106 ARTIFICIAL PARTHENOGENESIS AND FERTILIZATION 
hypertonic solution remains practically ineffective in the pres- 
ence of a minute quantity of potassium cyanide, but that the 
eggs are not in the least injured by the latter in so short a time. 
It can be directly shown that the hypertonic solution is 
effective only in the presence of free oxygen, by simply expelling 
the air from the hypertonic solution. But this experiment 
can very easily miscarry on account of the interference of a 
troublesome source of error. Usually the hypertonic solution 
can be freed as far as possible from oxygen by passing through 
it for several hours a current of scrupulously purified hydrogen. 
Then a drop or two of eggs are placed in the solution. And 
herein lies the source of error. On opening the flask, of course 
some oxygen enters it, and for a short time the hypertonic 
solution is acting, not in the absence, but in the presence, of 
some oxygen. Now obviously only a little oxygen is sufficient 
to maintain the processes of oxidation which underlie the 
development of the egg. I had already noticed this eighteen 
years ago in my first experiments upon the necessity of oxygen 
for normal cleavage. But, as already mentioned, the eggs 
need to remain only a short time—some thirty to fifty minutes 
—in the hypertonic solution after membrane formation, and it 
is clear that in so short a time the introduction of a little oxygen 
into the hypertonic solution may easily frustrate the whole 
experiment. I reduced this risk by opening the stopper of the 
flask with the aid of a skilled assistant for only about a second 
and for a distance of only a millimeter in order to introduce 
the eggs. Before, during, and immediately after the opening a 
very strong current of hydrogen was passed through the flask. 
Negative experiments, i.e., ones in which the hypertonic solu- 
tion caused a few or many of the eggs to develop, even after 
the passing of a current of hydrogen, do not prove much;! but 
1 They only show that too much oxygen was present inadvertently in the 
hypertonic solution. I indicated this source of error in my first note upon the 
subject: Loeb, ‘On the Necessity of the Presence of Free Oxygen in the Hyper- 
tonic Sea-Water for the Production of Artificial Parthenogenesis,’’ University of 
California Publications, Physiology, III, 39, 1906. 
