CHEMICAL STIMULATION OF GROWTH OF ASPERGILLUS NIGER 335 
(H+), i.e., the negative logarithm of the hydrogen-ion concentration in 
gram-ions.] Furthermore, they note the same phenomenon observed by 
Hoagland (20) with barley; the acidity of the medium is modified as a 
result of growth, increasing where the initial acidities were too low, de- 
creasing where the initial acidities were too high. Hoagland found pn = 5-2 
the optimum acidity for barley plants. 
There is also some evidence bearing out the view of Pfeffer (53, p. 487) 
that a specific limiting acidity is reached as the result of the growth, etc., 
of certain organisms. Michaelis and Marcora (42) have found it to hold 
true in the case of Bacterium coli; Clark and Lubs (10) with Aspergillus 
niger; and Ayers (2) with many streptococci. This fact is at the basis of 
the procedure used in the laboratory of the addition of CaCOs in lactic 
acid fermentation. As is similarly generally recognized (53) an analogous 
phenomenon occurs in acetic acid, and in alcoholic, fermentation; and 
probably also in butyric acid fermentation. It is, we may say, frequently 
the case that the growth of an organism is limited by the attainment of a 
definite concentration of some acid produced by its own metabolism. 
Experiments on the relation of the acidity of the Pfeffer nutrient solu- 
tion to the growth and fructification of Aspergillus niger under conditions 
known to exclude to a high degree the ''stimulative" ions, were undertaken 
in order to obtain cultures directly comparable with zinc cultures. 
Methods 
In the following experiments the Pfeffer nutrient solution was used 
exclusively. The compounds employed in the preparation of this solution 
were Merck's '* Reagent" ammonium nitrate and magnesium sulphate; 
and Baker's "Analysed" potassium phosphate and ferric sulphate; the 
water was redistilled through glass (the first and last quarters of the distillate 
are rejected). The cane sugar used is that sold under the proprietary name 
of "Crystal Domino" sugar. The use of other compounds is always noted 
in the description accompanying the experiments. Tests with A. niger 
showed that the results obtained with "Crystal Domino" sugar in nowise 
differed from those obtained with Merck's "Highest Purity" cane sugar. 
In both cases growth proceeded as well with as without the addition of a 
trace of iron salt; and in the presence of zinc a marked acceleration of 
growth occurred. The information was obtained on inquiry from the 
manufacturer that no coloring matter is added to this sugar in the refining 
process, and that this practice had gone out of use over thirteen years ago. 
The flasks used were 150 cc. pyrex Erlenmeyers. They were cleaned 
by rinsing with concentrated sulphuric acid (Baker's "Special"), tap- 
water, lastly with distilled water, and inverted to drain dry. It may be 
noted here that when zinc is present in the glass of the cultural flasks (Jena 
glass) sufficient amounts may go into solution to cause partial or maximum 
"stimulation," as I have shown elsewhere (64). 
