Experiments to show the factors required in photosynthesis (2) - light and carbon dioxide

This unit may be unduly censored by software installed by (well meaning??) network managers, resulting in safety advice being removed due to references to uncovered flames - perceived lack of clothing around sources of ignition being apparently seen as a greater moral risk than the physical risk of injury due to burns.

These experiments start with destarched potted plants - Geranium (Pelargonium) is often used.
These are then subjected to various treatments, left in the light for a few days then subjected to decolourising procedures and then finally tested for the presence of starch.

Destarching the plant:

The plant is placed in a dark place, e.g. a cupboard for 24 or 48 hours. It is essential that the plant is watered beforehand, and not forgotten about! This is so as to force the plant to use its reserve of starch, and not allow its replacement by photosynthesis.

Before proceeding to the next stage, a leaf is removed and checked for the presence of starch (See under the section "Testing a leaf for starch").
If no starch is present, then it is possible to proceed to the next stage. If it is, then it must be returned to the dark.

Treatments

A The effect of light

Part of the leaf may be covered with dark paper or card - possibly with perforations - and held in place with paper clips.

Alternatively, an envelope of aluminium foil (with a distinctively shaped hole in the centre) may be placed over both sides of the leaf, and the edges of the hole pinched in a little so that light only reaches the exposed part of the leaf.
Another method involves using a pair of thin cork or discs which are used to sandwich the leaf, held in place by a pin.

It is also possible to attach a photographic negative to a leaf using a special holder.

B The effect of carbon dioxide

Leaves of the plant are enclosed within glass flasks, supported at an angle by stands and clamps (250/350ml wide-mouthed flasks will usually do). Each flask will contain small amounts of various liquids, which should not be allowed to touch the leaves.
Corros B1 Sodium hydroxide solution or potassium hydroxide solution - CAUTION - CORROSIVE!
B2 Sodium hydrogencarbonate solution or simply carbonated water
B3 water
B4 no liquid

2flasks After the insertion of the leaf, 2 wads of absorbent cotton wool are placed in the neck of the flask, one above and one below the petiole (leaf stalk). These are then saturated with water (from a washbottle). This may need to be repeated a couple of times a day.
Alternatively, split corks or bungs can be used, together with lots of petroleum jelly (messy!).

The plant must be kept well watered and exposed to good light.
After a couple of days or more, the leaves are removed for testing. It may be useful to mark them with cuts to identify them later.

Decolourising the leaf

Strict safety precautions must be observed, because ethanol vapour is very flammable.
This procedure might be best demonstrated by the teacher.
Depending on the facilities available in the lab, there are two main methods. Each has 2 steps (a & b).

Using Bunsen burners:
Heat Noheat
a) A beaker containing water (supported by a tripod and gauze) is brought to the boil by heating it with a bunsen burner set to a non-luminous flame.
Check that the beaker is stable on the tripod, then:
Place the leaf in the boiling water for 1-2 minutes or until soft and flabby.

b) Turn off your burner and ensure that all burners on your bench are out before moving to the next step.
Transfer the leaf to another container of Industrial Methylated Spirit (initially colourless). Do not use purple meths or pure ethanol.

The container is then heated gently - preferably by floating it in the hot water in the same beaker as before, and left for several minutes.
Do not re-warm the beaker using a lit Bunsen burner.

Using electrically heated waterbaths:

a) Place the leaf directly into a waterbath at 100 °C/maximum setting.

b) Transfer to a beaker of IMS in a waterbath set to 78 °C.
The same precautions apply as above - no naked flames.

Procedure after decolourisation - testing for the presence of starch

Immerse the whole leaf briefly in cold water - tap water will do.

Spread out the leaf in a plastic (Petri) dish.

Add a few drops of iodine solution (I/KI), and wait a few minutes for colour to develop.

Analysis

Grey-blue-black areas represent starch (e.g. B2 above).

Yellow-orangey-red areas represent no starch (e.g. B1 above).

It may be necessary to hold the dishes (together?) over white paper to see any differences.

It is necessary to make comparisons of :
- areas where light fell and areas where light did not fall in experiment A

- whole leaves exposed to high levels of CO₂ and leaves exposed to low/no CO₂.