METHODS OF INVESTIGATION 15 



the study of invertebrates, the former group has been 

 used primarily to elucidate the problems of the depress- 

 ing action exerted on oxygen consumption (by cyanide 

 in particular), to gain knowledge concerning the aerobic 

 enzymatic chain, to prove the presence of an axial grad- 

 ient, or to study the importance of the respiratory pig- 

 ments to the organism. Perhaps with a combination of 

 both groups of poisons one might make a quantitative 

 approach to the question of how far anaerobic can re- 

 place aerobic processes, a question that up to now has 

 been primarily attacked by exposing animals to varying 

 oxygen tensions. 



B. Removal of oxygen. The second way in which 

 anaerobic conditions can be established consists in re- 

 moving the oxygen from the environment in which the 

 experimental animals are kept. This can be achieved by 

 the following means : (a) evacuation, (b) absorption of 

 oxygen, (c) replacement of the atmosphere with an inert 

 gas, like nitrogen or hydrogen. A combination of the 

 various procedures is of course possible and in some in- 

 stances advisable or necessary. We shall consider briefly 

 the advantages and disadvantages of each method. 



(a) Evacuation. Small animals are placed in 

 a glass container provided with an opening which can 

 be drawn out to a capillary. By means of a high vacuum 

 pump the atmosjjheric pressure is then lowered to such 

 an extent (to 10'^ mm. Hg for example, in Becquerel's ex- 

 periments, 1936) that the remaining oxygen will play 

 no significant role after the capillary has been fused. 

 This method can be used effectively with organisms that 

 resist desiccation, e.g., cysts of protozoa, many rotatoria, 

 moss nematodes and others. In active terrestrial ani- 

 mals the evaporation will be so high under these condi- 

 tions that a lethal effect could hardly be ascribed to lack 

 of oxygen alone. It is moreover questionable whether 

 the lowering of pressure itself may not affect some ani- 

 mals. 



