1

Laboratory Schedule The experiments have been designed such that they can be completed in the allotted time. It is expected, however, that you arrived well-prepared. You must leave the laboratory at 11:25 at the latest. Your final cleanup of glassware and work area must begin no later than 11:15.

Month Monday (A) Tuesday (B) Thursday (D) Friday (E)

September 5 Labor day

6 —

8 —

9 —

September 12 Check-in

13 Check-in

15 (Quiz #1) Check-in

16 Check-in

September 19 Exp. 1

20 Exp. 1

22 (Quiz #2) Exp. 1

23 Exp. 1

September/ October

26 Exp. 2

27 Exp. 2

29 (Quiz #3) Exp. 2

30 Exp. 2

October 3 Exp. 3

4 Exp. 3

6 (Quiz #4) Exp. 3

7 Exp. 3

October 10 NO LAB!

11 EXP 4

13 (Quiz#5) EXP 4

14 EXP 4

October 17 Exp. 5

18 Exp. 5

20 Exp. 5

21 Exp. 5

October Reading week

24 NO LABS!

25 NO LABS!

27 (Quiz #6) NO LABS!

28 NO LABS!

October/ November

31 Exp. 6

1 Exp. 6

3 (Quiz #7) Exp. 6

4 Exp. 6

November 7 Exp. 7

8 Exp. 7

10 (Quiz #8) Exp. 7

11 Exp. 7

November 14 Exp. 8

15 Exp. 8

17 (Quiz #9) Exp. 8

18 Exp. 8

November 21 Exp. 9

22 Exp. 9

24 (Quiz #10) Exp. 9

25 Exp. 9

November/ December

28 Exp. 10

29 Exp. 10

1 Exp. 10

2 Exp. 10

December Section A, F (MONDAY students) must perform Exp 4. Wed, Dec 7th.

2 Quizzes (pre-lab) and Lab Reports

The lab experiments are typically done in groups of two. The schedule below tells

you if your lab report must be done individually or with your partner.

A quiz will be posted on virtual campus every Thursday. You are responsible for

printing out and answering the quiz. The quiz is due (in your TA’s box in the lab) on the

following Monday before 4 pm.

As part of each pre-lab, you must prepare the following in your lab notebook: a

table of reactants and solvents, as well as a table of observations. The table of reactants

must contain the following information: reactant/solvent name, molar mass, quantity,

density (if a pure liquid), number of moles and number of equivalents (equivalents are

not required for solvents). You will fill the table of observations throughout the lab

period. The lab notebook component of your pre-lab will be evaluated by your TA.

Experiment 1. Report: handed in by each student

Experiment 2. Report: One report per group of two

Experiment 3. Report: handed in by each student

Experiment 4. Report: One report per group of two

Experiment 5. Report: handed in by each student

Experiment 6. Report: No lab report

Experiment 7. Report: One report per group of two for both experiments 6 and 7.

Experiment 8. Report: handed in by each student

Experiment 9. Report: handed in by each student

Experiment 10. Report: One report per group of two

3

EXPERIMENT 1

A. EXTRACTION AND RECRYSTALLIZATION

B. THIN LAYER CHROMATOGRAPHY (TLC)

Required reading: Solomons & Fryhle, Organic Chemistry, 10th Ed., Section 2.13 Annex I ………….. Melting point apparatus

Annex II …………. Re-crystallization Annex III ………… Extraction Annex IV ………… Chromatography Annex V ………….. Example of a lab report

INTRODUCTION The purpose of this experiment is to re-acquaint you with two important

techniques learned last year, extraction and re-crystallization, and introduce the

technique of analysis of reaction mixtures and qualitative determination of product

purities using thin layer chromatography [TLC].

In this experiment you are asked to separate a mixture of benzoic acid and benzil

into the two components, purify each by re-crystallization, verify the purity of each of the

separated components by melting point and TLC, and determine the composition of the

mixture. Each of the samples given to students has a composition, varying from 25% to

75% benzoic acid, the remainder being benzil.

- colourless plates, mp = 122 °C - yellow prisms, 95 °C

- natural product found in a variety - a synthetic compound that has been of fruit and berries used in the synthesis of heterocyclic

- useful as food preservative compounds

You will take advantage of the acid-base properties of the benzoic acid in order to

separate it from the benzil. The two components of the mixture, when in their neutral

form, are soluble with common organic solvents but have a low solubility in water.

CO2HO

OBenzilBenzoic acid

4 Benzoic acid will react with bases such as sodium hydroxide, sodium carbonate or

bicarbonate to form sodium benzoate, an ionic compound that is highly soluble in water

but insoluble in common organic solvents (such as methylene chloride or ether). We can

therefore separate the two compounds by first treating the mixture, dissolved in an

organic solvent, with aqueous NaOH, then extracting. The sodium benzoate will be found

in the aqueous phase, while the benzil, which does not undergo acid-base reactions, will

remain in the organic phase. In order to recuperate the benzoic acid, acidification of the

aqueous phase is all that is required. The benzoic acid, being insoluble in water, will

precipitate from this acidic aqueous environment.

- very soluble in CH2Cl2 - very soluble in water - sparingly soluble in water - sparingly soluble in CH2Cl2

Once the constituents have been separated, you will use the technique of

recrystallization to purify each compound (see annex 2). This technique is based on the

differences in solubility of the two compounds in hot and cold solvents. The impure

compound is first dissolved in a minimum of hot solvent, the hot solution is filtered to

remove any insoluble impurities, then the hot solution is allowed to cool slowly, and

lastly, the solution is filtered to obtain the pure crystals.

The efficiency of your separation will be analyzed by thin layer chromatography

(TLC). See Annex IV for details about this technique. You will use plastic plates covered

with a thin layer of silica gel (SiO2•x H2O), a very polar compound. The silica gel is the

stationary phase. A small amount of the compound or mixture to be analyzed (a few mg

dissolved in a small amount of organic solvent) is spotted on the plate (~ 1 cm from the

base), using thin capillary tubes.

O

ONa

OH

O

benzoic acid sodium benzoate

NaOH

5 To properly follow and analyze the progress of a reaction by TLC, it is standard to

have 3 points on the TLC plate: 1) starting material(s), 2) a co-spot containing the starting

material(s) and the final reaction mixture (or product), and 3) the final reaction mixture

(or product). The plate is eluted and the compounds migrate up the plate at different rates

depending on their polarity. The rate of migration depends on the interaction of each

compound with two phases: the stationary phase, silica gel in this case (the compound is

adsorbed1 onto the silica gel), and the mobile phase (an organic solvent or mixture of

solvents), which passes over/through the stationary phase. Generally, more polar

compounds interact most strongly with the polar silica gel and therefore migrate more

slowly that less polar compounds. Less polar compounds generally migrate faster than

more polar compounds.

In summary: (1) The silica gel, the stationary phase, is polar: more polar compounds interact more

strongly with this phase than less polar compounds.

(2) Increasing the polarity of the mobile phase (solvent) causes ALL compounds to

migrate further on the plate.

(3) The compounds, once separated, can be visualized on the TLC plate using ultra-

violet light or a staining method.

In this lab, you will use an ultra-violet lamp to visualize the compounds that have

separated on TLC.

1 Adsorb: superficial accumulation of a compound onto a surface

start line

Finish line of solvent front

Compound a

Compound b

0 mm

x mmdistance of migration of a

distance of migration of solventRf(a) =

distance of migration of b

distance of migration of solventRf(b) =

6

The TLC results are recorded for each compound by calculating the ratio of the

distance of migration of a particular component to that of the solvent. This is known as

the Rf (retention factor). The Rf is characteristic for each compound under a specific set

of conditions (stationary phase and elution solvent).

Distance traveled by the compound from the origin Rf = -------------------------------------------------------------- Distance traveled by the solvent from the origin

In the annex you will find a more complete explanation of chromatography. You

are responsible for reading this annex before coming to the laboratory.

7 REMINDER OF KEY CONCEPTS

Extraction • Technique employed to separate components of a mixture.

• Based on differences in solubility of the compounds in two

immiscible solvents.

• Extraction generally implies movement of a desired

compound from one phase into another, while washing

generally means keeping the desired compound in one phase

and forcing the impurities into the other.

• Generally used at the end of a reaction to separate organic

compounds, which are typically quite soluble in organic

solvents, from inorganic compounds, which are typically

much more soluble in water than in organic solvents.

• We can increase the quantity of a compound transferred from

solvent A to solvent B by using more of solvent B. It is,

however, move efficient to perform multiple extractions with

a smaller volume of solvent B than to do a single extraction

with a larger volume of B (3 × 10 mL of solvent B transfers

more of the desired compound into B than 1 × 30 mL).

• Acid-base properties of a compound are frequently used to

render a compound more/less soluble in a given solvent.

Recrystallization

• An important purification technique, especially in organic

chemistry.

• Based on the differences in solubility of a compound and

impurities in hot and cold solvents.

• Typically, to make the re-crystallization technique effective,

the product needs to have undergone a first purification to

remove major impurities.

Solvent The ideal solvent satisfies the following criteria:

8 • Non reactive

• Boiling point lower than the melting point of the crystal

• Dissolves a large quantity of product when hot

• Dissolves a small quantity of product when cold

• Easily removed by evaporation

• Non toxic, non-flammable, and cheap

• Either readily dissolves impurities or does not dissolve them

at all

• When a mixture of solvents is used, they must be miscible

Crystal formation

The rate of crystal formation is an important factor in the

determination of purity. Fast cooling results in pockets of solvent

containing impurities inside the crystals. This is termed

precipitation instead of crystallization. Slow cooling leaving the

flask undisturbed is recommended.

It’s important to chose the solvent carefully and to use a minimum

amount of hot solvent to dissolve the compound. Using too much

solvent will result in a low yield, because a significant quantity of

the desired compound will remain dissolved in the solvent even

when cold.

Melting Points

The melting points of your isolated products will assist you in

analyzing their purity. The melting point must always be reported

as a range: the first temperature represents the start of the

transition from solid to liquid, and the second point represents the

end of this transition. A large melting point range indicates the

presence impurities, as does a range that is lower than the

literature value.

9 MANIPULATION

All transfers or liquids and solids should be as quantitative as possible. For

example, in the transfer of the original solution to the separatory funnel some of it

invariably remains in the small Erlenmeyer flask. To recover most of this, rinse the flask

with a small amount of dichloromethane, and add this extra volume to the separatory

funnel.

Use the right size of equipment! In this laboratory course, this aspect is not

always possible because you do not have all sizes of each type of equipment.

Nevertheless do not to re-crystallize a sample from 10–20 mL of solvent using a 500 mL

flask; a 50 mL or 25 mL flask would be much more appropriate.

An Erlenmeyer flask is the appropriate vessel for re-crystallization (not a beaker).

SAFETY

Evaporation of all organic solvents must be done in the fumehood.

Read the instructions in the annex on the proper use of the extraction funnel

before starting the experiment. Do not hesitate to ask your TA for help.

Before the experiment, wash your glassware with water and then acetone in order

to avoid contamination. If you spill acid/base or reactants on yourself, wash immediately

with water. You should always wash your hands before leaving the lab.

Work carefully!

EXPERIMENTAL PROCEDURES

Isolation of benzoic acid

1. Preheat 900 mL of water in a 1 L beaker. You will need it for the re-crystallization

steps.

10 2. Dissolve 1 g (record the precise mass used) of the benzoic acid and benzil mixture

(provided by your TA) in 10 mL of CH2Cl2 in a 50 mL Erlenmeyer. Record the

label of your vial.

3. Keep a small quantity (a few milligrams) of the mixture for your TLC (done later).

4. Use the wooden support to hold the extraction funnel. Never fill the extraction

funnel more than ¾ full—this head-space is needed to allow mixing when shaking

the funnel. Before filling the funnel, make sure it is not leaking (test with a small

volume of distilled water).

5. Transfer the solution into the extraction funnel. Rinse the Erlenmeyer with 5 mL of

CH2Cl2 to maximize the transfer of the mixture—transfer this to the extraction

funnel also.

6. Extract the solution with 10 mL of 2M NaOH. Carefully separate the two phases.

Use 50 mL Erlenmeyers (preferred) or 50 mL beakers to hold the aqueous and

organic phases obtained.

7. Extract the organic phase again with 5 mL of 2 M NaOH. Keep the aqueous phase!

***Reminder: The solvent with the highest density will be on the bottom.

Remember to record your observations!

8. Combine the basic aqueous phases and set aside the organic phase for now.

9. Acidify the aqueous phase by adding concentrated HCl dropwise. The benzoic acid

will precipitate from the aqueous phase. Use pH paper to verify the pH of the

aqueous mixture. Don’t add too much acid or NaCl will start to precipitate from the

solution.

10. Cool the acidic mixture in an ice bath (this should actually be an ice-water bath) to

complete the precipitation of the benzoic acid. Use a 150 mL beaker to make the ice

bath.

11. Filter the product using a Buchner funnel. Wash the precipitate with a small amount

of cold water to remove any traces of NaCl and dry the solid by suction.

11 Re-crystallization of benzoic acid

12. Dissolve the crude product in a minimum of hot distilled water in a 50 mL

Erlenmeyer. Use the hot water bath to warm up the distilled water. There might be

some insoluble impurities (see step 13).

13. If there are any insoluble impurities, filter the hot solution using a glass funnel

lightly plugged with cotton.

14. Once the solution has cooled to room temperature, place in an ice-bath to complete

the re-crystallization.

15. Filter the product by suction filtration using a Buchner funnel and dry by suction

filtration for at least 20 minutes. Note your observations. Dry the crystals as much

as possible. You can begin the re-crystallization of the benzil during this time.

16. Determine the melting point (record as a range) and mass of the crystals

obtained. Using the melting point apparatus, you will determine the melting point

range for your purified product. Recall that the crystals must be dry for acceptable

results. It is sometimes necessary to let the crystals dry for a while to allow

complete solvent evaporation.

Isolation of benzil

1. Add approximately 1 g of sodium sulfate to the organic phase, CH2Cl2. Swirl

regularly over a period of about 2 minutes to allow complete drying of the solution.

At this point no droplets of water should be visible and some of the sodium sulfate

should still be a mobile granular solid.

2. Filter the solution into a 50 mL Erlenmeyer using a funnel that is lightly plugged

with some cotton. Rinse the flask containing the Na2SO4 with a further 5 mL of

CH2Cl2 solution.

3. Evaporate the solvent (b.p. = 42 °C) on a steam bath. Swirl the flask fairly

continually (holding it securely with tongs) to prevent uneven boiling or splattering.

Discontinue the evaporation when you have a paste or a solid.

12

Re-crystallization of benzil

4. In a water bath, heat a solution containing 5 mL ethanol and 1 mL H2O in a small

beaker.

5. Dissolve the solid or paste in a minimum amount of a hot mixture of ethanol-

water (5:1). Do this by adding the solvent mixture in small portions and swirling

regularly. It is crucial to do this step well in order to get good results. You should

need approximately 1 mL / 100 mg of benzil.

6. Allow the solution to cool to room temperature. At this point, the crystallization

should have begun.

7. Once the solution is at room temperature, complete the crystallization by letting the

flask stand quietly for a few minutes in an ice-bath (clamp your flask!). If the

crystallization is allowed to proceed slowly the product should appear as yellowish

needles. If crystallization is very rapid then a yellow powder is obtained. Note

your observations.

8. Filter and dry the product by suction filtration using a Buchner funnel.

9. Allow to dry for at least 10 minutes, then weigh, and determine its mp (reported as

a range).

In your report, indicate the mass of each product isolated, as well as the percent

composition (% mass) of the original mixture.

THIN LAYER CHROMATOGRAPHY

Take the time to read the annex on chromatography before starting this part.

1. Use 3 TLC plates (about 65 by 20 mm each). These are made of a sheet of plastic

covered with a thin layer of silica gel.

2. Practice your spotting technique by using small pieces of TLC plates before you run

your experimental samples. Your TA will explain how to do this.

13 3. Lightly pencil in lines about 1 cm from each end of the plate.

4. Use a 4:6 mixture of hexanes to ethyl acetate (v/v) as the developing mixture. Add a

small amount of developing mixture, about 10 mL, into a developing jar (so that the

solvent level will be below the starting line of your plate).

5. Dissolve about 1 mg of the initial benzoic acid and benzil mixture in a minimum

amount of CH2Cl2.

6. Dissolve about 1 mg of the benzoic acid obtained after re-crystallization in 1 mL of

CH2Cl2 in a small test tube.

7. Repeat the above step with benzil, in a second test tube.

8. Spot 1–2 drops of the benzoic acid solution on the left, 1–2 drops of benzil on the

right, and 1–2 drops of the initial mixture in the middle of the TLC plate (see

diagram).

9. Carefully place the TLC plate into the jar while avoiding any splashing, and be sure

the plate is as upright as possible. Cap the jar. The starting line of the compounds

must be above the solvent level.

10. Allow solvent to develop to about 5 mm from the top of the plate and mark the

finish line with a pencil.

11. Allow the solvent to evaporate off the TLC plate and then visualize the developed

plate with a UV Lamp and use a pencil to mark the spot migrations. Record the

results in your lab book. (NEVER LOOK DIRECTLY AT A UV LAMP).

12. Place the plate into a closed container containing a few iodine crystals. Each brown

spot that appears after a few minutes represents at least one chemical compound.

14 13. Record your results in your lab book (draw the TLC’s and record the Rf values).

Effect of the solvent on Rf

1. Develop a TLC plate spotted as described above with benzoic acid, the mixture, and

benzil in a 9:1 mixture of hexanes:ethyl acetate as the developing solvent.

2. Dissolve about 1-2 mg of benzoic acid in about 1 mL of dichloromethane and then

add 3 drops of triethylamine. Spot this mixture on a TLC plate and develop this

along side a sample of benzoic acid, the original mixture, and benzil in a 1:9

hexanes:ethyl acetate mixture developing solvent.

The results section of your report should include drawings of all the TLCs and the

Rf values in a table, as well as typical requirements for a results section. In the discussion,

briefly explain each of your observations. Refer to the description of requirements for a

lab report and consult the example of a report in the annex.

15 QUESTIONS

1. Why is ethyl acetate-H2O not a suitable solvent combination for re-

crystallization?

2. Draw a flow chart to explain how the following pairs can be separated. Draw the

structures of all compounds involved and the mechanisms of all reactions that

take place.

(a)

(b) NaBr + cholesterol

(c) CH3(CH2)8CO2H + CH3(CH2)8CH2NH2

(decanoic acid) (1-aminodecane)

(d) cyclohexanecarboxylic acid + cyclohexanol

3. In the following sets, which compound would be expected to have the smallest Rf

in a silica gel TLC using a 1:1 mixture of hexanes:ethyl acetate as eluant? Use a

sentence to explain your reasoning. Draw the Lewis structure of each compound.

a) Benzyl alcohol, benzaldehyde, benzyl acetate

b) PhCH2OH or PhCH(CH3)OH;

c) aniline, N,N-dimethylaniline or naphtalene;

d) cyclodecanone, cyclodecene or cyclodecane.

4. Draw a TLC plate that corresponds to the following description:

Compound A, Rf 0.6;

Compound B, Rf 0.1 and a mixture of 2:1 of A:B

NH2

aniline naphtalene

16 5. Indicate how you would separate sodium chloride from benzoic acid. Be

practical!

6. In the following sets, which compound would be expected to have the smallest Rf

in a silica gel TLC using a 4:1 mixture of hexane and ethyl acetate as eluant.

Which would have the smallest Rf if the solvent were changed to 1:4 hexane ethyl

acetate? Use a phrase to explain your reasoning.

1) cyclohexane; cyclohexanol

2) 1-octanol; 3-octanol

3)

4)

7. In silica gel chromatography, alcohols are typically more polar than ketones.

There are some exceptions to this, however, as in the following case below (A is

less polar than B). Propose a reason for this.

8. A student performs the following reaction:

What will be the product?

NH2 NCH3

CH3;

H

H

CO2CH3H

H

CO2H;

OHO

A B

O

NaBH4

17

Looking at the TLC below, what advice would you give the student concerning the status of the reaction?

9. Draw a mechanism (arrows) and give the products of each of the following

reactions.

a)

b)

c)

CO2H CH3O Na

CO2H2SO4

Na

NCH3

CH3 HCl