• Introduction to chromatography and spectroscopy

• Using chromatography and spectroscopy to solve a mystery

• Seeing in 3D

Used to investigate the purity and identity of samples– Stationary phase: silica plate– Mobile phase: solvent

As the solvent rises up the plate, different compounds move up the plate at different speeds– Polar compounds move slowly– Lipophilic compounds move quickly

UV lamp often used to visualise the compounds on the plate

Thin-layer chromatography (TLC)

Assessing purity

Pure compound 1Impure compound 1

impurity

Identifying unknown samples

Compound A Compound B Unknown

How would you confirm that the unknown was compound B?

Identifying unknown samples

Compound AMixed

UnknownCompound B

MixedUnknown

Consistent with unknown being compound A

How does spectroscopy work?

• Light is a mixture of different colours (wavelengths/frequencies)

Ultra-violet

Infra-red

How does spectroscopy work?• Different materials absorb different

parts of the spectrum

Ultra-violet

Infra-red

Ultra-violet

Infra-red

Absorbs everything but blue and red

This reflected light tells us something about the material

Infra-Red (IR) spectroscopy

• In IR spectra, regions of light are absorbed by specific parts of the molecule.

1715cm-1

The C=O bond of a ketone absorbs light in the region of 1715cm-1 so that the emerging IR light has this

piece of its spectrum missing

IR light C

O

For example

Infra-Red (IR) spectroscopy

• Every compound has a unique IR spectrum that tells us something about the structure

IR light

Unknown compound

http://www.le.ac.uk/spectraschool/sias/UV%20poster.pdf

Solving a mystery – Part 1

• A dead body has been found• 2 empty bottles of 3 painkillers found at the

scene– Aspirin – Paracetamol

• It is suspected that the victim took an overdose of one of these

• Using the Thin-Layer Chromatography (TLC) and Infra-Red (IR) spectroscopy, can you work-out which painkiller is most likely to have led to the death of the victim?

SafetySafety glasses and gloves to be worn throughout

PainkillersAspirin and paracetamol are active medicines. Do not ingest.

TLC tank solvent: diethyl etherdichloromethanetetrahydrofuran

Solvent for dissolving the painkillers:ethanol

Solvent for washing the IR sodium chloride platesAcetone

Use of UV lamp to observe TLC plates

} Do not inhale the solvent or allow it to contact your skinDiethyl ether and tetrahydrofuran are highly flammable

} Ethanol is highly flammable

Acetone is highly flammable

UV light is harmful to eyes and skin. Do not look directly at the light or place your hands under the light

Which one could have killed the victim?

Aspirin Paracetamol

Solving a mystery – Part 2

• We now know that aspirin was present in the victim’s blood.

• What we don’t know is whether there was enough in the blood to kill the victim.

• Therapeutic concentrations: <300mg/L• Severe overdose: >750mg/L

• We are going to use visible spectroscopy to reach a conclusion

http://www.le.ac.uk/spectraschool/sias/UV%20poster.pdf

Solving a mystery – Part 2

• How much aspirin was present in the victim’s blood?

• What was the likely cause of death?

Colour filters and 3D visualisation

• In IR spectra, regions of light are absorbed by specific parts of the molecule.

1715cm-1

The C=O bond of a ketone absorbs light in the region of 1715cm-1 so that the emerging IR light has this

piece of its spectrum missing

IR light C

O

• A colour filter on a camera works in a similar way

white light

It absorbs all the colours in white light apart from red, making everything look red

As a result, red/orange objects are difficult to see with a red filter

Colour filters and 3D visualisation

Here is a light orange circle and a light blue square

What do you expect to see when you look at this through a red filter?What do you expect to see when you look at this through a blue filter?

(e.g. 3D specs, Quality Street wrappers)

What about a red filter for one eye and a blue filter for the other?

Colour filters and 3D visualisation

When you do the same thing with shapes that are similar and are almost on top of one another,

something strange happens

What do you think is happening?

How can this help us think about the shape of molecules?

Colour filters and 3D visualisation

http://www.dddesign.com

We draw them flat but they do have 3D shape

Aspirin Paracetamol

http://www.3dchem.com/molecules.asp?ID=157 http://www.3dchem.com/moremolecules.asp?ID=9&othername=Paracetamol

Aspirin

Paracetamol