CP11 Guide Lines Biology A Level Tutor
Activity 5.5
Guide Lines for Core practical 11: Investigating Photosynthesis
To investigate the link between the light-dependent and independent reactions
Core practical instructions given by Pearson can be found at this link:
https://qualifications.pearson.com/content/dam/pdf/A%20Level/biology-a/2015/teaching-and-learning-
materials/A_level_Biology_A_Core_Practical_11_-_Investigating_Photosynthesis.pdf
Content:
1. Basic information on photosynthesis and on Hills reaction
2. Planning and Experimental Design
3. Health and Safety
4. Make a plan
5. Perform the experiment
6. Analysis and interpretation of data
7. Conclusion and evaluation
8. Write the report!
9. Sample of Data (Analysis and Explanations)
10. Answers to questions given by Pearson regarding the practical.
1/16
,CP11 Guide Lines Biology A Level Tutor
Point 1: Research the background science
Photosynthesis is a two-stage procedure to uses light energy to produce food
for plants, the energy from light is incorporated into organic molecules, into glucose.
Traditionally, in order to investigate photosynthesis, the production of oxygen and
starch are used. However, in this core practical, the light dependent stage is
investigated. During this stage NADPH is produced, which also links the light
dependent stage to the light independent stage, thus in order to investigate the
factors affecting the light dependent stage, one must investigate the production of
NADPH. However, this is not possible in a school laboratory, thus, DCPIP is used,
which is an electron acceptor as well. Using DCPIP is an easy way to monitor redox
reactions that take place during the light dependent stage. Normally DCPIP has a
blue colour (when oxidised), and when it picks up electrons, it becomes colourless
(reduced). The loss of colour in the DCPIP is due to reducing agent produced by light-
dependent reactions in the extracted chloroplasts.
The light dependent stage occurs on the thylakoid membranes of
chloroplasts. On the thylakoid membranes there are chlorophyll pigments, called
photosystems, which absorb light energy. During the light dependent stage, the light
excites and releases electrons from the chlorophyll pigments, and are then picked up
by the electron carriers and pass along the electron transport chain via redox
reactions. As the electrons pass along the electron transport chain, they lose energy,
which energy is used to pump hydrogen ions into the thylakoid space. This results in
hydrogen ion accumulation, thus, hydrogen ion concentration increases within the
space, this causes they hydrogen ions to flow through the ATP synthase enzyme (also
found on the thylakoid space), as the ions diffuse through the enzyme (down the
concentration gradient) back into the stroma, the energy produced as they pass
through the enzyme is used to produce ATP, by phosphorylation of ADP
(photophosphorylation). Whereas the electrons that flow through the electron
transport chain are picked up NADPH, however during this experiment the electron
are picked up by DCPIP. During this stage, apart from photophosphorylation and
reduction of NADP, photolysis occurs as well, that is water breaks down, releasing
2/16
, CP11 Guide Lines Biology A Level Tutor
hydrogen ions, oxygen and electrons, which electrons are used to replace the
electrons lost from the chlorophyll pigment.
AS already discussed, this experiment investigates the light dependent reactions
of photosynthesis, and it is called Hill reaction (from the scientist Robert Hill who
developed the procedure). This procedure depends on electrons released during the
light dependent stage and being picked up by DCPIP, the electron acceptor, which
has a blue colour when oxidized and it is colourless when it picks up the electrons
(reduced). Thus, the change is colour is used to indicate if the DCPIP has accepted an
electron or not, to monitor redox reactions. This reaction can only occur if the
thylakoid membranes are illuminated. In fact, the DCPIP takes the place of NADP,
allowing photolysis to continue even if the supply of NADP has stopped because the
DCPIP will continue to accept the electrons from the ETC (electron transport chain).
DCPIP Blue (oxidized form) DCPIP Colorless (reduced form)
During the experiment, in the process of isolating the chloroplasts, they might be
damaged, thus, the tubes will contain a mixture of intact chloroplasts and thylakoid
membranes without the surrounding stroma and outer cell membranes. This is
actually an advantage because it means that the DCPIP can access the thylakoid
membranes directly, without having to pass through the outer membranes, to
accept electrons directly from the electron transport chain.
This investigation depends on working quickly and keeping everything on ice;
thus, the pestle and mortar are kept in the freezer before using them in order to
prevent protein denaturation. Even though the light dependent stage does not have
enzymes, a higher temperature during the experiment will denature the electron
transport chain proteins thus affect the results. So, using ice cold buffer and
solutions, along with the frozen pestle and mortar are necessary to prevent damage
to the chloroplasts during this procedure.
3/16
Activity 5.5
Guide Lines for Core practical 11: Investigating Photosynthesis
To investigate the link between the light-dependent and independent reactions
Core practical instructions given by Pearson can be found at this link:
https://qualifications.pearson.com/content/dam/pdf/A%20Level/biology-a/2015/teaching-and-learning-
materials/A_level_Biology_A_Core_Practical_11_-_Investigating_Photosynthesis.pdf
Content:
1. Basic information on photosynthesis and on Hills reaction
2. Planning and Experimental Design
3. Health and Safety
4. Make a plan
5. Perform the experiment
6. Analysis and interpretation of data
7. Conclusion and evaluation
8. Write the report!
9. Sample of Data (Analysis and Explanations)
10. Answers to questions given by Pearson regarding the practical.
1/16
,CP11 Guide Lines Biology A Level Tutor
Point 1: Research the background science
Photosynthesis is a two-stage procedure to uses light energy to produce food
for plants, the energy from light is incorporated into organic molecules, into glucose.
Traditionally, in order to investigate photosynthesis, the production of oxygen and
starch are used. However, in this core practical, the light dependent stage is
investigated. During this stage NADPH is produced, which also links the light
dependent stage to the light independent stage, thus in order to investigate the
factors affecting the light dependent stage, one must investigate the production of
NADPH. However, this is not possible in a school laboratory, thus, DCPIP is used,
which is an electron acceptor as well. Using DCPIP is an easy way to monitor redox
reactions that take place during the light dependent stage. Normally DCPIP has a
blue colour (when oxidised), and when it picks up electrons, it becomes colourless
(reduced). The loss of colour in the DCPIP is due to reducing agent produced by light-
dependent reactions in the extracted chloroplasts.
The light dependent stage occurs on the thylakoid membranes of
chloroplasts. On the thylakoid membranes there are chlorophyll pigments, called
photosystems, which absorb light energy. During the light dependent stage, the light
excites and releases electrons from the chlorophyll pigments, and are then picked up
by the electron carriers and pass along the electron transport chain via redox
reactions. As the electrons pass along the electron transport chain, they lose energy,
which energy is used to pump hydrogen ions into the thylakoid space. This results in
hydrogen ion accumulation, thus, hydrogen ion concentration increases within the
space, this causes they hydrogen ions to flow through the ATP synthase enzyme (also
found on the thylakoid space), as the ions diffuse through the enzyme (down the
concentration gradient) back into the stroma, the energy produced as they pass
through the enzyme is used to produce ATP, by phosphorylation of ADP
(photophosphorylation). Whereas the electrons that flow through the electron
transport chain are picked up NADPH, however during this experiment the electron
are picked up by DCPIP. During this stage, apart from photophosphorylation and
reduction of NADP, photolysis occurs as well, that is water breaks down, releasing
2/16
, CP11 Guide Lines Biology A Level Tutor
hydrogen ions, oxygen and electrons, which electrons are used to replace the
electrons lost from the chlorophyll pigment.
AS already discussed, this experiment investigates the light dependent reactions
of photosynthesis, and it is called Hill reaction (from the scientist Robert Hill who
developed the procedure). This procedure depends on electrons released during the
light dependent stage and being picked up by DCPIP, the electron acceptor, which
has a blue colour when oxidized and it is colourless when it picks up the electrons
(reduced). Thus, the change is colour is used to indicate if the DCPIP has accepted an
electron or not, to monitor redox reactions. This reaction can only occur if the
thylakoid membranes are illuminated. In fact, the DCPIP takes the place of NADP,
allowing photolysis to continue even if the supply of NADP has stopped because the
DCPIP will continue to accept the electrons from the ETC (electron transport chain).
DCPIP Blue (oxidized form) DCPIP Colorless (reduced form)
During the experiment, in the process of isolating the chloroplasts, they might be
damaged, thus, the tubes will contain a mixture of intact chloroplasts and thylakoid
membranes without the surrounding stroma and outer cell membranes. This is
actually an advantage because it means that the DCPIP can access the thylakoid
membranes directly, without having to pass through the outer membranes, to
accept electrons directly from the electron transport chain.
This investigation depends on working quickly and keeping everything on ice;
thus, the pestle and mortar are kept in the freezer before using them in order to
prevent protein denaturation. Even though the light dependent stage does not have
enzymes, a higher temperature during the experiment will denature the electron
transport chain proteins thus affect the results. So, using ice cold buffer and
solutions, along with the frozen pestle and mortar are necessary to prevent damage
to the chloroplasts during this procedure.
3/16
