Unit 2 Assignment C Anonymous
Chromatography is a valuable analytical technique utilized in scientific
laboratories to identify and determine the composition of mixtures. It is based on
the principle that materials have varying affinities for the stationary and mobile
phases, therefore moving at varying rates and thus separating over time.
Chromatography finds extensive application in chemical, biological, and
environmental science for qualitative and quantitative analyses.
In addition to chromatography, several other techniques are commonly used in
laboratories to separate and identify components in mixtures:
Filtration:
Filtration is used to separate an insoluble solid from a liquid. A common example
is separating sand from water. The mixture is passed through filter paper, which
traps the solid particles while allowing the liquid to pass through. This method is
based on particle size.
Distillation:
Distillation separates mixtures based on differences in boiling points. For
example, ethanol can be separated from water by heating the mixture until
ethanol evaporates, then condensing it back into a liquid. This method is
especially useful for purifying liquids or separating liquid-liquid mixtures.
Centrifugation:
Centrifugation separates components of a mixture based on density. The sample
is spun at high speed in a centrifuge, causing denser substances to settle at the
bottom. It is widely used in biology for separating blood components or isolating
cell parts.
In the present lab report, various chromatographic techniques were performed to
separate and identify the components of plant pigments and amino acids.
Techniques employed included paper chromatography and thin layer
chromatography (TLC). Both the techniques included appropriate observation of
separation patterns and determination of retention factor (Rf) values, which are
used to identify unknown compounds using known standards for comparison.
The objective of this experiment was to demonstrate proficient practical skills in
chromatography, illustrate safe working practice, interpret the resulting
chromatograms, and evaluate the efficacy of each procedure. The results and
analysis are meant to determine the suitability of each technique and suggest
improvements where necessary.
Equipment:
- Chromatography paper
- TLC plates
- Solvent mixtures
- Mortar and pestle.
- Fresh spinach
- Capillary tubes
- Pencil and ruler
- Beakers (100-250mL)
- Lid or cling film.
, Unit 2 Assignment C Anonymous
- Micropipettes
- Amino acid solutions
- Ninhydrin spray
- UV lamp
- Protective gloves and safety goggles
- Lab coat
- Distilled water
- Scissors
- Hairdryer
Thin Layer Chromatography (TLC) of Plant Pigments
Purpose: To separate and identify pigments extracted from plant material
using TLC.
Procedure:
1. Grind fresh plant leaves in a mortar with a small amount of propanone to
extract pigments.
2. Draw a pencil line about 1–2 cm from the bottom of the TLC plate (this is
the baseline).
3. Using a capillary tube, apply a small drop of pigment extract onto the line.
Let it dry and repeat to concentrate the spot.
4. Prepare a solvent mixture in a beaker to a depth of 0.5–1 cm.
5. Place the TLC plate in the beaker, ensuring the spot is above the solvent
level.
6. Cover the beaker with a lid or cling film to reduce solvent evaporation.
7. Allow the solvent to rise up the plate until it is close to the top.
8. Remove the plate and immediately mark the solvent front with a pencil.
9. Allow the plate to dry.
10.Observe and record the separated pigment bands using visible or UV light.
11.Measure distances and calculate Rf values for each pigment.
Paper Chromatography of Amino Acids
Purpose: To separate and identify amino acids using paper chromatography.
Procedure:
1. Cut a strip of chromatography paper and draw a pencil line 2 cm from the
bottom.
2. Use a capillary tube to apply small spots of known amino acids and an
unknown sample along the line.
3. Let the spots dry and reapply if needed to concentrate them.
Chromatography is a valuable analytical technique utilized in scientific
laboratories to identify and determine the composition of mixtures. It is based on
the principle that materials have varying affinities for the stationary and mobile
phases, therefore moving at varying rates and thus separating over time.
Chromatography finds extensive application in chemical, biological, and
environmental science for qualitative and quantitative analyses.
In addition to chromatography, several other techniques are commonly used in
laboratories to separate and identify components in mixtures:
Filtration:
Filtration is used to separate an insoluble solid from a liquid. A common example
is separating sand from water. The mixture is passed through filter paper, which
traps the solid particles while allowing the liquid to pass through. This method is
based on particle size.
Distillation:
Distillation separates mixtures based on differences in boiling points. For
example, ethanol can be separated from water by heating the mixture until
ethanol evaporates, then condensing it back into a liquid. This method is
especially useful for purifying liquids or separating liquid-liquid mixtures.
Centrifugation:
Centrifugation separates components of a mixture based on density. The sample
is spun at high speed in a centrifuge, causing denser substances to settle at the
bottom. It is widely used in biology for separating blood components or isolating
cell parts.
In the present lab report, various chromatographic techniques were performed to
separate and identify the components of plant pigments and amino acids.
Techniques employed included paper chromatography and thin layer
chromatography (TLC). Both the techniques included appropriate observation of
separation patterns and determination of retention factor (Rf) values, which are
used to identify unknown compounds using known standards for comparison.
The objective of this experiment was to demonstrate proficient practical skills in
chromatography, illustrate safe working practice, interpret the resulting
chromatograms, and evaluate the efficacy of each procedure. The results and
analysis are meant to determine the suitability of each technique and suggest
improvements where necessary.
Equipment:
- Chromatography paper
- TLC plates
- Solvent mixtures
- Mortar and pestle.
- Fresh spinach
- Capillary tubes
- Pencil and ruler
- Beakers (100-250mL)
- Lid or cling film.
, Unit 2 Assignment C Anonymous
- Micropipettes
- Amino acid solutions
- Ninhydrin spray
- UV lamp
- Protective gloves and safety goggles
- Lab coat
- Distilled water
- Scissors
- Hairdryer
Thin Layer Chromatography (TLC) of Plant Pigments
Purpose: To separate and identify pigments extracted from plant material
using TLC.
Procedure:
1. Grind fresh plant leaves in a mortar with a small amount of propanone to
extract pigments.
2. Draw a pencil line about 1–2 cm from the bottom of the TLC plate (this is
the baseline).
3. Using a capillary tube, apply a small drop of pigment extract onto the line.
Let it dry and repeat to concentrate the spot.
4. Prepare a solvent mixture in a beaker to a depth of 0.5–1 cm.
5. Place the TLC plate in the beaker, ensuring the spot is above the solvent
level.
6. Cover the beaker with a lid or cling film to reduce solvent evaporation.
7. Allow the solvent to rise up the plate until it is close to the top.
8. Remove the plate and immediately mark the solvent front with a pencil.
9. Allow the plate to dry.
10.Observe and record the separated pigment bands using visible or UV light.
11.Measure distances and calculate Rf values for each pigment.
Paper Chromatography of Amino Acids
Purpose: To separate and identify amino acids using paper chromatography.
Procedure:
1. Cut a strip of chromatography paper and draw a pencil line 2 cm from the
bottom.
2. Use a capillary tube to apply small spots of known amino acids and an
unknown sample along the line.
3. Let the spots dry and reapply if needed to concentrate them.
