1. Looking at leaf stomata and plant pigments

2. Bloody Chlorophyll
Sunlight is white light that is actually a mixture of different wavelengths of light from the visible light spectrum. Photosynthetic plants gather energy from sunlight through the use of light-absorbing pigments. The plants use this energy to power the synthesis of organic molecules. Chlorophyll is the green pigment in the chloroplasts of most plants. Chlorophyll absorbs light very well in the red and blue-violet regions of the visible spectrum, but it does not absorb green light very well. Green light is instead reflected, which is why chlorophyll, and the leaves of plants where it is found, appear green. When chlorophyll absorbs energy in the form of light, most of that energy transfers directly to the electrons in the chlorophyll molecule. This raises the electrons to higher energy levels. These high-energy electrons that do the work of photosynthesis pass on to carrier molecules such as NADP+ to form NADPH in the electron transport chain. Thus the light energy is trapped in a chemical form. In this demonstration we remove chlorophyll from the chloroplasts of plant cells and place it in a solution of ethanol. When we excite the electrons of the chlorophyll molecules with the black light (ultraviolet light), in the absence of the electron transport chain the electrons release their energy in the form of red light as they return to their ground state.

3. Things to consider…
You can use chlorophyll fluorescence to measure photosynthesis.
You can also use chlorophyll fluorescence to measure plant stress, since these stresses typically impact the plant’s metabolism and create an imbalance between the amount of energy absorbed by the chlorophyll and the use of this energy in photosynthesis.

4. Are there other pigments in plants besides chlorophyll?
Paper chromatography is a process that uses special filter paper to separate and identify the different substances in a mixture. Chromatography means “to write with color.” The substances in a mixture dissolve in alcohol and move up the paper. Heavier substances move up the paper more slowly, while lighter substances move up the paper more quickly. Because of this, heavy and light substances get separated from one another on the paper.
From:

5. Leaf Pigment Chromatography
Obtain a strip of chromatography paper.
Use a ruler and draw a light PENCIL line 2 cm above the bottom of the paper strip.
Wrap a leaf around a coin with the waxy side of the leaf facing outward. Now rub the leaf along the pencil line on the paper strip until you make a dark green line. DO NOT RUB THE LEAF ABOVE OR BELOW THE LINE!

6. Leaf Pigment Chromatography
Wrap the top of the paper strip around a pencil so that the end of the strip with the green line hangs down. The pencil should be able to sit across the top of the beaker with the bottom of the paper strip just touching the bottom of the beaker. Cut off any excess paper from the TOP of the strip if it is too long. DO NOT CUT THE BOTTOM OF THE STRIP WITH THE GREEN LINE!
Remove the pencil and paper strip from the beaker.

7. Leaf Pigment Chromatography
Carefully add isopropyl alcohol to the beaker until it reaches a depth of 1 cm in the beaker.
Lay the pencil across the top of the beaker with the paper strip extending into the alcohol. MAKE SURE THAT THE LEVEL OF ALCOHOL IS BELOW THE GREEN LINE ON YOUR STRIP!
Observe as the alcohol gets absorbed and travels up the paper. This may take up to 20 minutes. Do not touch your experiment during this time.

8. Leaf Pigment Chromatography Observations
Using colored pencils, draw your results in your research notebook:
Before paper chromatography
After paper chromatography

9. Questions to consider…
Why is paper chromatography an appropriate technique to use to determine if different pigments are present in a leaf?
How does paper chromatography work?
Did the leaf you tested contain different pigments? How do you know?
Why do you think leaves tend to change color in the fall?
Leaves in New England change color in the fall. However leaves in Florida do not. Why do you think this happens?
What are the names of other pigments that plants contain and what color are they? (You might need to Google this one!)

10. What is the role of stomata in leaves?
From:
Plants have special pores called stomata to allow passage of material. The stomata pores are surrounded on both sides by jellybean shaped cells called guard cells. Unlike other plant epidermal cells, the guard cells contain chlorophyll to do photosynthesis. This allows the cells to expand/ contract to open or close the stomata. Guard cells also close when dehydrated. This keeps water in the plant from escaping. The opening or closing of guard cells can be viewed in a microscope by adding different water concentration to the leaf tissue. Most stomata are on the lower epidermis of the leaves on plants (bottom of the leaf). The number of stomata on the epidermal surface can tell you a lot about a plant. Usually, a high concentration of stomata indicates fast growth and wet climate. Lower concentrations of stomata indicate lower rates of photosynthesis and growth or adaptations for dry weather.

11. Leaf Stomata Obtain a spinach leaf.
Paint a thick patch that is about 1 square centimeter of clear nail polish on the under side of the leaf surface.
Allow the nail polish to dry completely.

12. Leaf Stomata
Tape a piece of clear tape to the dried nail polish patch.
Gently peel the nail polish patch from the leaf by pulling on a corner of the tape and “peeling” the fingernail polish off the leaf. This is the leaf impression you will examine.
Place your impression to a clean microscope slide.
Examine the leaf impression under a microscope at high power.

13. Leaf Stomata Observations
Search for areas where there are numerous stomata, and where there are no dirt, thumb prints, damaged areas, or large leaf veins.  Draw the leaf surface with stomata.
Count all the stomata in one microscopic field. Record the number in a data table.
Repeat counts for at least three other distinct microscopic fields. Record all the counts. Determine an average number per microscopic field.
From the average number/400X microscopic field, calculate the stomata per mm2 by multiplying by 8.

14. Questions to consider
Explain, in detail, how guard cells open and close stomata?
2. At what time of day would stomata be closed and why?
3. Why does the lower epidermis have more stomata than the upper epidermis of a leaf?
4. What is transpiration?
5. What two gases move in and out of the leaf stomata?
6. What does a larger number of leaf stomata indicate about the growing climate of that plant?