Thin Layer and Column Chromatography
Experimental Procedure for Part 2: Column Chromatography of a Simple
Mixture
Preparation:
1.
Weigh a 25-mL side-arm filtering flask and a 10-mL Erlenmeyer
flask. Record both weights in your notebook.
2.
Obtain a glass column, a plastic funnel, cotton (or glass wool),
sand, alumina, and stopcock. Check to make sure the stopcock is
not lose.
3.
Fill the column about 2/3’s full with alumina. Then pour the alumina
into a 10-mL Erlenmeyer flask.
4.
Pack the bottom of the column with cotton (or glass wool), followed
by a small amount of sand. Make sure the sand level creates an
even surface. Place the column in a clamp in the fume hood and
close the valve.
5.
Gently pour hexane (Caution! Flammable) into the column until
the liquid level reaches the top of the glass.
6.
Add about 8-mL of hexane to the alumina in the flask and stir the
mixture with a thin glass strirring rob to eliminate any bubbles.
7.
Swirl the flask to suspend the alumina in the hexane, and quickly
pour the entire slurry into the funnel at the top of the column. (This
is the tricky part!)
8.
Place the flask that contained the alumina under the column, open
the valve and drain some of the hexane out. Do not allow the
level of hexane to drop below the top of the alumina! The
alumina should settle within the column.
9.
If a significant amount of alumina was left behind in the flask, close
the valve, (add a little more hexane if necessary) and finish
transferring the alumina to the column. Do not worry if a small
amount of alumina was left behind in the flask.
10.
Once you have sufficient alumina in the column and it is packed
down, add a small amount of sand. Again, you want to create an
even surface. Then open the valve and drain enough hexane into an
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empty flask until the level of liquid in the column is just at the sand.
It is very important when performing column
chromatography that the column never be allowed to run
dry! Never let the level of liquid fall below the top of the
alumina!
Running the Column:
11. A 1:1 ferrocene:acetylferrocene mixture is provided for you. In a
10-mL Erlenmeyer flask, dissolve 0.090-g of this mixture in 1-mL of
dichloromethane.
12.
Obtain a silica gel TLC plate, and spot this solution of the mixture
on the left-hand side of the plate, allowing enough room on the
right-hand side for two more spots. Set the plate aside in a place
where it will not be disturbed.
13.
Add 0.30 g of alumina to the ferrocene:acetylferrocene mixture and
swirl to mix.
14.
Stir the mixture over a steam bath and completely evaporate the
dichloromethane. You must stir the mixture continuously as
the solvent evaporates or the mixture will bump. Once the
alumina is completely dry (it will be a bright orange color at this
point), remove the flask from the steam bath.
15.
Pour the alumina which has the ferrocene:actylferrocene mixture
adsorbed onto it through the funnel into the column of alumina.
16.
Using a Pasteur pipet, wash down any alumina that adheres to the
funnel or was with a few drops of hexane. Tap the column to
remove air bubbles from this layer of adsorbent-solute mixture.
Then add a small amount of sand.
17.
Place the tared 25-mL side-arm flask under the valve of the
column.
18.
Fill a pipet with hexane. Open the valve on the column, then place
the tip of the pipet close to the surface of the sand in the column,
and carefully add the hexane in such a manner that the top of the
sand is not disturbed. When the hexane has been applied in a
narrow band at the top of the column, continue filling the column to
the top of the glass with hexane.
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19.
As the column is eluted with hexane, ferrocene will travel down as a
yellow band. Collect this yellow solution in the side-arm flask. Add
hexane to the column as necessary so that it does not run dry. If
you see crystalline material at the tip of the valve, wash it into the
flask with a drop or two of hexane.
20.
When the yellow band has been completely eluted, let the level of
hexane lower until it is about 1 cm above the top of the sand, then
fill the column to the top of the glass with a 50:50 mixture of
hexane and t-butyl methyl ether.
21.
Elute the acetylferrocene, which will be seen as an orange band
with the 50:50 hexane:t-butyl methyl ether mixture. Collect this
orange solution into the tared 10-mL flask. Add 50:50 hexane:tbutyl methyl ether solvent system to the column as necessary so
that it does not run dry.
22.
When the elution of both bands is complete, shake out the contents
of the column into a beaker. Rinse out the glass column with water,
adding the rinses to the beaker. The mixture of alumina, sand, and
liquid must be disposed of in the appropriate waste jar.
23.
Spot the right-hand side of the silica gel plate with the solutions
from the yellow and orange bands (the plate should have 3 spots in
all). Elute the plate with 30:1 toluene:absolute ethanol. Measure
the distances of the solvent front and all spots on the TLC plate.
Sketch the TLC plate in your notebook.
O
C
CH3
Fe
Ferrocene
Melting Point = 174-176 oC
Fe
Acetylferrocene
Melting Point = 81-83 oC
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