124 PHOTOSYNTHESIS 



the living cell, at least with respect to the hydrogen accep- 

 tors so far investigated; or (2) the reduction of carbon 

 dioxide requires more than the four equivalents of hydrogen 

 chemically necessary (see Chapter 8). The second interpreta- 

 tion vi^ould be analogous v^ith other biochemical systems 

 but v^rould necessitate some modification of the mechanism 

 suggested by van Niel. 



Summarizing, we see that illuminated chloroplasts cata- 

 lyse the production of oxygen from water in the presence of 

 certain hydrogen acceptors in a reaction which shows many 

 of the characteristics of the oxygen-producing reaction in 

 photosynthesis. So far, however, the reducing properties of 

 illuminated chloroplasts are not of sufficient potential to 

 account for a direct reduction of carbon dioxide by reaction 

 with the minimum number of equivalents of hydrogen. 



COUPLING OF THE CHLOROPLAST REACTION 

 WITH CARBON DIOXIDE FIXATION 



In the preceding chapter, mechanisms of carbon dioxide 

 fixation using reduced TPN were discussed and it was seen 

 that fixation would only occur at moderate concentrations 

 of carbon dioxide if the ratio of reduced to oxidized co- 

 enzyme was maintained very high. Attempts to determine 

 whether DPN or TPN was reduced by illuminated chloro- 

 plasts were at first unsuccessful; this is in agreement with 

 the view that the maximum observed potential of the chloro- 

 plast system is about zero whilst that of the coenzyme is 

 -0*3 volt. If however the concentration of oxygen is main- 

 tained very low then the reduction of TPN can be demon- 

 strated using a carboxylating reaction system such as py- 

 ruvic acid, carbon dioxide, manganese salts, and 'malic 

 enzyme'. In this reaction system malic acid is formed when 

 the chloroplasts are illuminated, since TPN is reduced to 

 at least a limited extent and the reduced TPN is removed 

 sufficiently rapidly by the reaction between pyruvic acid and 

 carbon dioxide. Vishniac and Ochoa (1951) demonstrated 

 this coupled reaction between illuminated chloroplasts and 

 'malic' enzyme obtained from pigeon liver. With 7-5 io~^M. 

 pyruvate, 2-25 io~^M. TPN and with iq-^M. bicarbonate 



