IB Biology / Year 1
Rise and Shine!
Calculating gross rate of photosynthesis from floating and sinking measurements of vacuum-infiltrated leaf disks
Adapted from Dr. L. Zeman, Department of Biology, UW. Used with permission . Revised 12/08
Introduction
Photosynthesis is the process by which light energy is captured and used to manufacture carbohydrates that are the food for all life on earth. In this lab on photosynthesis, you will learn
• a technique for measuring the rates of photosynthetic oxygen production and respiratory oxygen
consumption in spinach leaf disks,
• the relationship between the rates of gross photosynthesis, respiration, and net photosynthesis.
Materials per team
Consumables: 0.2% bicarbonate solution*, wet paper towels, small square of Parafilm, drinking straw, turgid spinach leaf, sheet of tinfoil or cardboard box
*Recipe for 0.2% bicarb solution: Measure 2.0 g of NaHCO3, add water up to 1 L, and
add one drop of dilute soap solution.
Equipment: 10-mL syringe (1 per pair), strong light source, heat sink (square bottle, skinny aquarium, etc.)
Methods
Preparing the syringe and disks
1. Obtain and label a 10-mL syringe.
2. Remove the plunger and close off the opening (where the needle would go) with a small piece of Parafilm.
3. Pour 10 ml of 0.2% bicarbonate solution into the syringe.
4. Get a large, turgid spinach leaf and put it on a wet paper towel.
5. Use a straw to punch out 10 leaf disks. Avoid cutting through areas with large veins.
6. As each disk is punched, put it into the bicarb solution in the syringe.
Vacuum infiltration (VI) of the disks
7. Insert the plunger a small distance into the syringe and invert the syringe (stand it up on its plunger).
8. Remove the Parafilm and push in the plunger until liquid just oozes out the tiny opening.
9. All the spinach disks you punched are floating due to intercellular air. To remove this air and replace it with water, place a finger tightly on the opening and pull back on the plunger for about 10 sec.
10. Release the plunger to let off the vacuum. Tap the syringe sharply with your fingernail. Invert it quickly a few times. The disks should begin to sink as bicarbonate solution infiltrates their air spaces.
11. If air collected at the needle end, push in the plunger until liquid just oozes out.
12. If all the disks don't sink, repeat the procedure 1-2 more times. If that doesn’t work, seek help. You might have to start over because too much VI can irreparably damage the disks.
In the light
13. Set up a strong light source with a heat sink directly in front of the light source.
14. Place the syringe on the other side of the heat sink, in the light’s path. standing up on its plunger.
15. Note the time. At two-minute intervals for 20 minutes, or until most/all disks float, quickly invert the syringe to mix the contents. Then, stand it on its plunger again. Record the time at which each disk floats in Table 1.
16. Sum up the total number of minutes required for each disk to float. Then, calculate the average time it takes a disk to float (TF). Show your calculations.
In the dark (Note: the longer it took for steps #14-16, the longer it will take the
disks to sink. Be patient.)
17. When several disks are floating (it is not necessary to wait for all 10), remove the syringe from the lighted area and bring it to your table.
18. Lay the syringe on its side and loosely cover it with foil or a box.
19. Note the time again. At two-minute intervals, quickly invert the syringe to mix its contents, stand it up, and record the time at which each disk sinks in Table 2. DO THIS QUICKLY to minimize the disks’ light exposure. Then, lay the syringe on its side again and recover it.
20. Sum up the total number of minutes required for each disk to sink. Then, calculate the average time it takes a disk to sink (TS). Show your calculations.
Clean up
· Bicarbonate solution can be poured down the sink
· Leftover spinach should be composted
· Straw and paper towels should be discarded
· Rinse and return the syringe
Table 1: Float time for spinach disks in the light
LIGHT Starting time: ______Disk / Clock time / Min. for this disk to float
1
2
3
4
5
6
7
8
9
10
Sum (min)
Average time per disk (Tfloat)
Table 2: Sink time for spinach disks in the dark
DARK Starting time: ______Disk / Clock time / Min. for this disk to float
1
2
3
4
5
6
7
8
9
10
Sum (min)
Average time per disk (Tsink)
Post-Lab Data Analysis Questions
1. During which part(s) of the experiment were the spinach leaves performing photosynthesis? Respiration?
2. The relative rate of respiration is inversely proportional to the average time it takes a disk to sink in the dark (1/Tsink). Calculate the rate of respiration for your spinach leaf disks.
3. The relative rate of oxygen production in the light is inversely proportional to the average time it takes a disk to rise in the light (1/Tfloat). Calculate the rate of oxygen production in your spinach leaf disks.
4. The rate of oxygen production in # 3 is equal to the NET rate of photosynthesis. (Your answer to # 1
should tell you why.) From your answers to the previous questions, calculate the GROSS rate of photosynthesis for the spinach leaves.
5. How does the rate of respiration compare to the gross photosynthetic rate? Show your work!
6. When hiking in the forests of Washington state, you will find that many of the conifers are extremely tall and reach up into the clouds while other plants like ferns work much harder to grow on the forest floor. Use your understanding about the ratio of photosynthetic rate to respiration rate to predict how the ratios for conifers and ferns might compare to spinach. Use the back of this page for your answer.