Objective

The objective of this experiment was to measure the rate of photosynthesis at different concentrations of CO₂. To determine this, leaf discs were placed in different solutions of bicarbonate and their flotation times were measured.

Procedure

To begin, 30 leaf discs were extracted from spinach leaves using hole punchers. It was made sure that only photosynthetic tissue was extracted by avoiding the veins with the hole puncher. At the same time, three solutions were prepared, all of which contained water and dilute soap concentration. The solutions differed in their bicarbonate concentration: solution 1 contained 4% (12.0 grams) bicarbonate, solution 2 contained 2% bicarbonate (6.0 grams), and solution 3 was a control which contained 0% bicarbonate. 10 leaf discs were then placed each of the different solutions, 10 leaf discs per solution. The soap wets the hydrophobic leaf surface and allows solution to infiltrate the leaf. The bicarbonate served as a source of CO₂, a reactant in photosynthesis.

Under most circumstances, reactant concentration is proportional to reaction rate. Therefore, it was hypothesized that, because of this proportional relationship between reactant concentration and reaction rate, the leaf discs in the solutions with greater bicarbonate concentrations would photosynthesize at a greater rate than those with lesser or zero bicarbonate concentration.

When the leaf disks contain O₂, they maintain a density that allows them to float in water. Because a product of photosynthesis is O₂, a way of measuring photosynthesis rate would be to measure the flotation time. As the leaves photosynthesize and internally produce O₂, they float to the surface. However, before the solutions with the 10 leaf discs can be placed under a light source, O₂ must be removed from the leaves.

In order to remove O₂ from the leaves, three syringes without a needle were prepared. Each solution was placed in their respective syringe with the leaf discs. The plunger within the syringe was then pulled back, creating a vacuum that removed O₂ from the mesophyll tissue. The leaf disks in the solution were then placed back into their respective cups and placed at equal distances from a light source. After 5 of the 10 leaves in a solution reached the surface of the solution, the time was recorded (see table 1). This is the 𝐸₅₀ time, which is the time it takes for half of the leaf disks to reach the surface.

Data

Table 1: Rates of Photosynthesis for Each Solution

	Solution 1 (4% bicarbonate)	Solution 2 (2% bicarbonate)	Solution 3 (control/0% bicarbonate)
𝐸50 (minutes)	37.27	38.42	N/A

Discussion

30 leaf disks were extracted from spinach leaves and placed, 10 each, in three solutions of water, dilute soap, and differing bicarbonate concentration. Solution 1 contained 4% bicarbonate, solution 2 contained 2% bicarbonate, and solution 3 was the control which contained no bicarbonate. O₂ was then removed from the leaves using a syringe. Each of the solutions with their ten leaves were placed under a light source and their 𝐸₅₀ was calculated.

The results showed that the solutions containing a source of CO₂ were able to photosynthesize. The control group never rose from the bottom of the cup during the time of the experiment. This is due to the fact that it was not producing O₂ as a result of a lack of CO₂. CO₂ is an essential reactant in the process of photosynthesis as it is necessary in the production of glucose and the fixation step of the Calvin Cycle.

The hypothesis that increasing the concentration of bicarbonate increased the rate of photosynthesis was correct. Solution 1, which contained a 4% bicarbonate concentration, was able to photosynthesize at an 𝐸₅₀ that was approximately 1.15 minutes faster than the solution

containing only 2% bicarbonate solution. It is unknown how much time it would take for the control to produce O₂, but it can be concluded that it would be a much slower time than either bicarbonate solution.

In this experiment, the bicarbonate variable was changed in order to determine the effect of CO₂ concentration on photosynthesis rate. A bicarbonate solution of 4% was prepared along with the 2% bicarbonate solution. The results showed that doubling the bicarbonate concentration increased the rate of photosynthesis by 1.15 minutes. This is consistent with a fundamental rule of kinetics: that increasing reactant concentration increases reaction rate, provided that the reaction is greater than zero order with respect to the reactants. This same law can be applied to enzyme regulated reactions, which photosynthesis is.