BASIC ANALYTICAL CHEMISTRY CHM

256
LAB CASE STUDY REPORT

TITLE OF CASE STUDY (VIDEO): PLANAR CHROMATOGRAPHY

NO NAME OF MEMBERS STUDENT ID
1 ADRIANA FARIDA BINTI ABDULLAH 2020871438
2 NOOR NADA HAZIRAH BINTI 2020851882
ABDUL RAHMAN
3 NUR ALIA AYUNI BINTI ABDUL 2020882586
RAHMAN
4 NUR FARAHAN NAZIHA BINTI NOR 2020892478
HISHAM

GROUP : A4AS120413 (GROUP 2)

LECTURER’S NAME : MADAM NORSAKINA ZURINA BINTI

ZULKIFLI, MADAM ADIBATUL HUSNA BINTI FADZIL

DATE OF SUBMISSION : 24 JULY 2022

Declaration of Academic Honesty

Academic honesty or academic integrity is a very important virtue that all students should
uphold at all times.
I/We declare that the lab report submitted is not plagiarised and is entirely my/our own work,
and that no part of it has been copied from any work produced by other person(s)/ source(s)
or provided by any other student(s).
I/We understand that issuing a false declaration can result in severe penalties and I/we
am/are willing to be penalized if any form of copying is found valid.
TABLE OF CONTENT

No. Contents Pages

1. Objectives 1

2. Introduction 1

3. Analysis 2-3

4. Results and discussion 4-5

5. Conclusion 6

6. References 7
1.0 Objective

i. To separate the mixture of compound of spinach by planar chromatography (Thin

Layer Chromatography)
ii. To observe the distance travelled of the pigments of spinach.
iii. To find the Rf value of each pigment.

2.0 Introduction

Planar chromatography is a separation technique of the components of a mixture based

on the differences in the rate. This technique separates the mobile phase and stationary phase
of a component. Planar chromatography’s mobile phase moves through the capillary of action
while the stationary phase is supported on a flat plate. (Argekar, 2006) This class of
chromatography is divided into two main branches which are paper chromatography and thin-
layer chromatography (TLC). The thing that differences these two is the material used as the
plane for the stationary phase of the process. Paper chromatography uses paper while TLC
chromatography uses glass plate.

In this study, the chromatography that will be observed is the thin-layer chromatography
(TLC). The compounds in the mixture will travel through the flat plate and the distances will
tell how strong they interact with the stationary phase compared to the mobile phase. The
stationary of this chromatography will be in a solid phase while the mobile phase will be in a
liquid phase. Therefore, the type of separation of TLC chromatography is by absorption (solid-
liquid).

As the mixture travels up the flat plane, the components will be separated where polar
compound will move slowly with mobile phase as it is strongly bonded to the stationary phase
while the non-polar compound will moves faster as it is weakly bonded to the stationary phase.

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3.0 Analysis

Apparatus

1. TLC developing jar or a 150 mL beaker.

2. TLC silica gel plate
3. Mortar and pestle
4. Test tube
5. 50 mL beaker
6. Capillary tube
7. Pencil
8. Hair dryer

Chemical reagents

1. Sample of fresh leaves

2. Chromatography solvent
3. Acetone

PROCEDURE

Extraction of the Spinach Pigment

1. A pieces of fresh spinach were obtained and were cut into smaller pieces. The leaves
were grinded together with a few drops of acetone by using pestle and mortar. The leaves
were mixed well then.

2. The liquid extract was transferred into a small test tube.

3. All the extract was filtered into a 50 mL beaker. (If the extract is still diluted, the

extract should be concentrated by immersing the beaker into boiling water until the

final volume is about 2-3 mL).

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Preparation of Developing Solvent

1. The mobile phase (developing solvent) was prepared by mixing acetone and hexane in
3:7 ratio.

2. The developing solvent was poured into a chromatographic jar to a depth of

approximately 0.5 cm.

3. The jar was closed and shook slowly to equilibrate the solvent.

4. The jar was covered.

Preparation of the Extract to the TLC Plate

1. A line was draw approximately 1.5 cm from the bottom of the plate by using a pencil.

(Be careful not to scrap the coating material).

2. The leaf extract was deposited onto and along the line drawn by using a capillary

tube. The step was repeated until a very dark green dot is obtained. (Must allow the

extract to dry first before each addition).

Developing the TLC Plate

1. The loaded TLC plate was carefully placed into the TLC jar containing the developing

solvent.

2. The level of the solvent was placed below the line drawn/extract line.

3. The TLC jar was covered and was left undisturbed.

4. The position of the solvent was marked front when the solvent front has reached ¾ of

the length plate and was removed the TLC plate from the developing jar.

5. The Rf value were calculated using the formula below:

Calculations of Rf value: Distance travelled by the solute spot, (b)

Distance travelled by the solvent, (a)

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4.0 Results and Discussion

RESULTS:

Solvent front

(a)

5.1cm

(b)

Start line

Figure 1: Results of pigments travelled

Pigments Travelled distance, (b), Rf value

(cm)
Pale yellow 0.3 0.059
Yellow 1.0 0.196
Yellow green to bright green 1.7 0.333
Blue green 2.2 0.431

Table 1: Distance travelled of the pigments in spinach and the Rf value of each pigment

Calculations of Rf value: Distance travelled by the solute spot, (b)

Distance travelled by the solvent, (a)

The objective of this experiment is to observe the distance travelled of the pigments of
spinach and find the Rf value of each pigment. This experiment is conducted with prepared
the extraction of spinach pigment with grind the leaves that has been cut into smaller pieces
in pestle mortar then, prepared the developing solvent which is mixing acetone and hexane

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into 3:7 ratio, prepared the extract to the TLC plate with drop the extract using capillary tube
onto the drawn line of TLC plate and developed the TLC plate which is put the TLC plate into
the jar that contain developing solvent and close the lid. After doing all of the steps, the results
have been recorded and these recorded results used to calculate the Rf value. Calculates the
Rf value by using this formula:

Distance travelled by the solute spot,(b)

Rf value:
Distance travelled by the solvent,(a)

The results that we got from this experiment, there are four different pigments that travelled
through the TLC plate, which is pale yellow, yellow, yellow green to bright green and blue
green. The starting line is 0 cm, and the solvent front are 5.1 cm. The distance between start
line and solvent front are 5.1 cm which is known as (a). The distance travelled of the pigments
or label as (b) are pale yellow is 0.3 cm, yellow is 1.0 cm, yellow green to bright green is 1.7
cm and blue green is 2.2 cm. Then, the Rf value that we got from the pigments by using the
formula above are pale yellow is 0.059, yellow is 0.196, yellow green to bright green is 0.333
and blue green is 0.431.

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5.0 Conclusion

This experiment proved to distance travelled of the pigments of spinach and

find the Rf value of each pigment. It is also helped to widen our knowledges and understand
more about planar chromatography. Spinach pigment can be successfully processed using
planar chromatography in a very broad range of applications. Spinach pigment have more
than one photosynthetic pigment and have different rate of absorption. Thin-layer
chromatography has the flexibility of a few detection methods and the capacity to separate
mixtures of molecules with similar structures. It also has the benefits of being inexpensive
and simple to execute.

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6.0 References

1. Argekar, A. (2006). Planar Chromatography in Pharmaceutical Analysis. Encyclopedia of

Analytical Chemistry: Applications, Theory and Instrumentation.

2. Chromatography Today, (2015, March 19). What is planar chromatography. Retrieved

from https://www.chromatographytoday.com/news/solid-phase-extraction-
spe/34/breaking-news/what-is-planar-chromatography/33875

3. Libretexts. (2021, August 15). Thin layer chromatography. Chemistry LibreTexts.

Retrieved from
https://chem.libretexts.org/Ancillary_Materials/Demos_Techniques_and_Experiments/Ge
neral_Lab_Techniques/Thin_Layer_Chromatography
4. Science Direct (2022) Principles and Practice of Modern Chromatographic Methods
(Second Edition)Retrieved from https://www.sciencedirect.com/topics/chemistry/planar-
chromatography