Thomas Dunk

Rp8
Hypothesis - Rate of reaction is going to decrease.

Dehydrogenase enzyme – An enzyme in the chloroplast involved in NADP accepting electrons

DCPIP – A redox indicator which is blue normally, and colourless when it is oxidised with electrons

Ammonium hydroxide – Is alkaline, so could denature the dehydrogenase enzyme. It can also accept
electrons, so could pick up electrons instead of DCPIP or NADP.
Thomas Dunk

Summarised Steps
1. Prepare tubes
2. Add spinach leaves to isolation medium (controls pH and makes isotonic, so chloroplasts
don’t burst or shrivel).
3. Blend spinach leaves to separate chloroplasts into solution. Cool with ice so slow enzymes
which could damage chloroplasts.
4. Filter the spinach solution through a muslin cloth to remove debris. You are left with the
chloroplasts and isolation medium.
5. Add tubes into ice-cold bath and place in front of light source.
6. Time how long it takes the 2 experimental tubes to turn the same colour as tube C.
7. Once tube X has turned the same colour as tube C, wait 5 minutes for tube Y to change.

Limitations
 Subjective (could use colorimeter)
 Uneven distribution of light (could use more light sources)
 Foil does not block all light (cover top of tube with foil)

Questions

1. Reduced
2. It would contain a pH buffer solution to control the pH. It needs to be an isotonic solution
3. Slows enzyme activity
4. Releasing the chloroplasts from the cells
5. Re-filter
6. A (control 1) – To prove decolourisation will not occur without light
B (control 2) – To prove that chloroplasts are needed
C (standard) – A standard to compare with the others, to determine how ‘decolourisation,
looks.
X (experiment tube 1) – To investigate the time taken for DCPIP to turn colourless
Y (experiment tube 2) – To investigate the time taken for DCPIP to turn colourless with
ammonium hydroxide on dehydrogenase activity
7. To remove anomalies and get averages
8. Colorimeter gives an exact value
Thomas Dunk