Separation of the pigments of a leaf Introduction: We know that there is usually more than one pigment in a leaf.

We can separate these pigments using a process called chromatography. This process depends on the fact that different pigments have different molecular weights and have different solubility in certain chemicals Aim: To separate the pigments in a leaf Apparatus: 1 piece of Chromatography paper 1 boiling tube 1 cork with a paperclip inserted into it 1 mortar and pestle A few pinches of Sand 1 blunt toothpick Materials: Chromatography solution (92% petroleum and 8% acetone) Acetone Spinach leaves Method: 1. Cut piece of chromatography paper to fit a boiling tube without touching the sides. 2. Place chromatography fluid in boiling tube and cork so the atmosphere inside tube can become saturated with the vapour 3. Tear up spinach leaves and place in mortar and pestle. Add a little sand to help break up the tissue and acetone to extract the pigments. Grind mixture to a pulp 4. Let it settle- make sure you take the extract of pigment off the top of the mixture so it will be more concentrated. 5. Lightly mark a distance from the bottom of the paper- about 30mm- with a pencil 6. Place the paper on a raised ledge so that it pokes out into the air. Put a spot onto this overlap, so that the spot is not spread out or contaminated with dirt. Use the blunt toothpick to do this. 7. Keep putting a spot (or line) on, letting it dry in between, until the spot of extract is very concentrated. 8. Use the paper clip to place the paper in the boiling tube so that the bottom of the paper just touches the fluid. Make sure that the pigment spot is above the fluid line 9. Stand the tube upright and watch until the chromatography fluid has cleared the pigments Results: Pigments Carotene Xanthophyll Chlorophyll A Chlorophyll B Colour Yellow Yellow Blue-green Yellow-green

Conclusion: Which was the most soluble pigment? Carotene was the most soluble pigment as it travelled the furthest, as shown in above results. Which of the pigment would you always find in a leaf? Chlorophyll because the green pigment in leaves is chlorophyll. It absorbs red and blue light from the sunlight and reflects green, and the leaf therefore appears green. What is the advantage to the plant of having more than one pigment? Other pigments: carotene and xanthophyll, are able to capture energy from a light source that chlorophyll A or B cannot.