144 REporRT OF DEPARTMENT OF BACTERIOLOGY OF THE 
To ten broth tube cultures, 10 ¢. c. each, formalin was added 
as follows and the tubes then thoroughly shaken: Two tubes 
(1 & 1’), 0.57% formalin; two tubes (2 & 2’), 0.46%; two 
tubes (8 & 3’), 0.83%; two tubes (4 & 4’), 0.18%; two tubes 
(5 & 5’) 0.095%. Two tubes (6 & 6’) containing sterile 
broth were included to serve as controls. On the third day 
transfers made from these to sterile broth proved the sterility 
of all. On the fifth day, a sterile cube 5 mm. in diameter, 
cut from the interior of fresh carrot was inserted into each 
tube. On the eleventh day the carrot tissues in tubes 1, 1, 
2, 2’, 3, 3’ 6, 6’, were not softened; those in 4, 4’, 5, 5’, were 
completely softened. On the fourteenth day 3, 3’, were sof- 
tened somewhat; others unchanged. On the twenty-third day 
3, 3’, further softened; 2, 2’ softened somewhat but less than 
3, 3’. -On the thirty-third day 1, 1’, and 6, 6’, showed no 
softening; others completely softened. Final transfers to 
sterile broth showed all tubes sterile at the close of the trial. 
As a result of similar series, repeating the above and using 
other strengths of formalin, the following conclusions have 
been reached. The addition of 0.1% formalin is sufficient to 
sterilize a beef broth culture of B. carotovorus one to ten days 
old, providing the tube is thoroughly shaken. More formalin, 
0.2% or even more, may be needed if not thoroughly shaken. 
The presence of 0.6% or more formalin completely inhibits 
enzym action; amounts as low as 0.38% retard to a marked 
degree. There was perceptible retardation from 0.06% for- 
malin, although this amount was too slight to sterilize with 
certainty. In all the above trials the formalin acted on the 
broth several days before its relation to the enzymic activity 
was determined. 
Trials were also made to determine the effect of formalin 
additions to solutions of the enzym obtained from broth cul- 
tures by precipitation with alcohol. The activity in these 
cases was determined by trial on razor sections.of carrot roots. 
The results were in accord with those just discussed, viz., a 
slight but appreciable retardation from 0.05% addition, and 
almost complete inhibition where 0.5% was added. In these 
latter trials the formalin was added to the enzym solution 
some time before the cytolytic activity of the mixture was 
