82 PROBLEMS IN PHOTOSYNTHESIS 



The addition of pentavalent vanadium has an immediate influence on the 

 photosynthetic quotient. It seems that the reduction of CO2 is primarily 

 affected. The differences found in the quantum requirement are impressive. 



§ 33 The Hyposulfite Method 



Warburg and Krippahl (51) worked out a manometric method for the 

 determination of Oo in the presence of COo. They used sodium hyposulfite, 

 Na2S204, which, in aqueous solution, absorbs O2 



Na2S904 + H2O + O2 -^ NaHS04 + NaHSOs 



The side-arm of the manometer vessel contains dry sodium hyposulfite and 

 dry calcium chloride to protect against water vapor. When a liquid, i.e., 

 H2O2 and catalase, or a Chlorella suspension, produces Oo in the vessel, the 

 contents of the side-arm will absorb all the O2 developed when tipped into 

 the vessel. The application of this method in photosynthesis is, according 

 to Warburg and Schroder (63), as follows : two manometer vessels of the same 

 size each contain 3.5 ^ul cells suspended in 7 ml of the usual culture medium 

 (pH 3.8). The gas phase contains 10 vol % COo, 2.5 vol % Oo and argon. 

 The temperature is 20 ° C. The side-arms each contain 20 mg NaoSo04 

 and 20 mg CaClo. After tipping the salts into one vessel at time and into 

 the other vessel after 60 min illumination, two negative pressures //q and h 

 are recorded, h being greater than /zq, owing to the production of O2 during 

 illumination. The vessels are shaken rapidly so that after 10 min constant 

 values of /?o and h can be noted. We then have 



^o, = {ho — h)Ko^ 



To check the equipment, the determination of Vq, without cells has to give 

 the value 0. If the absorbed light intensity is known, the quantum require- 

 ment can be calculated. 



The hyposulfite method has some disadvantages. When the salts are tipped 

 into the vessel some COo is developed, due to the presence of traces of bicar- 

 bonate in the salts or to the production of sodium sulfite driving out CO2 

 from the cells. Corrections are therefore necessary when dense cell suspen- 

 sions are used. Thus, the hyposulfite method is not of the same high scientific 

 standing as the two-vessel method, which, because of its simplicity, reliability 

 and precision, must be considered to be the standard method of biochemical 

 manometry. 



Numerical example: 



W^e have for both vessels 



IV = 7.0 ml Vo = 26.212 ml T = 293° ^o^ = 2.47 mm^ 



Cell suspension: 0.5 /xl cells/ml 



Measured light: green. Incident intensity 24.8 /xl quanta/min of which 3.3% is 



absorbed. 

 Blue-green light: 4700 A. Incident intensity 1.0 jxl quanta/min of which 19% is 



absorbed 



