1636 CHEMICAL PATH OF CARBON DIOXIDE REDUCTION CHAP. 36 



part is "respiratory" in origin, and that the latter part is particularly large in starved 

 Chlorclla cells. 



Subsequently, CaKdn and co-workers achieved substantial progress by 

 distinguishing clearly between three types of C*02 uptake; in the dark 

 without preillumination, in the dark immediately after preillumination, 

 and in light. (A further distinction is indicated by the work of Gaffron, 

 Fager et al. — between preillumination in the absence of CO2, and that in 

 the presence of CO2.) 



Most revealing are the results obtained by first illuminating cells in 

 ordinary carbon dioxide with strong light long enough to establish a steady 

 state of photosynthesis, then rapidly supplying them with tagged C*02, 

 and killing them as quickly as possible after a very brief continuation of the 

 light exposure. In some experiments of Calvin and co-workers, photo- 

 synthesis in tagged C*02 lasted only 0.4 second. 



The results obtained at Berkeley in this study — which is being con- 

 tinued — have been presented, between 1948 and 1954, in about twenty 

 papers under the general title "Path of Carbon Dioxide in Photosynthesis," 

 and in several reviews, e. g., by Benson and Calvin (1948, 1950) and Calvin 

 (1952^-2), cf. also the reviews by Gaffron and Fager (1951), Brown and 

 Frenkel (1953), and Holzer (1954). 



Brown and Frenkel paid particular attention to the question of whether all the 

 Berkeley results were obtained in a steady state of photosynthesis, as was assumed in 

 their interpretation. A source of uncertainty in this respect was the flushing of the 

 vessel with air prior to the introduction of the tracer. The criterion of the steady state 

 is the linearity of the C* uptake with time, which was observed in many, but not in all, 

 of the Berkeley experiments. 



2. C*02 Fixation in Darkness with and without Preillumination. 



A Surviving Reductant? 



Benson and Calvin (1947) began by resuming Ruben and Kamen's 

 study of the C*02 uptake in darkness. They used two Chlorella suspen- 

 sions; suspension I was kept in the dark for eight hours in 4% carbon di- 

 oxide, while suspension II was strongly illuminated for one hour in CO2- 

 free nitrogen. Both were then exposed to C*02 in the dark for 5 min. and 

 the cells killed by acid (HCl -f CH3COOH). The purpose was to see 

 whether preillumination of cells in the absence of carbon dioxide creates 

 in them a chemical agent ("reducing power") able to cause subsequent up- 

 take (which Calvin and Benson presumed must mean reduction), of carbon 

 dioxide in the dark. The C* uptake was found to be five times greater in 

 the preilluminated algae. The chemical distribution of radiocarbon also 

 was different in the two samples: as much as 70% of total activity in 

 sample I was found in succinic acid (about 85% of it in the two carboxyl 



