1104 THE LIGHT FACTOR. II. QUANTUM YIELD CHAP. 29 



pressure changes is due entirely to different relative volumes of liquid 

 and gas. Even a very small difference in the actual amounts of gas pro- 

 duced (or consumed) in the two vessels, or in the speed with which this gas 

 is transferred into the manometer, can lead to large errors in calculation. 



Several reasons for such variations can be anticipated. Small physio- 

 logical differences may exist between samples taken at different times from 

 the same stock solution or may arise in the course of the experiment. 

 C'Onsiderable discrepancies of light absorption can bo caused by different 

 position of the two vessels in the light beam or by dilforcnces in their 

 shape and wall material. Shaking thins out the liquid layer in the middle 

 of the vessel, and the consequent incompleteness of absorption depends on 

 the total amount of the liquid present and on the shape of the vessel. 



In 1948-1949, an unsuccessful attempt was made to settle the quantum 

 yield controversy by a combined effort of Warburg and Emerson in the 

 latter's laboratory. Subsequently, in the summer of 1949, Warburg, Burk 

 and co-workers (1949''^, 1950^-^) carried out quantum yield measurements 

 by the two vessel method (this time with equal liquid volumes) at the Na- 

 tional Cancer Institute in Bethesda and at the Woods Hole Marine Bio- 

 logical Laboratory. In these experiments, single yields as high as I/70 = 

 0.44 (70 = 2.3) were observed; even the average yield was markedly above 

 0.25. The conditions under which these high jnelds have been obtained 

 were quite different from and, in some respects, opposite to those which had 

 been recommended bj'^ Warburg and Negelein in 1923. 



Cell Culture. The reduction of light intensity in the last day of culti- 

 vation, recommended by Warburg and Negelein to adapt Chlorella cells 

 to weak light, was discarded by Warburg and Burk. Another precaution, 

 called unimportant by Warburg and Negelein, was now found to be es- 

 sential: fast bubbhng of the carbon dioxide-bearing gas through the 

 culture bottle, preventing the cells from settling out, and thus assuring 

 adequate supply of oxygen and carbon dioxide to all of them. 



Very concentrated suspensions were used: 0.3 cc. cells in 7 cc. culture 

 medium (phosphate buffer, p}l 4.9, saturated with 5% CO2 in air). (In 

 1948, Warburg used only 0.1 cc. cells in 5 or 9 cc. solution.) This was done 

 to ensure complete light absorption, despite an increased rate of shaking, 

 which created a more pronounced "hole" in the center of the reaction vessel; 

 but the respiration correction — the main source of uncertainty in measure- 

 ments of this type — was thus made even larger than before. 



Response of the Manometer. Compared to the 1923 experiments, 

 the efficiency of shaking was increased (horizontal, back-and-forth motion 

 of a rectangular vessel with an amplitude of 2 cm., 150 times per min.). 

 It was asserted that stirring was thus made so effective that the response 

 of the manometer to gas production (or absorption) in the liquid was 



