448 RADIATION BIOLOGY 



sugar. This means that carbohydrates are necessary for the reduction, 

 apart from their importance as sources of carbon for the ensuing or 

 simultaneous synthesis of organic nitrogen compounds. If the break- 

 down of sugar is written 



2CH2O + 2H2O -> 2CO2 + 8H, (8-2) 



Eqs. (8-1) and (8-2) can be summed up to 



2CH.2O + HNO3 -^ 2CO2 + NH3 + H2O. (8-3) 



The presence of carbohydrates and the respiratory breakdown of sugars 

 will under all circumstances in darkness become prerequisites for the 

 reduction of nitrate; hence the supply of carbohydrates must always 

 be considered an important factor regulating the rate of the process. 

 Irrespective of the details of the connection between nitrate reduction 

 and respiration, the result according to Eq. (8-3) implies a production of 

 carbon dioxide without a corresponding uptake of oxygen, or a formation 

 of "extra carbon dioxide," according to Warburg and Negelein (1920). 

 Their discovery of an extra carbon dioxide formation in Chlorella in the 

 presence of nitrate showed that the reduction takes place according to 

 the general principles expressed in formula (8-3), and this has been cor- 

 roborated in later investigations (Ruhland and Ullrich, 1929; Tamiya, 

 1932; Yamagata, 1934; Myers and Cramer, 1948). 



Other results have been obtained in the light, but their interpretation 

 requires a consideration of the significance of the dark reaction, Eq. (8-3). 

 The first question is whether this formula actually holds true. 



The weakest point in this discussion is that in no instance have both 

 the consumption of nitrate and the production of extra carbon dioxide 

 been determined in the same short-time experiment. Warburg and 

 Negelein determined the production of ammonia and put it in relation 

 to the production of extra carbon dioxide. Theoretically the ammonia/ 

 carbon dioxide ratio should be 1/2, but this value was observed only 

 occasionally. The carbon dioxide evolution was initially much higher 

 than was expected; this was reasonably explained by assuming an assimi- 

 lation of part of the ammonia formed in the reduction. In the course of 

 time the formation of extra carbon dioxide decreased and that of ammonia 

 increased, but every series of experiments was cut off at the point where a 

 carbon dioxide/ammonia ratio of 2/1 was attained. Yamagata and 

 Tamiya computed in experiments of long duration the extra carbon 

 dioxide production in relation to consumed nitrate and found agreement 

 with the theory. This is inevitable, however, according to Yamagata's 

 deductions, if the average elementary composition of the cell matter does 

 not change through the assimilation of nitrate, so that the consistency 

 does not prove the strict validity of Eq. (8-3). The occurrence of an 

 extra carbon dioxide formation is established beyond doubt, but it is an 



