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label (ingredients) the spots in the chromatogram, including as many of the spots in the unknown as possible. Also label the solvent front! (c) From the number, position and appearance of the spots in the unknown, and the composition of the possible unknowns, identify your unknown analgesic. You should use the table below to aid the identification of your unknown. ie: If Midol is your unknown, you will see 2 spots, that match up with the reference compounds Acetaminophen and Caffeine Drug (Brand Name) Possible Unknowns Ingredients Acetaminophen Aspirin Caffeine Ibuprofen Advil 200 mg Anacin 400 mg 32 mg Excedrin 250 mg 250 mg 65 mg Midol 500 mg 60 mg Tylenol 325 mg Bayer 325 mg The reference compounds are: Acetaminophen [4-acetamidophenol], Aspirin [acetylsalicylic acid], Caffeine, Ibuprofen [2-(4-Isobutylphenyl)-propionic Acid] Part B: The Separation of Fluorene and Fluorenone by Column Chromatography Begin Part B after you have begun Part A. Work in PAIRS for Part B only. Safety and Disposal Data for Compounds used in Part B. Compound Mol. Wt. (g/mol) Safety and Disposal Data Alumina Diethyl Ether Fluorene Fluorenone Ligroin (Petroleum Ether) 101.96 74.12 166.22 180.20 --- May cause irritation. May be harmful if inhaled. Avoid all contact. Wash thoroughly after use. Dispose in empty pail in Waste Fumehood. Irritant. Extremely flammable. May form explosive peroxides. Dispose in Organic Waste. Irritant. Dispose in Organic Waste. Irritant. Dispose in Organic Waste. Extremely flammable. Dispose in Organic Waste.
1. Preparing the Column a) You will be provided with a `long' Pasteur pipette that has a small bend in the closed tip. b) Fill the pipette at lea least half-full with pure Ligroin (petroleum ether) NOT the 4:1 Ligroin:ether solution. Place a small wad of glass wool into the bottom of the pipette (3mm glass rod is ideal to push the glass wool down to the bottom or another long pipette), ), pour in some san sand d (0.3 cm). Tap the column to try and remove all air bubbles. c) Pack the `column' with alumina (5 cm), using a micro spatula, by slowly adding the alumina to the solvent in the column and tapping the column gently. Be sure to keep the solvent level above the level of the alumina. Note: The tapping promotes even settling and mixing in the column, and frees air bubbles. Try and keep the pipette vertical while doing this so the alumina settles evenly. d) Then pour sand (0.3 cm) on top of the alumina and clam clamp p the pipette carefully with a microclamp to the scaffolding at the back of your fumehood. It is important that the column be mounted as close to vertical as possible your column is ready for use. Microclamp 2. Separation and Collection of Fluorene and Fluorenone NOTE: Do not break the pipette tip until you have everything ready! DO NOT let the column run dry! a) Take 1 mL of the fluorene/fluorenone solution provided, which has been dissolved in pet-ether. ether. Break off the tip of the `colum `column' n' (hold the pipette near the bend when breaking) with a piece of paper towel and when the Ligroin just about disappears through the sand add the solution of fluorene/fluorenone to the top of the column drop wise until the sample has been loaded on the column. b) When the final volume of fluorene/fluorenone solution disappears through the sand at the top, add elution solvent (4:1 Ligroin/ether) to the top of the column drop wise (about 10 drops), then fill the column to the top. (Do not let the solvent lev level el drop below the sand at the top of the column. Channels will develop in the alumina resulting in a poor separation of the compounds.) You will likely need to add more elution solvent while you are collecting, have a small amount ready BEFORE you break the he column tip! c) Collect at least five (5) one mL fractions in test tubes. If the second compound has not appeared by fraction 5 (light yellow colour) collect 2 - 3 more 1 mL fractions. 3. Analysis of the Fractions a) Spot the fractions on Silica Gel TTLC LC strips along with the reference fluorene and fluorenone. b) Develop the strips in a chromatography jar containing a 4:1 mixture of pet-ether/ether ether/ether. c) Visualize the chromatograms with the ultraviolet lamp to determine which fractions are pure fluorene and which are pure fluorenone. Draw the chromatogram in your lab notebook (identifying and labeling the spots in the chromatogram). Calculation of RRf values is not needed, but may aid you in identifying the correct spots.
Page 1 and 2: Experiment 1. Thin in Layer Chromat
Page 3 and 4: power Diethyl Ether Ethyl Acetate A
Page 5 and 6: Part B: The Separation of Fluorene
Page 7: other end of the capillary tube or

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