The first step is to run a series of TLCs of the mixture, each in a different-polarity solvent. The goal is to find a solvent system that gives an Rf of 0.25 - 0.35 for the faster-moving compound (the ketone). The rationale behind this is that an Rf in the above range indicates that the solvent is "ideal" for eluting that particular compound from a flash chromatography column (Still, W. C.; Kahn, M.; Mitra, A. J. Org. Chem. 1978, 43, 2923-2925).
Seven different solvent systems of varying polarity will be available. Each is a mix of hexanes and ethyl acetate; #7 is the least polar at 95:5 hexanes/ethyl acetate, #1 is the most polar at 80:20 hexanes/ethyl acetate. The rule is:
if you want to have a compound run further up a plate (higher Rf), go to a more polar solvent
TLC Procedure:
Note: To see a larger view of each image, click on it and it will open in a new window.
Weigh out about 0.02 g of your unknown mixture and place it in a small vial. Label the vial "standard-unknown". Add about 1 mL of acetone (estimate the volume knowing that a Pasteur pipet holds about 1.5 mL). Stopper the vial. Mark the solvent level (in case the solvent evaporates). Use this solution for the TLC part of the experiment and save it (capped and labeled) for the next lab period. (You will use it as a standard when you run TLC's of the column fractions in the second part of this laboratory experiment.) Prepare for TLC: Set up a developing chamber (a beaker, lined with filter paper , filled with eluting solvent to a depth of less than 0.5 cm, covered with a beaker) for each solvent that you plan to test (the photo shows solvents 1, 2, and 3). These chambers need to sit for awhile for the solvent vapors to saturate the atmosphere. In the meantime, cut several TLC plates, mark the origin, and mark each with the number of a TLC solvent that you plan to try. Use the standard-unknown solution to spot the prepared TLC plates. (See the TLC page if you have forgotten how to spot/run TLC plates.) Run (develop) each plate in a different solvent system. The example at the right shows 3 plates being developed in solvents 1, 2, and 3. As soon as you take a plate out of the beaker, mark the solvent front - before the solvent dries. Visualize the developed plates under a UV lamp. Safety note: Wear your goggles whenever you use a UV lamp to protect your eyes from UV light.
For the sake of demonstration, we ran the standard unknown in the other 4 solvents too. The picture to the right shows the unknown run in all 7 solvents. While viewing under the UV lamp, circle all of the spots on the TLC plates. Note: If you see more than two spots on your unknown-mixture TLCs, the darkest spots are always the ones that you want. Measure the Rf values. You are looking for the system that gives an Rf of 0.25-0.35 for the faster-moving spot (this is the ideal system). The table linked at the right shows our values for distances traveled and Rf values for the standard unknown in all 7 solvent systems. When you find the "ideal" system, make sure that the slower-moving spot moves significantly slower in the same system. (Subtract the Rf of the slower-moving spot from that of the faster-moving spot and make use that the resulting number is at least 0.1.)
Table of distances and Rf values. Our choice of solvent system is solvent #3. This solvent will be used to elute the faster-moving spot from the column in the microscale flash column chromatography step.
