6 Livingston: Chemical stimulation of a green alga 



in order to simplify the otherwise almost hopelessly complicated 

 methods, the problem was further restricted by confining attention 

 to the cations * alone. In order to do this it was necessary to use 

 as stimulating agents salts whose anions were already present in 

 the control solution described above. Soluble nitrates and sulfates 

 were chosen for this purpose. Normal or decinormal solutions of 

 these salts were made up as accurately as possible, and these were 

 diluted to the concentrations needed for the cultures. Since the 

 addition of the poison to the nutrient solution would necessarily 

 dilute both solutions to some degree, the following method of 

 mutual dilution was adopted. The nutrient solution was made up 

 to nine-tenths of its required volume, while the poison solution for 

 each culture was prepared ten times its required concentration, or 

 one tenth of its required volume. Thus, for control culture, 9 c.c. 

 of stock nutrient solution plus 1 c.c. of water was used, and for any 

 concentration of poison (say kn, k being any traction of normal), 

 the culture medium was composed of 9 c.c. of nutrient solution plus 

 1 c.c. of 10 kn poison. An example will illustrate this : Suppose 

 that it was desired to test the effect of n\ 10,000 H 2 SO v 1 .5 c.c. of 

 nil Ca (N0 3 ) 2 was taken, together with 0.4 c.c. each of ;// 1 KNO s 

 and nji MgS0 4 , and 1.0 c.c. of ///ioo Fe (NO s ) 2 . After mixing, 

 these were diluted to a volume of 899.6 c.c. and then there was 

 added 0.4 c.c. of «/i K 2 HP0 4 . A further addition of 100.0 c.c. 

 of water would have produced the control medium. To 9.0 c.c. 

 of this " 9/10 dilution," was added 1.0 c.c. of ;//ioooH 2 SO r It 

 is readily seen that through mutual dilution both sets of ions come 

 to be of the desired concentration, the H 2 S0 4 being now ;// 10,000. 

 The slight increase in the osmotic pressure of the experiment 

 solution over that of the control, due to the addition of the poison 

 itself, is, in most cases, physically negligible (because of the 

 extreme dilutions used), and is always physiologically so (because 

 of the comparatively high concentration limits for the response of 

 this form, as previously f determined). Poisons of as high concen- 

 tration as u/100 have been used in only a few cases. 



The poison salts added to the nutrient solutions were also 



*The terminology of the ionic theory is here used, merely because it is most 

 convenient. 



t Livingston, B. E. 1900 and 1901, loc. (it. 



