Unit: 2 Practical Scientific Procedures and Techniques
Separate To Identify
Chromatography
Introduction
Chromatography is an experiment that is utilized to separate components in a mixture. The mixture
that is made up of different components is carried by the mobile phase, which is the solvent, and it is
carried through the stationary phase, which is the type of chromatography paper in most of the
cases. Chromatography is used for many reasons such as creating vaccines, food testing, beverage
testing, drug testing and forensic testing. There are many different types of chromatography such as
paper chromatography, thin-layer chromatography (TLC), high performance liquid chromatography
and gas chromatography.
In this experiment, the aim is to successfully carry out paper chromatography of extracted plant
pigments, thin layer chromatography (TLC) of extracted plant pigments, and paper chromatography
of amino acids. Additionally, the different components of the different mixtures will move through
the stationary phase at different speeds causing them to separate. The Rf value will be worked out
using the distance travelled by a given component and dividing it by the distance travelled by the
solvent front to find out what the components in the mixtures are.
Paper Chromatography – Extracted Plant Pigments
Paper chromatography was carried out on extracted plant pigments to visualize the different
components present in the plant, which in this case was grass.
Equipment List
Chromatography paper – This is utilized as the stationary phase of the practical being carried
out
Isopropyl alcohol – This is added into the mortar while grinding the grass because it makes
the separation of pigments easier
Sand – This is added into the mortar while grinding the grass because it breaks the plant cells
of the grass
Grass – This is the solution whose components will be separated
Pestle and Mortar – This is used to grind the grass to get a paste with the pigment
Pencil and Ruler – These are utilized to draw the pencil lines (solvent front and origin) on the
chromatography paper.
Solvent (Cyclohexane, propanone, and petroleum ether in a ratio of 3:2:1) – This is used as
the mobile phase of the practical that would be carried out
Capillary tube – This is used to spot the extracted plant pigment from the grass on the
chromatography paper.
Gas jar – This is the container utilized to separate the components of the plant pigment
Watch glass – This is used as the lid of the container to avoid the saturation of the solvent to
decrease as it may be affected by the air
Hair dryer – This is for the drying process of spot on the chromatography paper to become
swifter
Marker pen – This is utilized to mark the pencil line on the gas jar to make sure that the
solvent isn’t poured over the pencil line
Method
1. Firstly, some grass was teared in small pieces and placed in a mortar. Some Isopropyl alcohol
was poured inside the mortar as well as a pinch of sand granules. The sand granules were
added into the mortar while grinding the grass so that the plant cells could be broken down.
, 2. The pestle and mortar were used to extract the pigment from the grass by grinding the grass
for several minutes until a green coloured paste was seen in the mortar.
3. A piece of chromatography paper was taken, and a pencil line was drawn at the bottom to
mark the origin, which is where a spot of the pigment will be placed later. To make the origin
line, a pencil must have been used as it doesn't dissolve in the solvent meaning that it is
insoluble.
4. The chromatography paper with the pencil line was placed inside the gas jar to mark where
the pencil line was drawn on the paper with a marker pen so that when the solvent was
poured inside the gas jar, it wouldn’t be above it as it will cause the experiment to be started
again.
5. A capillary tube was lightly dipped into the pigment to pick up some of the pigment and
making a small spot on the pencil line (origin) previously drawn. The aim was to make a
small but concentrated spot and to do this a hair dryer was used to dry the spot on the
pencil line and then repeating the whole process of spotting the pigment on the origin 3 to 5
times.
6. The solvent was poured inside the gas jar below the marked line.
7. The top of the chromatography paper with the pigment spot was folded in a zig zag pattern
to make sure that it would fit inside the gas jar, and it was placed inside the gas jar while
being careful that as soon as the paper would touch the solvent it would not be moved as it
would lead to the components to move up the paper diagonally.
8. A watch glass was placed on top of the gas jar to make sure that the gas jar is saturated with
the vapour of the solvent and as minimum air as possible because it would affect the rate of
the solvent while it would be moving up the paper.
9. The gas jar was left untouched until the solvent reached the top of the chromatography
paper at about 1cm or 2cm away from the end of the paper.
10. The watch glass was then removed, and the chromatography paper was removed carefully.
11. A pencil line was drawn across the paper where the solvent got up to before it would dry to
display the end of the journey of the solvent, and this was called the solvent front.
Thin Layer Chromatography – Extracted Plant Pigments
Thin layer chromatography was carried out on extracted plant pigments although it was already
carried out on chromatography paper because it would show a more precise and accurate image of
the different components present in the grass.
Equipment List
Silica gel – This is utilized as the stationary phase of the practical being carried out
Isopropyl alcohol – This is added into the mortar while grinding the grass because it makes
the separation of pigments easier
Sand – This is added into the mortar while grinding the grass because it breaks the plant cells
of the grass
Grass – This is the solution whose components will be separated
Pestle and Mortar – This is used to grind the grass to get a paste with the pigment
Pencil and Ruler – These are utilized to draw the pencil lines (solvent front and origin) on the
silica gel paper.
Solvent (Cyclohexane, propanone, and petroleum ether in a ratio of 3:2:1) – This is used as
the mobile phase of the practical that would be carried out
Capillary tube – This is used to spot the extracted plant pigment from the grass on the silica
gel paper
Beaker – This is the container utilized to separate the components of the plant pigment
Watch glass – This is used as the lid of the container to avoid the saturation of the solvent to
decrease as it may be affected by the air
Hair dryer – This is for the drying process of spot on the silica gel paper to become swifter
Separate To Identify
Chromatography
Introduction
Chromatography is an experiment that is utilized to separate components in a mixture. The mixture
that is made up of different components is carried by the mobile phase, which is the solvent, and it is
carried through the stationary phase, which is the type of chromatography paper in most of the
cases. Chromatography is used for many reasons such as creating vaccines, food testing, beverage
testing, drug testing and forensic testing. There are many different types of chromatography such as
paper chromatography, thin-layer chromatography (TLC), high performance liquid chromatography
and gas chromatography.
In this experiment, the aim is to successfully carry out paper chromatography of extracted plant
pigments, thin layer chromatography (TLC) of extracted plant pigments, and paper chromatography
of amino acids. Additionally, the different components of the different mixtures will move through
the stationary phase at different speeds causing them to separate. The Rf value will be worked out
using the distance travelled by a given component and dividing it by the distance travelled by the
solvent front to find out what the components in the mixtures are.
Paper Chromatography – Extracted Plant Pigments
Paper chromatography was carried out on extracted plant pigments to visualize the different
components present in the plant, which in this case was grass.
Equipment List
Chromatography paper – This is utilized as the stationary phase of the practical being carried
out
Isopropyl alcohol – This is added into the mortar while grinding the grass because it makes
the separation of pigments easier
Sand – This is added into the mortar while grinding the grass because it breaks the plant cells
of the grass
Grass – This is the solution whose components will be separated
Pestle and Mortar – This is used to grind the grass to get a paste with the pigment
Pencil and Ruler – These are utilized to draw the pencil lines (solvent front and origin) on the
chromatography paper.
Solvent (Cyclohexane, propanone, and petroleum ether in a ratio of 3:2:1) – This is used as
the mobile phase of the practical that would be carried out
Capillary tube – This is used to spot the extracted plant pigment from the grass on the
chromatography paper.
Gas jar – This is the container utilized to separate the components of the plant pigment
Watch glass – This is used as the lid of the container to avoid the saturation of the solvent to
decrease as it may be affected by the air
Hair dryer – This is for the drying process of spot on the chromatography paper to become
swifter
Marker pen – This is utilized to mark the pencil line on the gas jar to make sure that the
solvent isn’t poured over the pencil line
Method
1. Firstly, some grass was teared in small pieces and placed in a mortar. Some Isopropyl alcohol
was poured inside the mortar as well as a pinch of sand granules. The sand granules were
added into the mortar while grinding the grass so that the plant cells could be broken down.
, 2. The pestle and mortar were used to extract the pigment from the grass by grinding the grass
for several minutes until a green coloured paste was seen in the mortar.
3. A piece of chromatography paper was taken, and a pencil line was drawn at the bottom to
mark the origin, which is where a spot of the pigment will be placed later. To make the origin
line, a pencil must have been used as it doesn't dissolve in the solvent meaning that it is
insoluble.
4. The chromatography paper with the pencil line was placed inside the gas jar to mark where
the pencil line was drawn on the paper with a marker pen so that when the solvent was
poured inside the gas jar, it wouldn’t be above it as it will cause the experiment to be started
again.
5. A capillary tube was lightly dipped into the pigment to pick up some of the pigment and
making a small spot on the pencil line (origin) previously drawn. The aim was to make a
small but concentrated spot and to do this a hair dryer was used to dry the spot on the
pencil line and then repeating the whole process of spotting the pigment on the origin 3 to 5
times.
6. The solvent was poured inside the gas jar below the marked line.
7. The top of the chromatography paper with the pigment spot was folded in a zig zag pattern
to make sure that it would fit inside the gas jar, and it was placed inside the gas jar while
being careful that as soon as the paper would touch the solvent it would not be moved as it
would lead to the components to move up the paper diagonally.
8. A watch glass was placed on top of the gas jar to make sure that the gas jar is saturated with
the vapour of the solvent and as minimum air as possible because it would affect the rate of
the solvent while it would be moving up the paper.
9. The gas jar was left untouched until the solvent reached the top of the chromatography
paper at about 1cm or 2cm away from the end of the paper.
10. The watch glass was then removed, and the chromatography paper was removed carefully.
11. A pencil line was drawn across the paper where the solvent got up to before it would dry to
display the end of the journey of the solvent, and this was called the solvent front.
Thin Layer Chromatography – Extracted Plant Pigments
Thin layer chromatography was carried out on extracted plant pigments although it was already
carried out on chromatography paper because it would show a more precise and accurate image of
the different components present in the grass.
Equipment List
Silica gel – This is utilized as the stationary phase of the practical being carried out
Isopropyl alcohol – This is added into the mortar while grinding the grass because it makes
the separation of pigments easier
Sand – This is added into the mortar while grinding the grass because it breaks the plant cells
of the grass
Grass – This is the solution whose components will be separated
Pestle and Mortar – This is used to grind the grass to get a paste with the pigment
Pencil and Ruler – These are utilized to draw the pencil lines (solvent front and origin) on the
silica gel paper.
Solvent (Cyclohexane, propanone, and petroleum ether in a ratio of 3:2:1) – This is used as
the mobile phase of the practical that would be carried out
Capillary tube – This is used to spot the extracted plant pigment from the grass on the silica
gel paper
Beaker – This is the container utilized to separate the components of the plant pigment
Watch glass – This is used as the lid of the container to avoid the saturation of the solvent to
decrease as it may be affected by the air
Hair dryer – This is for the drying process of spot on the silica gel paper to become swifter
