Question

Experiment 2 – Light Independent Reaction

After the light dependent reaction, the light independent takes place. During this phase, also known as the Calvin Cycle, the energy generated in the light independent phase is used to fixate the carbon atoms from carbon dioxide and glucose is produced.

In this experiment you will use baby spinach leaves to demonstrate the utilization of carbon dioxide during the light independent reaction of photosynthesis. Phenol red is an organic dye that undergoes a color change according to pH . The color of this compound will appear as follows:

Because phenol red can be used to detect changes in pH, it can also be used as an indirect method of detecting changes in the amount of CO2 dissolved in a solution. When CO2 is added to a solution, some of it reacts with water to produce an acid called carbonic acid. In turn, some of the carbonic acid dissociates to increase the [H⁺] in the solution. Therefore, if the amount of CO2 in a solution is increased, the [H⁺] increases (pH is lowered) and the solution becomes more acidic. It also follows that if the amount of CO2 in a solution is lowered (e.g., by removal of CO2 from the solution), the [H⁺] will decrease (pH increases) and the solution becomes more basic. The chemical reactions involved are shown below.

To detect the process of carbon fixation (CO2 reduction), you will use this pH indicator to detect changes in the CO2 level.

Materials

Fresh spinach leaves

2 glass test tubes

2 - 250 ml beakers

Phenol red solution

Permanent marker or wax pencil

Water *

Sunshine or bright light source *

Stopwatch/timer

Metric ruler

Procedure

Add 20 ml of phenol red solution to 2 glass test tubes.

With your straw , blow bubbles over the top of the solution until it turns yellow. The color change occurs when the water in the phenol red solution and the CO2 are combined forming carbonic acid . Make sure that both tubes are the same shade of yellow.

Add a small leaf of baby spinach to tube 2 only

Place the 2 test tubes into sunshine or a bright light at time for 2 hours.

Record the color every ½ hour.

Table 2: Change is Color (phenol red)
	0 time	½ hour	1 hours	1 ½ hours	2 hours
Tube 1
Tube 2

Questions

What is the independent variable in this experiment?

What is the dependent variable in this experiment?

What happened to the color of the solution over time?

What is the process that is occurring to make the phenol red solution change colors?

What is the function of the following components in the process of photosynthesis (this may differ than how this compounds were used in the experiment)

CO₂:

Light:

H₂O:

O₂:

Chloroplasts:

Answer 1

What is the independent variable in this experiment?

In this case the independent variable is the bright light because is a variable that the student choose for the experiment and It does not depends of any variable. In light is constant and there is not any variable in this experiment which could modify it.

What is the dependent variable in this experiment?

The dependent variable is the change of color of the phenol red because It will depend of the independent variable (light). Depending on light, the plant will be active to perform the Calvin cycle, consuming the CO2 (previously added by the student) and changing the color.

What happened to the color of the solution over time?

The most probable situation is that the tube turns red, because the plant is consuming the CO2, decreasing the [H+] and turning the pH more basic.

What is the process that is occurring to make the phenol red solution change colors?

When you add CO2 by blowing bubbles, you are promoting a chemical reaction between CO2 and water, producing carbonic acid. This compound will change the pH into an acid environment which will promote the change of color from red to yellow. On the other hand, when the plant is consuming the CO2, there is less CO2 and the pH will turn basic and you will observe a red coloration.