F. G. GUSTAFSON 183 



mental tube was then matched with that of a buffer tube of the same 

 size (and having the same concentration of indicator), using a con- 

 stant source of light ("Daylight" lamp).^ 



The alkalinity of the water was brought (by the means mentioned 

 above) to a point a little above pH 7.60; after adding the material it 

 was allowed to stand until it fell to pH 7.60, which was taken as the 

 starting point in all experiments. The time was noted and the tube 

 put aside for a minute or two, when it was gently shaken to dis- 

 tribute the CO2 throughout the solution. The tube was again 

 matched, this time with another buffer tube having a pH value of 

 7.25. If the experimental tube had not yet reached this, it was re- 

 peatedly examined at very short intervals until the two matched. 

 The change in pH was from 7.60 to 7.25, which was the standard 

 unit of measurement in all cases. 



In starting an experiment the time required for this change was 

 noted and the material was rejected unless this time was practically 

 constant for at least three periods before any anesthetic was added. 

 The time varied with the amount of material used, but was made as 

 near to 3 minutes as possible in each experiment. When the end- 

 point was reached, the material was taken out and rinsed in tap water 

 before starting a new measurement with a new solution. 



When a practically constant rate of respiration had been obtained, 

 the material was placed in a solution of the desired concentration of 

 the anesthetic. After the addition of the anesthetic the respiration 

 was measured in the same manner as before. The experiments with 

 anesthetics lasted from half an hour to 2 hours. 



On account of the short periods used it was found impossible to carry 

 on a control simultaneously with the experiment. Therefore a large 

 number of control experiments were made from a number of cultures 

 and at different times during the period of investigation. These con- 

 trol experiments agreed very closely, so they were averaged to make 

 up a control curve, which has been used in all figures. As will be 

 noticed from the figures, the control shows a gradual decrease in 

 rate of respiration. This is probably due to a decrease in oxidizable 



^ For other details see the introductory article by Osterhout, W. J. V., /. Gen. 

 Physiol., 1918, i, 171. 



