526 



SELMAN A. WAKSMAN 



microorganisms. Two liters of the filtrate from an eight-day-old 

 culture of A. niger grown on the Peptone-Czapek solution were 

 used for this work; this filtrate was strongly acid due to the 

 large quantities of oxalic and citric acid produced by the or- 

 ganism. One liter of the fluid was left acid and the other was 

 neutralized to phenolphthalein by means of n NaOH. To each 

 liter of the fluid 20 cc. of a saturated solution of safraninwere 

 added. A flocculent colored precipitate appeared in both in- 

 stances, but was much heavier in the neutral solution. The 

 precipitates were allowed to settle for twenty-four hours, then 

 filtered off, washed with alcohol, and dried over sulfuric acid. 

 The precipitate from the liter of medium left acid weighed 60 

 mgm. and that from the liter of neutral medium weighed 500 



TABLE 10 



Influence of filtration through a porcelain filter upon the action of exoenzymes of 



A. niger 



mgm. The precipitates and the filtrates left, after their removal 

 were now tested for their proteolytic activities. About one- 

 half of each precipitate and 20 cc. of the filtrate left were used 

 as the sources of enzymes; as substrata 100 cc. of 1 per cent 

 peptone and casein solutions were used. The cultures were in- 

 cubated for forty-eight hours at 37°C. 



It is seen from table 11 that the safranin does not precipitate 

 the enzyme from the culture medium. One liter of medium, or 

 just 50 times as much as is ordinarily used for the inoculation of 

 100 cc. of the 1 per cent substratum, did not give, on precipi- 

 tation with safranin, enough enzyme to decompose as much 

 of the protein, as 20 cc. of the solution usually does. We might 

 therefore conclude that the enzyme is not of the nature of trypsin. 



