373 



C.P. Whittingham and P.M. Bishop 



d) ferricyanide with catalytic amounts of TPIP - in which 0.01 

 pmoles of TPIP (2: 3-6-trichlorophenol-indophenol) were added to 

 the basic medium. 



e) TPIP - in which the potassium ferricyanide in the basic med- 

 ium was replaced by 0.1 pmoles of TPIP. 



Oxygen output from the chloroplasts was determined by a flow 

 system, using a modified version (11, c.f. also for details of 

 materials and methods) of that described by Whittingham (12). The 

 method consists essentially of the use of a carrier gas (oxygen 

 free nitrogen) to sweep continually through the chloroplast sus- 

 pension and then over an oxygen measuring galvanic cell. The out- 

 put from this cell was amplified and recorded, and the yield of 

 oxygen from the flash illuminated chloroplasts determined from the 

 area under the recorded curve. 



In experiments using white light, short flashes of 100 psec. 

 were generated by condenser discharge through a xenon discharge 

 tube. Long flashes of 35 msec, duration and of sufficient inten- 

 sity to achieve flash saturation, were produced from Photograph- 

 ers' flash bulbs. For monochromatic flashes, a flash bulb was 

 used in conjunction with an interference filter with transmission 

 either at 697±7 mp or 644- 13 mp. Continuous background light 

 was passed through a small grating monochromator with slit widths 

 adjusted to give a beam of 5 mp band width. Over the wavelength 

 range used there was a slight variation in light intensity from 

 7.4 X lO"'^ watts/cm2 at 645 mp to 6.0 x 10"^ watts/cm^ at 700 mp. 



In the arrangement of an experiment using white light flashes, 

 a single long flash was always fired first, and then as every 

 third illumination type thereafter. The yield plotted against 

 time showed the rate of decay of chloroplast activity. 



When assessing the effect of continuous supplementary light, 

 the chloroplasts were illuminated by a monochromatic beam until 

 steady state photosynthesis was reached, as shown by the plateau 

 on the record chart. A flash was then superimposed on this back- 

 ground; the supplementary light was not turned off until the 

 steady state background level of oxygen production was again 

 reached. After a dark interval, during which the oxygen was 

 flushed out of the system, another monochromatic background wave- 

 length was used for illumination and the process was repeated. 



In the third type of experiment, the monochromatic background 

 light was separated in time from a monochromatic flash. Here, 

 only two different wavelengths were used, a 653 mp background 



