2: Chromatography

Question 1

How could we separate photosynthetic pigments?

Answer 1

with chromatography

• paper chromatography used in class
• thin layer chromatography is better

Question 2

Describe method for chromatography (first 4 steps).

Answer 2

1. tear up leaf into small fragments
2. grind pieces of leaf with sharp sand and propanone to extract leaf pigments
3. transfer sample of extract to a watch glass
4. evaporate to dryness with hot air from hair-dryer

Question 3

Describe method for chromatography (second 4 steps).

Answer 3

5. add a few drops of propanone to dissolve the pigments
6. build up a concentrated spot of pigment 10mm from the end of the strip of paper/TLC strip
7. suspend the strip in a tube with the base dipping into running solvent
8. remove the strip from the tube when the solvent has nearly reached the top (of strip)

Question 4

What is the Rf value? Describe method for calculating Rf value.

Answer 4

Rf = ratio of the distance traveled by a compound to that of the solvent front

Rf = distance moved by spot (a pigment) / distance moved by solvent

Question 5

Why do pigments separate in chromatography?

Answer 5

because of their varying solubility in the running solvent

Question 6

State 4 main pigments and their approximate Rf values.

Answer 6

1. carotene (0.9)
2. chlorophyll a (0.65)
3. chlorophyll b (0.6)
4. xanthophylls (0.3-0.5)