Exercise 8.8 - Photosynthesis by Respirometry

LEVEL III

Table 8.1 Characteristics of isolated chloroplasts

Materials

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:

    Tube
    Side arm
    Main Vessel
    1-2

    2.0 ml Chloroplast buffer
    3-4
    0.5 ml NaHCO_3
    0.5 ml buffer
    1.0 ml Chloroplast Suspension
    5-6
    0.8 ml NaHCO_3
    0.2 ml buffer
    1.0 ml Chloroplast Suspension
    7-8
    0.5 ml NaHCO_3
    0.3 ml Sodium Azide
    0.2 ml buffer
    1.0 ml Chloroplast Suspension
    9-10
    0.5 ml NaHCO_3
    0.3 ml Inhibitor 1
    0.2 ml buffer
    1.0 Chloroplast Suspension
    11-12
    0.5 ml NaHCO_3
    0.3 ml Inhibitor 2
    0.2 ml buffer
    1.0 ml Chloroplast Suspension
    13-14
    0.5 ml NaHCO_3
    0.5 ml buffer
    1.0 ml Boiled Chloroplasts

  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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Cell Biology Laboratory Manual
Dr. William H. Heidcamp, Biology Department, Gustavus Adolphus College,
St. Peter, MN 56082 -- cellab@gac.edu