CHEMICAL CRITERIA OF ANAEROBIOSIS 



33 



Neither heating the' oil in the boiUng water bath nor boiling 

 (about 300°C.) over the free flame appeared to affect the density 

 of color in the superficial colored layers of alkahne, 2 per cent 

 agar containing methylene blue 1:100,000. This experiment 

 negatives the suggestion that de-aerated oil is able to abstract 

 oxygen and carbon dioxide from media containing them. Also, 

 whereas melted alkaline agar colored with methylene blue and 

 poured without further heating to decolorize into tubes, loses 

 its color to within 1 to 2 cm. of the surface in a few hours, the 

 immediate layering of such a solution with mineral oil makes 

 only a barely measurable difference in the thickness of the band 

 of color at the top. 



TABLE 6 

 A comparison of mineral oil and paraffine as seals against the recoloration of alkaline 



methylene blue agar 



Depth of agar, 7 cm. 



Depth of parafine and oil, 2.5 cm. 



An equal amount of paraffine in a similar tube, however, 

 reduced the band of undecolorized medium to 1 cm. in sixteen 

 hours. In a comparison of these three conditions where boiling 

 was used as a means of immediate decolorization followed by 

 rapid cooling and incubation at 37°C., the measurements of the 

 blue band indicated (in centimeters) in table 6 were made. 



An analogous comparison of paraffine and vaseline of approxi- 

 mately equal melting points (50°C.) and boiling points (about 

 300°C.) showed vaseline to be superior even to paraffine. • Table 

 7 shows the actual measurements in centimeters (alkaline meth- 

 ylene blue agar 7 cm. deep). 



The recession of the band under paraffine and vaseline was 

 not peculiar to this particular experiment; it has frequently been 



JOURNAL OF BACTERIOLOGY, VOL. VI, NO. 1 



