Energy Transfers in and Between Organisms: Photosynthesis - Required Practical 7, Chromatography

Question 1

What is chromatography?

Answer 1

• Chromatography is used to separate out different components in a sample

Question 2

What components will separate if we use grounded leaves in chromatography?

Answer 2

• The photosynthetic pigments of a plant sample are separated into bands of colour by paper chromatography

Question 3

What are the factors affecting the rate of migration of different pigments?

Answer 3

• Solubility
• Mass
• Affinity to the paper

Question 4

What does affinity mean?

Answer 4

• Affinity describes the degree to which a substance tends to combine with another
• ‘Affinity to the paper’ means how well the pigment binds to the paper

Question 5

What are the two different phases of chromatography?

Answer 5

• Mobile phase

- Stationary phase

Question 6

What is the mobile phase?

Answer 6

• Phase where the molecules can move

- In this experiment, the mobile phase is a liquid solvent (propanone and petroleum spirit)

Question 7

What is the stationary phase?

Answer 7

• Phase where the molecules can’t move

- In this experiment, this is the chromatography paper

Question 8

Describe what happens during the different stages of chromatography, including the phases

Answer 8

• The mobile phase moves through the stationary phase
• The components in the mixture spend different amounts of time in the mobile phase and the stationary phase
• The components that spend longer in the mobile phase travel faster or further

Question 9

How can you identify the pigments in leaves?

Answer 9

• Calculate a Rf value

- Each pigment has a specific Rf value, under specific conditions, which can be looked up in a database

Question 10

What is a Rf value?

Answer 10

• An Rf value is the distance a substance has moved through the stationary phase in relation to the solvent
• Distance substance moved / distance solvent moved

Question 11

Describe the method used

Answer 11

• Draw a horizontal pencil line 1cm above bottom of chromatography paper
• Grind leaves with pestle and mortar. Add propanone to stop drying
• Extract drop with capillary tube and blot on chromatography paper. Dry and repeat again, 9 more times
• Add the chromatography solvent (propanone and petroleum spirit) to boiling tube
• Suspend chromatography paper in boiling tube, line must be above solvent
• Wait for solvent to rise and separate pigments
• Remove paper before solvent reaches the top and mark the top of solvent and each pigment with pencil
• Measure distance moved by solvent and each pigment
• Repeat with a different leaf
• Calculate Rf values

Question 12

How does affinity affect the rate of mobility?

Answer 12

• The pigments with lower affinities will travel further up the paper

Question 13

How does solubility affect the rate of mobility?

Answer 13

• Pigments that are more soluble, travel faster up the paper and will end up closer to the top at the solvent front