Chromatography

TLC

• Thin layer chromatography

• Stationary phase• Mobile phase

Separation AND Characterization

Chromatography Basics• Based on different affinities for stationary and

mobile phases• Silica gel: polar, water-covered surface– Compound(s)• Polar: _______ affinity for plate, travels _______• Nonpolar: _______ affinity for plate, travels ______

– Developing solvent• Polar: higher affinity for plate, travels slower, displaces

compound more (compound travels __________)• Nonpolar: lower affinity for plate, travels faster,

displaces compound less (compound travels ________)

Test your Understanding

• Which spot represents a more polar compound?

• What would happen to each spot if a less polar solvent were used?

• Why should you ALWAYS report your developing solvent with any TLC data?

Quantitative Characterization

• Retention factor• Distance traveled/

solvent front distance

• Unitless• For silica gel TLC,

based on polarity of the compound(s)

Must report solvent!

Solvent effect on Rf

• Polar solvents outcompete compounds, drive them up plate

Choosing a Developing Solvent

• Adjust solvent to give Rf values around 0.4

• Common mixtures– Ether/Hex– EtOAc/Hex– CH2Cl2/methanol

• Determined experimentally

Visualization

• Most compounds are invisible on TLC

• UV lamp• Stains• Iodine chamber

Application of TLC

• Purity• Identity• Reaction Progress

What can we determine about the identity of the unknown?

Column 1 is your target compound; column 2 is an expected impurity. What can you determine about your reaction (column 3)?

Common Problems

• Overspotting• Underspotting• Wrong solvent

Column Chromatography

• Similar to TLC in separation• Preparative process– 0.1g to 5 g scale

• Purify small quantities of liquids/solids (contrast recrystallization)

• Mobile phase: eluent similar to TLC

• Stationary phase: silica gel similar to TLC

• Column is upside down from TLC, so a larger Rf for a compound means it comes out ____

Practical Considerations

• Preparing the column• Loading the sample• Choosing the solvent• Separation capacity

Practical Considerations

• Preparing the column• Loading the sample• Choosing the solvent• Separation capacity

Practical Considerations

• Preparing the column• Loading the sample• Choosing the solvent• Separation capacity

Most common: Hexane/ethyl acetate CH2Cl2/methanol

Practical Considerations

• Preparing the column• Loading the sample• Choosing the solvent• Separation capacity– Effect of diameter– Effect of length– Effect of Silica gel grade

Flash Chromatography

• Faster separation• Tighter

separations

HPLC