12 STEINBERG: GROWTH OF ASPERGILLUS NIGER 

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In all the cases given, therefore, there is no evidence of an in- 
crease in the yield on the standard liquid Pefffer medium as a 
result of carrying the organism for three generations preceding 
inoculation of this medium on agar media containing various min- 
eral salts—including zinc in some instances. This would indicate 
that the assumption made in attempting to account for the marked 
decrease in yields observed in the course of this investigation, 
namely, that the spores become impoverished in some essential 
ash constituent as a result of prolonged cultivation on peptone- 
sucrose agar, is improbable. The same conclusion is indicated 
by the result of Exp. XII, where about six months after the origi- 
nal separation of the two strains, cultures prepared by using spores 
of the original tubes (W1 and Y1) are compared with cultures 
prepared by using spores which had been on the peptone-sucrose 
agar for seven generations. The yields are practically identical. 
The thought of course lies close at hand that changes occurred 
in the cultural conditions as the experiments progressed which are 
responsible for the changes in the yields obtained. Unusual pre- 
cautions were taken, however, to maintain uniform conditions 
(see Steinberg, 6). By consulting the data given in the Appendix 
it will be noted that the individual cultures of an experiment (as 
a rule five flasks of a kind were prepared) are very uniform. It 
is also to be noted that throughout the maximum yield of about 
I gram per 50 c.c. of medium was obtained, provided the proper 
amount of zinc was added, showing that the capacity for growth 
as such did not decrease. The fact that the decrease in yield is 
most conspicuous in cultures to which no zinc or to which only a 
small amount of zinc has been added suggests that the supposedly 
zinc-free media are actually not entirely free of this element. 
Zinc is an extremely troublesome element to eliminate completely 
from the chemicals and utensils employed. It is perhaps ques- 
tionable whether thus far anyone has grown Aspergillus niger in 
the complete absence of zinc. In my own experiments it is con- 
ceivable that the pyrex flasks employed contain and give off to 
