Exercise 4: Thin layer chromatography of organic compounds

Transcription

1 Chemistry 162 Exercise 4: Thin layer chromatography of organic compounds Objective: Use thin layer chromatography to separate and characterize the polarity of a mixture of benzene derivatives. Introduction: The most widely used methods of separating components of organic chemical mixtures involve some form of chromatography. At its simplest, chromatography is the separation of two or more compounds by distributing them unequally between two phases, one of which is moving (called the mobile phase ) and the other of which is not ( the stationary phase ). The unequal distribution comes about due to the different solubilities and adsorptivities these compounds have for the two phases. In thin layer chromatography (TLC), the stationary phase is adsorbent silica bound to a thin flexible plastic sheet, called a TLC plate. Silica (silicon dioxide) is considered to be a polar substance. The mobile phase is an organic solvent system (in other words, a mixture of one or more organic solvents), which, by capillary action, will move up the stationary phase. Solvent systems are considered to be non-polar compared to the silica, though there are degrees of non-polarity. The sample mixture is applied as a small spot near one edge of the TLC plate and allowed to dry and thus adsorb to the silica; this is called spotting. The plate is then put vertically into a solvent system reservoir such that the spotted edge is placed down (but keeping the spots above the level of the reservoir) and the solvent system will ascend the plate. As the solvent system goes up, the compounds in the sample mixture will (ideally) separate; some of the compounds should stick to the stationary phase and some should dissolve and be carried up the plate along with the mobile phase. This process is called developing the TLC plate. To get an idea of why compounds in the mixture separate, consider a mixture that contains both a polar and a non-polar compound. The polar compound will favor the adsorbent silica (the stationary phase) because the silica is highly polar (following the rule of like dissolves like ). The non-polar compound will favor dissolving in the non-polar solvent system and travel upward with the solvent front. Thus, each compound will ascend the plate at different rates, with more polar compounds tending not to rise quickly. Separation is achieved! You will examine the TLC mobility of various benzene derivatives. Materials and methods 5% (w/v) solution of benzaldehyde in methylene chloride 5% (w/v) solution of benzoic acid in methylene chloride 5% (w/v) solution of benzyl alcohol in methylene chloride

2 5% (w/v) solution of methyl benzoate in methylene chloride 5% (w/v) solution of benzamide in methylene chloride 0.5% (v/v) glacial acetic acid in ethyl acetate hexane one 2 by 4 inch standard silica TLC plate capillary tubes one 20 ml Erlenmeyer flasks two 10 ml graduated cylinders ultraviolet viewing boxes developing jar pencil, ruler, spatula, forceps, transfer pipets Procedure 1. Obtain a 2-inch wide TLC plate and six capillary tubes for spotting the plate. Handle the plate by its edges, taking care not to bend the plate excessively or to otherwise cause the silica to flake off. 2. Using a soft pencil (not a pen!) and a ruler, draw a straight line lightly across the width of the plate, about 2 cm from what will be the bottom of the plate. Along this line, leaving some space at the edges, draw lightly five evenlyspaced tick marks; this is where you will be spotting the solutions. 3. Dip the end of a capillary tube into one of the five solutions (note which one) and watch it fill (it won t fill to the top). Spot the plate by lightly touching it to the TLC plate at one of the tick marks. Note the spreading circle of moistness as the capillary empties; make sure this spot does not exceed 2 mm in diameter (if it seems to be getting bigger, merely lift the capillary tube off of the plate, wait for the wet spot to dry and respot the plate at the same location repeat this until the tube is empty). 4. Draw an exact replica sketch of your TLC plate on the next page, labeling each spot as you finish making it. It is an excellent visual aid, especially since you can sketch in what the TLC plate looks like after developing. 5. Repeat step 3 with the four other solutions, taking care to use a fresh capillary tube for each different solution. 6. A jar is used for the development chamber. Make the solvent system in the glassware provided: a 50/50 mixture of hexane and 0.5% acetic acid in ethyl acetate. 10 ml should be plenty of solvent; carefully pour the solvent into the jar. The bottom of the jar should be completely covered, but the level must be shallow enough so the spots will be completely above the level of the solvent when the plate is in the jar.

3 7. Develop the plate by placing it in the jar such that it leans across the jar, facing upward. Make sure the spots are not immersed in solvent. If that happens, spotted material will dissolve in the pool of liquid rather than climbing up the silica on the plate. Be careful not to slosh liquid; that will interfere with the liquid climbing evenly across the plate and will give unreliable results. Cap the jar and let the solvent rise (you can see an ascending solvent front as time goes on). 8. When the solvent front is about 2 to 3 cm from the top of the plate, remove the plate from the jar and quickly mark the solvent front with the pencil. This is critical! The line of the solvent front will disappear quickly as it evaporates, and the distance from the spotting line to the highest point reached by the solvent is used to calculate all the Rf ( retention factor ) values. You will have no numerical data without it! Let the plate dry in the hood. 9. Place the dry TLC plate in the UV (ultraviolet) box; the compounds should appear as dark purple spots. Use a pencil to outline all the spots, including any irregularities. Make a sketch of the plate below, preserving all of the different distances and shape irregularities. The reason for this is that the silica will flake off the plate too readily for it to be a permanent record. Sketch of TLC plate (steps 4 and 9)

4 10. Enter your data into the table below: TLC plate lane number Distance traveled by compound (mm) Distance traveled by solvent front (mm) Rf value 11. Measurements should be made on the plate, not the drawing in the notebook. Measure the distance from the tick mark to the solvent front in each lane and the distance from the tick mark to the compound spot in each lane; enter the measurements (in millimeters) in the appropriate row in the table, and also mark it on the plate drawing. 12. Calculate the retention factor (Rf) value by dividing the distance traveled by the compound by the distance traveled by the solvent front, and enter that value into the table. Questions 1. All other things being equal, should a more polar substance have a higher or lower Rf than a less polar one? Explain your answer.

5 2. Predict the order of polarity of the five different benzene derivatives, from least to most polar. Indicate why you think the order is so. 3. Using your TLC results, rank the order of polarity of the five different benzene derivatives, from least to most polar. How did you obtain this order?

6 4. Describe the differences between your predicted order and your observed order. 5. What else besides overall molecular polarity might be going on here? Hint: consider other intermolecular forces. Come up with a new ranking based on your answer. Does this match the TLC results? 6. Benzoic acid and benzamide are solids at room temperature, while the rest of the benzene derivatives are liquids. Is the Rf value of a compound (which you know is already affected by the compound s polarity) affected by whether it is a liquid or solid at room temperature? Using observations from the lab, explain why or why not.