Photosynthesis by Respirometry

Materials

 

• Gilson respirometer equipped with photoflood lamps
  
• Chloroplast suspension
  
• Chloroplast suspension buffer
• 0.1 M NaHCO
  
• Inhibitor 1 (1 x 10 DCMU)
  
• Inhibitor 2 (5 x 10 DCMU)

Procedure

 

1. In general, use the procedure listed in Exercise 8.7 for mitochondria respiration, except that the reactions are run at 25° C and a bank of photoflood lights must be used to ensure photosynthesis.

    

2. Set up the reactions vessels in the following manner:

    

   <center> <table> <tbody> <tr> <td> Tube</td> <td> Side arm</td> <td> Main Vessel</td> </tr> <tr> <td> 1-2</td> <td>  </td> <td> 2.0 ml Chloroplast buffer</td> </tr> <tr> <td> 3-4</td> <td> 0.5 ml NaHCO</td> <td> 0.5 ml buffer<br /> 1.0 ml Chloroplast Suspension</td> </tr> <tr> <td> 5-6</td> <td> 0.8 ml NaHCO</td> <td> 0.2 ml buffer<br /> 1.0 ml Chloroplast Suspension</td> </tr> <tr> <td> 7-8</td> <td> 0.5 ml NaHCO<br /> 0.3 ml Sodium Azide</td> <td> 0.2 ml buffer<br /> 1.0 ml Chloroplast Suspension</td> </tr> <tr> <td> 9-10</td> <td> 0.5 ml NaHCO<br /> 0.3 ml Inhibitor 1</td> <td> 0.2 ml buffer<br /> 1.0 Chloroplast Suspension</td> </tr> <tr> <td> 11-12</td> <td> 0.5 ml NaHCO<br /> 0.3 ml Inhibitor 2</td> <td> 0.2 ml buffer<br /> 1.0 ml Chloroplast Suspension</td> </tr> <tr> <td> 13-14</td> <td> 0.5 ml NaHCO</td> <td> 0.5 ml buffer<br /> 1.0 ml Boiled Chloroplasts</td> </tr> </tbody> </table> </center>

    

3. Measure the amount of oxygen evolved every 10 minutes until a steady rate is reached. Plot results and compare the effects of the selected inhibitors.

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