Lab practical #2 review

Photosynthesis: Chlorophyll Extraction

1. Photosynthesis: the conversion of light energy into useful biological energy (ATP)

and reducing power that drives the conversion of Carbon dioxide (CO2) and water

into sugar.

2. Absorption of visible light photons by chlorophyll (pigments makes plants green).

3. The energy associated with a photon is inversely related to its wavelength.

(wavelength+, energy-.)

4. The photon energy absorbed by chlorophyll permits the removal of electrons from

water (producing water and protons.)

5. The movement of electrons and protons drive the generation of ATP and reduce CO2.

6. The operating principle:

- (1) Light from the source enters the monochromator

- (2) Split into its various wavelengths.

- (4) Light passes through the test tube, which may contain a colored solution.

- Light is not absorbed by solution falls on a detector, which convert light signal into an

electrical one. The electrical signal is then amplified and its magnitude is displayed to

you by way of a needle on the readout scale.

7. Separation of chlorophyll from carotenoids.

- Boiled spinach solution

- lower layer: ethanol layer (carotenoids.) [use 95% ethanol as the blank]

- upper layer

- Take the upper layer of the first step, and add 92% methanol

- lower layer: methanol layer (chlorophyll b) [use the methanol as the blank]

- upper layer: ether layer (chlorophyll a) [use the petroleum ether as the blank]
photosynthesis: Oxidation of water

1. The chlorophyll-based photosynthetic machinery of chloroplasts is able to oxidize

water. As a result of that, oxygen gas is produced, as well as electrons and protons.

2. Later in photosynthesis the energy associated with the electrons and protons is used to

help make ATP. And the combination of electrons with the protons from water are

used to reduce CO2.

3. The goal of this lab is to measure the rate at which water is oxidized during

photosynthesis when we vary the amount of chloroplasts participating in the reaction.

4. The independent variable: the amount of chloroplasts.

5. The dependent variable: the rate of water oxidation, which is measure as the rate of

the color change of DPIP from blue to clear.

6. The operating principle:

- The redox reaction is the loss of electrons

- The oxidation is the gain of electrons.

- The water oxidation accompany with the reduction of DPIP

- When DPIP is reduced, the water accept electrons, and it is oxidation, and the color of
 DPIP change from blue to clear.

7. The use of buffer: stabilize pH, avoid to have multiple independent variable.

8. The use of bisulfite

- To remove blue color from blank.

- Reduced DPIP. (add electrons to DPIP)

9. Most effective wavelength: blue and red.

10. Two aspects of reduction of DPIP by chloroplast

- The effect of chloroplast concentration on the rate of dye reduction by illuminated

chloroplast

- The effect of light on chloroplast function.

11. Controls groups

- No light, no heat

- No light, heat

- Light, heat.

- (light, no heat)

- draw the conclusion that heat is not an essential factor, light has the major effect.

12. Use the blank to zero the spectrophotometer EACH TIME.

13. The controls are to avoid the DPIP lose color because (1) spontaneously (2) simply
 due to heat (3) due to the combination of light and heat.

Photosynthesis: Production of Oxygen.

1. Oxygen allows many organisms, plants included, to fully extract the potential energy

of glucose.

2. The purpose of this lab is to study the effect of light quantity on the rate of oxygen

production by Chlorella. (Unicellular)

3. The independent variable is the light quantity, which is measured as the PFR (photon

fluence rate), to measure PFR, use a quantum. To vary PER, vary the predetermined

distances from the chlorella to the light source.

4. The dependent variable is the rate of producing oxygen. Can be measure with the aid

of a device which can produce Voltage that is proportional to dissolved oxygen

concentration.

5. Calibration
- First calibration point represents water that contains no dissolved oxygen.

- in this experiments, sulfate take about 100 sec to completely remove dissolved

oxygen from the water sample

- when the oxygen is gone, set the computer read-out to zero

- The second calibration point represents water that us saturated with atmospheric

oxygen.