Thin Layer Chromatography Lab Report

Thin Layer Chromatography Objective: This experiment uses thin-layer chromatography to determine the solvent polarity effects and to identify an unknown substance. Retention factors will be calculated to determine the identity of the unknown compounds. Materials: Breakers, flasks, stir rod, mini capillaries tubes, graduated cylinder, TLC plates, filter paper, Aluminum foil top, watch glass, tweezers, UV light, developing chamber, ruler, pencil, way paper, metal spatula, Acetaminophen, Aspirin, Caffeine, Ibuprofen (TBD), Ethanol, Dichloromethane, Acetic Acid, Ethyl Acetate, Hexanes. Table: Sample name Chemical Formula Molar Mass Density Melting Point Solubility in Ethanol Ethanol C2H5OH 46.07 g/mol 789 kg/m3 -144.1°C soluble Acetaminophen C8H9NO2 151.163 g/mol 1.26 g/cm³ 169°C Slightly soluble Caffeine C8H10N4O2 194.19 g/mol 1.23 g/cm³ 227-228 °C Slightly soluble Ibuprofen C13H18O2 206.29 g/mol 1.03 g/cm3 75-78 °C Soluble Aspirin C₉H₈O₄ 180.158 g/mol 1.4 g/cm³ 135°C Slightly Soluble Dichloromethane CH2Cl2 84.93 g/mol 1.33 g/cm³ -96.7°C Slightly Soluble Acetic Acid CH₃COOH 60.052 g/mol 1.05 g/cm³ 16.6°C Slightly Soluble Ethyl Acetate C4H8O2 88.11 g/mol 902 kg/m³ -83.6°C Slightly Soluble Paprika C40H56O3 584.87 g/mol 0.1 g/cm3 181–182 °C Slightly Soluble Hexanes C6H14 86.18 g/mol 655 kg/m³ 69°C Slightly Soluble Procedures: 1. Transfer a small amount of each solution onto paper and make sure to crush it into a powder before transferring it into a flask. 2. Add 5ml of ethanol into the acetaminophen (50mg), aspirin(30mg), caffeine (50mg), and the unknown (pre-made) solution. 3. Use glass rods for each solution to stir until fully dissolved 4. Place filter paper and place it in the bottom of a 250ml beaker 5. Measure out .2ml of acetic acid and 3.8ml of ethyl acetate into the 250ml beaker. (add 1.5-2ml of hexanes if needed) 6. Now, cover the beaker with a watch glass or aluminum foil. 7. Get a TLC plate and make three lines using a pencil labeling acetaminophen, caffeine, and aspirin. 8. Place a drop of each solution on the starting line of the TLC plate and where they are labeled. 9. Wait for the spots to dry and use the UV light and check to see if the spots are getting darker. 10. Add TLC plate into the beaker with the solution and wait until it starts to go up. 11. Remove the TLC plates and mark them using a pencil to the part where it went up. 12. Using UV light to mark the areas of the spots using a pencil 13. Measure the distance how far each solution traveled. For note, if needed: Unknown Sample preparation: Crush part of the unknown tablet, transfer to a test tube, mix with a bit of ethanol 65 mg of Unknown A in 6.5 mL of ethanol 50 mg of Unknown B in 5 mL of ethanol. 50 mg of Unknown C in 5 mL of ethanol. Use the same steps above to identify the unknown sample. Paparika color seperation: 1.) Prepare TLC plates like in the steps above (divide into two lines) 2.)Prepare a developing chamber by using a 250-mL beaker and adding 3.4 mL heptane, 0.6 mL diethyl ether(or 10ml), Filter paper, Aluminum foil, or Watch glass. 3.) Spot the plate with two spots of the paprika 4.) place the plate in the developing chamber until the solvent reaches 1cm from the top 5.) remove the plate from the chamber and mark with a pencil, and let dry 6.) Note the color and polarity and calculate the Rf values of the different spots Pre-Lab: 1.) Can you fill the solvent (mobile phase) above the origin line? What problems would you foresee? The solvent should not be filled over the origin line. The reason why is because the solvent will dissolve the compound into the reservoir and not allow the capillary action to proceed. Meaning that the spots will not be observed after the plate is developed. 2.) Sketch the movement of hypothetical samples with 2 carbon atoms [A-C2-OH (an alcohol); B-C-COOH (an acid); C-COOCH3 (an ester); D: C2H6 (an alkane)] on a TCL plate. Label clearly. 3.) Quote a hypothetical example when TLC and Recrystallization are employed in research (1-2 lines only). A hypothetical example would be if substances needed to be purified and identified their polarity and Rf values for research. Observation and Date/Results: Compound names Rf values (cm) Polarity Acetaminophen 2.7/3.3 = .818 Second least polar Caffeine .9/4= .225 Most polar Aspirin 3.1/3.3= .939 Least polar Unknown C 3.1/3.3= .939 or 2.7/4= .675 Least polar The Unknown C is Aspirin because of the matching Rf values. Conclusion: Thin-layer chromatography is a lab technique that can help identify compounds. Along with identifying the polarity of the compound. The more polar the compound is, the more absorption will happen, making the compound move at a lower rate, unlike nonpolar compounds that will move of the TLC plate at rapids rates, making them have higher Rf values. So the higher the Rf values are, the more nonpolar the compounds are. With the data that was collected, it can be assumed that Aspirin is the most nonpolar (Rf ), Acetaminophen is the second most nonpolar substance (Rf ), and caffeine is the least nonpolar (Rf ). Experimenting and calculating the Rf values can visualize which compounds make up the unknown substance, concluding that Unknown compound C was Aspirin. Post-Lab: 1.) Why is it necessary to run TLC in a closed container and have the interior vapor saturated with the solvent? The working principle of TLC separation is to move chemicals up to the plate through a solvent. Since the solvents used in TLC are usually very volatile, if the solvent starts to evaporate, the Rf value will change according to the amount of solvent evaporated. Due to evaporation, there will be inconsistent Rf values between TLC plates. Therefore, if the chamber is closed, all the air in the chamber will be saturated with solvent vapor. Since the air is saturated, the solvent on the plate will not evaporate, thus providing a more consistent Rf value. 2.) Explain what will be the result of applying too much or too little compound to a TLC plate? A- Adding too little of the compound to a TLC plate will cause no appearance after the elution making it hard to identify the compound. B- Applying too much of the compound to a TLC plate can cause the compound to mix with others, making it challenging to analyzed accurate reading. Also, too much compound can cause a tail to form cause smears on the plate. Bonus: 1.) What is the reason we start with a nonpolar solvent? Nonpolar solvents will force nonpolar compounds to the top of the plate because the compounds dissolve well and do not interact with the polar stationary phase. 2.) What will happen to the TLC plate if we start with a polar solvent? The polar and nonpolar compounds will run with polar solvents simultaneously. Then there will be no separation in the TLC plate, so there will be no movement up the plate and mix together.