Thin-Layer Chromatography

Question 1

What is the first stage/step in Thin Layer Chromatography? Describe the process for this stage. Why do you not want the lines to flake off the absorbent?

Answer 1

1. ) Spotting Stage
   - Preparation for spotting: Draw a line 1 cm away from the bottom of the TLC plate in PENCIL (this is the spotting line). Then draw a 5 mm line from the top of the TLC plate (this is the solvent front).
   - You want the lines to be drawn lightly.
   - You do not want to flake off the absorbent because if flaked, a gap may form that may stop the flow of eluent or cause the eluent to flow crookedly.

Question 2

What is eluent?

Answer 2

• The eluent is the liquid solvent that travels up the TLC plate.

Question 3

Why is pencil always used to mark a plate while ink is not?

Answer 3

• You use pencil because it is almost 100% graphite and will not be separated by the eluent solvent (no interaction), while ink is made up of many dyes and/or pigments, solvents and other liquids that will be separated by the eluent solvent. The separation of ink interferes with the results of your TLC plate.

Question 4

Describe Spotting.

Answer 4

1. ) Use capillary tubes to take up a sample.
2. ) LIGHTLY touch it to the TLC plate to apply a spot of the sample onto the spotting line (1 cm line). Try not to flake of absorbent.
3. ) Label the spots with a pencil.

Question 5

How to avoid streaking spots?

Answer 5

1. ) By touching lightly to the TLC plate and allowing time for your initial spot to completely dry before applying again to the same spot.
   - If this does not occur then the spot will become too large.
   - The sample should NOT be too concentrated.

Question 6

What problem could streaking spots lead to?

Answer 6

• Streaking may lead to an overlap in two or more compounds, skewing the results.

Question 7

What happens if a sample contains too many components?

Answer 7

• Then spots may just run together and appear as a streak.

Question 8

Why should you not put spots too close together on the spotting line?

Answer 8

• They may bleed into each other.

Question 9

Why should you not put spots too close to the edge?

Answer 9

• Leads to inaccurate Rf’s because there is not enough solvent or absorbent surrounding the spot (unequal forces for each spot).