When a pigment absorbs light, electrons of certain atoms in the pigment molecules are boosted to a higher energy level. The energy of an absorbed photon is converted to the potential energy of the electron that has been raised to an excited state. In most pigments, the excited electron drops back to its ground-state, or normal orbit, and releases the excess energy as heat. Some pigments, including chlorophyll, emit light as well as heat after absorbing photons.
In the chloroplast, these excited electrons jump from the chlorophyll molecule to a protein molecule in the thylakoid membrane, and are replaced by electrons from the splitting of water. The energy thus transferred, is used in carbohydrate production.
This release of light is called fluorescence. Chlorophyll will fluoresce in the red part of the spectrum, and also give off heat. In this lab, you will observe this fluorescence by separating the chlorophyll from the thylakoid membrane.
|Spinach leaves||Flashlight or small lab light|
|Mortar and pestle||Test tube|
|25-mL graduated cylinder||Funnel|
|Ring stand or funnel rack||Safety goggles|
1. Grind the spinach leaves using a mortar and pestle.
2. Add acetone to the ground leaves, using enough acetone and spinach leaves to get between 10 and 15 mL of extract.
3. Set up your filtering apparatus, and using proper filtering technique, filter the extract to a test tube. NOTE: Use a small amount of acetone to wet the filter paper, to hold it into place, instead of water.
4. Shine a flashlight, or other similar light source, through the test tube and extract.
5. Observe the fluorescence of the chlorophyll at a 90 degree angle to the flashlight.