Chromatography

Learning objectives: To describe and explain the

method and use of Chromatography.

KW: chromatography, retention values, chromatogram.

Success Criteria:
By the end of the lesson you will be able to:
•State that chromatography is a method for separating a mixture of soluble
substances
•Describe some of the uses of chromatography.
•Explain separation by chromatography in terms of the solutes being more or
less soluble in the solvent.
Chromatography - Definition
A method of separation and analysis of a
mixture of soluble chemical substances.
 How does chromatography work?
• Chromatography separates components of a
mixture so that we can identify them.

• You get a series of spots on the paper

• The position of the spot helps you to

determine what the substance is.
 What can chromatography be
used for?
- identify additives in foods
- compare fibres found at a crime scene
- identify drugs and alcohol
- test water samples for pollution
- detect bombs in airports
- detecting pesticides or insecticides in food
- fingerprinting
 Paper Chromatography – how does
it work?
Chromatography lets us separate inks
and dyes according to the size of
their particles.

As the solvent (water) rises through

the paper it dissolves the sample
mixture, which will then travel up
the paper.

Small particles travel further than

large particles, due to the
differences in solubility and their
attractions with the paper.
Molecules in dyes and inks
Many common dyes and inks are water
soluble and spread apart into the component
dyes using water as a solvent. If the dye or ink
you are testing does not spread out using
water, it may be “permanent” ink. In this case,
you will have to use a different solvent such as
alcohol.
 What are food additives and why are
they put in our food?
• We need to make food last longer – so we find
ways to preserve it.

• A substance used to preserve food is known as a

food additive. Food additives can also be used to
improve taste or appearance of food.

• Additives that have been approved for use in

Europe have been given E numbers to identify
them.
 Detecting Additives
• Food additives can be detecting by a technique known
as chromatography.

• This technique separates out the different components

within the food additives based on how well they
dissolve in a particular solvent.

• Their solubility determines how far they travel across a

surface (chromatography paper).

• Once the compounds have been separated out using

chromatography, they can be identified by comparing
with known substances. Alternatively a mass
spectrometer can be used.
 Chromatography
TASK
Write step-by-step instructions on how to carry out the separation of
food dyes using this technique.

Copy this picture into your books to help you explain the steps.

Beaker

Separated
ink
particles
Spot of dye to
be separated

Solvent
 Retention factor (Rf) values
The Rf factor is used to compare the
components of various samples. The
Rf values of suspect samples can be
compared with known samples.
Rf = distance from the base line to the spot
distance from the base line to the solvent front

If two substances Solvent front

have the same Rf the point at which the water
value, they are likely stopped moving up the paper
(but not necessarily)
the same compound.
Spot
If they have different
the point at which a band
Rf values, they are
or spot of colour is Base line
definitely different
compounds. the line where the original
sample was placed
 Chromatography - Improving results
SINGLE DEVELOPMENT
Usual method but not ideal for the
separation of components in
very small samples
x x

DOUBLE DEVELOPMENT
Run solvent up to 5mm.
Dry plate and re-run up to
10mm.
•Improves sensitivity 10mm
5mm
•Improves resolution x x x x
 ANALYSIS OF FIBRE DYES
• Soham murders – a high profile case in August 2002
• Two 10 yr old girls, Holly Wells and Jessica Chapman.
• Ian Huntley – charged with 2 counts of murder and sentenced to life
imprisonment.
Forensic Scientists found fibres from the
girls’ tops on clothing found at Mr Huntley's
home, as well as on a bathmat, carpet, and
in the boot of Mr Huntley's Ford Fiesta.

Similarly, the remains of Holly and Jessica's

clothes bore fibres from clothing and
carpets from Mr Huntley's house and car.

The components of the red fibre dye from

The burnt and cut shirts the football shirts matched with the
separated component dyes from the fibres
were found in a bin at
obtained as evidence – what steps do you
Soham Village College think they took to obtain a match?
 Paper Chromatography

Start Finish

How many colours in a) red dye b) green dye c) purple dye d) overall
How many dyes were used in a) Black, b) Green?
How many dyes were used overall?
How many blues were used?
Are the yellows in brown and orange the same? How do you know?
Inks i, ii, iii and iv were made from colours A, B, C and D
Try to work out which colours were mixed for each of the inks (i-iv)
 Learning intentions
To copy a diagram showing a gas liquid
chromatograph with mass spectroscopy (GC-MS)

To investigate how a GC-MS machine works

Success criteria:
Good notes on GC including a diagram
To be able to answer an exam question about GC
GC-MS
 Gas chromatography
• The mobile phase is an unreactive gas known
as the carrier gas
• The stationary phase is held inside a long
column and is lots of pieces of inert solid
coated in high bp liquid.
• The column is coiled in an oven
• The sample to be analysed is injected into the
carrier gas stream at the start of the column.
 GC-MS analysis
• GC machine separates the components in the
column because of the time they take to come
through
• A detector monitors the compounds coming out
of the column
• The mass spectrometer tells us the relative
molecular mass of the components.
• A database can tell us the likely identity of the
compound
GLC Chromatograph
 Interpretation
• The number of peaks shows the number of
compounds present in the sample
• The position of each peak shows the retention
time for each compound
• Area under peak or relative heights can be
used to work out relative amounts of
substances
Exam Question
 Exam Question

any two from:

• faster

• more accurate

• detects smaller amounts

 Exam question

gas chromatography / GC-MS

any one from:

• fast / quick or comment about speed
• small amount
• sensitive / accurate / precise
• ease of automation
• reliable / efficient
• can be left to run / continuous
analysis