CHEM 253  Organic Chemistry Lab

 

Experiment #1  Isolation of the Active Ingredient in an Analgesic Drug

(Adapted from Introduction to Organic Laboratory Techniques  by Pavia, et.al.)

 

Techniques to be learned: Weighing and Making Transfers, Extraction, Evaporation, Vacuum Filtration, Column Chromatography, Drying Solids, Melting Point

 

Background:  Analgesic drugs reduce pain; antipyretic drugs reduce fever.  Some drugs, including aspirin and acetaminophen, do both. Many of the common over-the-counter analgesics-antipyretic drug preparations contain aspirin, acetaminophen, or combinations of these substances with other ingredients.  For example, Extra Strength Excedrin contains aspirin, acetaminophen, and caffeine. 

 

Understanding the Experiment:  Most natural products and many commercial preparations are mixtures containing a number of different substances.  To obtain a pure compound from such a mixture, the desired compounds are separated from the other components of the mixture by taking advantage of differences in their physical and chemical properties.  For example, substances having very different solubilities in a given solvent may be separated by extraction or filtration, and liquids with different boiling points can be separated by distillation.  Acidic or basic substances are often converted to water-soluble salts, which can then be separated from the water-insoluble components of a mixture.

In this experiment, you will extract (dissolve) the active ingredient of an analgesic drug by mixing the powered tablet with a solvent, methanol.  Two steps are required to remove the fine particles of binder, which remains suspended in the solvent.  First, you will use centrifugation to remove most of the binder.  The second step will be a filtration technique using a Pasteur pipet packed with alumina (finely ground Al2O3).  The solvent will then be evaporated to yield the solid analgesic, which will be collected by filtration on a Hirsch funnel.  Finally, you will test the purity of the drug by doing a melting point determination.

 

 

 

 

Experimental Procedure:

 

A. Extraction of Active Ingredient

Use 1 tablet containing the aspirin or acetaminophen.  Crush the tablet between two pieces of weighing paper using a pestle. Add all of the powered material to a 3-mL conical vial.  Using a calibrated Pasteur pipet, add about 2 mL of methanol to the vial.  Cap the vial and mix thoroughly by shaking.  Loosen the cap at least once during the mixing process in order to release any pressure that may build up in the vial.

Allow the undissolved portion of the powder to settle in the vial.  A cloudy suspension may remain even after 5 minutes of more.  Using a filter tip pipet, transfer the liquid phase to a centrifuge tube.  Add a second 2-mL portion of methanol to the conical vial and repeat the shaking process described previously.  After the solid has settled, transfer the liquid phase to the centrifuge tube containing the first extract.

Place the tube in a centrifuge along with another centrifuge tube of equal weight on the opposite side (blank or another lab group’s).  Centrifuge for 2-3 minutes.  The suspended solids should collect on the bottom of the tube leaving a clear or nearly clear supernatant liquid, the liquid above the solid. If the liquid is still cloudy, repeat the centrifugation for a longer period of time.  Being careful not to disturb the solid at the bottom of the tube, transfer the supernatant liquid with a Pasteur pipet to a clean test tube.

 

Column Chromatography

Prepare an alumina column using a Pasteur pipet, as demonstrated by lab instructor.

Insert a small (!!) ball of cotton into the top of the column.  Using a long thin object, push the cotton down so that it fits into the Pasteur pipet where the constriction begins.  Add about 0.5 g of alumina to the pipet and tap the column with your finger to pack the alumina.  Clamp the pipet in a vertical position so that the liquid can drain from the column into a small beaker or a 5-mL conical vial.  Place a small beaker under the column.  With a calibrated Pasteur pipet, add about 2.0 mL of methanol to the column and allow the liquid to drain until the level of the methanol just reaches the top of the alumina. Once the methanol has been added to the alumina, the top of the alumina in the column should not be allowed to run dry.  If necessary, add more methanol.

When the level of the methanol reaches the surface of the alumina, transfer the solution containing the drug from the test tube to the column using a Pasteur pipet.  Collect the liquid that passes through the column into a 5-mL conical vial.  When all the liquid from the test tube has been added to the column and has penetrated the alumina, add an additional 1.0 mL of methanol to the column and allow it to drain.  This ensures that all of the analgesic drug has been eluted from the column. 

 

Evaporation of Solvent

If you are isolating aspirin, it is essential that the following procedure be completed in 10-15 minutes; otherwise, the aspirin may partially decompose.

Using a Pasteur pipet, transfer about half the liquid in the 5-ml conical vial to a small test tube.  Evaporate the methanol in the 5-mL conical vial using a water bath at about 50oC.  (Will be set up for you)  Evaporate the solvent until the volume is less than 1 mL.  Then add the remainder of the liquid and continue evaporation.

When the solvent has completely evaporated or it is apparent that the remaining liquid is no longer evaporating, remove the vial from the water bath and allow it to cool to room temperature.  (The volume should be less than 0.5 mL when you discontinue evaporation.)   If liquid remains, place the cool vial in an ice-water bath for 10-15 minutes.  If the solid is very hard and clumped together, you should use a microspatula to break up the solid as much as possible before going on to the next step.

 

Vacuum Filtration

Set up a Hirsch funnel for vacuum filtration.  {Hirsch funnel, filter flask, rubber adapter} [Remember that you must use the heavy-walled rubber tubing and an aspirator that doesn’t spit water back at you!  ALWAYS BREAK THE VACUUM SEAL BEFORE TURNING OFF THE WATER.  I get really ugly when students don’t do this correctly!!!!]  Turn on the aspirator to the fullest extent.  Use a microspatula to transfer the material in the conical vial to the Hirsch funnel.  The vacuum will draw any remaining solvent from the crystals.  Allow the crystals to dry for 5-10 minutes while air is drawn through the crystals in the Hirsch funnel.  PULL THE HOSE OFF THE ASPIRATOR/FILTER FLASK.  TURN OFF THE WATER. 

Carefully remove the dried crystals from the Hirsch funnel onto a previously weighed watch glass or weighing boat.  If necessary, use a spatula to break up any remaining large pieces of solid.  Allow the crystals to air dry.  To determine when the crystals are dry, move them around with a dry spatula.  When the crystals no longer clump together or cling to the spatula, they should be dry.  Weigh the watch glass or weighing boat with the crystals to determine the weight of analgesic drug that you have isolated. 

 

Wrap-up

 

Compound

Molecular Weight

Your M.P. range

M.P. Range

(handbook)

Amount Recovered

Label Amount (theoretical)

Aspirin

180.16 mg/mmol

 

138-140oC

 

 

Acetaminophen

151.17 mg/mmol

 

169-172oC

 

 

 

Use the weight of the active ingredient specified on the label of the container as a basis for calculating the weight recovery.

Use a small sample of the crystals to determine the melting point of your crystals. 

(A handout of instructions will be given in the lab)

 

Questions:

1.  Why was the percentage recovery less than 100%?  Give 3 reasons.

2.  Why was the tablet crushed?

3.  What was the purpose of the centrifugation step?

4.  What was the purpose of the alumina column?

5.  Why was it important for the crystals to be dry before weighing and determining the melting point?