier should be boiling. Set the hot plate temperature control to a
medium setting. Use 5 clean small test tubes if working alone, or 6 test tubes if working in pairs.
1. Add 5 mL of Fehling's Solution A to a small beaker and mix 5 mL of Fehling's Solution B

with it. Notice the change in color of the Cu2+ ion when these solutions are mixed.
2. Add 1.5 mL of the mixture prepared in step 1 to each of 5 (or 6) clean small test tubes.
3. Add 5 drops of the following test compounds (aldehydes/ketones) to the Fehling's

reagent in each test tube. Acetone in tube 1; benzaldehyde in tube 2; butanal
(butyraldehyde) in tube 3; butanone in tube 4; and unknown in tube 5 (and 6).
112
4. Make sure the tubes are properly labeled before placing them in the boiling water bath.
5. After heating the mixtures in boiling water for 5 minutes, take note of any changes
in color or formation of a red precipitate at the bottom of the tube.
113
6. Record your observations on the Report
Sheet for this section.
7. Turn off the hot plate, the water bath is

no longer needed.
Dispose of these reagents in the Organic Liquid

Waste container in the hood.
E. Oxidation with Bordwell-Wellman reagent
CAUTION!! Concentrated sulfuric acid in the Bordwell-Wellman reagent is very

corrosive. Handle it with care.
1. Add 0.5 mL of each of the following test compounds (aldehydes/ketones) to 5 (or 6 if

working in pairs) clean test tubes: acetone in tube 1; benzaldehyde in tube 2; butanal
(butyraldehyde) in tube 3; butanone in tube 4; and unknown(s) in tube(s) 5 (and 6).
114
2. Add 2 drops of Bordwell-Wellman reagent solution to each of the test tubes
containing the test aldehydes, alcohol or ketone from step 1.
3. Mix well and note any color changes after a minute or two. If there is a reaction, the
color should change from yellow/orange to green or brown.
4. Record your observations on the Report Sheet for this section.
Dispose of these reagents in the "Liquid Waste" container in the hood.
V. GUIDE QUESTIONS
1. For methanol, 1-butanol, 2-butanol,t-buthyl alcohol, cyclohexanol;

i) Write the structural formulas for each of the alcohols:
ANSWER:
Methanol:
1-butanol:
2-butanol:
115
t-butyl alcohol:
cyclohexanol:
ii) Which of the above alcohol is the least soluble in water? Why?
ANSWER:
2. For the following reactions of alcohols with chromic acid, write the equations, if any
reactions happen.
ANSWER:
3. Among the following alcohols which is not oxidized in 5 second by the chromic acid:
1-Pentanol, Cyclohexanol, 2-Methyl-1-Propanol and 2-Methyl-2- Butanol. Explain
your answer.
ANSWER:
4. Write the equation for the reaction of 1-Butanol and 3-Methyl-1-Butanol with acetic
acid.
ANSWER
5. Describe the difference between alcohols and phenols.
ANSWER:
6. Describe the difference between an aldehyde and a ketone, and indicate how each
differs from an alcohol.
ANSWER:
7. What is the color of the ferric chloride solution before adding it to phenol?
ANSWER:
8. What is the color of the solution after the reaction between phenol and ferric chloride
takes place?
ANSWER:
9. From the results with ferric chloride, would you classify your unknown as a phenol?
ANSWER:
10. Describe what is meant by oxidation and reduction in relation to organic
compounds, giving one example of oxidation of an organic compound and one
example of reduction of an organic compound. The compound you use for the
example may be the same or different for the oxidation and the reduction reactions.
116
Be sure to indicate what oxidizing agent is used and what reducing agent is used for
each example.
ANSWER:
A-1 After looking at the results in the above table, would you conclude your unknown is a
primary, secondary or tertiary alcohol?
A-2 Show the chemical reactions for the Lucas test with each of the butyl alcohols (n-, sec-
and tert-butyl alcohol or 1-butanol, 2-butanol and 2-methyl-2-propanol) that react with this
reagent. You do not need to show cases where there is no reaction. Indicate whether the
reaction is fast or slow.
A-3 Show the chemical reactions for the Bordwell-Wellman test with each of the butyl alcohols
(1-butanol, 2-butanol and 2-methyl-2-propanol) that react with this reagent. You do not need to
show cases where there is no reaction.
117
D-1 After looking at the results in the above table, would you conclude your unknown is an
aldehyde, a ketone or an aromatic aldehyde (like benzaldehyde)?
D-2 Did Fehling's reagent and Bordwell-Wellman reagent react differently with
benzaldehyde compared to butyraldehyde?
D-3 Why would benzaldehyde react differently than other aldehydes such as butyraldehyde?
D-4 You used Bordwell-Wellman reagent to test for alcohols and for aldehydes and
ketones. Which of the alcohols reacted with this reagent and what compounds of the
aldehydes and ketones reacted with this reagent?
D-5 Are you able to determine whether your unknown is an alcohol, aldehyde, ketone or
phenol from the observations you made in each of the tests? If so, which category of these
functional groups is your unknown?
118
VI. REFERENCES
Gilbert, J.C., and Martib, S.M., Experimental Organic Chemistry: A Miniscale and Microscale
Approach, 4th Edition, Cengage Learning, Boston, MA, 2006.
Landrie, C.L., and McQuade, L.E., Organic Chemistry: Lab Manual and Course Materials,
5th Edition, Hayden-McNeil, LLC, Plymouth, MI, 2016.
Lehman, J. W. Operational. Organic Chemistry: AProblem-Solving Approach to the
Laboratory Course, 3rd ed.;Prentice Hall:Upper Saddle River, NJ,1999;pp529-572.
VIDEO REFERENCES:
FIRST EXPIREMENT: https://youtu.be/sjhoUkz4TPY
SECOND EXPERIMENT: https://youtu.be/mAZqGzFbGQE
THIRD EXPERIMENT: https://youtu.be/sjhoUkz4TPY
FOURTH EXPERIMENT: https://youtu.be/e6tyKZS82cQ
FIFTH EXPERIMENT: https://youtu.be/idzztTrFsE4
119
CARBOXYLIC ACID AND DERIVATIVES
120
I. INTRODUCTION:
PART A:
Carboxylic acids are an important class of organic compounds. Other important classes of
compounds called acid derivatives are related to acids but differ from them in that the
hydroxyl portion of the carboxylic function is replaced by other groups. Among these acid
derivatives are the acyl halides, anhydrides, esters and amides. Many synthetic pathways are
available for the synthesis of these derivatives as well as many hydrolysis reactions can be
performed on such compounds. These reactions offer great help in the pharmaceutical and
chemical industry.
(a) Salt Formation

Carboxylic acids may be neutralized by strong bases to give salts. Acids which may be only
slightly soluble in water can be extracted from their solutions in an organic solvent by
aqueous base, as the salts are usually water soluble: RCOOH + NaOH ---> RCOO-Na+ + H2O.
7. Salt Hydrolysis
Most carboxylic acids are only partially dissociated in aqueous solution. Consequently, when the
salt of a carboxylic acid is dissolved in water, it is partially hydrolyzed to the undissociated acid
and hydroxide ion: RCOO-Na+ + H2O ---> RCOOH + NaOH.
5. Acid Chlorides
The halide ion is a good leaving group making acid halides very reactive towards nucleophilic
acyl substitution. Acid halides are so reactive that they are not found in nature, since they even
react with water (R' = H): RCOCl + HOR' ---> RCOOR' + HCl.
Similar to carboxylic acids (R' = H), these esters products can then be converted to salts
in aqueous base.
5. Amides
Values of pKa for amides of carboxylic acids are in the range of 15-17, which means that they
are comparable in acidity to alcohols.
121
Amides can also be converted to salts in aqueous base, but by a slightly different mechanism:
RCONH2 + NaOH ---> RCOO-Na+ + NH3.
II. OBJECTIVES
To prepare succinic anhydride from succinic acid and acetic anhydride using reflux and
vacuum filtration techniques. Also, this session aims to produce methyl salicylate (oil of
wintergreen) from acid-catalyzed heating salicylic acid with methanol. Finally, this lab aims to
hydrolyze acetamide under both alkaline and acidic conditions and identify the different products
formed using litmus paper.
The second part consists of a couple of simple test tube reactions that illustrate some of the
properties of carboxylic acids and various derivatives.
III. MATERIAL & EQUIPMENT
A- Experimental Glassware:
- 3 Beakers B- Experimental Equipment:
- Round Bottom Flask (50 ml) - Spatula
- Graduated pipet - Mettler balance
- Glass rod - Ice bucket + ice
- Test tubes - Heating Mantle & Plate
- Suction flask - Wooden clamp
- Boiling stones - Plastic Dropper
- Watch glass - Plastic plate (for weighing)
- Reflex Condenser - Metallic stand/green clamps
- Filter Paper
- Litmus Paper
- Buchner Funn
122
C-CHEMICALS/REAGENTS:
Chemical Structure M.W. and Amount Hazards/Safety Data
Sodium 39.997/2.4 gm Non-flammable. Very hazardous in case of skin
Hydroxide contact (corrosive, irritant, permeator), of eye
contact (irritant, corrosive), of ingestion,
of inhalation. Eye contact can result in corneal
damage or blindness. Skin contact can produce
inflammation and blistering. Wear Splash goggles.
Synthetic apron. Vapor and dust respirator, and
gloves. In case of eye contact, immediately flush
eyes with plenty of water for at least 15 minutes.
Cold water may be used. In case of contact,
immediately flush skin with plenty of water for at
least 15 minutes while removing contaminated
clothing and shoes. Cover the irritated skin with an
emollient. If inhaled, remove to fresh air. If not
breathing, give artificial respiration.
Succinic 118.09/3 gm May be combustible at high temperature.

acid Hazardous in case of skin contact (irritant), of eye
contact (irritant), of ingestion, of inhalation.
Check for and remove any contact lenses.
Immediately flush eyes with running water for at
least 15 minutes, keeping eyelids
open. Cold water may be used. Do not use an
eye ointment. Splash goggles. Lab coat. Dust
respirator and gloves.
Acetic 102.09/4 ml Flammable if liquid and vapor
Anhydride Harmful if inhaled. Causes severe skin burn and
eye damage. Keep away from heat, open flames
and hot surfaces
Use under fume hood. Avoid contact with skin
and eyes by wearing
gloves, lab coat and safety glasses with side shields.
Salicylic 138.118/0.5 gm Irritating to skin Irritating to respiratory system.
Acid Serious damaging to eyes. Flammable. Avoid
contact with skin by wearing gloves and lab coat.
Use under fume hood to avoid inhalation Wear
safety glasses with side shields Keep away from
heat, open flames and hot surfaces
Methanol 32.04/5 ml Flammable. Hazardous in case of skin contact
(irritant), of eye contact (irritant), of ingestion, of
inhalation. Slightly hazardous in case of
skin contact (permeator). Eye Contact: Check for
and remove any contact lenses. Immediately flush
eyes with running water for at least 15 minutes,
keeping eyelids open. Cold water may be used.
123
Get medical attention.
Skin Contact:
In case of contact, immediately flush skin with
plenty of water for at least 15 minutes while
removing contaminated clothing and shoes. Cover
the irritated skin with an emollient. Cold water
may be used. Splash goggles. Lab coat. Dust
Respirator and gloves.
Sulfuric 98.08/4.5 ml Non-flammable. Very hazardous in case of skin
acid contact (corrosive, irritant, permeator), of eye
contact (irritant, corrosive), of ingestion,
of inhalation. Check for and remove any contact
lenses. In case of contact, immediately flush eyes
with plenty of water for at least 15
minutes. Cold water may be used. Face shield.
Full suit. Vapor respirator. Gloves. Boots.
Acetamide 59.07/1.5 gm May be combustible at high temperature.
Hazardous in case of skin contact (irritant,
permeator), of eye contact (irritant), of ingestion,
of inhalation. Check for and remove any contact
lenses. In case of contact, immediately flush eyes
with plenty of water for at least 15
minutes. Cold water may be used. Splash goggles.
Lab coat. Dust Respirator and gloves.
Water 18.00 N/A
IV. PROCEDURE AND METHOD
1- Preparation of Succinic Anhydride:
O
O O O O O
OH
+ + 2CH3COOH
O
OH
Succinic Anhydride
O Acetic Anhydride
Succinic Acid
124
 In a clean and dry 50 ml RBF, 3 gm of succinic acid was weighed and added, and then
using a graduated pipette, 4 ml of acetic anhydride was added in the same RB
 The reflex condenser system was set up properly with proper water flow system, and few
boiling stones were added to the RBF. Then, the solution was reflexed on a heating
mantle for approximately 15 minutes after boiling has begun.
 Next, the heating mantle was turned off and the reflex condenser was dismantled and the
RBF content was transferred to an Erlenmeyer flask. After that, it was allowed first to
cool at room temperature for a few minutes, and then it was placed at an ice bath and the
inside walls of the flask were scratched for a few minutes.
125
 Meanwhile, vacuum filtration apparatus was being set up and the top of Buchner funnel
was covered with a filter paper which was wetted with some drops of water. Next, using a
spatula, succinic anhydride crystals were transferred to the Buchner funnel and vacuum
filtration process was performed.
 Later, the filter paper, containing crystals, was placed on a watch glass and dried in the
oven, and then it became completely dry, it was weighed, and the difference between its
mass and the empty filter paper mass was calculated to obtain the final pure mass of
succinic acid crystals.
2- Preparation of Methyl Salicylate (Oil of Wintergreen):
126
 0.5 gm of salicylic acid was weighed and put in a test tube, above which 5 ml methanol
was put using graduated pipette, followed by 10 drops of sulfuric acid that
were put using plastic dropper.
 The solution inside the test tube was heated in a previously-prepared water bath (beaker
containing tab water on a heating plate). The solution inside the test tube was left for
approximately 2 minutes after boiling. Next, the content was poured over 10 gm of ice in
a small beaker and stirred well and the odor of the product was observed.
3-
Hydrolysis of Acetamide:
 Two test tubes were used in this experiment. One of them was filled with weighed 0.5 gm
of acetamide, and the other was filled with 1 gm of acetamide. For the first test tube,
using a graduated pipette, 4 ml of NaOH 10% were added, and for the second tube 4 ml
of H2SO4 10% were added.
127
Both test tubes were heated to boil in the previous water bath. The odor of the evolved gas was
noticed in the first tube and the litmus paper was hold on the mouth of this tube and the second
tube and the change in color was observed.
PROCEDURE/METHODS:
PART B:
(a) Salt Formation

 In each of two test tubes, place 0.1 g of benzoic acid. To one tube add 3 mL of cold
water, to the other add 3 mL of 10% sodium hydroxide solution.
To one tube add 3 mL

place 0.1 g of of cold water, to the
benzoic acid other add 3 mL of 10%
sodium hydroxide
solution
 Shake both tubes, observe, and record the results. Then acidify salt with diluted
hydrochloric acid.
128
(b) Salt Hydrolysis
Dissolve approximately 0.2 g of sodium acetate in 5 mL of distilled water, test the
resulting solution with litmus paper, and note the result.
(c) Acid Chlorides

 [These experiments MUST BE DONE IN THE HOOD.] Add drop by drop 1 mL of
acetyl chloride to 1 mL of butanol. Allow to stand for about 2 minutes and then pour it
carefully into 5 mL of water (acetyl chloride reacts vigorously with water). Note the
odour [Recall: Smelling solutions is done by wafting the smell towards your nose].
Remove a little of the insoluble layer with a dropper and test its solubility in 5% NaOH
solution.
VII. GUIDE QUESTIONS:
1. What is the chemical bases for the tests in our experiment ?

Answer:
2. Was the desired product successfully extracted/ synthesized?
Answer:
3. What is the percentage yield? low/high?
Answer:
129
4. What were the errors encountered during the experiment and how did we solved them?
Answer:
5. What did you learn from this experiment?
Answer:
6. Write out a mechanism for the formation of the benzocaine? (Hint: this reaction is
called a fischer esterification)
Answer:
7. A solid precipitate is formed on adding sulphuric acid. What would this solid be?
Answer:
8. Write an equation which accounts for the solubility of benzoic acid in aqueous base.
Answer:
9. Write out the structures of all compounds to be tested in Part B.
Answer:
VIII- REFERENCES:
 The Reactions of Carboxylic Acids and Their Derviatives lecture and PowerPoint
presentation.
 Laboratory Manual to accompany Organic Chemistry- A Short Course
(Hart/Craine/Hart/Hadad)- 13th edition
 http://www.sciencelab.com
 en.wikipedia.org
Video references:
https://youtu.be/ap7ySufGjFk
https://youtu.be/BUJ2q5-K3zs
130
SYNTHESIS OF ASPIRIN
131
I. INTRODUCTION
Dating back 2500 years ago Native Americans chewed on willow bark to reduce fever
symptoms. In the early 1800’s, a Scottish physician discovered that extracts of the willow bark
helped to alleviate the pain associated with rheumatism. Further studies illustrated that willow bark
extracts were not only analgesic, but also reduced fever and swelling. However, the active
ingredients were unknown. During the 1800s, chemists had isolated and identified the active
component to be salicylic acid. Salicylic acid is extremely bitter tasting and frequent use can cause
severe stomach irritation.
In 1893, German Chemist Felix Hoffman, an employee at Bayer produced a synthesized
form of acetylsalicylic acid. Derived from salicylic acid the term acetylsalicylic acid is the
chemical name for Aspirin. The company Baye patented aspirin in 1899, which has made aspirin
one of the most widely used and commercially available drugs. This new drug had the medicinal
benefits of both salicylic acid and sodium salicylate without the problems. Aspirin has proven to
be a safe, effective analgesic, fever-reducer, and anti-inflammatory. Nowadays the world
consumes over 15 billion aspirin tablets each year, making it the most popular commercially used
drug today.
Although aspirin has health benefits, it also has some health disadvantages. It can cause
internal bleeding, nausea, vomiting, rashes, interfere with platelet functioning and can cause Reyes
syndrome in some children. Caution most also be taken when performing this experiment. Aspirin
is a combustible chemical. Acetic anhydride and salicylic acid are both very toxic chemicals. These
toxic chemicals should never come not contact with bare skin. The fume hood should be used for
multiple parts of this experiment to protect the product from undesired results/ reactions and to
protect the experimenter.
132
Figure 1. Reaction of Aspirin.
Synthesis of aspirin can be completed through a process called esterification. In the process
of esterification an organic acid is combined with an alcohol to form an ester and water. The
reaction mechanism entails salicylic acid reacting with acetic anhydride in the presence of acid to
form acetylsalicylic acid. The hydroxyl group from the benzene ring of salicylic acid reacts with
the acetic anhydride to produce the functional group of an ester. Sulphuric acid is used to catalyze
this nonspontaneous reaction because its conjugate base is a made of a strong deprotonating group.
The structural formula of aspirin is C9H8O4.Aspirin has a high molecular weight and is not soluble
in water hence the solid can be separated by crystallization.
Verifying the identity of aspirin both the NMR and IR spectrums can be used to analyze
and identify the functional groups as well as hydrogen atoms present in the synthesized aspirin
product. The IR spectrum entails the transmittance percent of the compounds at different
wavelengths. To analyse an aspirin sample on a spectrum, the reactants (acetic acid and salicylic
acid) need to be analysed at the wavenumber of their peaks on the IR spectrum. Acetylsalicylic
acid’s (aspirin) structure indicates that there should be peaks for a carbonyl acid and carbonyl ester,
which should range from 1800 cm-1 to 1715 cm-1 and 1725 cm-1 to 1680 cm-1. Acetic acid has
a peak around the wavenumber 1700 cm-1 and salicylic acid has a peak around the wavenumber
1650 cm-1. Comparing the theoretical values of the wavenumbers of the reactants to values.
133
II. OBJECTIVES
The aim of the experiment was to enable us to synthesize aspirin, reinforce the skills of
recrystallization and the technique of melting point determination.
III. MATERIAL AND EQUIPMENT
 Googles
 Gloves
 Lab coat Vacuum filtration apparatus
 2.0 g of Salicylic acid

 Filter papers 7.0 cm
 125 mL and 50 mL Erlenmeyer flask
 A Spatula
 5.0 mL of acetic anhydride
 5 mL of Cold H2O
 4 DROPS OF CONCENTRATED
 Buchner Funnel
SULFURIC ACID
 A watch glass
 Hotplate
 7 mL of ethanol
 Thermometer
 50 degrees Celsius water bath For the ferric chloride test:
 10 mL graduate cylinder  0.5 mL of H2O
 Glass rod  A test tube
 Ice bath  Crystals from experiment
 A drop of Ferric chloride solution
134
CHEMICAL USED:
135
IV. PROCEDURE
1. To start the experiment, 2.0 g of salicylic acid was weighed into a 125 mL
Erlenmeyer flask
2. And 5.0 ml of acetic anhydride and four drops of concentrated sulfuric acid was
added.
3. The mixture was then heated gently in a 50o C water bath for 15 minutes being
swirled occasionally. The flask was then removed from the water and allowed
to cool to room temperature. As the mixture cooled, the product began to
crystallize.
136
4. Collected by vacuum filtration the solid product was rinsed with five milliliters
of cold water. The product was then allowed to dry for 10minutes in the
Buchner funnel and transferred to a watch glass to air dry for 10 minutes. The
crude weight of the product was then obtained for the calculation of the crude
yield and a small amount was saved in a small vial
5. Afterwards, the remainder of the solid was transferred to a 50 mL Erlenmeyer
flask and recrystallized from ethanol/water. Seven milliliters of ethanol was
added to the flask and it was heated until the solid was dissolved.
Approximately 20 mL of warm H2O was added to the flask and mixed
completely. After the solution cleared up, it was placed in the hood and allowed
to slowly cool. After the crystals formed, the flask was cooled in an ice bath so
that the yield may be maximized. The crystals were collected by Vacuum
filtration again, rinsed with cold H2O and allowed to dry completely. The final
weight was obtained and the m.p.s for both crude and pure samples were
collected.
137
6. In the final analysis, a ferric chloride test was performed for the presences of
phenol groups. 0.5 ml of H2O was added to a small test tube and small
amount of crystals was added to the test sample with one drop of the ferric
chloride solution. This test was done for both the pure and crude product.
V. GUIDE QUESTION
Part I. Synthesis of Aspirin
1. Mass of Salicylic acid (g)
2. Odor after addition of water
3.Mass of dried product (g)
138
4. Calculate the maximum amount of aspirin(C9H8O4) in grams that can be made
from the amount of salicylic acid (C7H6O3) that you used. This is known as the
theoretical yield of the reaction. The chemical equation shown in the introduction
tells us that there is a 1:1 ratio between the moles of salicylic acid we start with
and the amount of aspirin produced. This means that for every one mole of
salicylic acid (C7H6O3) we start with, we should produce one mole of aspirin
(C9H8O4). In order to calculate the theoretical yield, first convert the grams of
salicylic acid (C7H6O3) to moles (Remember gram to mole conversions from
Chapter 3!). This amount of moles is equal to the amount of moles of
aspirin that should be produced (since it is a 1:1 ratio). Then convert the moles of
aspirin(C9H8O4) to grams of aspirin and this is your theoretical yield.
Theoretical Yield of aspirin ___________________(g)
4. Calculate the percent yield for your reaction by dividing the mass of the dried
product by the theoretical yield that you calculated above and multiplying by 100.
Show your work below.
Part II. Iron (III) Chloride Test for salicylic acid

Substance Color Presence of Salicylic
Acid (+/-)
Salicylic Acid
Store brand aspirin
Synthesized aspirin
139
POST LAB QUESTION
5. Draw the structure of aspirin below. Circle and name the functional groups present
in aspirin.
ANSWER:
6. Thinking about the three reagents (Look at procedure Step 3) you used to synthesize
aspirin, which one was present in order to catalyze the reaction?
ANSWER:
7. Taking more than 150mg/kg of aspirin can be fatal. For an adult who weighs 150
lbs, how many tablets (1 tablet = 325 mg of aspirin) is this? (Hint: 1 kg = 2.205 lbs)
ANSWER:
8. What was the percent yield of aspirin in your experiment? Describe an
experimental error that could explain why your yield was too high or too low.
ANSWER:
9. Discuss how the following instances would affect the results of this synthesis?
a. The product was washed with hot water during filtration instead of cold water.
Answer:
b. The aspirin was left out to dry for weeks, resulting in a vinegar-like smell. Write
the reaction responsible for this.
Answer:
10. We used the concept of limiting reagents in this experiment.
a. Why was salicylic acid chosen as the limiting reagent? Discuss several advantages
of this choice.
Answer:
b. A student performing this experiment misread the amount of reactants and added
0.50 mL of acetic anhydride instead of 5.0 mL. Did this procedural error affect
the theoretical yield? Explain and support your answer.
Answer:
140
11. In this experiment, we used spectroscopy to analyze the purity of your aspirin.
a. The upper range for abs. measurements is around 1.0. Why is this (what is
happening chemically)? What happens to the graph when the absorbance is
large?
Answer:
b. Why is it necessary to collect a calibration or standard curve when using the
absorption spectroscopy technique in lab?
Answer:
c. Discuss one advantage and one disadvantage of absorption spectroscopy.
Answer:
VI. REFERENCES
Lola. “Aspirin Synthesis Lab Analysis.” Odinity, 17 Dec.
2017, www.odinity.com/aspirin-synthesis-analysis/.
“NMR Spectrum of Aspirin.” Thermo Fisher Scientific - US,

www.thermofisher.com/bs/en/home/industrial/spectroscopy-elemental-
isotope-analysis/spectroscopy-elemental-isotope-analysis-learning-
center/spectroscopy-elemental-isotope-analysis-resource-library/nmr-
tech-talk/nmr-tech-talk-march-2015/nmr-spectrum-aspirin.html.
“Synthesis and Recrystallization of Aspirin: Lab Report.”

UKEssays.com, www.ukessays.com/essays/biology/the-synthesis-of-
aspirin-biology-essay.php.
VIDEO REFERENCES:
https://youtu.be/XqfKx63mZCQ
https://youtu.be/7_ozW4yX_F8
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ORGANIC CHEMISTRY

EXTRACTION: DETERMINATION OF ITS EFFICIENCY; 49-52

PREPARATION OF SYNTHETIC FOOD FLAVORS 53-60

ISOLATION OF CAFFEINE FROM TEA LEAVES 61-70

THIN LAYER CHROMATOGRAPHY OF ANALGESIC DRUGS 71-84

ANALYSIS OF ALCOHOLS/PHENOLS/ALDEHYDES/KETONES 102-118

1. Chemical hazards - The use of chemicals in research laboratories is inevitable,

IMPORTANT STANDARDS TO HELP MITIGATE THESE HAZARDS

LABORATORY HAZARDS SYMBOL

For Biological spill accidents:

Radioactive spill emergency:

What to do for Clothing on fire:

Hazardous material splashed in eye

Some other important lab safety rules:

BASIC LAB TECHNIQUES

When Pouring a liquid from a bottle:

Harvey (2013). “Gravity Filtration”. Retrieved from:

Mcleod (2011). “ Laboratory Hazards and Risks”. Retrieved from:

PURIFICATION OF AN IMPURE ACETANILIDE SAMPLE BY

Recrystallization (AKA: crystallization) is a purification technique that is based on the

Dissolution of the Impure Acetanilide

c. Continue to heat until no more solid appears to dissolve.

a. Place a 50 mL Erlenmeyer flask containing about 5 mL of distilled water on a

a. Cool the Erlenmeyer flask containing acetanilide solution to room temperature

Mass of recovered acetanilide =

Matt (2011), “Preparation/Recrystallization of Acetanilide”, Retrieved form :

Stoddard (2020). “CHM 242 Lab 1 Recrystallization of Acetanilide”. Retrieved from:

SEPARATION OF A BINARY MIXTURE BY SIMPLE AND

a) To purify a compound by separating it from a non-volatile or less-volatile material.

III. EQUIPMENT/ MATERIAL:

large test tubes (3)

test tube rack (1) ring stand

10-mL graduated cylinder glass adaptor

50- mL round bottom flask grease

clamp (1 or 2) rubber tubing (2)

heating mantle boiling chips

condenser (1 or 2) thermometer adaptor

thermometer 50- mL round bottom flask

Cyclohexane and toluene

Table of Chemicals Used:

a) the liquid is relatively pure to begin with (e.g., no more

b) essentially a pure material is separated from a non-

c) the liquid is contaminated by a liquid with a boiling

Figure 1. Cyclohexane Figure 2. Toluene.

Diagram 1. Summary of Simple and Fractional Distillation.

The second procedure was the fractional distillation.

The fractional column was connected with the distillation head,

SHOW THIS FOLLOWING DATA:

Table 3. Fractional Distillation data for Cyclohexane. Drops Temperature

Table 4. Fractional Distillation data for Toluene.

Calculate the percent recovery of both solvents for fractional distillation

% Table 5. Simple distillation for Cyclohexane.

Graph 2. Simple distillation curve for cyclohexane and toluene

Calculate the percent recovery of both solvents for simple distillation

Encyclopædia Britannica. http://www.britannica.com/EBchecked/topic/46765/azeotrope

Organic Chemistry at CU Boulder.

Snelling, C.R. Distillation-Simple vs. Fractional.