RP8 Dehydrogenase activity in chloroplasts Questions

Specification route: RP8
Question bank: 14 questions
Course stage: Year 13 / A-level only

Sample questions

QUESTION 1 · RP8 · LEVEL 3

In the experiment investigating the rate of dehydrogenase activity in chloroplasts, what is the role of the blue dye DCPIP?

It acts as an artificial electron acceptor, picking up electrons from the light-dependent reaction and turning colourless as it is reduced.
It acts as a substitute for carbon dioxide in the light-independent reaction.
It physically binds to the chloroplast envelope to make the organelles visible under a microscope.
It prevents the photolysis of water, allowing the rate of respiration to be measured instead.

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It takes the place of NADP in the electron transfer chain.

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Answer: It acts as an artificial electron acceptor, picking up electrons from the light-dependent reaction and turning colourless as it is reduced.

Required practical 8. Dehydrogenase enzymes in chloroplasts normally reduce NADP. DCPIP acts as an alternative electron acceptor. When reduced, it changes from blue to colourless, providing a measurable endpoint.

QUESTION 2 · RP8 · LEVEL 2

When extracting chloroplasts from leaves, the extraction medium must be cold, isotonic, and buffered. Why must it be isotonic?

To prevent water from moving into or out of the chloroplasts by osmosis, which could cause them to burst or shrivel.
To reduce the kinetic energy of hydrolytic enzymes, preventing them from digesting the chloroplasts.
To maintain a constant pH so that the dehydrogenase enzymes do not denature.
To provide a massive surplus of water to fuel the photolysis stage of photosynthesis.

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Isotonic means having the same water potential.

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Answer: To prevent water from moving into or out of the chloroplasts by osmosis, which could cause them to burst or shrivel.

Required practical 8. An isotonic solution has the same water potential as the organelles. This prevents osmotic lysis or shrinkage. (Cold reduces enzyme activity, buffered maintains pH).

QUESTION 3 · RP8 · LEVEL 2

In the experiment measuring dehydrogenase activity using DCPIP, one test tube containing chloroplasts and DCPIP is completely wrapped in aluminium foil. What is its purpose?

It acts as a control to prove that light energy is strictly required for the photoionisation of chlorophyll and the subsequent reduction of DCPIP.
It prevents the DCPIP from evaporating out of the test tube.
It acts as a control to prove that chloroplasts are the source of the dehydrogenase enzymes.
It simulates the conditions of the light-independent reaction (the Calvin cycle).

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Without light, the light-dependent reaction cannot happen.

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Answer: It acts as a control to prove that light energy is strictly required for the photoionisation of chlorophyll and the subsequent reduction of DCPIP.

Required practical 8. The foil-wrapped tube is a negative control for light. It demonstrates that the colour change (reduction of DCPIP) is driven by the light-dependent reaction, which requires illumination.

QUESTION 4 · RP8 · LEVEL 3

A student forgets to add a buffer to the chloroplast extraction medium. Why might the DCPIP fail to turn colourless during the experiment?

The pH may have fluctuated, denaturing the dehydrogenase enzymes in the chloroplasts and halting the electron transfer chain.
The absence of the buffer causes the chloroplasts to burst via osmosis.
The buffer provides the essential carbon dioxide required for the light-dependent reaction.
Without the buffer, the DCPIP molecule chemically degrades into a permanent blue state.

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Dehydrogenase is an enzyme. Enzymes are highly sensitive to their environment.

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Answer: The pH may have fluctuated, denaturing the dehydrogenase enzymes in the chloroplasts and halting the electron transfer chain.

Required practical 8. A buffer maintains pH. If pH changes significantly, the active sites of dehydrogenase enzymes denature. They can no longer remove electrons/hydrogen, so DCPIP cannot be reduced.

QUESTION 5 · RP8 · LEVEL 2

In the experiment investigating dehydrogenase activity in chloroplasts, how does DCPIP act as an indicator?

It is a redox indicator that takes the place of NADP; it turns from blue to colourless when it accepts electrons released from the photoionisation of chlorophyll.
It acts as a pH indicator, turning colourless as the chloroplasts absorb carbon dioxide and make the solution alkaline.
It binds physically to the oxygen produced by photolysis, turning colourless in the presence of $O_2$.
It acts as a competitive inhibitor of rubisco, stopping the Calvin cycle and turning blue.

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It 'steals' the electrons that are moving down the electron transfer chain in the light-dependent reaction.

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Answer: It is a redox indicator that takes the place of NADP; it turns from blue to colourless when it accepts electrons released from the photoionisation of chlorophyll.

Required practical 8. Dehydrogenase enzymes usually reduce NADP. DCPIP acts as an artificial electron acceptor. When it gains electrons (is reduced) during the light-dependent reaction, it changes from blue to colourless.

QUESTION 6 · RP8 · LEVEL 2

During the preparation of the chloroplast suspension, the leaf blending process is carried out in a cold, isotonic, buffered solution. Why must the solution be ice-cold?

To reduce the kinetic energy and activity of hydrolytic enzymes that could digest and damage the chloroplasts once the cells are broken open.
To prevent the water inside the chloroplasts from boiling under the friction of the blender.
To freeze the chloroplasts solid so they crack open to release the dehydrogenase enzymes.
To slow down the rate of photosynthesis so the experiment does not finish too quickly.

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When you destroy the cell, you release dangerous digestive chemicals from other organelles like lysosomes.

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Answer: To reduce the kinetic energy and activity of hydrolytic enzymes that could digest and damage the chloroplasts once the cells are broken open.

Required practical 8. Homogenisation releases lysosomal enzymes. Keeping the medium ice-cold lowers the kinetic energy, severely reducing enzyme activity and preventing the auto-digestion of the extracted chloroplasts.

QUESTION 7 · RP8 · LEVEL 2

A student sets up a control tube containing chloroplast suspension and DCPIP, but wraps it completely in aluminium foil. What is the expected result and what does it prove?

The DCPIP will remain blue. This proves that light energy is strictly required for the release of electrons to reduce the DCPIP.
The DCPIP will turn colourless very quickly. This proves that dark reactions (Calvin cycle) reduce the DCPIP.
The DCPIP will remain blue. This proves that the foil physically insulates the tube from heat.
The DCPIP will turn red. This proves that respiration takes over when photosynthesis stops.

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Foil blocks out a specific abiotic factor necessary for the first stage of photosynthesis.

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Answer: The DCPIP will remain blue. This proves that light energy is strictly required for the release of electrons to reduce the DCPIP.

Required practical 8. The dark control proves that the reduction of DCPIP is driven specifically by the light-dependent reaction (photoionisation and photolysis), which cannot occur without light energy.

QUESTION 8 · RP8 · LEVEL 3

Another control tube is set up containing DCPIP and an extraction medium that lacks any chloroplasts. What is the purpose of this specific control?

To prove that the reduction (colour change) of DCPIP is caused exclusively by the action of the chloroplasts, and does not occur spontaneously in the light.
To prove that the buffer solution is capable of reducing DCPIP on its own.
To act as a standard to calibrate the colorimeter to zero absorbance.
To demonstrate the maximum possible speed of DCPIP reduction.

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If the blue colour fades in a tube of just water and DCPIP sitting in the light, your entire experiment is flawed.

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Answer: To prove that the reduction (colour change) of DCPIP is caused exclusively by the action of the chloroplasts, and does not occur spontaneously in the light.

Required practical 8. A tube with DCPIP but no chloroplasts proves that light alone does not bleach the dye. It confirms the biological activity of the isolated chloroplasts is responsible for the electron transfer.

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