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).

AP Lab 4: Plant Pigments and Photosynthesis Prelab Discussion!

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

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

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

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.

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

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.

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! http://www.stolaf.edu/people/giannini/flashanimat/metabolism/photosynthesis.swf Fig. 7.12a, p. 121

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

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?

Part B: Photosynthesis Note setup!

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!

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

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

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

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

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

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

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