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Light, Dark, Does It Really Matter?
Light, Dark, Does It Really Matter?
Examining the Factors of the Light Reaction
Photosynthesis is a most wonderful process in which the chloroplasts of plants make sugar from carbon
dioxide and water. Glucose has more energy than carbon dioxide. This is because the glucose molecule
has hydrogen.
6CO2+6H2O
C6H12O6+6O2
Photosynthesis converts solar energy into the chemical energy needed to make glucose. This chemical
energy is temporarily stored in the bonds of ATP and NADPH through the events of the light reactions.
18ATP+12NADPH
6CO2+6H2O
18ADP+18P+12NADP
C6H12O6+12O2
In order for this reaction to continuously occur, the plant cell must regenerate both the ATP (put the
phosphate back on ADP) and NADPH (put hydrogen on NADP). This regeneration also requires
energy, but this energy comes from light and chlorophyll molecules found in the chloroplasts.
18ATP+12NADPH
6CO2+6H 2O
18ADP+18P+12NADP
C 6H12O6+12O 2
Since photons of light are required, this is called the light reaction. The purpose of the light reactions is
to make the chemical energy needed to make glucose in the Calvin cycle. The light reaction occurs in
the thylakoid of the chloroplast.
PURPOSE
The purpose of this lab is to investigate the factors that affect the light reaction of photosynthesis. These
factors include investigation of the importance of light and the importance the structure of the
chloroplasts. The light reaction is also simulated with a model.
To investigate this reaction, a dye DPIP is used. DPIP is blue but will turn clear if it is mixed with
chloroplasts and the light reaction is allowed to occur. This color change allows us to determine
whether or not the light reaction is indeed occurring.
Laying the Foundation in Biology
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Light, Dark, Does It Really Matter?
MATERIALS
4 test tubes 13 x 100 mm
phosphate buffer
5 cc syringe
aluminum foil
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test tube rack
distilled water
Beral disposable pipet
Safety Alert
CAUTION: DPIP and chlorophyll may stain skin or clothing.
# 1 Control
# 2 Light
Test Tube
Chloroplast
# 3 Dark
Test Tube
Chloroplast
# 4 Light
Test Tube
Chlorophyll
Phosphate Buffer
1 cc
1 cc
1 cc
1 cc
Distilled Water
3 cc
3 cc
3 cc
3 cc
DPIP
1 cc
1 cc
1 cc
1 cc
PROCEDURE
1. Formulate a hypothesis that will test the factors that affect photosynthesis. Record your hypothesis
in the space provided on the student answer page.
2. Obtain the materials needed above. Label the test tubes 1-4.
3. Put the solutions into each of the test tubes as indicated in the chart above. Compare the colors and
record the beginning colors on the student answer sheet.
4. Wrap test tube #3 it in foil and make a foil cap, so that no light is allowed into the tube.
5. In test tubes #2 and #3, add three drop of chloroplasts. Mix by swirling the test tubes. Observe the
color and compare it to test tube #1. Return the #3 test tube to the aluminum foil.
6. In test tube #4 add three drop of CHLOROPHYLL not chloroplasts. Swirl the test tubes. Observe
the color and compare it to test tube #1.
7. Every five minutes observe the color of tests tubes and compare them to test tube #1 for fifteen
minutes. Record the final color of each test tube on the student answer sheet.
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Light, Dark, Does It Really Matter?
The Light Reaction of Photosynthesis
Read about the steps of the light reaction in following text. On your student answer sheet is a diagram
of the light reaction. As you read about the light reaction, cut out the missing and place them in the
proper sequence. The thylakoid membrane has built into it collections of pigment molecules, which
include chlorophylls and carotenoids. This collection is called a photosystem. When a photon of light is
absorbed by one of the pigment molecules, electrons are excited and elevated to a higher energy level.
The energy is passed from pigment to pigment until it reaches an “antenna” molecule.
1. The electron transport chain captures excited electrons. The electrons move from one electron
carrier to another. Each time the electron is transferred, it loses a small amount of energy.
2. As the electrons move through the electron transport chain, one electron carrier in particular has
special properties. This carrier has the ability to “pump” hydrogen ions, H+, across the thylakoid
membrane from the outside to the inside of the thylakoid.
3. To finish the trip, the electron must be energized again at another photosystem with photons of light.
4. Once energized, the electron continues it trip down the electron transport chain. The ultimate
electron acceptor at the end of the electron transport chain is NADP. NADP will also combine with
H+ to form NADPH.
5. Once this trip is complete, the system needs replacement electrons. These replacement electrons
come from the splitting of water molecules. This reaction will produce 2 H+ ions, 2e-, and ½ of an
O2 molecule. The oxygen gas leaves the thylakoid, diffusing out the chloroplasts and eventually out
of the leaf.
6. The e- are used by the photosystem as replacement electrons.
7. The H+ ions accumulate, to make the interior of the thylakoid acidic relative to the stroma.
8. This separation of charge is used to phosphorylate ADP and make ATP as 3 H+ ions move through
the CF1 particle to make ATP.
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Laying the Foundation in Biology
Light, Dark, Does It Really Matter?
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Name _____________________________________
Period _____________________________________
Light, Dark, Does It Really Matter?
Examining the Factors of the Light Reaction
HYPOTHESIS
DATA AND OBSERVATIONS
# 1 Control
# 2 Light
Test Tube
Chloroplast
# 3 Dark
Test Tube
Chloroplast
# 4 Light
Test Tube
Chlorophyll
Beginning Color
Final Color
Light (+,-) Reaction
CONCLUSION QUESTIONS
1. Why are ATP and NADPH needed for photosynthesis?
2. Is light needed for photosynthesis? How do you know?
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3. Are intact chloroplasts needed for the light reaction or is chlorophyll by itself enough? How do you
know?
4. Explain how the thylakoid is analogous to a battery.
5. Explain how ATP is produced during the light reaction.
LIGHT REACTION DIAGRAM
Below are missing parts to the light reaction with a diagram of the thylakoid. Supply the missing parts
to the light reaction in the thylakoid.
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Light, Dark, Does It Really Matter?
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