Experiment 3.7 Solvent and Polarity Effects in Thin-layer Chromatography

1. Experiment 3.7Solvent and Polarity Effects in Thin-layer Chromatography

2. Purpose of Experiment • Determine factors that affect rate of elution (Rf) of organic compounds. • Relationship between polarities of compounds and their rates of elution (Rf) • Relationship between solvent polarity and rate of elution (Rf) • Select an appropriate solvent system to separate and identify components of a mixture.

3. Purpose of Experiment • Possible components of unknown mixture trans-stillbene 9-fluorenone benzoic acid

4. Chromatography “Color Writing”

5. Chromatography Stationary PhaseMobile Phase What does it do? stays put! flows around SP What is it? fine solid with fluid (liquid or gas) (physical form) lots of surface How does it effect retains sample by moves sample along sample movement? surface interaction Analogous to… an obstacle course “motivating force” silica gel (SiO2)x ethyl acetate

6. These exposed OH units give silica gel a relatively polar surface. surface Thin Layer Chromatography TLC plate silica gel - silicon dioxide (SiO2)x (a common, inexpensive stationary phase) 5 x 10 cm 250 mm silica gel layer impregnated with a fluorescent indicator, on a foil backing bulk (SiO2)x

7. Thin Layer Chromatography • Five Stages in TLC • Prepare Developing Chamber Saturate with solvent vapor. • Apply Samples Capillary used to spot solution of each sample. • Develop Plate This is when the separation actually occurs. • Visualize Developed Plate View under UV light. • Interpret Results Determine Rf values & identify components.

8. 1. Prepare Developing Chamber • 400mL beaker. • Place a piece of filter paper against side of beaker. • Pour 10mL of developing solvent (mobile phase) onto filter paper. • Cover beaker with a watch glass and allow to stand undisturbed.

9. 2. Apply Samples (spot the plate)  1 cm. “starting line”  1 cm. TLC plate Process A. Draw “guide lines” lightly with pencil B.Dissolve solid sample in CH2Cl2 C. Use TLC capillary to transfer and spot dissolved sample T-stillbene benzoic acid 9-fluorenone unknown

10. TLC plate 3. Develop TLC Plate • Place spotted TLC plate in developing chamber. • Developing solution is drawn up the plate by capillary action. • Remove TLC plate when solvent is ~ 1cm from top. • Mark solvent front position with a pencil NOTE: During this ~20 min. developing stage, compounds in the original spots are being pulled through the silica gel. Developing Chamber (400 mL beaker with 10mL solvent)

11. B. View results under UV light. Look for colored spots on the fluorescent green background   4. Visualize Results A. Allow solvent to evaporate from surface of TLC plate. UV C. Mark spots with a pencil while viewing under UV light.

12. X Y T Y Z Y distance spot has moved distance solvent has moved distance spot has moved distance solvent has moved distance spot has moved distance solvent has moved ____________ ___________ _______________________ _______________________ Rf= Rf= Rf= = = = 5. Interpret Results (Rf Values) T Y Z Z X

13. UNK 9-f ts ba std std std 5. Interpret Results (identify components) • Compare Rf values of components in sample to Rf values of standards.

14. Prelab Preparation • Read Technique K (pp. 92-100) • Read Experiment 3.7 A & B (pp. 210 – 216) • Prepare Notebook • Title, purpose, procedural reference, procedural changes • Table of reagents: • Structures and hazards of solvents (hexane, ethyl acetate and acetone) and compounds to be separated • Rank the solvents in order of increasing polarity. • Rank the compounds in order of increasing polarity. • Prepare a data table like Table 3.7-1 to record distances and Rf data.

15. Procedural Changes • Ethanol will not be evaluated as a solvent. • Do part A in groups of 3. • Do part B individually.

16. In Lab • Part A • Determine Rf of trans-stilbene, 9-fluorenone and benzoic acid in hexane, ethyl acetate and acetone. • Each person in group does 1 solvent. • Determine the best solvent to use to separate a mixture of trans-stilbene, 9-fluorenone and benzoic acid. • Review with lab instructor before doing part B.

17. Data Collection • Part A • Record the solvent used for each TLC plate. • Measure and record the distances that each compound and the solvent front traveled for each solvent used. • Calculate and record the Rf values for each compound in each solvent.

18. In Lab • Part B • Use the best solvent determined in part A to detemine components in an unknown mixture.

19. Data Collection • Part B • Record the unknown number. • Record the solvent used for the analysis of the unknown. • Measure and record the distances that each known compound, each spot in the unknown and the solvent front traveled. • Calculate and record the Rf for each known compound and each spot in the unknown. • Identify components in unknown.

20. Thin Layer Chromatography • TLC lab technique hints • Slide watch glass off beaker instead of lifting it off to maintain solvent vapor saturation in beaker. • Keep spots small. • Never double dip spotting pipette. • Put sample(s) to be analyzed near center of plate. • Separate spotting points evenly. • Do not disturb beaker during development.

21. Thin Layer Chromatography How does solvent polarity affect Rf? How does solvent polarity affect relative Rf of 2 or more components? How does component polarity affect Rf?