- Column Chromatography
Mohrig, Technique 17.1-17.6, pp. 206-219 or Technique 18.1-18.6 pp. 236-248 and Appendix.
the boiling points of the pure solvents used in this lab.
[Reference Mohrig, 2nd edition, Technique 4, pp. 34-38 or 3rd edition Technique 3.3 pp. 25-28 to find paper and online sources for obtaining physical property information (i.e.-boiling points) about organic compounds.]
(Please turn in the answers to these questions when you arrive to lab.)
1. Please describe the procedure you will use to create 50-mL of a 10% (by volume) methanol solution in ethyl acetate. (Note: since the solution concentration has only two sig. figs in it (10%), the precision with which you make this solution does not have to be very precise.)
2. What effect would a slight impurity (<10%) have on the melting point of a solid and why? (Note…check out the concept of “Freezing Point Depression” from intro. chem. stuff.)
3. What are the boiling points of all the individual solvents you will be using in this lab and list them in order of least polar to most polar? (Dielectric constants are a measure of a compound’s relative polarity…check Mohrig, Table 9.1.)
- To resolve a mixture
of unknown composition in reasonable quantity
- To identify the
components of the mixture by their melting points and comparison to
a list of possible unknowns.
You will be given
a mixture of two of the 4 compounds listed in Table 2. This mixture
be separated using Column
Chromatography. The separation will be monitored by Thin
Layer Chromatography. Once separated, the melting points of the unknowns
can be determined and the compounds can be identified using the information
in Table 2.
Table 2: Melting
Points for Possible Unknowns in Column Chromatography Lab
The unknowns listed in Table 2 are toxic and will stain skin and clothing; wear gloves when working with them. The eluting solvents are flammable and volatile. All work should be carried out in the hood.
and Identification of Binary Unknowns using Column Chromatography
Read this experiment all the way through and gather your material before
you begin to work.
The first eluting solvent you will need will be a 30% (by volume) solution
of Ethyl Acetate in Petroleum Ether. Create 100-mL of this eluting solvent.
Mix well. The second eluting solvent you will need will be a well-mixed
10% (by volume) solution of methanol in Ethyl Acetate. Make approximately
50 mL of this solvent. Mix well. Store in covered containers to prevent
absorption of moisture and/or evaporation.
chromatography (TLC) developing chamber and plate
Create a TLC developing chamber by placing a half-circle of filter paper
and about 30 mL eluting solvent (30% Ethyl
Acetate in Petroleum Ether)
in a 1500 mL beaker (your 'developing chamber.') Cover the 'chamber'
aluminum foil or plastic wrap. What's the purpose of the filter paper? Using a pencil, label your alumina-coated
TLC plate so that it has three lanes, one for your crude product, and
for each of the two elutions you will obtain from your column. Store
the plate in a clean place until needed.
of Binary Unknown for Column
Add approximately 0.5 mL of anhydrous ethyl acetate to your binary unknown
mixture. You may need to heat over a steam bath for complete dissolution
of the unknown mixture.
of the Column
Label three 50-mL beakers: Waste, Elution #1, and Elution #2.
Using a long stirring rod, place a small plug of glass wool in the bottom
of the column. What's the purpose of this plug of glass wool? Close the bottom of the column with a small piece of rubber
tubing and a screw clamp. Fill the column 3/4 full with your eluting
solvent. Slowly pour enough sand into the column to create an approximately
1-cm layer of sand on top of the glass wool. What's the purpose of this bottom layer of sand? Pour approximately 10 grams
of Alumina into the column on top of the sand. Lastly, create another
approximately 1-cm layer of sand by pouring the sand slowly on top of
the Alumina. What's the purpose of this top layer of sand? When you are ready to add your mixture to the column, place
your "Waste" beaker under the column and remove the tubing and
clamp from the tip of the column allowing the solvent to flow
freely for the rest of the experiment.
of Mixture to Column
Acetate in Petroleum Ether eluting
small amounts of eluting solvent until the solvent reservoir above
the top of the sand layer
is colorless. Why?
Let the eluting solvent drain
to approximately 1 cm above the top of the sand. Using a long pipet,
transfer the binary unknown/ethyl acetate mixture to the top of the
sand layer making sure not to squirt your mixture onto the inner sides
of the column. *Allow the mixture to adsorb into the top of the
Alumina before adding
Once you start adding solvent to the column, try to never let the
solvent go below the top of the sand layer. Why? Once the small solvent reservoir
is colorless, fill the column with solvent to help prevent
the column from going
for TLC analysis.
from the Column
Continue to add 30% Ethyl
Acetate in Petroleum Ether
to elute the first unknown into a labeled 50 mL beaker. After all of
the first unknown has been
from the column, switch to the second eluting solvent, 10% Methanol
in Ethyl Acetate,
and collect the second unknown into another labeled 50 mL beaker. Do
not collect more than 40 mL of the 2nd elution. Finally,
allow the column to drain into a waste beaker. Is it OK to mix the two eluting solvents when switching from one eluting solvent to the other eluting solvent? Why or why not?
Add 0.5 mL ethyl acetate to the residue in your vial. Spot your prepared
TLC plate with this solution of your unknown mixture, and with Elution
1 and Elution 2. If the spots are not visible, check that you have placed
enough of each compound on the plate by viewing the plate under a UV lamp.
Develop the plate in your developing chamber. Immediately mark the solvent
front when you remove your plate from the developing chamber. Examine
your plate and comment on the purity of each elution. Use the UV lamp,
if necessary. Calculate the Rf values for each compound.
and Identification of Solid Unknowns
Follow the instructions of your lab instructor to evaporate the solvent from your two elutions.
the solids by taking their melting points and comparing them
to the melting points in Table 2.
Waste Disposal and Clean-up
- Place used columns in the paper-lined tote in the hood.
- Rinse your glassware with acetone and dispose of the acetone rinse in the flammables waste bottle. Place developing beakers back in their original tote and place all other beakers in the dishwashing tote.
(Please turn in the answers to these questions when you arrive at your NEXT lab.)
- In a data table, please report your unknown number and what the two components of your binary unkown mixture were from the list of possible unknowns in Table 2. Please justify your answer. [Please report the voltage/power level that was used to obtain your melting points and any observations you took while recording the melting points.]
- In your TLC analysis, how many spots did you see under UV for your crude unknown mixture? How many spots did you see for each elution? What does this tell you about the purity of your eluted compounds?
- Knowing how the
two compounds in your binary unknown mixture separated on the column,
what do you know about how they differ chemically?
- Based on what you know about freezing point depression and wide mp ranges due to small amounts of impurities, do the melting points for each of your unknowns support or contradict your TLC results? Explain.
- How would you separate and isolate two colorless compounds using only column chromatography and TLC?
- Discuss any unexpected results
in your experiment. What you would do differently to improve your results if you were to
do this lab again.