1. Wednesday, January 14, 2015
Agenda: Photosynthesis/ Chromatography lab.
Learning Objective: Students will plan and carry out investigations of factors affecting photosynthetic rates.
HW: Finish lab (due next Wed.) and bring lab materials for Friday (optional).

2. AP Lab 4: Plant Pigments and Photosynthesis
Prelab Discussion!

3. AP Lab 4 Part A: Pigment Chromatography
What is chromatography?
A separation technique that separates molecules by size and solubility

4. AP Lab 4 Part A: Pigment Chromatography
What will our setup look like?
We will substitute a screw-top jar for a graduated cylinder!

5. Part A: Chromatography
Remember to make a dark pigment line!
Allow pigment to dry between applications.
Do not allow solvent above pigment line at start!
Be sure to mark solvent line at end for calculations!
solvent

6. AP Lab 4 Part A: Pigment Chromatography
What is Rf and how do you calculate it?
It is the relationship between the distance moved by a pigment and the distance moved by the solvent.

7. AP Lab 4 Part B: Photosynthesis
How could one measure photosynthetic rates?
Amount of O2 produced
Amount of glucose produced
Amount of ATP produced
Amount of NADPH created by the reduction reaction:
NADP+ + H+  NADPH

8. AP Lab 4 Part B: Photosynthesis
We will focus on the NADPH production.
We know that NADPH is the final electron and H receptor in the light dependent reactions.
We will be substituting the compound DPIP (a blue dye to be the final electron acceptor)
Therefore as photosynthesis progresses, the blue dye will become increasingly clear.

9. THYLAKOID COMPARTMENT first electron transport system
Light Dependent Reactions:
sunlight
photolysis
THYLAKOID COMPARTMENT
second electron transport system
H2O
NADP+
NADPH e–
ATP
ATP SYNTHASE
PHOTOSYSTEM I
PHOTOSYSTEM II
ADP + Pi
first electron transport system
STROMA
DPIP!
Fig. 7.12a, p. 121

10. Glucose made with ATP energy Sun = Energy Source CO2 reactant
Chlorophyll absorbs solar energy
ATP
NADPH drops off H
NADPH H H C C O
NADP+
H2O = reactant O O H O H
O2 waste
Light Dependent Rxns: Thylakoid Membrane
Light Independent Rxns: Stroma

11. AP Lab 4 Part B: Photosynthesis
We will measure the % absorbance with a spectrophotometer
What would a high % absorbance mean reagrding photosynthetic rate?
A low % absorbance?

12. Part B: Photosynthesis
Note setup!

13. Part B: Photosynthesis
Troubleshooting Tips:
Calibrate the spec before each reading
Remember to mix each cuvette prior to taking an absorbance reading!
Remember to handle cuvettes by the curved sides and load properly!

14. While you are waiting between readings
Answer analysis questions
Design a lab with your group members

15. Harmonizer Makes sure everyone understands the instructions
Asks Ms. Pham when instructions are unclear
Makes sure everyone in the group is participating and contributing
Keeps time

16. Recorder Reads the spectrophotometer
Records data and shares with the group
Writes data on board at end of class

17. Materials Manager (2 students)
Physically prepares the solutions and handles the pipettes
Prepares the cuvettes
Handles the spectrophotometer

18. Write names Harmonizer: Recorder: Materials Manager (2):
ALL students help clean and makes sure the lab looks as it did when it started.

19. Each group ONLY prepare 2 cuvettes:1
5 + 2 2
3 + 4 3
Blank + 2 4 5
Blank + 5 6
2 + 3 7 8
4 + 3

20. Transmittance (%) Cuvette 5 10 15 2 Unboiled/Dark 3 Unboiled/ Light5 10 15
2 Unboiled/Dark
3 Unboiled/ Light
4 Boiled/ Light
5 No Chloroplasts/Light