454 



M. Calvin iiud P. M.\ssl^•l: Tlic Path of Carbon iu Photosynthesis 



[F.xi'ER1f.niiaVol.V1II/121 



SUCROSE 



A 6 MALIC ACIO 



O-O GLUTAMIC ACIO 

 D- -Q CITRIC ACIO 



labU 11 



Steady State Concentrations of Some Compounds Involved iu the 



Photosynthesis Cycle. Scenedesmus, experimental conditions as in 



Figure Iu 



TIME (mtoj OF EXPOSURE TO C'*0, 



Fig. 12. — Behavior of radioactivity in specific compounds in the ex- 

 tract from an experiment done under conditions corresponding to 

 those of Figure 10. 



photosynthesis followed by 5 min of darkness. The 

 slope in the total fixation curve in the light corres- 

 ponds to a 13 cm^ COj assimilation (N.T.P.) per hour. 



Figure 11 shows the amount of radioactivity incor- 

 porated into sucrose and three phosphorus compounds 

 for the experiment of Figure 10. 



Figure 12 gives the number of counts in sucrose, 

 glutamic, malic and citric acid, for a different experi- 

 ment of 15 min photosynthesis, followed by 10 min 

 dark, and again 5 min of photosynthesis. 



Although the variation between experiments is quite 

 high, there are some striking features which are com- 

 mon to all : 



(1) The curves of some of the compounds show a 

 marked decrease in slope after 5 min of photo- 

 synthesis. This quite clearly indicates the presence of 

 rapidly turning-over reservoirs in the photosynthesis 

 cycle which are then thoroughly labeled and reach the 

 specific activity of the fed carbon dioxide : Diphosphate 

 area (mainly ribulose diphosphate) ; hexose-monophos- 

 phate area (50% glucose-, 26% sedoheptulose-, some 

 fructose- and mannose-monophosphate) ; phospho- 

 glyceric acid. The leveling off of these curves permits 

 the calculation of the concentration of the reservoirs of 

 those compounds in the photosynthesis cycle, by divid- 

 ing the measured amount of radioactivity per carbon 

 atom by the specific activity of the fed carbon dioxide'. 



Table II gives the steady state concentrations during 

 photosynthesis for some compounds determined by 

 this method. 



(2) The fact that the activity vs. time curves show 

 a definite yet low slope for as long as 30 min can be 

 taken to indicate that the breakdown of carbohydrates 



continues throughout the illumination, i.e. their for- 

 mation from photosynthetic intermediates is revers- 

 ible. Thus, there are two sources of the intermediates: 

 {a) the carbon dioxide fed; the amount of compound 

 formed from this source reaches the maximum specific 

 activity in 5 to 10 min ; (6) the carbohydrate pool of the 

 ceUs; the amount formed from this source is labeled 

 only slowly since the specific activity of the carbo- 

 hydrate pool rises slowly due to the large size of the pool . 

 (3) Other compounds show almost constant rate of 

 labeling during the whole period of photosynthesis; 

 sucrose, malic and glutamic acid. For this and other 

 reasons it is clear that these compounds are not in the 

 photosynthesis cycle, but are formed during the photo- 

 sjmthesis at a constant rate. Their large reservoirs in 

 the cells are labeled only slowly. 



Table 111 



Phosphatase Treatment of H.M.P. Area after au min Photosynthesis 

 and 30 min Photosynthesis Followed by 5 min Dark 



(4) When illumination is interrupted there appears 

 a sudden great increase in the concentration of phos- 

 phoglyceric acid (followed by a slow decrease after 2 

 min), and an almost complete depletion of the diphos- 

 phate area. Analysis of the monophosphate area 

 showed that the amount of sedoheptulose phosphate 

 decreased also (cf. Table III). The concentration of 



' The efficiency factor of the counting of spots on papers has been 

 determined by converting three cut out spots to barium carbonate 

 and measuring their activity in an ionization chamber. It is 19 

 disintegrations per count. 



^ Volume measured as wet packed cells 



- An appreciable fraction of this count is certainly hexosc so that 

 one may estimate a maximum value ot the heptose at around 800 

 counts/min. 



88 



