In this lesson, we will learn about paper and thin layer chromatography, both of which
are used in forensic science applications.
Chromatography is a separation technique, and this will be demonstrated using food colouring.
Place this in a beaker with water, ensuring that the water level does not touch the spots.
As the water is absorbed, it separates the multiple components of the food colouring.
We have now created a chromatogram.
The chromatography paper is called the stationary phase as it “stays put”.
Water is called the mobile phase as it “moves” up the chromatography paper.
Here is the key idea – a component that is more soluble in water travels a greater
distance up
the chromatography paper, and a less soluble component travels a shorter distance.
The retention factor, or Rf value,value, is a measure of the distance that a separated
component has
travelled compared to the distance that the solvent, in our example, water, has travelled.
Rf = distance travelled by a component / distance travelled by the solvent
These distances can be measured using a ruler.
Rf = distance travelled by a component / distance travelled by the solvent
Distance travelled by solvent = 6cm
Distance travelled by component A = 4cm
Distance travelled by component B = 2cm
Rf for component A = 4cm / 6cm = 0.67
Rf for component A = 2cm / 6cm = 0.33
Challenge:
What does the Rf value tell you about the solubility of a separated component in the
mobile phase solvent?
Please pause the video to think about this.
A larger Rf value means that the component is more soluble in the solvent.
A smaller Rf value means that the component is less soluble in the solvent.
Thin layer chromatography, or TLC, works in exactly the same manner.
Instead of chromatography paper, a glass plate with a thin layer of silica gel or any other
adsorbent material is used.
This thin layer of adsorbent material is the stationary phase.
Any suitable solvent may be used as the mobile phase.
If the separated components are colourless, the dried chromatogram may be sprayed with
ninhydrin, which can turn some components purple, or the chromatogram may be viewed
under UV light.
Ninhydrin is be used to detect fingerprints from a crime scene.
Both chromatography methods can be used to identify and compare samples of drugs, inks,
lipsticks, saliva, and explosives, just to name a few.
In conclusion, paper and thin layer chromatography are used to separate components of a
mixture based on their solubility.
The relative solubility of separated components can be determined by calculating their Rf
values.
More soluble components have a larger Rf value and less soluble components have a smaller
Rf value.