199, 



aerobic conditions. In these experiments Spitta and Weldert's Methy- 

 lene Blue Putrescibility Test was used. This test depends upon the 

 formation of a colorless leucobase as the oxygep in the sample becomes 

 exhausted. The technique is simple. One cc. Df o. i per cent, aqueous 

 solution of methylene blue is added to the sample, which is then kept 

 in an incubator either at 20 degrees C. or at 37 degrees C. and ob- 

 served frequently. The blue color of the sample remains practically 

 unchanged until the available oxygen contained in it has been con- 

 sumed and putrefactive conditions have been established. At this time 

 the dye is reduced and the color disappears. The time for the de- 

 coloration (reduction time) therefore indicates quite closely the time 

 at which the available oxygen is consumed. Phelps ('09, p. jy^ added 

 further value to the methylene blue test by putting it on a quanti- 

 tative working basis so that the putrescibility of a given sample can 

 be expressed in terms of relative stability. This makes it possible 

 to indicate the proportion of the oxygen present as compared with 

 the total amount required to oxidize a given sample. 



This test lends itself to this kind of experimentation, since be- 

 side making it an easy matter to determine the reduction time the 

 presence of the methylene blue in the sample has little or no deleterious 

 effect on the worms. At the time that the worms were transferred 

 to the bottle i cc. of a o. i per cent, aqueous solution of methylene blue 

 was added to each bottle and the time carefully noted. These test 

 bottles together with the checks were placed in a constant-tempera- 

 ture incubator at 20 degrees C, and careful watch was kept and the 

 reduction time of each noted. 



The worms in the sampling bottles were frequently observed in 

 order to determine whether or not any of them died while under these 

 conditions, since it is evident that the death of any of them would 

 constitute a source of error by increasing quantitatively the amount 

 of putrescible matter in the sample. Fortunately the mortality was 

 very low, so low that the writer feels confident that it did not vitiate 

 the results of the experiments. The following table indicates the 

 results of one of the series. 



