Fluorescence of Chlorophyll
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Chlorophylls
In reality, chlorophyll is not a single substance but an entire class of chemical compounds with similar structures and properties. Chlorophylls are organic compounds found in plants, algae, and photosynthetic bacteria. They are responsible for the characteristic green color of plants.
The role of chlorophylls in photosynthetic organisms is to capture photons and transfer their energy for use in later stages of photosynthesis. The green color of chlorophyll results from its strong absorption in the red and blue regions of the light spectrum and weak absorption in the green region.
There are several types of chlorophyll. The most common in nature are chlorophyll a and chlorophyll b, found in all plants that carry out photosynthesis. Chlorophylls c and d occur only in certain algae.
As an example, here is the structural formula of chlorophyll b:
Its molecular formula is C55H70O6N4Mg, which makes it a relatively complex molecule.
Today, we will use a simple method to isolate chlorophyll from plant tissues and observe its fluorescence under ultraviolet light.
Experiment
We need a source of chlorophyll. Any green plant material will work, though fleshy leaves give the best results. In winter, when fresh plants are scarce, spinach (even frozen) can be used.
The leaves should first be cut into small pieces. Since chlorophylls are insoluble in water, we must use a different solvent. Acetone C3H6O is suitable for this purpose.
Warning: Acetone is not highly toxic, but it is classified as an irritant and may cause temporary health issues. It is a very volatile liquid, so avoid inhaling its vapors. The author assumes no responsibility for any potential damage. Proceed at your own risk.
Grind the leaf fragments in a mortar with a small amount of acetone. During this process, the solvent extracts the chlorophyll, turning a characteristic green color. The solution should then be filtered to remove plant tissue fragments.
The resulting acetone solution of chlorophylls has a bright grass-green color, as shown below.
Is the solution truly green? Under visible light, it certainly is, but let’s try illuminating the sample with ultraviolet light. The result looks quite different:
Under UV light, the solution glows with a vivid carmine-red color. You can also observe this effect by illuminating the solution with a semiconductor laser that emits light at a wavelength of λ = 405 nm:
This simple experiment confirms the fluorescent properties of chlorophyll, one of the most common natural substances.
Explanation
Each chlorophyll molecule is based on a pheoporphyrin structure derived from porphyrin. Four of its rings are pyrrole rings, and the fifth consists entirely of carbon atoms. The bonds between the atoms in the rings alternate between single and double, forming a conjugated bonding system.
At the center of the porphyrin system is a magnesium atom that bonds with the nitrogen atoms of each ring. A porphyrin complex containing magnesium can absorb electromagnetic radiation in the visible range.
Because of this structure, a chlorophyll molecule can also be excited by ultraviolet light. An excited molecule has more energy than it does in its ground state. This excited state is unstable, and after a very short time, the molecule spontaneously returns to the ground state. According to the law of energy conservation, the difference between these energy levels is released as electromagnetic radiation, which appears as the red light we observe.
Further readings
- Berg J.M., Tymoczko J.L., Stryer L., Clarke N.D., Szweykowska-Kulińska Z., Jarmołowski A., Augustyniak H., Biochemia, Wydawnictwo Naukowe PWN, Warszawa, 2007, p. 530
- Kopcewicz J., Lewak S., Gabryś H., Fizjologia roślin, Wydawnictwo Naukowe PWN, Warszawa, 2005
- Muyskens M., Vitz E., The Fluorescence of Lignum nephriticum: A Flash Back to the Past and a Simple Demonstration of Natural Substance Fluorescence, Journal of Chemical Education, 2006, 83(5), p. 765
Marek Ples