Hands-on Chromatography
Presented by:
Sheryl K. Hoffmann, B Sc., Grad Dip. O. H., MSIA
Concordia College
shoffmann@concordia.sa.edu.au
CONASTA 60 – Darwin High School, 2011
Definition 1:
Chromatography is a technique for separating similar compounds from one another
by using differences in the strength of their adsorption on an inert material.
Adsorb: hold (molecules of a gas or liquid or solute) to its surface
Absorb: swallow up, incorporate, take in
Definition 2:
Chromatography is a technique which can be used to separate the components of a
mixture. The method depends upon
 differences in the abilities of the components to adhere to a porus solid such
as silica gel, called the stationary phase
 differences in the abilities of the components to be dissolved in a stream of
liquid or gas, called the mobile phase, which is passed over the solid.
Main Types
1. Paper Chromatography
2. Thin Layer Chromatography (TLC)
3. Column Chromatography
4. High Performance Liquid Chromatography (HPLC)
5. Gas-Liquid Chromatography
Basic Method for Paper Chromatography
1. Take a piece of chromatography paper and mark a pencil line 2 cm from the
bottom.
2. Mark the positions of the samples evenly across the paper with a cross,
leaving at least 2cm at each side.
3. Spot the samples on the positions marked, and allow to dry.
4. Pour < 1cm of solvent into a beaker, test tube or chromatography tank.
5. Lower the chromatography paper into the solvent in a beaker & support.
6. Allow the run until the solvent nearly reaches the top of the paper.
7. Mark final solvent height if Rf values are to be calculated.
Spotting Methods
To get good separation of bands, it is important to keep the spot as small as possible
& as concentrated as possible. Capillary Tubes, Toothpick & Pasteur Pipettes can all
be used to spot chromatography paper.
Tips
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If the spot is large, components start from different origins and will thus blur
any separation that would normally occur.
Try a quick dabbing motion, the longer the liquid is in contact with the paper
the larger the spot
If the spot is not very concentrated the resulting bands, once spread over a
wider area, will be even fainter.
If the solution to be spotted is not very strong, spot several times, being sure
to allow the spot to dry in-between, a hair drier will speed up the drying
process.
Suppliers
SCIENCE SUPPLY
AUSTRALIA
PO Box 465, Glen
Waverley, VIC 3150,
Phone : 1300 857 544
Fax : 1300 857 533
Email :
sales@ssapl.com.au
Web: www.ssapl.com.au
WESTLAB
PO Box 1680,
Ballarat, Vic 3354
Phone: 1800 358 101
Fax: (03) 5333 4144
Chromatography Paper
20 mm X 100m $44.15
30mm X 100m $60.30
50mm X 100m $87.15
TLC Sheets
200 X 200 mm box/25
200mm X 200mm
box/100 sheets $80.40
Chromatography
Column
355 X 14mm $35
Chromatography Paper
250mm X 250mm
box/100 sheets
LIVINGSTONE
106-116 Epsom Rd,
Rosebery, NSW 2018
Phone:1300 780 078
Fax: (02) 9313 6444
Email:
laboratory@livingstone.c
om.au
Web:
www.livingstone.com.au
Chromatography Paper
20 mm X 20mm $140.95
Capillary tubes
Plain 75mm 100/vial
$2.30
Different Styles & Support Methods for Paper Chromatography
There are numerous examples of paper chromatography. Below are some pictures
of various styles & different methods of supporting the paper.
NB: A lid is only needed if the solvent is not water
My Preferred Methods
Paper
Large battery tank, paperclips & skewer
TLC
25ml McCartney bottle