Practicals

Question 1

how can valid results be obtained with a potometer?

Answer 1

• use a healthy, fresh plant shoot
• cut plant shoot and assemble potometer underwater (avoid air bubbles)
• dry leaves (control humidity/initial Ψ around leaves)

Question 2

how to prepare a photosynthetic pigment extract? (and why)

Answer 2

• soak leaves in acetone (propanone) to act as a solvent
• add a pinch of sand (help grind)
• use a mortar and pestle to grind up leaves with acetone

Question 3

stationary phase in TLC

Answer 3

silica gel layer of TLC plate

Question 4

what solvent is used for photosynthetic pigment TLC?

Answer 4

• cyclohexane
• propanone (acetone)
• petroleum ether

Question 5

why must a lid be used on TLC?

Answer 5

• solvents are volatile
• evaporate quickly
• will not diffuse up the TLC plate and carry extract

Question 6

how and why must you be careful when handling TLC plates?

Answer 6

• avoid fingerprints
• avoid damaging fragile silica gel layer

Question 7

why must the TLC plate not touch the sides of the beaker/tube?

Answer 7

• solvent may evaporate off the plate, condense and drip down the side of the container
• can damage the silica gel
• can skew the movement of solvent (making it hard to measure Rf values)

Question 8

how can photosynthetic pigments be identified from TLC?

Answer 8

• measure Rf values and record colours
• compare to known data

Question 9

what can be used to add sample spots to TLC and why?

Answer 9

capillary tube for small spots

Question 10

how many spots should be applied in TLC?

Answer 10

• multiple (3/4) spots of EACH sample in the SAME spot (concentrated pigment)
• if using different samples, spot them in different places

Question 11

what is a TLC plate placed in?

Answer 11

developing chamber (tall beaker with lid)

Question 12

how can solvent evaporation be reduced in a developing chamber?

Answer 12

• add a lid
• place a layer of filter paper around inner walls of developing chamber, wet the filter paper with solvent to adhere to the sides of the chamber and saturate it (prevent solvent from TLC evaporating and condensing on the side of the chamber)

Question 13

how does the solvent travel through TLC?

Answer 13

capillary action

Question 14

why must the TLC plate be taken out before the solvent reaches the top?

Answer 14

• once the solvent reaches the top, Rf values may be invalid
• pigments may start to run into each other (ones at the bottom catch up with the ones at the top etc)

Question 15

why must you mark the solvent front immediately with a pencil?

Answer 15

• avoid solvent evaporating (will no longer be able to see the solvent front)
• pencil to avoid interference with the pigments and damage to the silica gel

Question 16

how are molecules separated in TLC?

Answer 16

separated by polarity (depending on the solvent)