Biology 1406

Photosynthesis Lab
Objectives:
 Observe oxygen produced by photosynthesis
 Determine pigments in plants that allow for photosynthesis

Terms:
 Chromatography
 Photosynthesis
 Autotroph

Materials Needed:
Exercise 1 Exercise 2

 4 large beakers or clear bowls/cups

 1 Tall beaker or glass
 4 small beakers or clear bowls/cups
 Pencil or pen
 Light source (lamp or window)
 Coin (quarter, nickel, dime or penny)
 Fresh spinach (or other soft) leaves
 Paper towel
 Baking soda
 Coffee filter
 Paper towel
 Fresh spinach (or other soft, dark green) leaf
 Rubbing alcohol
 Tape (or something else to secure filter)
 Plastic wrap (not necessary but helpful)

Introduction
Photosynthesis is the process by which autotrophic organisms make their own food in the form of glucose
using light energy from the sun and carbon dioxide. Autotrophic organisms include; plants, some bacteria
and some protists (eukaryotic organisms such as algae). Photosynthesis takes 6 carbon dioxides, 6 water
molecules and energy from sunlight to form a single glucose and 6 oxygen molecules (see figure 1).

Autotrophic organisms will then utilize the glucose produced for cellular respiration to get ATP from it. The
only difference in cellular respiration in autotrophs versus heterotrophs (organisms who consume other
plants and animals for energy) is where the glucose for cellular respiration comes from. Autotrophs get their
glucose from photosynthesis while heterotrophs must eat another organism to obtain a glucose for cellular
respiration.
 Figure 1: Photosynthesis equation. Showing the reactions uses 6 carbon dioxides, 6 water molecules and energy from the sun to
form glucose and 6 oxygen molecules.
(Image source: Professor Mello)

Exercise 1: Photosynthesis
First we will look at photosynthesis in plants. In this experiment you will compare the amount of photosyn-
thesis produced by spinach leaves under varying conditions such as water with baking soda added. Baking
soda when placed in water will release carbon dioxide molecules into the water.

Procedure 1—
1. Obtain 4 large beakers or clear bowls/cups. And 4 small beakers or clear cups/bowls. The key here is that
you want the smaller beakers/bowls to fit inside the larger ones and you need enough room that you can
turn the small beaker upside down while its inside the larger beaker. Meaning you want to be able to put
the small beaker inside the larger facing up. Then turn the small beaker upside down without pulling it
out of the larger beaker to do so.
2. In one of the large beakers fill it about 3/4 of the way full with regular water. You want the water lever in
the large beaker to be taller than the smaller beaker.
3. Fill the remaining 3 beakers with the same amount of water that you filled the first.
4. Label one beaker control light, and one control dark, set them both aside. These two beakers will just
have regular water in them.
5. To the next beaker add a tablespoon of baking soda and stir to dissolve.
6. Continue to add baking soda to the beaker one table spoon at a time, stirring in-between tablespoons,
until the water becomes slightly cloudy.
7. Add the same number of tablespoons of baking soda to the remaining beaker.
8. Label one beaker light, and one dark.
9. Take one of the control beakers, place it on a paper towel, towel, or in a sink, to prevent a mess
occurring for the next couple of steps as water may splash out.
10. Place a small beaker in one of the control beakers face up
11. Place 3 spinach leaves inside the submerged small beaker.
12. Carefully turn the smaller beaker upside down while keeping it submerged in water. You want to prevent
any air from getting inside the small beaker
13. If your spinach leaves float out carefully stuff them back up underneath the small beaker without getting
 any air bubbles in the smaller beaker. If air bubbles occur turn the small beaker right side up, remove the
air bubbles and try again. See figure 2 for an example of what the completed setup should look like.
14. Repeat this process for the remaining 3 beakers.
15. Place the control dark and dark beakers in a cabinet or under a cover to keep them in the dark.
16. Place the control light and light beakers in a window sill or by a bright light.
17. Observe the small beakers for bubbles once every 5 minutes for 20 minutes.
18. Rank the amount of bubbles on a scale of 0-5. 0 = no bubbles, 5 = lots of bubbles.
19. Record the results in the student handout portion of the lab.

Figure 2. Image A shows the setup using a measuring cup and a small bowl instead of beakers. Image B shows the setup using two
beakers. Image c shows that there are no bubbles in the smaller beaker/bowl with initial setup. Image D shows what the bubbles
will look like as they form over time.
(Image source: Professor Mello)
Exercise 2: Chromatography
Photosynthetic organisms capture the energy from sunlight utilizing pigments. Plant leaves tend to have
multiple pigement types in them so they can absorb multiple wavelengths of light. These pigments are what
give plant leaves their colors. Most plants have pigments that absorb all wavelengths except for green light.
Green light is instead reflected back or transmitted which is why most plants appear green. All things that we
see as color reflect back the wavelength of light you see and absorb or transmit (allow to pass through) all
other colors.

So if a shirt is red its either absorbing or transmitting all wavelengths of light except red. The red wavelength
is being reflected back and that is what your eyes see. Black and white colors are the exception to this. A
black tshirt absorbs all wavelengths of light so black is actually the absence of color wavelengths being
reflected into your eye. While a white shirt reflects all wavelengths of light, your brain process all
wavelengths being reflected together as white. This is why black shirts can be much hotter than other color
of shirts on a sunny day, the black shirt absorbs all of the light energy, a white shirt is generally the coolest
color of shirt to wear on a hot day as it reflects of all the sun’s energy.

In this procedure we will separate the pigments in spinach leaves so you can see how many pigments the
plant uses to perform photosynthesis.

Procedure:
1. Obtain a tall cup or beaker.

2. Obtain a coffee filter or piece of chromatography paper. Cut it into long rectangular strip that is an inch
wide and long enough to reach from the top of your beaker to the bottom (see figure 3 for an example
setup). Try to touch the strip you cut by the edges, oils on your hands can be absorbed by the paper and
mess up the experiment.

3. Measure half an inch up from the bottom of the strip and place a spinach leaf on it.

4. Use a coin to rub the spinach leave over the line multiple times until the green from the leaf has been
rubbed into the paper, see figure 3 for an example.

5. Tape the top of the strip (part furtherest away from your green spinach line) to a pencil or pen and hang
it inside the beaker (see figure 3 for an example of setup).

6. Carefully pour rubbing alcohol into the bottom of the beaker you want enough to get the bottom of the
strip wet but not enough to cover the green spinach line.

7. Place a plastic wrap over the top of the beaker to help prevent evaporation.

8. Allow the experiment to run until the alcohol is about an inch from the top of the strip.
9. Remove the strip from the container and observe the different pigment lines.

10. Identify which pigments you see using figure 3 as a reference.

Figure 3. Image A shows the chromatography strip with a quarter and spinach leaf ready to be used. Image B shows the spinach
leaf about an inch up on the paper and the quarter is about to be used to rub the spinach. Image c shows the quarter rubbing a line
along the spinach leaf to transfer pigment. Image D shows the green leaf pigment that has been transferred to the paper
successfully. Image E shows the complete final setup with the chromatography paper attached to a pen and hanging in a beaker.
Notice the bottom of the paper touches the bottom of the beaker.
(Image source: Professor Mello)
 Name:

Biology 1406
Lab 1: Student Handout
Exercise 1 Photosynthesis
Rank the amount of bubbles on a scale of 0-5. 0 = no bubbles, 5 = lots of bubbles. Record your observations
in the table below.

Treatment 0 min 5 min 10 min 15 min 20 min

Control Dark
Dark
Control light
Light

1. Which treatment(s) produced the most bubbles? Why?

2. Which treatment(s) produced the least bubbles? Why?

3. What was the purpose of the baking soda? Why was it necessary for photosynthesis?
Exercise 2 Chromatography
Draw your completed chromatography strip below and label the pigments. You may use colored pencils if
available or draw it in black and white labeling the color of each line as well as the pigment.

4. Why do plants utilize multiple pigments for photosynthesis? What is the benefit or drawback of using
multiple?

5. What color does each of the pigments you saw reflect back? What color(s) do they each absorb?