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Lab #1 Thin Layer Chromatography

Organic Chemistry I (University of Ottawa)

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Thin Layer Chromatography

Written by:

Dagan Doubad, 300063550

CHM1321 Section Z02

TA: Vimarsha Bogahawatta

Date: January 15th 2018

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Protocol

Refer to pages 5-6 of the Experiment 1: Thin Layer Chromatography document

Observations

1. Solvents containing acetates had a strong smell

2. Most liquids used were clear and lacked viscosity
3. The unknown compound #94 was white and powder-like
4. The o-bromonitrobenzene sample was yellow
5. The hexanes used smelled slightly of gasoline
6. The side of the TLC plate in which we placed the organic mixtures on were easily scraped

Thin Layer Chromatography

Part A: Identifying the Components of an Unknown Mixture using TLC

Figure 1 Figure 2

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Part B: Effect of Solvent on TLC

Solvent System: Hexanes

Figure 3 Figure 4

Solvent System: EtOAc

Figure 5 Figure 6

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Part C: Identification & Separation of Unknowns

Reference Compound: o-bromonitrobenzene

Solvent System: 9:1 Hexanes: Ethyl Acetate

Reference Compound: m-bromonitrobenzene

Solvent System: 9:1 Hexanes: Ethyl Acetate

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Reference Compound: p-bromonitrobenzene

Solvent System: 9:1 Hexanes: Ethyl Acetate

Sample Calculations

Calculating Rf Value

R f = d 1/ d 2

Sample Calculation for Part A: Biphenyl

R f = d 1/ d 2
= 4.1/4.4
= .93

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Discussion

Throughout this lab, thin layer chromatography (TLC) was used. This mechanism allows the separation
of organic compounds in correspondence with their polarity. The TLC paper itself is composed of 2
phases: the stationary phase and the mobile phase. The stationary phase is polar and made of silica
whereas the mobile phase which is made up of the organic solvent (the eluent) and is not polar.

In part A, TLC was used to identify the components of an unknown mixture. Both biphenyl
and benzophenone were used as reference compounds. As seen in Figure 1, the unknown
sample #94 and benzophenone have the same Rf value and is a match for the unknown sample.

In part B, the effect of the solvent in thin layer chromatography was investigated. In part A, a 2:8
mixture of Ethyl acetate and hexanes was used as a solvent. In part B, the 2 compounds were isolated
and separately used as organic solvents. When ethyl acetate alone, the more polar substance, was the
organic solvent, the components in the mixture migrated significantly in relation to the solvent which
also led to some overlapping and gave somewhat misleading results. This action is due to components in
the mixture undergoing hydrogen bonding with the oxygen molecules in ethyl acetate in the mobile
phase more readily than with the silica gel. When hexanes alone were used, the opposite was true. There
was less migration and low Rf values, as shown in Figures 3 and 4. The components in the mixture
undergo hydrogen bonding more readily with the silica gel in the mobile phase than the hexane solvent
and less migration occurs. The extra readings on the reference sample in the TLC with the hexanes
(Figure 3) as a solvent could be explained as being due to cross contamination during the experiment.

In part C, thin layer chromatography was used to identify which substances (o-bromonitrobenzene,
m-bromonitrobenzene or p-bromonitrobenzene) made up an unknown mixture. To investigate this,
3 TLC plates were analysed witch each isomer as a reference solution. When examining the 3 TLC
plates and the Rf values for the unknown compounds and the 3 isomers, we see that the unknown
mixture is made up of p-bromonitrobenzene and o-bromonitrobenzene. The lower Rf value
corresponds to o-bromonitrobenzene and the larger Rf value corresponds to p-bromonitrobenzene.
However, there is no Rf value in the components of the unknown sample that is similar to the R f
value of m-bromonitrobenzene and thus, the unknown sample is not made up of the latter.

It is also important to note the various sources of error within this experiment. First of all, the mixtures and
reference compounds used were all clear and when dropping them onto the TLC paper, it was hard to see
how much, if at all, had made it onto the paper. This, not only made it hard to analyze the paper under the
UV light due to accidentally putting too much onto the paper which lead to overlapping of the dots, but the
opposite was also seen to be true. The TLC plate in part C which used m-bromonitrobenzene as the reference
compound, seemed to have only one dot reading in the co-spot lane. This could have been due to not
unknowingly not putting enough of the unknown mixture in the co-spot lane.

It is also important to account for the slight discrepancies in the Rf values for the same compound when using
thin layer chromatography, specifically in part C of this experiment where it was shown that the mixture was
a mix of both p-bromonitrobenzene and o-bromonitrobenzene. When analysing the migration

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of the 2 compounds on the TLC plates, the TLC plate which used p-bromonitrobenzene showed the
same Rf values in the reference lane as in the sample lane (0.85). However, there is a slight difference
between the same 2 lanes in the TLC plate which used o-bromonitrobenzene where the reference spot
read Rf=0.25 and the same compound in the sample lane had an Rf value of 0.20. This difference can be
accounted for by the random error of the ruler used to measure d1 and d2 values.

Discussion Questions

1. Why is it important to make co-spots last?

a. Making co-spots last ensures that there is no cross contamination via the capillary.
2. How does increasing the polarity of the solvent system affect the results of the TLC?
a. Increasingly polar solvents, due to their larger potential for hydrogen bonding in
comparison with the silica gel, cause more migration. The components of the mixture interact with the
solvent through hydrogen bonding and follow the solvent in the mobile phase and migration is
significant which leads to a larger Rf value.
3. A student is monitoring a reaction using TLC. The chemical reaction converts compound A
into compound B and she uses molecule A as her reference. Compound A has an R f value of 0.45
while compound B has an Rf of 0.68 in the solvent system being used.
a. Draw a picture of the TLC plate at the beginning of the reaction

b. Draw a picture of the TLC after 50% completion.

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c. Draw a picture of the TLC plate at the end of the reaction.

d. Why is it better to use a sample of molecule A rather than molecule B to follow the reaction? i.
This makes it easier to monitor the progress of the reaction as we can measure the
merging of the 2 spots on the B column.

4. A student is monitoring a reaction involving compounds C and D using TLC. Compound C has
an Rf value of 0.43; Compound D has an Rf of 0.40

a. Draw the TLC plate at the beginning of the reaction

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b. Draw the TLC plate after 50% completion

c. Draw TLC plate at the end of the reaction

d. Why is it important to use a co-spot?

I. In this case, the Rf values for both compounds are very similar and a co-spot helps to
highlight the disparity between the two compounds when monitoring reaction progress that we may
miss without this as a reference.

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5.

Benzyl Alcohol Benzaldehyde Benzyl Acetate

a. Benzyl alcohol, due to its hydroxyl group will be the most polar. Benzaldehyde has more of
an ability to engage in hydrogen bonding than benzyl acetate, due to its hydrogen molecule, than
benzyl acetate.

Aniline N,N-dimethylaniline Naphthalene

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b. Aniline is the most polar due its distribution of dipoles being uneven, followed by N,N
dimethylaniline and naphthalene which has the most even distribution of dipoles.

Benzophenone Biphenyl

Benzoic Acid

c. Due to its carboxyl group and the OH molecule present, benzoic is the most polar, followed by
benzophenone and then biphenyl. Benzophenone retains some polarity due to it being a ketone
and containing a carbonyl group, however not as polar as benzoic acid. Biphenyl is the least
polar as it does not contain any polar functional groups.

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Raw Data

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