PHOTOCHEMICAL NITRATE REDUCTION 251 



demonstrate a really significant reduction of nitrate with algae in the 

 absence of CO2 have been unsuccessful (10-12). It is only when glu- 

 cose has been added in the light that after several hours a strong 

 evolution of oxygen due to nitrate reduction occurs (12). Also under 

 the extreme conditions of the "nitrate mixture" (O.l M NaNOj + 

 0.01 M HNO3), a two- to threefold increase in reduction in the light 

 without CO2 over the rate of the dark reaction has been observed (2). 

 The small evolution of oxygen in the presence of nitrate, reported by 

 Kok (11), can hardly be regarded as convincing proof of the existence 

 of a photochemical nitrate reduction. All these observations point 

 to an indirect role of light via the formation of photosynthetic car- 

 bon compounds rather than to the existence of a direct photochemical 

 reduction of nitrate. 



Our earlier observation that light very much accelerates the re- 

 duction of nitrite, the first intermediate of nitrate reduction in algae 

 (13,14), formed the basis of our new approach to this problem. 

 The nitrite accumulated by the green alga, Ankistrodesmus hraunii, 

 in an acid culture medium (pH<4) in the dark rapidly disappeared 

 upon illumination (13). In these experiments, performed in the 

 presence of CO2, glucose was not an effective substitute for light. 

 A similar influence of light on the reduction of nitrite by diatoms has 

 been observed by Harvey (15). 



The present experiments compare the rate of nitrate reduction 

 to that of nitrite reduction in the light in the absence of CO2 (Fig. 

 1). Upon addition of nitrate to a suspension of Ankistrodesmus 

 hraunii in N-free culture medium, only a very small amount of 

 oxygen is evolved. This result agrees very well with the previous ob- 

 servations of other authors (10-12). The rate of nitrate reduction 

 under these conditions is only about 10% of that observed in the 

 hght in the presence of CO2 or of glucose (12). On the other hand, an 

 addition of nitrite leads to a strong evolution of oxygen {cf. 16). 

 For every mole of nitrite reduced, 1.5 moles of oxygen are evolved. 

 In contrast to the reduction of nitrite in the dark, the rate of this re- 

 action in the light considerably increases with increasing hydrogen 

 ion concentration within the physiological range. The rate of oxygen 

 evolution with nitrite is comparable to that in the quinone Hill 

 reaction in whole Ankistrodesmus cells. 



Furthermore, we have studied the influence of low temperatures 

 on the reduction of nitrite under different conditions in the light and 



