392 DAVID H. TENNENT 
A. Chemically fertilized eggs 
The method of chemical fertilization that I used was that of 
Loeb (’09), with a very slight modification. The eggs were 
placed in a butyric acid mixture (6 cc. +4 butyric acid + 94 ce. 
sea water), for from one and one-half to two and one-half minutes, 
transferred from this to sea water to which an amount of +, NaOH 
solution calculated to be sufficient to neutralize the amount of 
butyric acid which had been carried over in the pipette, had been 
added; allowed to remain in this for twenty minutes; transferred 
to hypertonic sea water, (13 cc. 24 m. NaCl + 87 cc. sea water), 
for fifteen to twenty-five minutes; and finally transferred to sea 
water. 
A larger percentage of eggs formed membranes when placed 
in sea water plus NaOH, upon removal from the butyric acid 
mixture, than when placed in straight sea water. 
With this sea urchin, Toxopneustes, it is possible to determine 
the proper length of time that the eggs should remain in the hyper- 
tonic sea water by observing nuclear changes in the egg. Hindle, 
(10), in describing the parthenogenetic development of Stron- 
gylocentrotus purpuratus says, in one place, that no apparent 
changes ‘‘beyond a slight reduction of the clear zone of hyalo- 
plasm surrounding the nucleus’ may be observed while the eggs 
are immersed in the hypertonic solution, and in another place, 
‘during the treatment with hypertonic salt solution there is a 
slight increase in the size of the nucleus and the clear zone almost 
disappears.” 
In Toxopneustes the nucleus and the cytoplasm immediately 
surrounding it may be seen to be in a state of activity. From 
the surface of the nucleus, processes seem to push out and then 
retract, the nucleus meantime enlarging in volume, while in the 
cytoplasm, currents showing a movement of substance toward the 
nucleus may be seen. 
If the eggs be transferred from the hypertonic sea water at the 
time when, in the greatest number, the nucleus has reached its 
maximum size, just prior to the bursting of the membrane, it 
will be found that a larger percentage of regularly segmenting 
