Key Terminology
Term Definition
Chromatography Family of separation techniques which separates a mixture dissolved in a solvent
which moves over a stationary phase
Mobile Phase The solvent which moves the soluble components of the mixture
Stationary Phase Solid which holds back components in the mixture it is attracted to
Thin Layer
A development of paper chromatography which uses a metal or glass sheet
coated in silica gel or alumina
Use of a powder or resin as a stationary phase in a column with an eluent added
at the top.
A modern technique used to identify gas components based on their retention
times in a spiral tube containing the stationary phase
Rf Retention Factor – used to identify a component; calculated by distance moved
by spot ÷ distance moved by solvent
Chromatography uses the principle of a mobile phase, which carries a component further if it is more
soluble, and a stationary phase, which attracts certain components more strongly, to identify molecules
TLC uses a glass or metal sheet coated in a thin layer of silica gel (SiO2) or alumina (Al2O3) as a plate. It
is placed in a TLC tank and a solvent is added. Once the chromatogram has run, the position of spots
may need to be identified with a locating agent and UV light. Rf is then calculated and compared to a
database to identify the compound
Column Chromatography
A powder, such as silica, or resin, is used as the
stationary phase and is packed into a narrow
column. A solvent (eluent) is added at the top and
is allowed to run down the column. The
components of the mixture move through at
different rates and can be collected separately in
flasks at the bottom. Multiple eluents ensure better
separation. This method allows for large amounts
of mixture to be separated and collected.
Gas-Liquid Chromatography
The stationary phase is a powder coated in oil
which is packed into a long capillary tube, then
coiled up and placed in an oven. The mobile
phase is an unreactive gas, such as nitrogen.
After injection, the sample is carried by the gas
and the mixture separated as some of the
components are moved by the gas and some are
retained by the oil. Components leave the
column at different times, and have different
retention times. Detectors measure the
emerging gases.
Some GC systems have a mass spectrometer as a detector through which components flow. Abundance
can be calculated then the component can then be identified by fragmentation pattern or by
measurement of mass.
Core Practical 12: Separation of Species by Thin-Layer Chromatography
An investigation into the types of analgesic (pain killer) found in an
Anacin tablet.
1. Crush the Anacin tablet sample with a mortar and pestle. Produce
a dissolved sample. Repeat for each of the pure analgesics e.g.
aspirin, ibuprofen, caffeine
2. Draw a baseline on an aluminium TLC plate coated in silica gel
3. Use microcapillary tubes to spot each of the analgesics and the
tablet sample onto the baseline. Label each sample
4. Place into a beaker. Add a suitable solvent (e.g. ethanoic acid in ethyl acetate) and allow to run up.
Close the tank with a watch glass to ensure that the tank is saturated with solvent vapour
5. Once the solvent almost reaches the top of the plate, remove the plate and mark the positions of
all spots
6. Calculate Rf values and match the spots to identify the pain killers found in the tablet e.g. Anacin
contains aspirin and caffeine